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1
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Shelest A, Alaburda A, Vaiciuleviciute R, Uzieliene I, Bialaglovyte P, Bernotiene E. The Effect of TGF-β3 and IL-1β on L-Type Voltage-Operated Calcium Channels and Calcium Ion Homeostasis in Osteoarthritic Chondrocytes and Human Bone Marrow-Derived Mesenchymal Stem Cells During Chondrogenesis. Pharmaceutics 2025; 17:343. [PMID: 40143007 PMCID: PMC11945166 DOI: 10.3390/pharmaceutics17030343] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/09/2025] [Revised: 02/26/2025] [Accepted: 03/05/2025] [Indexed: 03/28/2025] Open
Abstract
Background: Transforming growth factor-β (TGF-β) and interleukin 1β (IL-1β) are key regulators of the chondrogenic differentiation, physiology and pathology of cartilage tissue, with TGF-β promoting chondrogenesis and matrix formation, while IL-1β exerts catabolic effects, inhibiting chondrogenesis and contributing to cartilage degradation. Both cytokines alter the intracellular calcium ion (iCa2+) levels; however, the exact pathways are not known. Objectives: This study aimed to evaluate the impact of TGF-β3 and IL-1β on calcium homeostasis in human bone marrow-derived mesenchymal stem cells (hBM-MSCs) and chondrocytes during chondrogenesis. Results: TGF-β3 increased iCa2+ levels in both hBM-MSCs and chondrocytes. Furthermore, TGF-β3 increased the functional activity of L-type voltage-operated calcium channels (L-VOCCs) in hBM-MSCs but not in chondrocytes. TGF-β3 and IL-1β reduced L-VOCCs subunit CaV1.2 (CACNA1C) gene expression in chondrocytes. In hBM-MSCs, TGF-β3 and IL-1β increased SERCA pump (ATP2A2) gene expression, while in chondrocytes, this effect was observed only with TGF-β3. Conclusions: TGF-β3 increases iCa2+ both in osteoarthritic chondrocytes and hBM-MSCs during chondrogenesis. In hBM-MSCs, TGF-β3-mediated elevation in iCa2+ is related to the increased functional activity of L-VOCCs. IL-1β does not change iCa2+ in osteoarthritic chondrocytes and hBM-MSCs; however, it initiates the mechanisms leading to further downregulation of iCa2+ in both types of cells. The differential and cell-specific roles of TGF-β3 and IL-1β in the calcium homeostasis of osteoarthritic chondrocytes and hBM-MSCs during chondrogenesis may provide a new insight into future strategies for cartilage repair and osteoarthritis treatment.
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Affiliation(s)
- Anastasiia Shelest
- Institute of Biosciences, Life Sciences Center, Vilnius University, LT-10257 Vilnius, Lithuania
| | - Aidas Alaburda
- Institute of Biosciences, Life Sciences Center, Vilnius University, LT-10257 Vilnius, Lithuania
| | - Raminta Vaiciuleviciute
- Department of Regenerative Medicine, State Research Institute Centre for Innovative Medicine, LT-08406 Vilnius, Lithuania; (R.V.); (I.U.); (P.B.); (E.B.)
| | - Ilona Uzieliene
- Department of Regenerative Medicine, State Research Institute Centre for Innovative Medicine, LT-08406 Vilnius, Lithuania; (R.V.); (I.U.); (P.B.); (E.B.)
| | - Paulina Bialaglovyte
- Department of Regenerative Medicine, State Research Institute Centre for Innovative Medicine, LT-08406 Vilnius, Lithuania; (R.V.); (I.U.); (P.B.); (E.B.)
| | - Eiva Bernotiene
- Department of Regenerative Medicine, State Research Institute Centre for Innovative Medicine, LT-08406 Vilnius, Lithuania; (R.V.); (I.U.); (P.B.); (E.B.)
- VilniusTech Faculty of Fundamental Sciences, Department of Chemistry and Bioengineering, Vilnius Gediminas Technical University, LT-10223 Vilnius, Lithuania
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Zhao F, Jia Z, Zhang L, Liu G, Li J, Zhao J, Xie Y, Chen L, Jiang H, He W, Wang A, Peng J, Zheng Y. A MnO 2 nanosheets doping double crosslinked hydrogel for cartilage defect repair through alleviating inflammation and guiding chondrogenic differentiation. Biomaterials 2025; 314:122875. [PMID: 39454507 DOI: 10.1016/j.biomaterials.2024.122875] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/07/2024] [Revised: 08/29/2024] [Accepted: 10/03/2024] [Indexed: 10/28/2024]
Abstract
The inflammatory microenvironment and inferior chondrogenesis are major symptoms after cartilage defect. Although various modifications strategies associated with hydrogels exhibit remarkable capacity of pro-cartilage regeneration, the adverse effect by prolonging inflammation is still formidable to hamper potential biomedical applications of different hydrogel implants. Herein, inspired by the repair microenvironment of articular cartilage defects, an injectable, immunomodulatory, and chondrogenic L-MNS-CMDA hydrogel is prepared through grafting vinyl and catechol groups to chitosan macromolecules using amide reaction, then further loading MnO2 nanosheets (MNS). The double crosslinking of photopolymerization and catechol oxidative polymerization endows L-MNS-CMDA hydrogel with preferable mechanical property, affording a suitable mechanical support for cartilage defect repair. Additionally, the robust tissue adhesion capability stemming from catechol groups guarantees the long-term retention of the hydrogel in the defect site. Meanwhile, L-MNS-CMDA hydrogel decomposes exogenous and intracellular H2O2 into O2 and H2O, to effectively alleviate cellular oxidative stress caused by long-term hypoxia. Under the synergies of catechol groups and MNS, L-MNS-CMDA hydrogel not only inhibits macrophages polarizing into M1 phenotype, but encourages them turn into M2 phenotype, thereby, reconstructing an immunization friendly microenvironment to ultimately enhance cartilage regeneration. Predictably, the hydrogel markedly induces rat bone marrow mesenchymal stem cells differentiating into chondrocytes by expressing abundant glycosaminoglycan and type II collagen. A cartilage defect model of rat knee joint indicates that L-MNS-CMDA hydrogel visually regulate the early inflammatory response of post-implantation, and facilitate cartilage regeneration and recovery of joint function after 12 weeks of post-implantation. All in all, this multifunctional L-MNS-CMDA hydrogel exhibits superior immunomodulatory and chondrogenic properties, holding immense clinical potential in the treatment of cartilage defects.
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Affiliation(s)
- Feilong Zhao
- School of Materials Science and Engineering, University of Science and Technology Beijing, Beijing, 100083, China
| | - Zhibo Jia
- Beijing Key Lab of Regenerative Medicine in Orthopaedics, Key Laboratory of Musculoskeletal Trauma &War Injuries, PLA Institute of Orthopedics, Chinese PLA General Hospital, Beijing, 100853, China
| | - Liyang Zhang
- School of Materials Science and Engineering, University of Science and Technology Beijing, Beijing, 100083, China
| | - Guodong Liu
- School of Materials Science and Engineering, University of Science and Technology Beijing, Beijing, 100083, China
| | - Junfei Li
- School of Materials Science and Engineering, University of Science and Technology Beijing, Beijing, 100083, China
| | - Jianming Zhao
- School of Materials Science and Engineering, University of Science and Technology Beijing, Beijing, 100083, China
| | - Yajie Xie
- School of Materials Science and Engineering, University of Science and Technology Beijing, Beijing, 100083, China
| | - Lu Chen
- School of Materials Science and Engineering, University of Science and Technology Beijing, Beijing, 100083, China
| | - Hongyu Jiang
- Beijing Key Lab of Regenerative Medicine in Orthopaedics, Key Laboratory of Musculoskeletal Trauma &War Injuries, PLA Institute of Orthopedics, Chinese PLA General Hospital, Beijing, 100853, China
| | - Wei He
- School of Materials Science and Engineering, University of Science and Technology Beijing, Beijing, 100083, China
| | - Aiyuan Wang
- Beijing Key Lab of Regenerative Medicine in Orthopaedics, Key Laboratory of Musculoskeletal Trauma &War Injuries, PLA Institute of Orthopedics, Chinese PLA General Hospital, Beijing, 100853, China
| | - Jiang Peng
- Beijing Key Lab of Regenerative Medicine in Orthopaedics, Key Laboratory of Musculoskeletal Trauma &War Injuries, PLA Institute of Orthopedics, Chinese PLA General Hospital, Beijing, 100853, China.
| | - Yudong Zheng
- School of Materials Science and Engineering, University of Science and Technology Beijing, Beijing, 100083, China.
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Sarkar A, Gallo MC, Bell JA, Mayfield CK, Ball JR, Ayad M, Lechtholz-Zey E, Chang SW, Sugiyama O, Evseenko D, Lieberman JR. Ex Vivo Regional Gene Therapy Compared to Recombinant BMP-2 for the Treatment of Critical-Size Bone Defects: An In Vivo Single-Cell RNA-Sequencing Study. Bioengineering (Basel) 2025; 12:29. [PMID: 39851303 PMCID: PMC11762083 DOI: 10.3390/bioengineering12010029] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/10/2024] [Revised: 12/22/2024] [Accepted: 12/26/2024] [Indexed: 01/26/2025] Open
Abstract
Ex vivo regional gene therapy is a promising tissue-engineering strategy for bone regeneration: osteogenic mesenchymal stem cells (MSCs) can be genetically modified to express an osteoinductive stimulus (e.g., bone morphogenetic protein-2), seeded onto an osteoconductive scaffold, and then implanted into a bone defect to exert a therapeutic effect. Compared to recombinant human BMP-2 (rhBMP-2), which is approved for clinical use, regional gene therapy may have unique benefits related to the addition of MSCs and the sustained release of BMP-2. However, the cellular and transcriptional mechanisms regulating the response to these two strategies for BMP-2 mediated bone regeneration are largely unknown. Here, for the first time, we performed single-cell RNA sequencing (10x Genomics) of hematoma tissue in six rats with critical-sized femoral defects that were treated with either regional gene therapy or rhBMP-2. Our unbiased bioinformatic analysis of 2393 filtered cells in each group revealed treatment-specific differences in their cellular composition, transcriptional profiles, and cellular communication patterns. Gene therapy treatment induced a more robust chondrogenic response, as well as a decrease in the proportion of fibroblasts and the expression of profibrotic pathways. Additionally, gene therapy was associated with an anti-inflammatory microenvironment; macrophages expressing canonical anti-inflammatory markers were more common in the gene therapy group. In contrast, pro-inflammatory markers were more highly expressed in the rhBMP-2 group. Collectively, the results of our study may offer insights into the unique pathways through which ex vivo regional gene therapy can augment bone regeneration compared to rhBMP-2. Furthermore, an improved understanding of the cellular pathways involved in segmental bone defect healing may allow for the further optimization of regional gene therapy or other bone repair strategies.
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Affiliation(s)
- Arijita Sarkar
- Department of Orthopaedic Surgery, Keck School of Medicine of the University of Southern California, Los Angeles, CA 90033, USA; (M.C.G.); (J.A.B.); (C.K.M.); (J.R.B.); (M.A.); (E.L.-Z.); (S.W.C.); (O.S.); (D.E.); (J.R.L.)
| | - Matthew C. Gallo
- Department of Orthopaedic Surgery, Keck School of Medicine of the University of Southern California, Los Angeles, CA 90033, USA; (M.C.G.); (J.A.B.); (C.K.M.); (J.R.B.); (M.A.); (E.L.-Z.); (S.W.C.); (O.S.); (D.E.); (J.R.L.)
| | - Jennifer A. Bell
- Department of Orthopaedic Surgery, Keck School of Medicine of the University of Southern California, Los Angeles, CA 90033, USA; (M.C.G.); (J.A.B.); (C.K.M.); (J.R.B.); (M.A.); (E.L.-Z.); (S.W.C.); (O.S.); (D.E.); (J.R.L.)
| | - Cory K. Mayfield
- Department of Orthopaedic Surgery, Keck School of Medicine of the University of Southern California, Los Angeles, CA 90033, USA; (M.C.G.); (J.A.B.); (C.K.M.); (J.R.B.); (M.A.); (E.L.-Z.); (S.W.C.); (O.S.); (D.E.); (J.R.L.)
| | - Jacob R. Ball
- Department of Orthopaedic Surgery, Keck School of Medicine of the University of Southern California, Los Angeles, CA 90033, USA; (M.C.G.); (J.A.B.); (C.K.M.); (J.R.B.); (M.A.); (E.L.-Z.); (S.W.C.); (O.S.); (D.E.); (J.R.L.)
| | - Mina Ayad
- Department of Orthopaedic Surgery, Keck School of Medicine of the University of Southern California, Los Angeles, CA 90033, USA; (M.C.G.); (J.A.B.); (C.K.M.); (J.R.B.); (M.A.); (E.L.-Z.); (S.W.C.); (O.S.); (D.E.); (J.R.L.)
| | - Elizabeth Lechtholz-Zey
- Department of Orthopaedic Surgery, Keck School of Medicine of the University of Southern California, Los Angeles, CA 90033, USA; (M.C.G.); (J.A.B.); (C.K.M.); (J.R.B.); (M.A.); (E.L.-Z.); (S.W.C.); (O.S.); (D.E.); (J.R.L.)
| | - Stephanie W. Chang
- Department of Orthopaedic Surgery, Keck School of Medicine of the University of Southern California, Los Angeles, CA 90033, USA; (M.C.G.); (J.A.B.); (C.K.M.); (J.R.B.); (M.A.); (E.L.-Z.); (S.W.C.); (O.S.); (D.E.); (J.R.L.)
| | - Osamu Sugiyama
- Department of Orthopaedic Surgery, Keck School of Medicine of the University of Southern California, Los Angeles, CA 90033, USA; (M.C.G.); (J.A.B.); (C.K.M.); (J.R.B.); (M.A.); (E.L.-Z.); (S.W.C.); (O.S.); (D.E.); (J.R.L.)
| | - Denis Evseenko
- Department of Orthopaedic Surgery, Keck School of Medicine of the University of Southern California, Los Angeles, CA 90033, USA; (M.C.G.); (J.A.B.); (C.K.M.); (J.R.B.); (M.A.); (E.L.-Z.); (S.W.C.); (O.S.); (D.E.); (J.R.L.)
| | - Jay R. Lieberman
- Department of Orthopaedic Surgery, Keck School of Medicine of the University of Southern California, Los Angeles, CA 90033, USA; (M.C.G.); (J.A.B.); (C.K.M.); (J.R.B.); (M.A.); (E.L.-Z.); (S.W.C.); (O.S.); (D.E.); (J.R.L.)
- Alfred E. Mann Department of Biomedical Engineering, Viterbi School of Engineering, University of Southern California, Los Angeles, CA 90089, USA
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Canalis E, Schilling L, Denker E. TNFα has differential effects on the transcriptome profile of selected populations in murine cartilage. OSTEOARTHRITIS AND CARTILAGE OPEN 2024; 6:100528. [PMID: 39494399 PMCID: PMC11530803 DOI: 10.1016/j.ocarto.2024.100528] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/30/2024] [Accepted: 10/04/2024] [Indexed: 11/05/2024] Open
Abstract
Objective To further our understanding of the role of tumor necrosis factor (TNF)α on the inflammatory response in chondrocytes. Design We explored the effects of TNFα on the transcriptome of epiphyseal chondrocytes from newborn C57BL/6 mice at the total and single cell (sc) resolution. Results Gene set enrichment analysis of total RNA-Seq from TNFα-treated chondrocytes revealed enhanced response to biotic stimulus, defense and immune response and cytokine signaling and suppressed cartilage and skeletal morphogenesis and development. scRNA-Seq analyzed 14,239 cells and 24,320 genes and distinguished 16 cell clusters. The more prevalent ones were constituted by limb bud and chondrogenic cells and fibroblasts comprising ∼73 % of the cell population. Genes expressed by joint fibroblasts were detected in 5 clusters comprising ∼45 % of the cells isolated. Pseudotime trajectory finding revealed an association between fibroblast and chondrogenic clusters which was not modified by TNFα. TNFα decreased the total cells recovered by 18.5 % and the chondrogenic, limb bud and mesenchymal clusters by 32 %, 27 % and 7 %, respectively. TNFα had profound effects on the insulin-like growth factor (IGF) axis decreasing Igf1, Igf2 and Igfbp4 and inducing Igfbp3 and Igfbp5, explaining an inhibition of collagen biosynthesis, cartilage and skeletal morphogenesis. Ingenuity Pathway Analysis of scRNA-Seq data revealed that TNFα enhanced the osteoarthritis, rheumatoid arthritis, pathogen induced cytokine storm and interleukin 6 signaling pathways and suppressed fibroblast growth factor signaling. Conclusions Epiphyseal chondrocytes are constituted by diverse cell populations distinctly regulated by TNFα to promote inflammation and suppression of matrix biosynthesis and growth.
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Affiliation(s)
- Ernesto Canalis
- Departments of Orthopaedic Surgery, UConn Health, Farmington, CT 06030, USA
- Departments of Medicine, UConn Health, Farmington, CT 06030, USA
- UConn Musculoskeletal Institute, UConn Health, Farmington, CT 06030, USA
| | - Lauren Schilling
- UConn Musculoskeletal Institute, UConn Health, Farmington, CT 06030, USA
| | - Emily Denker
- UConn Musculoskeletal Institute, UConn Health, Farmington, CT 06030, USA
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5
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Kitahara K, Ebata T, Liyile C, Nishida Y, Ogawa Y, Tokuhiro T, Shiota J, Nagano T, Takasuka TE, Endo T, Shimizu T, Alhasan H, Asano T, Takahashi D, Homan K, Onodera T, Kadoya K, Terkawi MA, Iwasaki N. Chondroprotective functions of neutrophil-derived extracellular vesicles by promoting the production of secreted frizzled-related protein 5 in cartilage. Cell Commun Signal 2024; 22:569. [PMID: 39604981 PMCID: PMC11603793 DOI: 10.1186/s12964-024-01953-8] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/28/2024] [Accepted: 11/21/2024] [Indexed: 11/29/2024] Open
Abstract
BACKGROUND Osteoarthritis (OA) is the most common degenerative joint disease characterized by cartilage degradation and various degrees of inflammation in the synovium. Growing evidence highlights that neutrophil extracellular vesicles (EVs) play a protective role in arthritic joints by promoting the resolution of inflammation and the synthesis of proteoglycans in cartilage. However, this homeostatic function is dependent on the activation state of neutrophils and the surrounding environment/tissues. Hence, we explored the chondroprotective functions of neutrophil-derived EVs under different stimulation conditions and the underlying molecular mechanism. METHODS Human blood-derived neutrophils, murine bone marrow-derived neutrophils, C-28I2 cells and primary chondrocytes were used. Neutrophils were stimulated with different cytokines, and their EVs were isolated for chondrocyte stimulation and further subjected to RNA-sequencing analysis. Two experimental murine OA models were used, and the treatment was performed by intraarticular injections. RESULTS Conditioned medium from neutrophils stimulated with TGF-β (N-β) had the greatest inhibitory effect on the expression of catabolic factors in stimulated chondrocytes. These protective effects were not impaired when conditioned medium of N-β from AnxA1-deficient mice was used. Consistent with these results, EVs isolated from N-β significantly reduced the expression of catabolic factors in stimulated chondrocytes. Bulk RNA-seq analysis revealed that secreted frizzled-related protein 5 (SFRP5) is upregulated in N-β-EV-stimulated chondrocytes. Furthermore, recombinant SFRP5 treatment significantly reduced the expression of catabolic factors in vitro and catabolic process in experimental murine OA models. CONCLUSIONS The current study emphasizes the potential therapeutic application of neutrophils in OA and provides new knowledge on the molecular mechanisms underlying their function.
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Affiliation(s)
- Keita Kitahara
- Department of Orthopedic Surgery, Faculty of Medicine, Graduate School of Medicine, Hokkaido University, Kita-15, Nish-7, Kita-ku, Sapporo, 060-8638, Japan
| | - Taku Ebata
- Department of Orthopedic Surgery, Faculty of Medicine, Graduate School of Medicine, Hokkaido University, Kita-15, Nish-7, Kita-ku, Sapporo, 060-8638, Japan
| | - Chen Liyile
- Department of Orthopedic Surgery, Faculty of Medicine, Graduate School of Medicine, Hokkaido University, Kita-15, Nish-7, Kita-ku, Sapporo, 060-8638, Japan
| | - Yoshio Nishida
- Department of Orthopedic Surgery, Faculty of Medicine, Graduate School of Medicine, Hokkaido University, Kita-15, Nish-7, Kita-ku, Sapporo, 060-8638, Japan
| | - Yuki Ogawa
- Department of Orthopedic Surgery, Faculty of Medicine, Graduate School of Medicine, Hokkaido University, Kita-15, Nish-7, Kita-ku, Sapporo, 060-8638, Japan
| | - Taiki Tokuhiro
- Department of Orthopedic Surgery, Faculty of Medicine, Graduate School of Medicine, Hokkaido University, Kita-15, Nish-7, Kita-ku, Sapporo, 060-8638, Japan
| | - Junki Shiota
- Department of Orthopedic Surgery, Faculty of Medicine, Graduate School of Medicine, Hokkaido University, Kita-15, Nish-7, Kita-ku, Sapporo, 060-8638, Japan
| | - Tatsuya Nagano
- Research Faculty of Agriculture, Hokkaido University, Kita-9, Nishi-9, Kitaku, Sapporo, 060-8589, Japan
| | - Taichi E Takasuka
- Research Faculty of Agriculture, Hokkaido University, Kita-9, Nishi-9, Kitaku, Sapporo, 060-8589, Japan
| | - Tsutomu Endo
- Department of Orthopedic Surgery, Faculty of Medicine, Graduate School of Medicine, Hokkaido University, Kita-15, Nish-7, Kita-ku, Sapporo, 060-8638, Japan
| | - Tomohiro Shimizu
- Department of Orthopedic Surgery, Faculty of Medicine, Graduate School of Medicine, Hokkaido University, Kita-15, Nish-7, Kita-ku, Sapporo, 060-8638, Japan
| | - Hend Alhasan
- Department of Orthopedic Surgery, Faculty of Medicine, Graduate School of Medicine, Hokkaido University, Kita-15, Nish-7, Kita-ku, Sapporo, 060-8638, Japan
| | - Tsuyoshi Asano
- Department of Orthopedic Surgery, Faculty of Medicine, Graduate School of Medicine, Hokkaido University, Kita-15, Nish-7, Kita-ku, Sapporo, 060-8638, Japan
| | - Daisuke Takahashi
- Department of Orthopedic Surgery, Faculty of Medicine, Graduate School of Medicine, Hokkaido University, Kita-15, Nish-7, Kita-ku, Sapporo, 060-8638, Japan
| | - Kentaro Homan
- Department of Orthopedic Surgery, Faculty of Medicine, Graduate School of Medicine, Hokkaido University, Kita-15, Nish-7, Kita-ku, Sapporo, 060-8638, Japan
| | - Tomohiro Onodera
- Department of Orthopedic Surgery, Faculty of Medicine, Graduate School of Medicine, Hokkaido University, Kita-15, Nish-7, Kita-ku, Sapporo, 060-8638, Japan
| | - Ken Kadoya
- Department of Orthopedic Surgery, Faculty of Medicine, Graduate School of Medicine, Hokkaido University, Kita-15, Nish-7, Kita-ku, Sapporo, 060-8638, Japan
| | - M Alaa Terkawi
- Department of Orthopedic Surgery, Faculty of Medicine, Graduate School of Medicine, Hokkaido University, Kita-15, Nish-7, Kita-ku, Sapporo, 060-8638, Japan.
| | - Norimasa Iwasaki
- Department of Orthopedic Surgery, Faculty of Medicine, Graduate School of Medicine, Hokkaido University, Kita-15, Nish-7, Kita-ku, Sapporo, 060-8638, Japan
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Wang S, Kurth S, Burger C, Wirtz DC, Schildberg FA, Ossendorff R. TNFα-Related Chondrocyte Inflammation Models: A Systematic Review. Int J Mol Sci 2024; 25:10805. [PMID: 39409134 PMCID: PMC11476358 DOI: 10.3390/ijms251910805] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/05/2024] [Revised: 09/26/2024] [Accepted: 10/04/2024] [Indexed: 10/20/2024] Open
Abstract
Tumor necrosis factor alpha (TNFα), as a key pro-inflammatory cytokine, plays a central role in joint diseases. In recent years, numerous models of TNFα-induced cartilage inflammation have been developed. However, due to the significant differences between these models and the lack of consensus in their construction, it becomes difficult to compare the results of different studies. Therefore, we summarized and compared these models based on important parameters for model construction, such as cell source, cytokine concentration, stimulation time, mechanical stimulation, and more. We attempted to analyze the advantages and disadvantages of each model and provide a compilation of the analytical methods used in previous studies. Currently, TNFα chondrocyte inflammation models can be categorized into four main types: monolayer-based, construct-based, explant-based TNFα chondrocyte inflammation models, and miscellaneous TNFα chondrocyte inflammation models. The most commonly used models were the monolayer-based TNFα chondrocyte inflammation models (42.86% of cases), with 10 ng/mL TNFα being the most frequently used concentration. The most frequently used chondrocyte cell passage is passage 1 (50%). Human tissues were most frequently used in experiments (51.43%). Only five articles included models with mechanical stimulations. We observed variations in design conditions between different models. This systematic review provides the essential experimental characteristics of the available chondrocyte inflammation models with TNFα, and it provides a platform for better comparison between existing and new studies in this field. It is essential to perform further experiments to standardize each model and to find the most appropriate experimental parameters.
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Lin CY, Naruphontjirakul P, Huang TY, Wu YC, Cheng WH, Su WT. The Exosomes of Stem Cells from Human Exfoliated Deciduous Teeth Suppress Inflammation in Osteoarthritis. Int J Mol Sci 2024; 25:8560. [PMID: 39201248 PMCID: PMC11354937 DOI: 10.3390/ijms25168560] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/04/2024] [Revised: 07/30/2024] [Accepted: 08/02/2024] [Indexed: 09/02/2024] Open
Abstract
Hyaluronic acid injection is commonly used clinically to slow down the development of osteoarthritis (OA). A newly developed therapeutic method is to implant chondrocytes/stem cells to regenerate cartilage in the body. The curative effect of stem cell therapy has been proven to come from the paracrine of stem cells. In this study, exosomes secreted by stem cells from human exfoliated deciduous teeth (SHED) and hyaluronic acid were used individually to evaluate the therapeutic effect in slowing down OA. SHED was cultured in a serum-free medium for three days, and the supernatant was collected and then centrifuged with a speed difference to obtain exosomes containing CD9 and CD63 markers, with an average particle size of 154.1 nm. SW1353 cells were stimulated with IL-1β to produce the inflammatory characteristics of OA and then treated with 40 μg/mL exosomes and hyaluronic acid individually. The results showed that the exosomes successfully inhibited the pro-inflammatory factors, including TNF-α, IL-6, iNOS, NO, COX-2 and PGE2, induced by IL-1β and the degrading enzyme of the extrachondral matrix (MMP-13). Collagen II and ACAN, the main components of the extrachondral matrix, were also increased by 1.76-fold and 2.98-fold, respectively, after treatment, which were similar to that of the normal joints. The effect can be attributed to the partial mediation of SHED exosomes to the NF-κB pathway, and the ability of exosomes to inhibit OA is found not inferior to that of hyaluronic acid.
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Affiliation(s)
- Chuang-Yu Lin
- Department of Biomedical Science and Environmental Biology, Kaohsiung Medical University, Kaohsiung 807378, Taiwan;
- Regenerative Medicine and Cell Therapy Research Center, Kaohsiung Medical University, Kaohsiung 807378, Taiwan;
| | - Parichart Naruphontjirakul
- Biological Engineering Program, Faculty of Engineering, King Mongkut’s University of Technology Thonburi, Bangkok 10140, Thailand;
| | - Te-Yang Huang
- Department of Orthopedic Surgery, Mackay Memorial Hospital, Taipei 104217, Taiwan;
| | - Yi-Chia Wu
- Regenerative Medicine and Cell Therapy Research Center, Kaohsiung Medical University, Kaohsiung 807378, Taiwan;
- Division of Plastic Surgery, Department of Surgery, Kaohsiung Medical University Hospital, Kaohsiung 807378, Taiwan
| | - Wei-Hsuan Cheng
- Department of Chemical Engineering and Biotechnology, National Taipei University of Technology, Taipei 106344, Taiwan;
| | - Wen-Ta Su
- Department of Chemical Engineering and Biotechnology, National Taipei University of Technology, Taipei 106344, Taiwan;
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Song K, Hu J, Yang M, Xia Y, He C, Yang Y, Zhu S. Pulsed electromagnetic fields potentiate bone marrow mesenchymal stem cell chondrogenesis by regulating the Wnt/β-catenin signaling pathway. J Transl Med 2024; 22:741. [PMID: 39107784 PMCID: PMC11301989 DOI: 10.1186/s12967-024-05470-7] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/24/2024] [Accepted: 07/03/2024] [Indexed: 08/10/2024] Open
Abstract
BACKGROUND Pulsed electromagnetic fields (PEMFs) show promise as a treatment for knee osteoarthritis (KOA) by reducing inflammation and promoting chondrogenic differentiation of bone marrow-derived mesenchymal stem cells (BMSCs). PURPOSE To identify the efficacy window of PEMFs to induce BMSCs chondrogenic differentiation and explore the cellular mechanism under chondrogenesis of BMSCs in regular and inflammatory microenvironments. METHODS BMSCs were exposed to PEMFs (75 Hz, 1.6/2/3/3.8 mT) for 7 and 14 days. The histology, proliferation, migration and chondrogenesis of BMSCs were assessed to identify the optimal parameters. Using these optimal parameters, transcriptome analysis was performed to identify target genes and signaling pathways, validated through immunohistochemical assays, western blotting, and qRT-PCR, with or without the presence of IL-1β. The therapeutic effects of PEMFs and the effective cellular signaling pathways were evaluated in vivo. RESULTS BMSCs treated with 3 mT PEMFs showed the optimal chondrogenesis on day 7, indicated by increased expression of ACAN, COL2A, and SOX9, and decreased levels of MMP3 and MMP13 at both transcriptional and protein levels. The advantages of 3 mT PEMFs diminished in the 14-day culture groups. Transcriptome analysis identified sFRP3 as a key molecule targeted by PEMF treatment, which competitively inhibited Wnt/β-catenin signaling, regardless of IL-1β presence or duration of exposure. This inhibition of the Wnt/β-catenin pathway was also confirmed in a KOA mouse model following PEMF exposure. CONCLUSIONS PEMFs at 75 Hz and 3 mT are optimal in inducing early-stage chondrogenic differentiation of BMSCs. The induction and chondroprotective effects of PEMFs are mediated by sFRP3 and Wnt/β-catenin signaling, irrespective of inflammatory conditions.
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Affiliation(s)
- Kangping Song
- Rehabilitation Medicine Center, Institute of Rehabilitation Medicine, West China Hospital, Sichuan University, #37 Guoxue Alley, Wuhou strict, Chengdu, Sichuan, 610041, PR China
- Key Laboratory of Rehabilitation Medicine in Sichuan Province, West China Hospital, Sichuan University, Chengdu, China
- School of Rehabilitation Sciences, West China School of Medicine, Sichuan University, Chengdu, China
| | - Jing Hu
- Rehabilitation Medicine Center, Institute of Rehabilitation Medicine, West China Hospital, Sichuan University, #37 Guoxue Alley, Wuhou strict, Chengdu, Sichuan, 610041, PR China
- Key Laboratory of Rehabilitation Medicine in Sichuan Province, West China Hospital, Sichuan University, Chengdu, China
- School of Rehabilitation Sciences, West China School of Medicine, Sichuan University, Chengdu, China
| | - Ming Yang
- The Lab of Aging Research, State Key Laboratory of Biotherapy, West China Hospital, National Clinical Research Center for Geriatrics, Sichuan University, Chengdu, China
| | - Yong Xia
- Rehabilitation Medicine Center, Institute of Rehabilitation Medicine, West China Hospital, Sichuan University, #37 Guoxue Alley, Wuhou strict, Chengdu, Sichuan, 610041, PR China
- Key Laboratory of Rehabilitation Medicine in Sichuan Province, West China Hospital, Sichuan University, Chengdu, China
- School of Rehabilitation Sciences, West China School of Medicine, Sichuan University, Chengdu, China
| | - Chengqi He
- Rehabilitation Medicine Center, Institute of Rehabilitation Medicine, West China Hospital, Sichuan University, #37 Guoxue Alley, Wuhou strict, Chengdu, Sichuan, 610041, PR China
- Key Laboratory of Rehabilitation Medicine in Sichuan Province, West China Hospital, Sichuan University, Chengdu, China
- School of Rehabilitation Sciences, West China School of Medicine, Sichuan University, Chengdu, China
| | - Yonghong Yang
- Rehabilitation Medicine Center, Institute of Rehabilitation Medicine, West China Hospital, Sichuan University, #37 Guoxue Alley, Wuhou strict, Chengdu, Sichuan, 610041, PR China.
- Key Laboratory of Rehabilitation Medicine in Sichuan Province, West China Hospital, Sichuan University, Chengdu, China.
- School of Rehabilitation Sciences, West China School of Medicine, Sichuan University, Chengdu, China.
| | - Siyi Zhu
- Rehabilitation Medicine Center, Institute of Rehabilitation Medicine, West China Hospital, Sichuan University, #37 Guoxue Alley, Wuhou strict, Chengdu, Sichuan, 610041, PR China.
- Key Laboratory of Rehabilitation Medicine in Sichuan Province, West China Hospital, Sichuan University, Chengdu, China.
- School of Rehabilitation Sciences, West China School of Medicine, Sichuan University, Chengdu, China.
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9
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Liu C, Zhang L, Zheng X, Zhu J, Jin L, Gao R. Pleiotrophin inhibited chondrogenic differentiation potential of dental pulp stem cells. Oral Dis 2024; 30:1439-1450. [PMID: 36840423 DOI: 10.1111/odi.14554] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/29/2022] [Revised: 02/13/2023] [Accepted: 02/22/2023] [Indexed: 02/26/2023]
Abstract
OBJECTIVE Studies have shown that the levels of pleiotrophin (PTN) are greatly elevated in the synovial fluid and cartilage in osteoarthritis. Therefore, the purpose of this study was to investigate the effect and mechanism of PTN on the chondrogenic differentiation of DPSCs in inflammatory and normal microenvironments. MATERIALS AND METHODS A lentiviral vector was used to deplete or overexpress PTN in DPSCs. The inflammatory microenvironment was simulated in vitro by the addition of IL-1β to the culture medium. The chondrogenic differentiation potential was assessed using Alcian Blue staining and the main chondrogenic markers. A dual-luciferase reporter assay was used to explore the relationship between miR-137 and PTN. RESULTS The results showed that 0.1 ng/mL IL-1β treatment during chondrogenic induction greatly impaired the chondrogenic differentiation of DPSCs. Supplementation with PTN and PTN overexpression inhibited chondrogenic differentiation of DPSCs, while PTN depletion promoted chondrogenic differentiation. MiR-137 negatively regulated the expression of PTN by binding to the 3'UTR of its mRNA. Moreover, miR-137 promoted chondrogenic differentiation of DPSCs in normal and inflammatory microenvironments. CONCLUSION Our results suggest that PTN may play an inhibitory role in the chondrogenic differentiation of DPSCs in normal and inflammatory microenvironments, which is regulated by miR-137.
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Affiliation(s)
- Chang Liu
- Department of Stomatology, Beijing Friendship Hospital, Capital Medical University, Beijing, China
| | - Lili Zhang
- Department of General Dentistry and Integrated Emergency Dental Care, Beijing Stomatological Hospital, Capital Medical University, Beijing, China
| | - Xiaoyu Zheng
- Laboratory of Molecular Signaling and Stem Cells Therapy, Beijing Key Laboratory of Tooth Regeneration and Function Reconstruction, Capital Medical University School of Stomatology, Beijing, China
| | - Jiaman Zhu
- Department of Stomatology, Beijing Friendship Hospital, Capital Medical University, Beijing, China
| | - Luyuan Jin
- Department of General Dentistry and Integrated Emergency Dental Care, Beijing Stomatological Hospital, Capital Medical University, Beijing, China
| | - Runtao Gao
- Department of Stomatology, Beijing Friendship Hospital, Capital Medical University, Beijing, China
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10
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Kaur G, Wu B, Murali S, Lanigan T, Coleman RM. A synthetic, closed-looped gene circuit for the autonomous regulation of RUNX2 activity during chondrogenesis. FASEB J 2024; 38:e23484. [PMID: 38407380 PMCID: PMC10981937 DOI: 10.1096/fj.202300348rr] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/24/2023] [Revised: 01/23/2024] [Accepted: 01/30/2024] [Indexed: 02/27/2024]
Abstract
The transcription factor RUNX2 is a key regulator of chondrocyte phenotype during development, making it an ideal target for prevention of undesirable chondrocyte maturation in cartilage tissue-engineering strategies. Here, we engineered an autoregulatory gene circuit (cisCXp-shRunx2) that negatively controls RUNX2 activity in chondrogenic cells via RNA interference initiated by a tunable synthetic Col10a1-like promoter (cisCXp). The cisCXp-shRunx2 gene circuit is designed based on the observation that induced RUNX2 silencing after early chondrogenesis enhances the accumulation of cartilaginous matrix in ATDC5 cells. We show that the cisCXp-shRunx2 initiates RNAi of RUNX2 in maturing chondrocytes in response to the increasing intracellular RUNX2 activity without interfering with early chondrogenesis. The induced loss of RUNX2 activity in turn negatively regulates the gene circuit itself. Moreover, the efficacy of RUNX2 suppression from cisCXp-shRunx2 can be controlled by modifying the sensitivity of cisCXp promoter. Finally, we show the efficacy of inhibiting RUNX2 in preventing matrix loss in human mesenchymal stem cell-derived (hMSC-derived) cartilage under conditions that induce chondrocyte hypertrophic differentiation, including inflammation. Overall, our results demonstrated that the negative modulation of RUNX2 activity with our autoregulatory gene circuit enhanced matrix synthesis and resisted ECM degradation by reprogrammed MSC-derived chondrocytes in response to the microenvironment of the degenerative joint.
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Affiliation(s)
- Gurcharan Kaur
- Department of Biomedical Engineering, University of Michigan, Ann Arbor, MI, USA
| | - Biming Wu
- Department of Biomedical Engineering, University of Michigan, Ann Arbor, MI, USA
| | - Sunjana Murali
- Program in Cellular and Molecular Biology, University of Michigan, Ann Arbor, MI, USA
| | - Thomas Lanigan
- Biomedical Research Vector Core, University of Michigan, Ann Arbor, MI, USA
| | - Rhima M. Coleman
- Department of Biomedical Engineering, University of Michigan, Ann Arbor, MI, USA
- Department of Mechanical Engineering, University of Michigan, Ann Arbor, MI, USA
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11
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Panos JA, Coenen MJ, Nagelli CV, McGlinch EB, Atasoy-Zeybek A, De Padilla CL, De la Vega RE, Evans CH. Segmental defect healing in the presence or absence of recombinant human BMP2: Novel insights from a rat model. J Orthop Res 2023; 41:1934-1944. [PMID: 36850029 PMCID: PMC10440238 DOI: 10.1002/jor.25530] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/31/2022] [Revised: 01/16/2023] [Accepted: 02/08/2023] [Indexed: 03/01/2023]
Abstract
This study defined and compared the course of native, impaired and growth factor-stimulated bone regeneration in a rat femoral defect model. A mid-diaphyseal defect with rigid internal fixation was surgically created in the right femur of male Fischer rats and serially analyzed over 36 weeks. Native bone regeneration was modeled using a sub-critical, 1 mm size defect, which healed uneventfully. Critical size defects of 5 mm were used to analyze impaired bone regeneration. In a third group, the 5 mm defects were filled with 11 µg of recombinant human bone morphogenetic protein 2 (rhBMP2) impregnated onto an absorbable collagen sponge, modeling its clinical use. Native bone regeneration was characterized by endochondral ossification with progressive remodeling to ultimately resemble intact femora. An endochondral response was also observed under conditions of impaired bone regeneration, but by week 8 medullary capping occurred with fibrofatty consolidation of the tissue within the defect, resembling an atrophic non-union. rhBMP2 treatment was associated with prolonged inflammatory cytokine expression and rapid intramembranous bone formation occurring with reduced expression of cartilage-associated collagens. Between weeks 4 and 36, rhBMP2-treated bones demonstrated decreased trabecular number and increased trabecular separation, which resulted in inferior mechanical properties compared with bones that healed naturally. Clinical Significance: Recombinant human bone morphogenetic protein 2 (rhBMP2) is used clinically to promote healing of long bones. Our data suggest that it drives intramembraneous ossification producing an inferior regenerate that deteriorates with time. Clinical outcomes would be improved by technologies favoring endochondral regenerative ossification.
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Affiliation(s)
- Joseph A. Panos
- Rehabilitation Medicine Research Center, Mayo Clinic; Rochester, Minnesota, USA
- Musculoskeletal Gene Therapy Research Laboratory, Mayo Clinic; Rochester, Minnesota, USA
- Graduate School of Biomedical Sciences, Mayo Clinic; Rochester, Minnesota, USA
- Medical Scientist Training Program, Mayo Clinic; Rochester, Minnesota, USA
| | - Michael J. Coenen
- Rehabilitation Medicine Research Center, Mayo Clinic; Rochester, Minnesota, USA
- Musculoskeletal Gene Therapy Research Laboratory, Mayo Clinic; Rochester, Minnesota, USA
| | - Christopher V. Nagelli
- Rehabilitation Medicine Research Center, Mayo Clinic; Rochester, Minnesota, USA
- Musculoskeletal Gene Therapy Research Laboratory, Mayo Clinic; Rochester, Minnesota, USA
| | - Erin B. McGlinch
- Rehabilitation Medicine Research Center, Mayo Clinic; Rochester, Minnesota, USA
- Musculoskeletal Gene Therapy Research Laboratory, Mayo Clinic; Rochester, Minnesota, USA
- Graduate School of Biomedical Sciences, Mayo Clinic; Rochester, Minnesota, USA
- Virology and Gene Therapy Graduate Program, Mayo Clinic; Rochester, Minnesota, USA
| | - Aysegul Atasoy-Zeybek
- Rehabilitation Medicine Research Center, Mayo Clinic; Rochester, Minnesota, USA
- Musculoskeletal Gene Therapy Research Laboratory, Mayo Clinic; Rochester, Minnesota, USA
| | - Consuelo Lopez De Padilla
- Rehabilitation Medicine Research Center, Mayo Clinic; Rochester, Minnesota, USA
- Musculoskeletal Gene Therapy Research Laboratory, Mayo Clinic; Rochester, Minnesota, USA
| | - Rodolfo E. De la Vega
- Rehabilitation Medicine Research Center, Mayo Clinic; Rochester, Minnesota, USA
- Musculoskeletal Gene Therapy Research Laboratory, Mayo Clinic; Rochester, Minnesota, USA
- Department of Cell Biology-Inspired Tissue Engineering, MERLN Institute; Maastricht, The Netherlands
| | - Christopher H. Evans
- Rehabilitation Medicine Research Center, Mayo Clinic; Rochester, Minnesota, USA
- Musculoskeletal Gene Therapy Research Laboratory, Mayo Clinic; Rochester, Minnesota, USA
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12
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Sonmez Kaplan S, Sazak Ovecoglu H, Genc D, Akkoc T. TNF-α, IL-1B and IL-6 affect the differentiation ability of dental pulp stem cells. BMC Oral Health 2023; 23:555. [PMID: 37568110 PMCID: PMC10422753 DOI: 10.1186/s12903-023-03288-1] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/26/2023] [Accepted: 08/05/2023] [Indexed: 08/13/2023] Open
Abstract
BACKGROUND This in vitro study examined the effect of the inflammatory cytokines (tumour necrosis factor-α (TNF-α), interleukin (IL)-1β, and IL-6) on osteogenic, chondrogenic, and adipogenic differentiation of dental pulp stem cells (DPSCs) which have significant relevance in future regenerative therapies. METHODS DPSCs were isolated from the impacted third molar dental pulp and determined with flow cytometry analysis. DPSCs were divided into into 5 main groups with 3 subdivisions for each group making a total of 15 groups. Experimental groups were stimulated with TNF-α, IL-1β, IL-6, and a combination of all three to undergo osteogenic, chondrogenic, and adipogenic differentiation protocols. Next, the differentiation of each group was examined with different staining procedures under a light microscope. Histological analysis of osteogenic, chondrogenic, and adipogenic differentiated pellets was assessed using a modified Bern score. Statistical significance determined using one-way analysis of variance, and correlations were assessed using Pearson's test (two-tailed). RESULTS Stimulation with inflammatory cytokines significantly inhibited the osteogenic, chondrogenic and adipogenic differentiation of DPSCs in terms of matrix and cell formation resulting in weak staining than the unstimulated groups with inflammatory cytokines. On contrary, the unstimulated groups of MSCs have shown to be highly proliferative ability in terms of osteogenic, chondrogenic, and adipogenic differentiation. CONCLUSIONS DPSCs have high osteogenic, chondrogenic, and adipogenic differentiation capabilities. Pretreatment with inflammatory cytokines decreases the differentiation ability in vitro, thus inhibiting tissue formation.
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Affiliation(s)
- Sema Sonmez Kaplan
- Department of Endodontics, Faculty of Dentistry, Biruni University, 10. Yıl Caddesi Protokol Yolu No: 45, 34010, Topkapı, Istanbul, Turkey.
| | - Hesna Sazak Ovecoglu
- Faculty of Dentistry Department of Endodontics, Marmara University, Istanbul, Turkey
| | - Deniz Genc
- Department of Pediatric Health & Diseases Faculty of Health Sciences, Muğla Sıtkı Koçman University, Mugla, Turkey
- Research Laboratories Center, Immunology and Stem Cell Laboratory, Muğla Sıtkı Koçman University, Mugla, Turkey
| | - Tunc Akkoc
- Immunology Department, Marmara University Medical Faculty, Istanbul, Turkey
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13
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Voskamp C, Koevoet WJLM, Van Osch GJVM, Narcisi R. Senescence during early differentiation reduced the chondrogenic differentiation capacity of mesenchymal progenitor cells. Front Bioeng Biotechnol 2023; 11:1241338. [PMID: 37609111 PMCID: PMC10441241 DOI: 10.3389/fbioe.2023.1241338] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/16/2023] [Accepted: 07/25/2023] [Indexed: 08/24/2023] Open
Abstract
Introduction: Mesenchymal stromal/progenitor cells (MSCs) are promising for cartilage cell-based therapies due to their chondrogenic differentiation capacity. However, MSCs can become senescent during in vitro expansion, a state characterized by stable cell cycle arrest, metabolic alterations, and substantial changes in the gene expression and secretory profile of the cell. In this study, we aimed to investigate how senescence and the senescence-associated secretory phenotype (SASP) affect chondrogenic differentiation of MSCs. Methods: To study the effect of senescence, we exposed MSCs to gamma irradiation during expansion or during chondrogenic differentiation (the pellet culture). Western blot analysis was used to evaluate MSCs response to the chondrogenic inductor TGF-β. Results: When senescence was induced during expansion or at day 7 of chondrogenic differentiation, we observed a significant reduction in the cartilage matrix. Interestingly, when senescence was induced at day 14 of differentiation, chondrogenesis was not significantly altered. Moreover, exposing chondrogenic pellets to the medium conditioned by senescent pellets had no significant effect on the expression of anabolic or catabolic cartilage markers, suggesting a neglectable paracrine effect of senescence on cartilage generation in our model. Finally, we show that senescent MSCs showed lower phosphorylated SMAD2 levels after TGFβ1 stimulation than control MSCs. Conclusion: Overall, these results suggest that the occurrence of senescence in MSCs during expansion or early differentiation could be detrimental for cartilage tissue engineering.
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Affiliation(s)
- Chantal Voskamp
- Department of Orthopaedics and Sports Medicine, University Medical Center Rotterdam, Rotterdam, Netherlands
| | - Wendy J. L. M. Koevoet
- Department of Otorhinolaryngology, University Medical Center Rotterdam, Rotterdam, Netherlands
| | - Gerjo J. V. M. Van Osch
- Department of Orthopaedics and Sports Medicine, University Medical Center Rotterdam, Rotterdam, Netherlands
- Department of Otorhinolaryngology, University Medical Center Rotterdam, Rotterdam, Netherlands
- Department of Biomechanical Engineering, Faculty of Mechanical, Maritime and Materials Engineering, Delft University of Technology, Delft, Netherlands
| | - Roberto Narcisi
- Department of Orthopaedics and Sports Medicine, University Medical Center Rotterdam, Rotterdam, Netherlands
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14
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Zhu C, Zhou Q, Wang Z, Zhang J, Xu C, Ruan D. Growth differentiation factor 5 inhibits lipopolysaccharide-mediated pyroptosis of nucleus pulposus mesenchymal stem cells via RhoA signaling pathway. Mol Biol Rep 2023; 50:6337-6347. [PMID: 37310547 DOI: 10.1007/s11033-023-08547-6] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/26/2023] [Accepted: 05/23/2023] [Indexed: 06/14/2023]
Abstract
BACKGROUND Degenerative disc disease(DDD)is one of the most important causes of low back pain (LBP). Programmed death of human nucleus pulposus mesenchymal stem cells (NPMSCs) plays an important role in the progression of DDD. Growth differentiation factor-5 (GDF-5) is a protein that promotes chondrogenic differentiation, and has been reported to slow the expression of inflammatory factors in nucleus pulposus cells. Compared with those in normal rats, MRI T2-weighted images show hypointense in the central nucleus pulposus region of the intervertebral disc in GDF-5 knockout rats. METHODS AND RESULTS We aimed to evaluate the role of GDF-5 and Ras homolog family member A (RhoA) in NPMSCs. We used lipopolysaccharide (LPS) to simulate the inflammatory environment in degenerative disc disease, and performed related experiments on the effects of GDF-5 on NPMSCs, including the effects of pyroptosis, RhoA protein, and the expression of extracellular matrix components, and the effects of GDF-5, on NPMSCs. In addition, the effect of GDF-5 on chondroid differentiation of NPMSCs was included. The results showed that the addition of GDF-5 inhibited the LPS-induced pyroptosis of NPMSCs, and further analysis of its mechanism showed that this was achieved by activating the RhoA signaling pathway. CONCLUSION These findings suggest that GDF-5 plays an important role in inhibiting the pyroptosis of NPMSCs and GDF-5 may have potential for degenerative disc disease gene-targeted therapy in the future.
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Affiliation(s)
- Chao Zhu
- The Second School of Clinical Medicine, Southern Medical University, Guangzhou, Guangdong, 510515, China
- Department of Orthopedic Surgery, The Sixth Medical Centre of PLA General Hospital, Beijing, 100048, China
| | - Qing Zhou
- Department of Orthopedic Surgery, The Sixth Medical Centre of PLA General Hospital, Beijing, 100048, China
- Department of Orthopedic Surgery, Navy Clinical College of Anhui Medical University, Beijing, 100048, China
| | - Zuqiang Wang
- Department of Orthopedic Surgery, The Sixth Medical Centre of PLA General Hospital, Beijing, 100048, China
| | - Junyou Zhang
- The Second School of Clinical Medicine, Southern Medical University, Guangzhou, Guangdong, 510515, China
- Department of Orthopedic Surgery, The Sixth Medical Centre of PLA General Hospital, Beijing, 100048, China
| | - Cheng Xu
- Department of Orthopedic Surgery, The Sixth Medical Centre of PLA General Hospital, Beijing, 100048, China
| | - Dike Ruan
- The Second School of Clinical Medicine, Southern Medical University, Guangzhou, Guangdong, 510515, China.
- Department of Orthopedic Surgery, The Sixth Medical Centre of PLA General Hospital, Beijing, 100048, China.
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15
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Gasparella M, Cenzi C, Piccione M, Madia VN, Di Santo R, Tudino V, Artico M, Taurone S, De Ponte C, Costi R, Di Liddo R. Effects of Modified Glucosamine on the Chondrogenic Potential of Circulating Stem Cells under Experimental Inflammation. Int J Mol Sci 2023; 24:10397. [PMID: 37373540 DOI: 10.3390/ijms241210397] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/24/2022] [Revised: 06/05/2023] [Accepted: 06/16/2023] [Indexed: 06/29/2023] Open
Abstract
Glucosamine (GlcN) is a glycosaminoglycan (GAGs) constituent in connective tissues. It is naturally produced by our body or consumed from diets. In the last decade, in vitro and in vivo trials have demonstrated that the administration of GlcN or its derivates has a protective effect on cartilage when the balance between catabolic and anabolic processes is disrupted and cells are no longer able to fully compensate for the loss of collagen and proteoglycans. To date, these benefits are still controversial because the mechanism of action of GlcN is not yet well clarified. In this study, we have characterized the biological activities of an amino acid (AA) derivate of GlcN, called DCF001, in the growth and chondrogenic induction of circulating multipotent stem cells (CMCs) after priming with tumor necrosis factor-alpha (TNFα), a pleiotropic cytokine commonly expressed in chronic inflammatory joint diseases. In the present work, stem cells were isolated from the human peripheral blood of healthy donors. After priming with TNFα (10 ng/mL) for 3 h, cultures were treated for 24 h with DCF001 (1 μg/mL) dissolved in a proliferative (PM) or chondrogenic (CM) medium. Cell proliferation was analyzed using a Corning® Cell Counter and trypan blue exclusion technique. To evaluate the potentialities of DCF001 in counteracting the inflammatory response to TNFα, we measured the amount of extracellular ATP (eATP) and the expression of adenosine-generating enzymes CD39/CD73, TNFα receptors, and NF-κB inhibitor IκBα using flow cytometry. Finally, total RNA was extracted to perform a gene expression study of some chondrogenic differentiation markers (COL2A1, RUNX2, and MMP13). Our analysis has shed light on the ability of DCF001 to (a) regulate the expression of CD39, CD73, and TNF receptors; (b) modulate eATP under differentiative induction; (c) enhance the inhibitory activity of IκBα, reducing its phosphorylation after TNFα stimulation; and (d) preserve the chondrogenic potentialities of stem cells. Although preliminary, these results suggest that DCF001 could be a valuable supplement for ameliorating the outcome of cartilage repair interventions, enhancing the efficacy of endogenous stem cells under inflammatory stimuli.
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Affiliation(s)
- Marco Gasparella
- Local Health Unit Treviso, Department of Pediatric Surgery, 31100 Treviso, Italy
| | - Carola Cenzi
- Department of Pharmaceutical and Pharmacological Sciences, University of Padova, 35131 Padova, Italy
| | - Monica Piccione
- Department of Pharmaceutical and Pharmacological Sciences, University of Padova, 35131 Padova, Italy
| | - Valentina Noemi Madia
- Department of Drug Chemistry and Technology, University of Rome "La Sapienza", 00185 Rome, Italy
| | - Roberto Di Santo
- Department of Drug Chemistry and Technology, University of Rome "La Sapienza", 00185 Rome, Italy
| | - Valeria Tudino
- Department of Drug Chemistry and Technology, University of Rome "La Sapienza", 00185 Rome, Italy
| | - Marco Artico
- Department of Sensory Organs, University of Rome "La Sapienza", 00185 Rome, Italy
| | - Samanta Taurone
- Department of Movement, Human and Health Sciences-Division of Health Sciences, University of Rome "Foro Italico", 00185 Rome, Italy
| | - Chiara De Ponte
- Department of Sensory Organs, University of Rome "La Sapienza", 00185 Rome, Italy
| | - Roberta Costi
- Department of Drug Chemistry and Technology, University of Rome "La Sapienza", 00185 Rome, Italy
| | - Rosa Di Liddo
- Department of Pharmaceutical and Pharmacological Sciences, University of Padova, 35131 Padova, Italy
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16
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Guo J, Ye W, Wu X, Huang H, Li B, Sun Z, Ren Z, Yang Z. TNF-α activates RELA expression via TNFRSF1B to upregulate OPA1 expression and inhibit chondrogenic differentiation of human adipose stem cells. J Orthop Surg Res 2023; 18:430. [PMID: 37312126 DOI: 10.1186/s13018-023-03846-x] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/08/2023] [Accepted: 05/09/2023] [Indexed: 06/15/2023] Open
Abstract
BACKGROUND Tumor necrosis factor-alpha (TNF-α), one of the pro-inflammatory cytokines mediating the local inflammatory process in joints, inhibits cartilage formation and has a detrimental effect on stem cell-based cartilage regeneration for the treatment of osteoarthritis (OA). However, the mechanisms behind this inhibitory effect are still poorly understood. Mitochondrial morphological changes mediated by mitochondrial fusion and fission are highly plastic, are quite sensitive to environmental stimuli and play a crucial role in maintaining cell structure and function. In our study, chondrogenic differentiated human adipose stem cells (hADSCs) were exposed to TNF-α and the effect of TNF-α on the ability of hADSCs to chondrogenic differentiate and on mitochondrial fusion and fission was observed and analyzed. The aim was to investigate the role and mechanisms of mitochondrial fusion and fission regulation in the chondrogenic differentiation of hADSCs under normal conditions and under exposure to TNF-α. METHODS We used flow cytometry to identify hADSCs immunophenotypes CD29, CD44, CD34, CD45, and HLA-DR. Alcian blue staining and Sirius red staining were used to observe the formation of proteoglycans and collagen during the chondrogenic differentiation of hADSCs, respectively. The mRNA and protein expression levels of the cartilage formation marker SOX9, type II collagen (COL2A1), and Aggrecan were measured by real-time fluorescent quantitative PCR (RT-qPCR) and western blot, respectively. The fluorescent probes MitoTracker® Red CMXRos and JC-1 were used to visualize mitochondria morphology and detect mitochondrial membrane electricity (MMP). Affymetrix PrimeView™ chips were used for gene expression profiling. RESULTS The results showed that the chondrogenic differentiation of hADSCs was inhibited in the presence of TNF-α that optic atrophy 1 (OPA1) expression was significantly upregulated and mitochondria were prolonged and interconnected during this process. Gene microarray and RT-qPCR data showed that the presence of TNF-α led to increased expression of TNFα receptor 2 (TNFRSF1B) and RELA during chondrogenic differentiation of hADSCs. CONCLUSIONS TNF-α inhibits chondrogenic differentiation of human adipose stem cells by activating RELA expression through TNFRSF1B upregulating OPA1 expression thereby increasing mitochondrial fusion.
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Affiliation(s)
- Jiajia Guo
- Medical College of Guizhou University, Guiyang, 550025, Guizhou, China
| | - Wang Ye
- Department of Orthopedics, Guizhou Provincial People's Hospital, Guiyang, 550002, Guizhou, China
| | - Xinglin Wu
- Department of Orthopedics, Guizhou Provincial People's Hospital, Guiyang, 550002, Guizhou, China
| | - Haifeng Huang
- Department of Orthopedics, Guizhou Provincial People's Hospital, Guiyang, 550002, Guizhou, China
| | - Bo Li
- Department of Orthopedics, Guizhou Provincial People's Hospital, Guiyang, 550002, Guizhou, China
| | - Zeyu Sun
- Department of Orthopedics, Guizhou Provincial People's Hospital, Guiyang, 550002, Guizhou, China
| | - Zhijing Ren
- Department of Clinical Laboratory, Guizhou Provincial People's Hospital, Guiyang, 550002, Guizhou, China.
| | - Zhen Yang
- Department of Orthopedics, Guizhou Provincial People's Hospital, Guiyang, 550002, Guizhou, China.
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17
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Yamaura K, Nelson AL, Nishimura H, Rutledge JC, Ravuri SK, Bahney C, Philippon MJ, Huard J. The effects of losartan or angiotensin II receptor antagonists on cartilage: a systematic review. Osteoarthritis Cartilage 2023; 31:435-446. [PMID: 36586717 DOI: 10.1016/j.joca.2022.11.014] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/01/2022] [Revised: 11/06/2022] [Accepted: 11/28/2022] [Indexed: 12/29/2022]
Abstract
OBJECTIVE The aim of this study is to analyze the latest evidence on the effects of losartan or Ang II receptor antagonists on cartilage repair, with a focus on their clinical relevance. DESIGN The PubMed, Embase, and Cochrane Library databases were searched up to November 12th 2021 to evaluate the effects of losartan or Ang II receptor antagonists on cartilage repair in in vitro studies and in vivo animal studies. Study design, sample characteristics, treatment type, duration, and outcomes were analyzed. The risk of bias and the quality of the eligible studies were assessed using the Systematic Review Centre for Laboratory Animal Experimentation (SYRCLE) risk of bias assessment tool and Collaborative Approach to Meta-Analysis and Review of Animal Data from Experimental Studies (CAMARADES). RESULTS A total of 12 studies were included in this systematic review. Of the 12 eligible studies, two studies were in vitro human studies, three studies were in vitro animal studies, one study was an in vitro human and animal study, and six studies were in vivo animal studies. The risk bias and quality assessments were predominantly classified as moderate. Since meta-analysis was difficult due to differences in treatment type, dosage, route of administration, and method of outcome assessment among the eligible studies, qualitative evaluation was conducted for each study. CONCLUSIONS Both in vitro and in vivo studies provide evidence to demonstrate beneficial effects of Ang II receptor antagonists on osteoarthritis and cartilage defect models across animal species.
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Affiliation(s)
- K Yamaura
- Center for Regenerative and Personalized Medicine, Steadman Philippon Research Institute, Vail, CO, USA; Department of Orthopaedic Surgery, Kobe University Graduate School of Medicine, Kobe, Japan.
| | - A L Nelson
- Center for Regenerative and Personalized Medicine, Steadman Philippon Research Institute, Vail, CO, USA.
| | - H Nishimura
- Center for Regenerative and Personalized Medicine, Steadman Philippon Research Institute, Vail, CO, USA; Department of Orthopaedic Surgery, University Hospital of Occupational and Environmental Health, Fukuoka, Japan.
| | - J C Rutledge
- Center for Regenerative and Personalized Medicine, Steadman Philippon Research Institute, Vail, CO, USA.
| | - S K Ravuri
- Center for Regenerative and Personalized Medicine, Steadman Philippon Research Institute, Vail, CO, USA.
| | - C Bahney
- Center for Regenerative and Personalized Medicine, Steadman Philippon Research Institute, Vail, CO, USA; The Orthopaedic Trauma Institute, University of California, San Francisco (UCSF), San Francisco, CA, USA.
| | - M J Philippon
- Center for Regenerative and Personalized Medicine, Steadman Philippon Research Institute, Vail, CO, USA; The Steadman Clinic, Vail, CO, USA.
| | - J Huard
- Center for Regenerative and Personalized Medicine, Steadman Philippon Research Institute, Vail, CO, USA.
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18
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Panos JA, Coenen MJ, Nagelli CV, McGlinch EB, Atasoy-Zeybek A, De Padilla CL, Coghlan RF, Johnstone B, Ferreira E, Porter RM, De la Vega RE, Evans CH. IL-1Ra gene transfer potentiates BMP2-mediated bone healing by redirecting osteogenesis toward endochondral ossification. Mol Ther 2023; 31:420-434. [PMID: 36245128 PMCID: PMC9931547 DOI: 10.1016/j.ymthe.2022.10.007] [Citation(s) in RCA: 8] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/23/2022] [Revised: 09/14/2022] [Accepted: 10/14/2022] [Indexed: 11/05/2022] Open
Abstract
An estimated 100,000 patients each year in the United States suffer severe disability from bone defects that fail to heal, a condition where bone-regenerative therapies could provide substantial clinical benefits. Although recombinant human bone morphogenetic protein-2 (rhBMP2) is an osteogenic growth factor that is clinically approved for this purpose, it is only effective when used at exceedingly high doses that incur substantial costs, induce severe inflammation, produce adverse side effects, and form morphologically abnormal bone. Using a validated rat femoral segmental defect model, we show that bone formed in response to clinically relevant doses of rhBMP2 is accompanied by elevated expression of interleukin-1 (IL-1). Local delivery of cDNA encoding the IL-1 receptor antagonist (IL-1Ra) achieved bridging of segmental, critical size defects in bone with a 90% lower dose of rhBMP2. Unlike use of high-dose rhBMP2, bone formation in the presence of IL-1Ra occurred via the native process of endochondral ossification, resulting in improved quality without sacrificing the mechanical properties of the regenerated bone. Our results demonstrate that local immunomodulation may permit effective use of growth factors at lower doses to recapitulate more precisely the native biology of healing, leading to higher-quality tissue regeneration.
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Affiliation(s)
- Joseph A Panos
- Rehabilitation Medicine Research Center, Mayo Clinic, Rochester, MN, USA; Musculoskeletal Gene Therapy Research Laboratory, Mayo Clinic, Rochester, MN, USA; Graduate School of Biomedical Sciences, Mayo Clinic, Rochester, MN, USA; Medical Scientist Training Program, Mayo Clinic, Rochester, MN, USA
| | - Michael J Coenen
- Rehabilitation Medicine Research Center, Mayo Clinic, Rochester, MN, USA; Musculoskeletal Gene Therapy Research Laboratory, Mayo Clinic, Rochester, MN, USA
| | - Christopher V Nagelli
- Rehabilitation Medicine Research Center, Mayo Clinic, Rochester, MN, USA; Musculoskeletal Gene Therapy Research Laboratory, Mayo Clinic, Rochester, MN, USA
| | - Erin B McGlinch
- Rehabilitation Medicine Research Center, Mayo Clinic, Rochester, MN, USA; Musculoskeletal Gene Therapy Research Laboratory, Mayo Clinic, Rochester, MN, USA; Graduate School of Biomedical Sciences, Mayo Clinic, Rochester, MN, USA; Virology and Gene Therapy Graduate Program, Mayo Clinic, Rochester, MN, USA
| | - Aysegul Atasoy-Zeybek
- Rehabilitation Medicine Research Center, Mayo Clinic, Rochester, MN, USA; Musculoskeletal Gene Therapy Research Laboratory, Mayo Clinic, Rochester, MN, USA
| | - Consuelo Lopez De Padilla
- Rehabilitation Medicine Research Center, Mayo Clinic, Rochester, MN, USA; Musculoskeletal Gene Therapy Research Laboratory, Mayo Clinic, Rochester, MN, USA
| | - Ryan F Coghlan
- Research Center, Shriners Hospitals for Children, Portland, OR, USA
| | - Brian Johnstone
- Research Center, Shriners Hospitals for Children, Portland, OR, USA; Department of Orthopedics and Rehabilitation, Oregon Health & Science University, Portland, OR, USA
| | - Elisabeth Ferreira
- Center for Musculoskeletal Disease Research, Departments of Internal Medicine and Orthopedic Surgery, University of Arkansas for Medical Sciences, Little Rock, AR, USA
| | - Ryan M Porter
- Center for Musculoskeletal Disease Research, Departments of Internal Medicine and Orthopedic Surgery, University of Arkansas for Medical Sciences, Little Rock, AR, USA
| | - Rodolfo E De la Vega
- Rehabilitation Medicine Research Center, Mayo Clinic, Rochester, MN, USA; Musculoskeletal Gene Therapy Research Laboratory, Mayo Clinic, Rochester, MN, USA; Department of Cell Biology-Inspired Tissue Engineering, MERLN Institute, Maastricht, the Netherlands
| | - Christopher H Evans
- Rehabilitation Medicine Research Center, Mayo Clinic, Rochester, MN, USA; Musculoskeletal Gene Therapy Research Laboratory, Mayo Clinic, Rochester, MN, USA.
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19
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Stoddart MJ, Della Bella E, Armiento AR. Cartilage Tissue Engineering: An Introduction. Methods Mol Biol 2023; 2598:1-7. [PMID: 36355280 DOI: 10.1007/978-1-0716-2839-3_1] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 06/16/2023]
Abstract
Once damaged, cartilage has limited healing capability. This has led to a huge body of research that aims to repair or regenerate this important tissue. Despite the progress made, significant hurdles still need to be overcome. This chapter highlights some of the progress made, while elaborating on areas that need further research. The concept of translation and the route to clinical translation must be kept in mind if some of the promising preclinical research is to make it to routine clinical application.
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Affiliation(s)
| | | | - Angela R Armiento
- AO Research Institute Davos, Davos Platz, Switzerland
- UCB Pharma, Slough, UK
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20
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Li W, Liu Q, Shi J, Xu X, Xu J. The role of TNF-α in the fate regulation and functional reprogramming of mesenchymal stem cells in an inflammatory microenvironment. Front Immunol 2023; 14:1074863. [PMID: 36814921 PMCID: PMC9940754 DOI: 10.3389/fimmu.2023.1074863] [Citation(s) in RCA: 31] [Impact Index Per Article: 15.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/20/2022] [Accepted: 01/24/2023] [Indexed: 02/09/2023] Open
Abstract
Mesenchymal stem cells (MSCs) are pluripotent stem cells with multidirectional differentiation potential and strong immunomodulatory capacity. MSCs have been widely used in the treatment of injured, inflammatory, and immune-related diseases. Resting MSCs lack differentiation and immunomodulatory ability. Instead, they rely on microenvironmental factors to: 1) stimulate and regulate their expression of specific cell growth factors, chemokines, immunomodulatory factors, or receptors; or 2) direct their differentiation into specific tissue cells, which ultimately perform tissue regeneration and repair and immunomodulatory functions. Tumor necrosis factor (TNF)-α is central to the creation of an inflammatory microenvironment. TNF-α regulates the fate and functional reprogramming of MSCs, either alone or in combination with a variety of other inflammatory factors. TNF-α can exert opposing effects on MSCs, from inducing MSC apoptosis to enhancing their anti-tumor capacity. In addition, the immunomodulation and osteogenic differentiation capacities of MSCs, as well as their exosome or microvesicle components vary significantly with TNF-α stimulating concentration, time of administration, or its use in combination with or without other factors. Therefore, this review discusses the impact of TNF-α on the fate and functional reprogramming of MSCs in the inflammatory microenvironment, to provide new directions for improving the immunomodulatory and tissue repair functions of MSCs and enhance their therapeutic potential.
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Affiliation(s)
- Weiqiang Li
- Department of Stem Cell & Regenerative Medicine, State Key Laboratory of Trauma, Burn and Combined Injury, Daping Hospital, Army Medical University, Chongqing, China.,Department of Research and Development, Ankerui (Shanxi) Biological Cell Co., Ltd., Shanxi, China
| | - Qianqian Liu
- Department of Research and Development, Ankerui (Shanxi) Biological Cell Co., Ltd., Shanxi, China
| | - Jinchao Shi
- Department of Research and Development, Ankerui (Shanxi) Biological Cell Co., Ltd., Shanxi, China
| | - Xiang Xu
- Department of Stem Cell & Regenerative Medicine, State Key Laboratory of Trauma, Burn and Combined Injury, Daping Hospital, Army Medical University, Chongqing, China.,Department of Biochemistry and Molecular Biology, College of Basic Medical Science, Army Medical University, Chongqing, China
| | - Jinyi Xu
- State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University, Guangzhou, China
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21
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Hechavarria ME, Richard SA. Elucidating the Focal Immunomodulatory Clues Influencing Mesenchymal Stem Cells in the Milieu of Intervertebral Disc Degeneration. Curr Stem Cell Res Ther 2023; 18:62-75. [PMID: 35450531 DOI: 10.2174/1574888x17666220420134619] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/19/2021] [Revised: 03/21/2022] [Accepted: 03/29/2022] [Indexed: 11/22/2022]
Abstract
The intervertebral discs (IVDs) are a relatively mobile joint that interconnects vertebrae of the spine. Intervertebral disc degeneration (IVDD) is one of the leading causes of low back pain, which is most often related to patient morbidity as well as high medical costs. Patients with chronic IVDD often need surgery that may sometimes lead to biomechanical complications as well as augmented degeneration of the adjacent segments. Moreover, treatment modalities like rigid intervertebral fusion, dynamic instrumentation, as well as other surgical interventions are still controversial. Mesenchymal stem cells (MSCs) have exhibited to have immunomodulatory functions and the ability to differentiate into cartilage, making these cells possibly an epitome for IVD regeneration. Transplanted MSCs were able to repair IVDD back to the normal disc milieu via the activation of the generation of extracellular matrix (ECM) proteins such as aggrecan, proteoglycans and collagen types I and II. IVD milieu clues like, periostin, cluster of differentiation, tumor necrosis factor alpha, interleukins, chemokines, transforming growth factor beta, reactive oxygen species, toll-like receptors, tyrosine protein kinase receptor and disialoganglioside, exosomes are capable of influencing the MSCs during treatment of IVDD. ECM microenvironment clues above have potentials as biomarkers as well as accurate molecular targets for therapeutic intervention in IVDD.
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Affiliation(s)
| | - Seidu A Richard
- Department of Medicine, Princefield University, P. O. Box MA 128, Ho-Volta Region, Ghana, West Africa
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22
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Zelinka A, Roelofs AJ, Kandel RA, De Bari C. Cellular therapy and tissue engineering for cartilage repair. Osteoarthritis Cartilage 2022; 30:1547-1560. [PMID: 36150678 DOI: 10.1016/j.joca.2022.07.012] [Citation(s) in RCA: 32] [Impact Index Per Article: 10.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/23/2022] [Revised: 07/01/2022] [Accepted: 07/05/2022] [Indexed: 02/02/2023]
Abstract
Articular cartilage (AC) has limited capacity for repair. The first attempt to repair cartilage using tissue engineering was reported in 1977. Since then, cell-based interventions have entered clinical practice in orthopaedics, and several tissue engineering approaches to repair cartilage are in the translational pipeline towards clinical application. Classically, these involve a scaffold, substrate or matrix to provide structure, and cells such as chondrocytes or mesenchymal stromal cells to generate the tissue. We discuss the advantages and drawbacks of the use of various cell types, natural and synthetic scaffolds, multiphasic or gradient-based scaffolds, and self-organizing or self-assembling scaffold-free systems, for the engineering of cartilage constructs. Several challenges persist including achieving zonal tissue organization and integration with the surrounding tissue upon implantation. Approaches to improve cartilage thickness, organization and mechanical properties include mechanical stimulation, culture under hypoxic conditions, and stimulation with growth factors or other macromolecules. In addition, advanced technologies such as bioreactors, biosensors and 3D bioprinting are actively being explored. Understanding the underlying mechanisms of action of cell therapy and tissue engineering approaches will help improve and refine therapy development. Finally, we discuss recent studies of the intrinsic cellular and molecular mechanisms of cartilage repair that have identified novel signals and targets and are inspiring the development of molecular therapies to enhance the recruitment and cartilage reparative activity of joint-resident stem and progenitor cells. A one-fits-all solution is unrealistic, and identifying patients who will respond to a specific targeted treatment will be critical.
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Affiliation(s)
- A Zelinka
- Lunenfeld Tanenbaum Research Institute, Sinai Health, Dept. Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Canada
| | - A J Roelofs
- Arthritis and Regenerative Medicine Laboratory, Aberdeen Centre for Arthritis and Musculoskeletal Health, University of Aberdeen, Aberdeen, UK
| | - R A Kandel
- Lunenfeld Tanenbaum Research Institute, Sinai Health, Dept. Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Canada.
| | - C De Bari
- Arthritis and Regenerative Medicine Laboratory, Aberdeen Centre for Arthritis and Musculoskeletal Health, University of Aberdeen, Aberdeen, UK.
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23
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Glycogen Synthase Kinase 3β inhibits BMSCs Chondrogenesis in Inflammation via the Cross-Reaction between NF-κB and β-Catenin in the Nucleus. Stem Cells Int 2022; 2022:5670403. [PMID: 36132167 PMCID: PMC9484947 DOI: 10.1155/2022/5670403] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/14/2022] [Accepted: 08/17/2022] [Indexed: 11/17/2022] Open
Abstract
Inflammation can influence the pluripotency and self-renewal of mesenchymal stem cells (MSCs), thereby altering their cartilage regeneration ability. Sprague-Dawley (SD) rat bone marrow mesenchymal stem cells (BMSCs) were isolated and found to be defective in differentiation potential in the interleukin-1β- (IL-1β-) induced inflammatory microenvironment. Glycogen synthase kinase-3β (GSK-3β) is an evolutionarily conserved serine/threonine kinase that plays a role in numerous cellular processes. The role of GSK-3β in inflammation may be related to the nuclear factor-κB (NF-κB) signaling pathway and the Wnt/β-catenin signaling pathway, whose mechanism remains unclear. In this study, we found that GSK-3β can inhibit chondrogenesis of IL-1β-impaired BMSCs by disrupting metabolic balance and promoting cell apoptosis. By using the inhibitors LiCl and SN50, we demonstrated that GSK-3β regulates the chondrogenesis via the NF-κB and Wnt/β-catenin signaling pathways and possibly mediates the cross-reaction between NF-κB and β-catenin in the nucleus. Given the molecular mechanisms of GSK-3β in chondrogenic differentiation in inflammation, GSK-3β is a crucial target for the treatment of inflammation-induced cartilage disease.
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24
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Chen R, Li X, Sun Z, Yin J, Hu X, Deng J, Liu X. Intra-bone marrow injection of magnesium isoglyrrhizinate inhibits inflammation and delays osteoarthritis progression through the NF-κB pathway. J Orthop Surg Res 2022; 17:400. [PMID: 36045373 PMCID: PMC9429748 DOI: 10.1186/s13018-022-03294-z] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/27/2022] [Accepted: 08/18/2022] [Indexed: 11/28/2022] Open
Abstract
Objective Osteoarthritis (OA) presents cartilage damage in addition to chronic inflammation. However, self-recovery of damaged cartilage in an inflammatory environment is not possible. Mesenchymal stem cells (MSCs) in the bone marrow are a source of regenerative repair of damaged cartilage. To date, whether intra-luminal administration of the bone marrow can delay the progression of OA is still unknown. This study, therefore, aimed to explore the role of intra-bone marrow injection of Magnesium isoglycyrrhizinate (MgIG) in delaying the OA progression and to investigate the underlying mechanism. Methods Rabbit OA models were established using the anterior cruciate ligament transection method while a catheter was implanted into the bone marrow cavity. 1 week after surgery, MgIG treatment was started once a week for 4 weeks. The cartilage degradation was analyzed using hematoxylin–eosin staining, Masson’s trichrome staining and Alcian blue staining. Additionally, the pro-inflammatory factors and cartilage regeneration genes involved in the cartilage degeneration and the underlying mechanisms in OA were detected using enzyme-linked immunosorbent assay, quantitative real-time PCR (qRT-PCR) and Western blotting. Results The results of histological staining revealed that intra-bone marrow injection of MgIG reduced degeneration and erosion of articular cartilage, substantially reducing the Osteoarthritis Research Society International scores. Furthermore, the productions of inflammatory cytokines in the bone marrow cavity and articular cavity such as interleukin-1β(IL-1β), IL-6, and tumor necrosis factor-α (TNF-α) were inhibited upon the treatment of MgIG. At the same time, the expression of alkaline phosphate, tartrate-resistant acid phosphatase-5b (TRAP-5b) and C-telopeptides of type II collagen (CTX-II) in the blood also decreased and was positively correlated. On the contrary, cartilage-related genes in the bone marrow cavity such as type II collagen (Col II), Aggrecan (AGN), and SRY-box 9 (SOX9) were up-regulated, while matrix metalloproteinase-3 (MMP-3) was down-regulated. Mechanistically, MgIG was found to exert an anti-inflammatory effect and impart protection to the cartilage by inhibiting the NF-κB pathway. Conclusion Intra-bone marrow injection of MgIG might inhibit the activation of the NF-κB pathway in the progression of OA to exert an anti-inflammatory effect in the bone marrow cavity and articular cavity, thereby promoting cartilage regeneration of MSCs in the bone marrow, making it a potential new therapeutic intervention for the treatment of OA.
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Affiliation(s)
- Rong Chen
- Department of Traumatic Orthopedics, Renmin Hospital, Hubei University of Medicine, Shiyan, 442000, Hubei, China
| | - Xiangwei Li
- Department of Traumatic Orthopedics, Renmin Hospital, Hubei University of Medicine, Shiyan, 442000, Hubei, China
| | - Zhibo Sun
- Department of Traumatic Orthopedics, Renmin Hospital, Hubei University of Medicine, Shiyan, 442000, Hubei, China
| | - Junyi Yin
- Hubei Key Laboratory of Embryonic Stem Cell Research, Department of Anatomy, School of Basic Medical Sciences, Hubei University of Medicine, No. 30 Renmin South Road, Maojian District, Shiyan, 442000, Hubei, China
| | - Xiaowei Hu
- Hubei Key Laboratory of Embryonic Stem Cell Research, Department of Anatomy, School of Basic Medical Sciences, Hubei University of Medicine, No. 30 Renmin South Road, Maojian District, Shiyan, 442000, Hubei, China
| | - Jingwen Deng
- Hubei Key Laboratory of Embryonic Stem Cell Research, Department of Anatomy, School of Basic Medical Sciences, Hubei University of Medicine, No. 30 Renmin South Road, Maojian District, Shiyan, 442000, Hubei, China
| | - Xinghui Liu
- Department of Traumatic Orthopedics, Renmin Hospital, Hubei University of Medicine, Shiyan, 442000, Hubei, China. .,Hubei Key Laboratory of Embryonic Stem Cell Research, Department of Anatomy, School of Basic Medical Sciences, Hubei University of Medicine, No. 30 Renmin South Road, Maojian District, Shiyan, 442000, Hubei, China.
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25
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Neefjes M, Housmans BAC, Thielen NGM, van Beuningen HM, Vitters EL, van den Akker GGH, Welting TJM, van Caam APM, van der Kraan PM. An improved diagnostic tool to predict cartilage formation in an osteoarthritic joint environment. Tissue Eng Part A 2022; 28:907-917. [PMID: 35943880 DOI: 10.1089/ten.tea.2022.0023] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/13/2022] Open
Abstract
Osteoarthritis (OA) is a degenerative joint disease with progressive articular cartilage loss. Due to the chondrogenic potential of human mesenchymal stromal cells (MSCs), MSC-based therapies are promising treatment strategies for cartilage loss. However, the local joint microenvironment has a great impact on the success of cartilage formation by MSCs. This local joint environment is different between patients and therefore the outcome of MSC therapies is uncertain. We previously developed gene promoter-based reporter assays as a novel tool to predict the effect of a patient's OA joint microenvironment on the success of MSC-based cartilage formation. Here we describe an improved version of this molecular tool with increased prediction accuracy. For this, we generated fourteen stable cell lines using transcription factor (TF) binding elements (AP1, ARE, CRE, GRE, ISRE, NFAT5, NFκB, PPRE, SBE, SIE, SOX9, SRE, SRF, TCF/LEF) to drive luciferase reporter gene expression, and evaluated the cell lines for their responsiveness to an osteoarthritic microenvironment by stimulation with OA synovium-conditioned medium (OAs-cm; n=31). To study the effect of this OA microenvironment on MSC-based cartilage formation, MSCs were cultured in a three-dimensional pellet culture model while stimulated with OAs-cm. Cartilage formation was assessed histologically and by quantifying sulfated glycosaminoglycan (sGAG) production. Six TF reporters correlated significantly with the effect of OAs-cm on cartilage formation. We validated the predictive value of these TF reporters with an independent cohort of OAs-cm (n=22) and compared the prediction accuracy between our previous and the current new tool. Furthermore, we investigated which combination of reporters could predict the effect of the OA microenvironment on cartilage repair with the highest accuracy. A combination between the TF (NFκB) and the promoter-based (IL6) reporter proved to reach a more accurate prediction compared to the tools separately. These developments are an important step towards a diagnostic tool that can be used for personalized cartilage repair strategies for OA patients.
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Affiliation(s)
- Margot Neefjes
- Radboudumc, Experimental Rheumatology, Geert Grooteplein 28, Nijmegen, Netherlands, 6500 HB;
| | - Bas A C Housmans
- Maastricht University, Department of Orthopedic Surgery, Maastricht, Limburg, Netherlands;
| | | | | | - Elly L Vitters
- Radboudumc, Experimental Rheumatology, Nijmegen, Netherlands;
| | - Guus G H van den Akker
- Maastricht University, Department of Orthopedic Surgery, Maastricht, Limburg, Netherlands;
| | - Tim J M Welting
- University Hospital Maastricht, Department of Orthopaedic Surgery, P Debyelaan 25, Maastricht, Limburg, Netherlands, 6202 AZ;
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26
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Guilak F, Estes BT, Moutos FT. Functional tissue engineering of articular cartilage for biological joint resurfacing-The 2021 Elizabeth Winston Lanier Kappa Delta Award. J Orthop Res 2022; 40:1721-1734. [PMID: 34812518 PMCID: PMC9124734 DOI: 10.1002/jor.25223] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/23/2021] [Revised: 11/11/2021] [Accepted: 11/20/2021] [Indexed: 02/04/2023]
Abstract
Biological resurfacing of entire articular surfaces represents a challenging strategy for the treatment of cartilage degeneration that occurs in osteoarthritis. Not only does this approach require anatomically sized and functional engineered cartilage, but the inflammatory environment within an arthritic joint may also inhibit chondrogenesis and induce degradation of native and engineered cartilage. Here, we present the culmination of multiple avenues of interdisciplinary research leading to the development and testing of bioartificial cartilage for tissue-engineered resurfacing of the hip joint. The work is based on a novel three-dimensional weaving technology that is infiltrated with specific bioinductive materials and/or genetically-engineered stem cells. A variety of design approaches have been tested in vitro, showing biomimetic cartilage-like properties as well as the capability for long-term tunable and inducible drug delivery. Importantly, these cartilage constructs have the potential to provide mechanical functionality immediately upon implantation, as they will need to replace a majority, if not the entire joint surface to restore function. To date, these approaches have shown excellent preclinical success in a variety of animal studies, including the resurfacing of a large osteochondral defect in the canine hip, and are now well-poised for clinical translation.
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Affiliation(s)
- Farshid Guilak
- Department of Orthopaedic Surgery, Washington University, St. Louis, MO, USA,Shriners Hospitals for Children – St. Louis, St. Louis, MO, USA,Center of Regenerative Medicine, Washington University, St. Louis, MO, USA,Cytex Therapeutics, Inc., Durham, NC, USA
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27
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Nagelli CV, De La Vega RE, Coenen M, De Padilla CL, Panos JA, Tovar A, Müller SA, Evans CH. Expedited gene delivery for osteochondral defect repair in a rabbit knee model: A one-year investigation. OSTEOARTHRITIS AND CARTILAGE OPEN 2022; 4. [PMID: 36338933 PMCID: PMC9635382 DOI: 10.1016/j.ocarto.2022.100257] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/03/2022] Open
Abstract
Objective: To evaluate a single-step, gene-based procedure for repairing osteochondral lesions. Design: Osteochondral lesions were created in the patellar groove of skeletally mature rabbits. Autologous bone marrow aspirates were mixed with adenovirus vectors carrying cDNA encoding green fluorescent protein (Ad.GFP) or transforming growth factor-β1 (Ad.TGF-β1) and allowed to clot. The clotted marrow was press-fit into the defects. Animals receiving Ad.GFP were euthanized at 2 weeks and intra-articular expression of GFP examined by fluorescence microscopy. Animals receiving Ad.TGF-β1 were euthanized at 3 months and 12 months; repair was compared to empty defects using histology and immunohistochemistry. Complementary in vitro experiments assessed transgene expression and chondrogenesis in marrow clots and fibrin gels. In a subsequent pilot study, repair at 3 months using a fibrin gel to encapsulate Ad.TGF-β1 was evaluated. Results: At 2 weeks, GFP expression was seen at variable levels within the cartilaginous lesion. At 3 months, there was no statistically significant improvement (p > 0.05) in healing of lesions receiving Ad.TGF-β1 and variability was high. At 12 months, there were still no significant difference (p > 0.05) between the empty defects and those receiving Ad.TGF-β1 in the overall, cartilage, and bone scores. Variability was still high. In vitro experiments suggested that variability reflected variable transduction efficiency and chondrogenic activity of the marrow clots; using fibrin gels instead of marrow may address this issue but more research is needed. Conclusions: This approach to improving the repair of osteochondral lesions needs further refinement to reduce variability and provide a more robust outcome.
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28
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Wesdorp MA, Capar S, Bastiaansen-Jenniskens YM, Kops N, Creemers LB, Verhaar JA, Van Osch GJ, Wei W. Intra-articular Administration of Triamcinolone Acetonide in a Murine Cartilage Defect Model Reduces Inflammation but Inhibits Endogenous Cartilage Repair. Am J Sports Med 2022; 50:1668-1678. [PMID: 35315287 PMCID: PMC9069659 DOI: 10.1177/03635465221083693] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/31/2023]
Abstract
BACKGROUND Cartilage defects result in joint inflammation. The presence of proinflammatory factors has been described to negatively affect cartilage formation. PURPOSE To evaluate the effect and timing of administration of triamcinolone acetonide (TAA), an anti-inflammatory drug, on cartilage repair using a mouse model. STUDY DESIGN Controlled laboratory study. METHODS A full-thickness cartilage defect was created in the trochlear groove of 10-week-old male DBA/1 mice (N = 80). Mice received an intra-articular injection of TAA or saline on day 1 or 7 after induction of the defect. Mice were euthanized on days 10 and 28 for histological evaluation of cartilage defect repair, synovial inflammation, and synovial membrane thickness. RESULTS Mice injected with TAA had significantly less synovial inflammation at day 10 than saline-injected mice independent of the time of administration. At day 28, the levels of synovitis dropped toward healthy levels; nevertheless, the synovial membrane was thinner in TAA- than in saline-injected mice, reaching statistical significance in animals injected on day 1 (70.1 ± 31.9 µm vs 111.9 ± 30.9 µm, respectively; P = .01) but not in animals injected on day 7 (68.2 ± 21.86 µm vs 90.2 ± 21.29 µm, respectively; P = .26). A thinner synovial membrane was moderately associated with less filling of the defect after 10 and 28 days (r = 0.42, P = .02; r = 0.47, P = .01, respectively). Whereas 10 days after surgery there was no difference in the area of the defect filled and the cell density in the defect area between saline- and TAA-injected knees, filling of the defect at day 28 was lower in TAA- than in saline-injected knees for both injection time points (day 1 injection, P = .04; day 7 injection, P = .01). Moreover, there was less collagen type 2 staining in the filled defect area in TAA- than in saline-injected knees after 28 days, reaching statistical significance in day 1-injected knees (2.6% vs 18.5%, respectively; P = .01) but not in day 7-injected knees (7.4% vs 15.8%, respectively; P = .27). CONCLUSION Intra-articular injection of TAA reduced synovial inflammation but negatively affected cartilage repair. This implies that inhibition of inflammation may inhibit cartilage repair or that TAA has a direct negative effect on cartilage formation. CLINICAL RELEVANCE Our findings show that TAA can inhibit cartilage defect repair. Therefore, we suggest not using TAA to reduce inflammation in a cartilage repair setting.
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Affiliation(s)
- Marinus A. Wesdorp
- Department of Orthopaedic Surgery and Sports Medicine, Erasmus MC, University Medical Center, Rotterdam, the Netherlands
| | - Serdar Capar
- Department of Orthopaedic Surgery and Sports Medicine, Erasmus MC, University Medical Center, Rotterdam, the Netherlands
| | | | - Nicole Kops
- Department of Orthopaedic Surgery and Sports Medicine, Erasmus MC, University Medical Center, Rotterdam, the Netherlands
| | - Laura B. Creemers
- Department of Orthopedic Surgery, UMC Utrecht, University Medical Center, Utrecht, the Netherlands
| | - Jan A.N. Verhaar
- Department of Orthopaedic Surgery and Sports Medicine, Erasmus MC, University Medical Center, Rotterdam, the Netherlands
| | - Gerjo J.V.M. Van Osch
- Department of Orthopaedic Surgery and Sports Medicine, Erasmus MC, University Medical Center, Rotterdam, the Netherlands,Department of Otorhinolaryngology, Erasmus MC, University Medical Center, Rotterdam, the Netherlands,Department of Biomechanical Engineering, Faculty of Mechanical, Maritime, and Materials Engineering, Delft University of Technology, Delft, the Netherlands,Gerjo J.V.M. Van Osch, PhD, Department of Orthopaedic Surgery and Sports Medicine and Department of Otorhinolaryngology, Erasmus MC, University Medical Center, Room Ee16.55c, Dr Molewaterplein 40, Rotterdam, 3015 GD, the Netherlands ()
| | - Wu Wei
- Department of Orthopaedic Surgery and Sports Medicine, Erasmus MC, University Medical Center, Rotterdam, the Netherlands,Department of Orthopedic Surgery, Elisabeth-Tweesteden Ziekenhuis, Tilburg, the Netherlands
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Bhogoju S, Khan S, Subramanian A. Continuous Low-Intensity Ultrasound Preserves Chondrogenesis of Mesenchymal Stromal Cells in the Presence of Cytokines by Inhibiting NFκB Activation. Biomolecules 2022; 12:434. [PMID: 35327626 PMCID: PMC8946190 DOI: 10.3390/biom12030434] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/01/2022] [Revised: 02/21/2022] [Accepted: 03/03/2022] [Indexed: 02/04/2023] Open
Abstract
Proinflammatory joint environment, coupled with impeded chondrogenic differentiation of mesenchymal stromal cells (MSCs), led to inferior cartilage repair outcomes. Nuclear translocation of phosphorylated-NFκB downregulates SOX9 and hinders the chondrogenesis of MSCs. Strategies that minimize the deleterious effects of NFκB, while promoting MSC chondrogenesis, are of interest. This study establishes the ability of continuous low-intensity ultrasound (cLIUS) to preserve MSC chondrogenesis in a proinflammatory environment. MSCs were seeded in alginate:collagen hydrogels and cultured for 21 days in an ultrasound-assisted bioreactor (5.0 MHz, 2.5 Vpp; 4 applications/day) in the presence of IL1β and evaluated by qRT-PCR and immunofluorescence. The differential expression of markers associated with the NFκB pathway was assessed upon a single exposure of cLIUS and assayed by Western blotting, qRT-PCR, and immunofluorescence. Mitochondrial potential was evaluated by tetramethylrhodamine methyl ester (TMRM) assay. The chondroinductive potential of cLIUS was noted by the increased expression of SOX9 and COLII. cLIUS extended its chondroprotective effects by stabilizing the NFκB complex in the cytoplasm via engaging the IκBα feedback mechanism, thus preventing its nuclear translocation. cLIUS acted as a mitochondrial protective agent by restoring the mitochondrial potential and the mitochondrial mRNA expression in a proinflammatory environment. Altogether, our results demonstrated the potential of cLIUS for cartilage repair and regeneration under proinflammatory conditions.
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Affiliation(s)
| | | | - Anuradha Subramanian
- Department of Chemical and Materials Engineering, The University of Alabama in Huntsville, Huntsville, AL 35899, USA; (S.B.); (S.K.)
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Li M, Yin H, Yan Z, Li H, Wu J, Wang Y, Wei F, Tian G, Ning C, Li H, Gao C, Fu L, Jiang S, Chen M, Sui X, Liu S, Chen Z, Guo Q. The immune microenvironment in cartilage injury and repair. Acta Biomater 2022; 140:23-42. [PMID: 34896634 DOI: 10.1016/j.actbio.2021.12.006] [Citation(s) in RCA: 189] [Impact Index Per Article: 63.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/07/2021] [Revised: 12/01/2021] [Accepted: 12/05/2021] [Indexed: 02/07/2023]
Abstract
The ability of articular cartilage to repair itself is limited because it lacks blood vessels, nerves, and lymph tissue. Once damaged, it can lead to joint swelling and pain, accelerating the progression of osteoarthritis. To date, complete regeneration of hyaline cartilage exhibiting mechanical properties remains an elusive goal, despite the many available technologies. The inflammatory milieu created by cartilage damage is critical for chondrocyte death and hypertrophy, extracellular matrix breakdown, ectopic bone formation, and progression of cartilage injury to osteoarthritis. In the inflammatory microenvironment, mesenchymal stem cells (MSCs) undergo aberrant differentiation, and chondrocytes begin to convert or dedifferentiate into cells with a fibroblast phenotype, thereby resulting in fibrocartilage with poor mechanical qualities. All these factors suggest that inflammatory problems may be a major stumbling block to cartilage repair. To produce a milieu conducive to cartilage repair, multi-dimensional management of the joint inflammatory microenvironment in place and time is required. Therefore, this calls for elucidation of the immune microenvironment of cartilage repair after injury. This review provides a brief overview of: (1) the pathogenesis of cartilage injury; (2) immune cells in cartilage injury and repair; (3) effects of inflammatory cytokines on cartilage repair; (4) clinical strategies for treating cartilage defects; and (5) strategies for targeted immunoregulation in cartilage repair. STATEMENT OF SIGNIFICANCE: Immune response is increasingly considered the key factor affecting cartilage repair. It has both negative and positive regulatory effects on the process of regeneration and repair. Proinflammatory factors are secreted in large numbers, and necrotic cartilage is removed. During the repair period, immune cells can secrete anti-inflammatory factors and chondrogenic cytokines, which can inhibit inflammation and promote cartilage repair. However, inflammatory factors persist, which accelerate the degradation of the cartilage matrix. Furthermore, in an inflammatory microenvironment, MSCs undergo abnormal differentiation, and chondrocytes begin to transform or dedifferentiate into fibroblast-like cells, forming fibrocartilage with poor mechanical properties. Consequently, cartilage regeneration requires multi-dimensional regulation of the joint inflammatory microenvironment in space and time to make it conducive to cartilage regeneration.
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Liu Y, Shah KM, Luo J. Strategies for Articular Cartilage Repair and Regeneration. Front Bioeng Biotechnol 2022; 9:770655. [PMID: 34976967 PMCID: PMC8719005 DOI: 10.3389/fbioe.2021.770655] [Citation(s) in RCA: 66] [Impact Index Per Article: 22.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/04/2021] [Accepted: 12/01/2021] [Indexed: 12/19/2022] Open
Abstract
Articular cartilage is an avascular tissue, with limited ability to repair and self-renew. Defects in articular cartilage can induce debilitating degenerative joint diseases such as osteoarthritis. Currently, clinical treatments have limited ability to repair, for they often result in the formation of mechanically inferior cartilage. In this review, we discuss the factors that affect cartilage homeostasis and function, and describe the emerging regenerative approaches that are informing the future treatment options.
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Affiliation(s)
- Yanxi Liu
- Shanghai Key Laboratory of Regulatory Biology, Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai, China
| | - Karan M Shah
- Department of Oncology and Metabolism, The Medical School, The University of Sheffield, Sheffield, United Kingdom
| | - Jian Luo
- Shanghai Key Laboratory of Regulatory Biology, Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai, China.,Shanghai Yangzhi Rehabilitation Hospital (Shanghai Sunshine Rehabilitation Centre), Tongji University School of Medicine, Shanghai, China
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Wesdorp MA, Bastiaansen-Jenniskens YM, Capar S, Verhaar JA, Narcisi R, Van Osch GJ. Modulation of Inflamed Synovium Improves Migration of Mesenchymal Stromal Cells in Vitro Through Anti-Inflammatory Macrophages. Cartilage 2022; 13:19476035221085136. [PMID: 35306879 PMCID: PMC9137323 DOI: 10.1177/19476035221085136] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/09/2021] [Revised: 01/25/2022] [Accepted: 01/26/2022] [Indexed: 01/15/2023] Open
Abstract
OBJECTIVE Inflammation is known to negatively affect cartilage repair. However, it is unclear how inflammation influences the migration of mesenchymal stromal cells (MSCs) from the underlying bone marrow into the defect. We therefore aimed to investigate how synovial inflammation influences MSC migration, and whether modulation of inflammation with triamcinolone acetonide (TAA) may influence migration. DESIGN Inflamed human osteoarthritic synovium, M(IFNγ+TNFα) pro-inflammatory macrophages, M(IL4) repair macrophages, M(IL10) anti-inflammatory macrophages, or synovial fibroblasts were cultured with/without TAA. Conditioned medium (CM) was harvested after 24 hours, and the effect on MSC migration was studied using a Boyden chamber assay. Inflammation was evaluated with gene expression and flow cytometry analysis. RESULTS Synovium CM increased MSC migration. Modulation of synovial inflammation with TAA further increased migration 1.5-fold (P < 0.01). TAA significantly decreased TNFA, IL1B, and IL6 gene expression in synovium explants and increased CD163, a gene associated with anti-inflammatory macrophages. TAA treatment decreased the percentage of CD14+/CD80+ and CD14+/CD86+ pro-inflammatory macrophages and increased the percentage of CD14+/CD163+ anti-inflammatory macrophages in synovium explants. Interestingly, MSC migration was specifically enhanced by medium conditioned by M(IL4) macrophages and by M(IL10) macrophages treated with TAA, and unaffected by CM from M(IFNγ+TNFα) macrophages and synovial fibroblasts. CONCLUSION Macrophages secrete factors that stimulate the migration of MSCs. Modulation with TAA increased specifically the ability of anti-inflammatory macrophages to stimulate migration, indicating that they play an important role in secreting factors to attract MSCs. Modulating inflammation and thereby improving migration could be used in approaches based on endogenous repair of full-thickness cartilage defects.
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Affiliation(s)
- Marinus A. Wesdorp
- Department of Orthopaedics and Sports Medicine, Erasmus MC, Rotterdam, The Netherlands
| | | | - Serdar Capar
- Department of Orthopaedics and Sports Medicine, Erasmus MC, Rotterdam, The Netherlands
| | - Jan A.N. Verhaar
- Department of Orthopaedics and Sports Medicine, Erasmus MC, Rotterdam, The Netherlands
| | - R. Narcisi
- Department of Orthopaedics and Sports Medicine, Erasmus MC, Rotterdam, The Netherlands
| | - Gerjo J.V.M. Van Osch
- Department of Orthopaedics and Sports Medicine, Erasmus MC, Rotterdam, The Netherlands
- Department of Otorhinolaryngology, Erasmus MC, Rotterdam, The Netherlands
- Department of Biomechanical Engineering, Faculty of Mechanical, Maritime, and Materials Engineering, Delft University of Technology, Delft, The Netherlands
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Ekram S, Khalid S, Salim A, Khan I. Regulating the fate of stem cells for regenerating the intervertebral disc degeneration. World J Stem Cells 2021; 13:1881-1904. [PMID: 35069988 PMCID: PMC8727226 DOI: 10.4252/wjsc.v13.i12.1881] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/04/2021] [Revised: 07/12/2021] [Accepted: 11/15/2021] [Indexed: 02/06/2023] Open
Abstract
Lower back pain is a leading cause of disability and is one of the reasons for the substantial socioeconomic burden. The etiology of intervertebral disc (IVD) degeneration is complicated, and its mechanism is still not completely understood. Factors such as aging, systemic inflammation, biochemical mediators, toxic environmental factors, physical injuries, and genetic factors are involved in the progression of its pathophysiology. Currently, no therapy for restoring degenerated IVD is available except pain management, reduced physical activities, and surgical intervention. Therefore, it is imperative to establish regenerative medicine-based approaches to heal and repair the injured disc, repopulate the cell types to retain water content, synthesize extracellular matrix, and strengthen the disc to restore normal spine flexion. Cellular therapy has gained attention for IVD management as an alternative therapeutic option. In this review, we present an overview of the anatomical and molecular structure and the surrounding pathophysiology of the IVD. Modern therapeutic approaches, including proteins and growth factors, cellular and gene therapy, and cell fate regulators are reviewed. Similarly, small molecules that modulate the fate of stem cells for their differentiation into chondrocytes and notochordal cell types are highlighted.
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Affiliation(s)
- Sobia Ekram
- Dr. Panjwani Center for Molecular Medicine and Drug Research, International Center for Chemical and Biological Sciences, University of Karachi, Karachi 75270, Sindh, Pakistan
| | - Shumaila Khalid
- Dr. Panjwani Center for Molecular Medicine and Drug Research, International Center for Chemical and Biological Sciences, University of Karachi, Karachi 75270, Sindh, Pakistan
| | - Asmat Salim
- Dr. Panjwani Center for Molecular Medicine and Drug Research, International Center for Chemical and Biological Sciences, University of Karachi, Karachi 75270, Sindh, Pakistan
| | - Irfan Khan
- Dr. Panjwani Center for Molecular Medicine and Drug Research, International Center for Chemical and Biological Sciences, University of Karachi, Karachi 75270, Sindh, Pakistan.
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Li M, Li H, Ran X, Yin H, Luo X, Chen Z. Effects of adenovirus-mediated knockdown of IRAK4 on synovitis in the osteoarthritis rabbit model. Arthritis Res Ther 2021; 23:294. [PMID: 34863246 PMCID: PMC8643028 DOI: 10.1186/s13075-021-02684-8] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/17/2021] [Accepted: 11/20/2021] [Indexed: 01/15/2023] Open
Abstract
BACKGROUND The use of interleukin-1 receptor-associated kinase 4 (IRAK4) inhibitor as a treatment for the inflammatory joint disease is a promising method. However, its underlying mechanism in osteoarthritis (OA) remains unclear. The purpose of this study is to look into the effects of adenovirus-mediated knockdown of IRAK4 on synovitis in the OA rabbit model. METHODS Ad-shIRAK4 was injected two weeks after anterior cruciate ligament resection. Six weeks later, the rabbits were killed. The expression of IRAK4, TNFR-associated factor 6(TRAF6), TGF-activated kinase 1(TAK1), p-IKB kinase (p-IKK), p-nuclear factor kappa-B (p-NFκB), p38, and p-p38 in the synovial membrane was detected by western blot, qRT-PCR, and immunohistochemistry analysis. Immunohistochemistry was to detect the expression of IRAK4 proteins in articular cartilage. H&E staining was to assess the pathological changes of synovium and cartilage. The levels of interleukin (IL)-1β, tumor necrosis factor-α(TNF-α), and MMP-13 in the synovial fluid were measured by ELISA. X-ray and micro-computerized tomography (μCT) scans were used to assess knee joint conditions and microstructure of subchondral bone. RESULTS IRAK4 expression levels in synovial tissues of the OA model group exhibited a significant upward trend. Ad-shIRAK4 significantly reduced IRAK4 mRNA expression in synovium tissues. Notably, Ad-shIRAK4 suppressed the Toll-like receptor/interleukin-1 receptor (TLR/IL-1R) signaling. In addition, in the Ad-shIRAK4 treatment group, we can see less inflammatory cell infiltration and reduced hyperplasia and angiogenesis. The levels of IL-1β, TNF-α, and MMP-13 in the synovial fluid in the OA model group were significantly higher than that in the control group, which were reduced by Ad-shIRAK4 treatment. Finally, Results of HE stains, immunohistochemistry, and μCT showed that Ad-shIRAK4 treatment has a protective effect on cartilage damage. CONCLUSIONS IRAK4 is significantly upregulated in the synovium from the osteoarthritis rabbit model. In addition, Ad-shIRAK4 reduced the expression of IRAK4 and suppressed TLR/IL-1R signaling in the synovium from the osteoarthritis rabbit model. Ad-shIRAK4 could alleviate synovitis and cartilage degradation in the osteoarthritis rabbit model, and thus alleviate the symptoms of OA and prevent the progression of OA.
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Affiliation(s)
- Muzhe Li
- Department of Orthopedic, The First Affiliated Hospital, Hengyang Medical School, University of South China, No. 69, Chuanshan Road, Hengyang City, 421001, Hunan Province, China
| | - Huiyun Li
- Department of Orthopedic, The First Affiliated Hospital, Hengyang Medical School, University of South China, No. 69, Chuanshan Road, Hengyang City, 421001, Hunan Province, China
| | - Xun Ran
- Department of Orthopedic, The First Affiliated Hospital, Hengyang Medical School, University of South China, No. 69, Chuanshan Road, Hengyang City, 421001, Hunan Province, China
| | - Han Yin
- Institute of Orthopedics, The First Medical Center, Chinese PLA General Hospital, Beijing Key Lab of Regenerative Medicine in Orthopedics, Key Laboratory of Musculoskeletal Trauma and War Injuries PLA, No. 28 Fuxing Road, Haidian District, Beijing, 100853, China
| | - Xuling Luo
- Department of Orthopedic, The First Affiliated Hospital, Hengyang Medical School, University of South China, No. 69, Chuanshan Road, Hengyang City, 421001, Hunan Province, China.
| | - Zhiwei Chen
- Department of Orthopedic, The First Affiliated Hospital, Hengyang Medical School, University of South China, No. 69, Chuanshan Road, Hengyang City, 421001, Hunan Province, China.
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Abraham DM, Herman C, Witek L, Cronstein BN, Flores RL, Coelho PG. Self-assembling human skeletal organoids for disease modeling and drug testing. J Biomed Mater Res B Appl Biomater 2021; 110:871-884. [PMID: 34837719 DOI: 10.1002/jbm.b.34968] [Citation(s) in RCA: 17] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/20/2021] [Revised: 11/02/2021] [Accepted: 11/13/2021] [Indexed: 01/09/2023]
Abstract
Skeletal conditions represent a considerable challenge to health systems globally. Barriers to effective therapeutic development include a lack of accurate preclinical tissue and disease models. Most recently, work was attempted to present a novel whole organ approach to modeling human bone and cartilage tissues. These self-assembling skeletal organoids mimic the cellular milieu and extracellular organization present in native tissues. Bone organoids demonstrated osteogenesis and micro vessel formation, and cartilage organoids showed evidence of cartilage development and maturation. Skeletal organoids derived from both bone and cartilage tissues yielded spontaneous polarization of their cartilaginous and bone components. Using these hybrid skeletal organoids, we successfully generated "mini joint" cultures, which we used to model inflammatory disease and test Adenosine (A2A ) receptor agonists as a therapeutic agent. The work and respective results indicated that skeletal organoids can be an effective biological model for tissue development and disease as well as to test therapeutic agents.
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Affiliation(s)
- Diana M Abraham
- Department of Biomaterials, New York University College of Dentistry, New York, New York, USA
| | - Calvin Herman
- Department of Biomaterials, New York University College of Dentistry, New York, New York, USA
| | - Lukasz Witek
- Department of Biomaterials, New York University College of Dentistry, New York, New York, USA.,Department of Biomedical Engineering, New York University Tandon School of Engineering, Brooklyn, New York, USA
| | - Bruce N Cronstein
- Department of Medicine, New York University Grossman School of Medicine, New York, New York, USA
| | - Roberto L Flores
- Hansjörg Wyss Department of Plastic Surgery, New York University Grossman School of Medicine, New York, New York, USA
| | - Paulo G Coelho
- Department of Biomaterials, New York University College of Dentistry, New York, New York, USA.,Hansjörg Wyss Department of Plastic Surgery, New York University Grossman School of Medicine, New York, New York, USA.,Department of Mechanical and Aerospace Engineering, New York University Tandon School of Engineering, Brooklyn, New York, USA
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Genç D, Sezer Kürkçü M, Yiğittürk G, Günaydın B, Elbe H, Aladağ A, Çöl B, Tarhan EF. Synovial fluid niche promoted differentiation of dental follicle mesenchymal stem cells toward chondrogenesis in rheumatoid arthritis. Arch Rheumatol 2021; 37:94-109. [PMID: 35949879 PMCID: PMC9326389 DOI: 10.46497/archrheumatol.2022.8891] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/03/2021] [Accepted: 05/31/2021] [Indexed: 11/30/2022] Open
Abstract
Objectives
In this study, we aimed to investigate the differentiation potential of dental follicle mesenchymal stem cells (MSCs) in the synovial fluid (SF) niche of early-onset or end-stage rheumatoid arthritis (RA). Patients and methods
Between May 2020 and January 2021, six patients (1 male, 5 females; mean age: 57.5±11.2 years; range, 49 to 65 years) who were diagnosed with RA with the indication of SF aspiration were included in the study. The third passage dental follicle stem cells (DFSCs) were cocultured with fresh SF samples of end-stage or early-onset RA patients in micromass culture system for 21 days. SF samples were analyzed for secreted cytokines. Chondrogenic markers (CD49e, CD49f) were analyzed in DFSCs, gene expression analysis was performed for the expressions of Col I, Col II, Aggrecan and Sox-9, and histochemical analysis was performed by staining three-dimensional pellets with anti-collagen II antibody. The neutralization assay was performed with anti-interleukin (IL)-6, anti-interferon-gamma (IFN-g), and anti-IL-1beta(b). Results
The high levels of IL-1b and IL-6 were observed in end-stage RA patients’ SF samples compared to the early-onset patients (p<0.05). The CD49e and CD49f expressions in DFSCs were significantly higher in the SF samples of end-stage RA patients (p<0.05). Also, the Col II, Sox-9 and Aggrecan messenger ribonucleic acid (mRNA) expressions increased in the DFSCs, when cultured with end-stage RA patients’ SF samples (p<0.01). Collagen-II expression in histochemical analysis of micromass pellets was higher in the DFSCs cultured with end-stage RA patients’ SF samples. The neutralization of IL-6 significantly decreased the CD49e and CD49f expressions (p<0.05). Conclusion
The high levels of IL-6 in SF niche of end-stage RA patients were found to differentiate DFSCs toward chondrogenesis. Based on these findings, DFSCs can be used as a new cell-based treatment in RA patients for the cartilage damage.
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Affiliation(s)
- Deniz Genç
- Department of Pediatric Health and Diseases Nursing, Muğla Sıtkı Koçman University, Faculty of Health Sciences, Muğla, Turkey
| | - Merve Sezer Kürkçü
- Muğla Sıtkı Koçman University, Research Laboratories Center, Muğla, Turkey
| | - Gürkan Yiğittürk
- Department of Histology and Embryology, Muğla Sıtkı Koçman University, Faculty of Medicine, Muğla, Turkey
| | - Burcu Günaydın
- Department of Histology and Embryology, Muğla Sıtkı Koçman University, Institute of Health Sciences, Muğla, Turkey
| | - Hülya Elbe
- Department of Histology and Embryology, Muğla Sıtkı Koçman University, Faculty of Medicine, Muğla, Turkey
| | - Akın Aladağ
- Muğla Sıtkı Koçman University, Faculty of Dentistry, Muğla, Turkey
| | - Bekir Çöl
- Department of Biology, Muğla Sıtkı Koçman University, Faculty of Science, Muğla, Turkey
| | - Emine Figen Tarhan
- Department of Rheumatology, Muğla Sıtkı Koçman University, Faculty of Medicine, Muğla, Turkey
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Lu KH, Lu PWA, Lu EWH, Tang CH, Su SC, Lin CW, Yang SF. The potential remedy of melatonin on osteoarthritis. J Pineal Res 2021; 71:e12762. [PMID: 34435392 DOI: 10.1111/jpi.12762] [Citation(s) in RCA: 18] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/28/2021] [Revised: 08/16/2021] [Accepted: 08/21/2021] [Indexed: 01/15/2023]
Abstract
Osteoarthritis (OA), the most common arthritis worldwide, is a degenerative joint disease characterized by progressive cartilage breakdown, subchondral remodeling, and synovial inflammation. Although conventional pharmaceutical therapies aimed to prevent further cartilage loss and joint dysfunction, there are no ideal strategies that target the pathogenesis of OA. Melatonin exhibits a variety of regulatory properties by binding to specific receptors and downstream molecules and exerts a myriad of receptor-independent actions via intracellular targets as a chondrocyte protector, an anti-inflammation modulator, and a free radical scavenger. Melatonin also modulates cartilage regeneration and degradation by directly/indirectly regulating the expression of main circadian clock genes, such as transcriptional activators [brain and muscle aryl hydrocarbon receptor nuclear translocator-like protein (Bmal) and circadian locomotor output cycles kaput (Clock)], transcriptional repressors [period circadian regulator (Per)1/2, cryptochrome (Cry)1/2, and Dec2], and nuclear hormone receptors [Rev-Erbs and retinoid acid-related orphan receptors (Rors)]. Owing to its effects on cartilage homeostasis, we propose a potential role for melatonin in the prevention and therapy of OA via the modulation of circadian clock genes, mitigation of chondrocyte apoptosis, anti-inflammatory activity, and scavenging of free radicals.
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Affiliation(s)
- Ko-Hsiu Lu
- Department of Orthopedics, Chung Shan Medical University Hospital, Taichung, Taiwan
- School of Medicine, Chung Shan Medical University, Taichung, Taiwan
| | | | | | - Chih-Hsin Tang
- School of Medicine, China Medical University, Taichung, Taiwan
- Department of Medical Laboratory Science and Biotechnology, College of Medical and Health Science, Asia University, Taichung, Taiwan
| | - Shih-Chi Su
- Whole-Genome Research Core Laboratory of Human Diseases, Chang Gung Memorial Hospital, Keelung, Taiwan
- Department of Dermatology, Drug Hypersensitivity Clinical and Research Center, Chang Gung Memorial Hospital, Taipei, Linkou and Keelung, Taiwan
| | - Chiao-Wen Lin
- Institute of Oral Sciences, Chung Shan Medical University, Taichung, Taiwan
- Department of Dentistry, Chung Shan Medical University Hospital 402, Taichung, Taiwan
| | - Shun-Fa Yang
- Institute of Medicine, Chung Shan Medical University, Taichung, Taiwan
- Department of Medical Research, Chung Shan Medical University Hospital, Taichung, Taiwan
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Bi R, Chen K, Wang Y, Luo X, Li Q, Li P, Yin Q, Fan Y, Zhu S. Regulating Fibrocartilage Stem Cells via TNF-α/Nf-κB in TMJ Osteoarthritis. J Dent Res 2021; 101:312-322. [PMID: 34515572 DOI: 10.1177/00220345211037248] [Citation(s) in RCA: 21] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/05/2023] Open
Abstract
In this study, we investigate harnessing fibrocartilage stem cell (FCSC) capacities by regulating tumor necrosis factor α (TNF-α) signaling for cartilage repair in temporomandibular joint osteoarthritis (TMJOA). Stem cell specifics for FCSCs were characterized in the presence of TNF-α. Etanercept as a TNF-α inhibitor and BAY 11-7082 as an Nf-κB inhibitor were used to study TNF-α regulation of FCSCs. Lineage tracing was performed in Gli1-CreERT+;Tmfl/fl mice when etanercept (1 mg/kg, every 3 d) or isometric vehicle was subcutaneously injected to trace specific changes in FCSCs. Surgically induced TMJOA Sprague-Dawley rats were generated with BAY 11-7082 (5 mg/kg, every 3 d) or vehicle subcutaneous injection to investigate the functional role of TNF-α/Nf-κB in TMJOA. Anterior disc displacement (ADD) rabbits were used to analyze the therapeutic effect of etanercept as a TMJOA intra-articular treatment with etanercept (0.02 mg in 100 μL, every 2 wk) or isometric vehicle. In vitro, TNF-α inhibited proliferation of FCSCs and increased FCSC apoptosis. TNF-α activation interfered with osteogenic and chondrogenic differentiation of FCSCs, while etanercept could partially recover FCSC specificity from TNF-α. FCSC lineage tracing in Gli1-CreERT+;Tmfl/fl mice showed that the chondrogenic capacity of Gli1+ cell lineage was markedly suppressed in osteoarthritis cartilage, the phenotype of which could be significantly rescued by etanercept. Specifically blocking the Nf-κB pathway could significantly weaken the regulatory effect of TNF-α on FCSC specificity in vitro and in TMJOA rats in vivo. Finally, intra-articular etanercept treatment efficiently rescued TMJ cartilage degeneration and growth retardation in ADD rabbits. Inhibition of TNF-α signaling reduced Nf-κB transcripts and recovered FCSC specificities. In vivo, etanercept treatment effectively rescued the osteoarthritis phenotype in TMJOA mice and ADD rabbits. These data suggest a novel therapeutic mechanism whereby TNF-α/Nf-κB inhibition promotes FCSC chondrogenic capacity for cartilage transformation in TMJOA.
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Affiliation(s)
- R Bi
- State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, Department of Orthognathic and TMJ Surgery, West China Hospital of Stomatology, Sichuan University, Chengdu, China
| | - K Chen
- State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, Department of Orthognathic and TMJ Surgery, West China Hospital of Stomatology, Sichuan University, Chengdu, China
| | - Y Wang
- State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, Department of Orthognathic and TMJ Surgery, West China Hospital of Stomatology, Sichuan University, Chengdu, China
| | - X Luo
- State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, Department of Orthognathic and TMJ Surgery, West China Hospital of Stomatology, Sichuan University, Chengdu, China
| | - Q Li
- State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, Department of Orthognathic and TMJ Surgery, West China Hospital of Stomatology, Sichuan University, Chengdu, China
| | - P Li
- State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, Department of Orthognathic and TMJ Surgery, West China Hospital of Stomatology, Sichuan University, Chengdu, China
| | - Q Yin
- State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, Department of Orthognathic and TMJ Surgery, West China Hospital of Stomatology, Sichuan University, Chengdu, China
| | - Y Fan
- State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, Department of Operative Dentistry and Endodontics, West China Hospital of Stomatology, Sichuan University, Chengdu, China
| | - S Zhu
- State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, Department of Orthognathic and TMJ Surgery, West China Hospital of Stomatology, Sichuan University, Chengdu, China
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Abstract
» Orthopaedics pioneered the expansion of gene therapy beyond its traditional scope of diseases that are caused by rare single-gene defects. Orthopaedic applications of gene therapy are most developed in the areas of arthritis and regenerative medicine, but several additional possibilities exist. » Invossa, an ex vivo gene therapeutic for osteoarthritis, was approved in South Korea in 2017, but its approval was retracted in 2019 and remains under appeal; a Phase-III clinical trial of Invossa has restarted in the U.S. » There are several additional clinical trials for osteoarthritis and rheumatoid arthritis that could lead to approved gene therapeutics for arthritis. » Bone-healing and cartilage repair are additional areas that are attracting considerable research; intervertebral disc degeneration and the healing of ligaments, tendons, and menisci are other applications of interest. Orthopaedic tumors, genetic diseases, and aseptic loosening are additional potential targets. » If successful, these endeavors will expand the scope of gene therapy from providing expensive medicines for a few patients to providing affordable medicines for many.
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Neefjes M, Housmans BAC, van Beuningen HM, Vitters EL, van den Akker GGH, Welting TJM, van Caam APM, van der Kraan PM. Prediction of the Effect of the Osteoarthritic Joint Microenvironment on Cartilage Repair. Tissue Eng Part A 2021; 28:27-37. [PMID: 34039008 DOI: 10.1089/ten.tea.2021.0051] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/14/2023] Open
Abstract
Osteoarthritis (OA) is characterized by progressive articular cartilage loss. Human mesenchymal stromal cells (MSCs) can be used for cartilage repair therapies based on their potential to differentiate into chondrocytes. However, the joint microenvironment is a major determinant of the success of MSC-based cartilage formation. Currently, there is no tool that is able to predict the effect of a patient's OA joint microenvironment on MSC-based cartilage formation. Our goal was to develop a molecular tool that can predict this effect before the start of cartilage repair therapies. Six different promoter reporters (hIL6, hIL8, hADAMTS5, hWISP1, hMMP13, and hADAM28) were generated and evaluated in an immortalized human articular chondrocyte for their responsiveness to an osteoarthritic microenvironment by stimulation with OA synovium-conditioned medium (OAs-cm) obtained from 32 different knee OA patients. To study the effect of this OA microenvironment on MSC-based cartilage formation, MSCs were cultured in a three-dimensional pellet culture model, while stimulated with OAs-cm. Cartilage formation was assessed histologically and by quantifying sulfated glycosaminoglycan (sGAG) production. We confirmed that OAs-cm of different patients had significantly different effects on sGAG production. In addition, significant correlations were obtained between the effect of the OAs-cm on cartilage formation and promoter reporter outcome. Furthermore, we validated the predictive value of measuring two promoter reporters with an independent cohort of OAs-cm and the effect of 87.5% of the OAs-cm on MSC-based cartilage formation could be predicted. Together, we developed a novel tool to predict the effect of the OA joint microenvironment on MSC-based cartilage formation. This is an important first step toward personalized cartilage repair strategies for OA patients.
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Affiliation(s)
- Margot Neefjes
- Experimental Rheumatology, Department of Rheumatology, Radboud University Medical Center, Nijmegen, the Netherlands
| | - Bas A C Housmans
- Laboratory for Experimental Orthopedics, Department of Orthopedic Surgery, Maastricht University Medical Center, Maastricht, the Netherlands
| | - Henk M van Beuningen
- Experimental Rheumatology, Department of Rheumatology, Radboud University Medical Center, Nijmegen, the Netherlands
| | - Elly L Vitters
- Experimental Rheumatology, Department of Rheumatology, Radboud University Medical Center, Nijmegen, the Netherlands
| | - Guus G H van den Akker
- Laboratory for Experimental Orthopedics, Department of Orthopedic Surgery, Maastricht University Medical Center, Maastricht, the Netherlands
| | - Tim J M Welting
- Laboratory for Experimental Orthopedics, Department of Orthopedic Surgery, Maastricht University Medical Center, Maastricht, the Netherlands
| | - Arjan P M van Caam
- Experimental Rheumatology, Department of Rheumatology, Radboud University Medical Center, Nijmegen, the Netherlands
| | - Peter M van der Kraan
- Experimental Rheumatology, Department of Rheumatology, Radboud University Medical Center, Nijmegen, the Netherlands
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41
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Schneider C, Dungel P, Priglinger E, Danzer M, Schädl B, Nürnberger S. The impact of photobiomodulation on the chondrogenic potential of adipose-derived stromal/stem cells. JOURNAL OF PHOTOCHEMISTRY AND PHOTOBIOLOGY B-BIOLOGY 2021; 221:112243. [PMID: 34217028 DOI: 10.1016/j.jphotobiol.2021.112243] [Citation(s) in RCA: 15] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 12/30/2020] [Revised: 04/23/2021] [Accepted: 06/16/2021] [Indexed: 01/11/2023]
Abstract
Due to their capacity to differentiate into the chondrogenic lineage, adipose-derived stromal/stem cells (ASC) are a promising source of therapeutically relevant cells for cartilage tissue regeneration. Their differentiation potential, however, varies between patients. In our study, we aim to stimulate ASC towards a more reliable chondrogenic phenotype using photobiomodulation (PBM). LED devices of either blue (475 nm), green (516 nm) or red (635 nm) light were used to treat human ASC from donors of varying chondrogenic potential. The treatment was applied either once during the 2D expansion phase or repeatedly during the 3D differentiation phase. Chondrogenic differentiation was assessed via pellet size, GAG/DNA content, histology and gene expression analysis. Reactions to PBM were found to be wavelength-dependent and more pronounced when the treatment was applied during expansion. Donors were assigned to responder categories according to their response to the treatment during expansion, whereby good responders were mainly donors with low intrinsic chondrogenic potential. Exposed to light, they revealed a particularly high relative increase in pellet size (more than twice the size of untreated controls after red light PBM), intense collagen type II immunostaining (low/absent in untreated controls) and activation of otherwise absent COL2A1 expression. Conversely, on a donor with high intrinsic chondrogenic potential, light had adverse effects. When applied with shorter wavelengths (blue, green), it led to reduced pellet size, GAG/DNA content and collagen type II immunostaining. However, when PBM was applied in 3D, the same donor was the only one to react with increased differentiation to all three wavelengths. We were able to demonstrate that PBM can be used to enhance or hamper chondrogenesis of ASC, and that success depends on treatment parameters and intrinsic cellular potential. The improvement of chondrogenesis in donors with low intrinsic potential highlights PBM as potent tool for cell-based cartilage regeneration. Its cost-effectiveness and ease of use make for an attractive treatment option to enhance the performance of ASC in cartilage tissue engineering.
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Affiliation(s)
- C Schneider
- Ludwig Boltzmann Institute for Experimental and Clinical Traumatology, AUVA Trauma Research Center, Vienna, Austria; Austrian Cluster for Tissue Regeneration, Vienna, Austria
| | - P Dungel
- Ludwig Boltzmann Institute for Experimental and Clinical Traumatology, AUVA Trauma Research Center, Vienna, Austria; Austrian Cluster for Tissue Regeneration, Vienna, Austria.
| | - E Priglinger
- Ludwig Boltzmann Institute for Experimental and Clinical Traumatology, AUVA Trauma Research Center, Vienna, Austria; Austrian Cluster for Tissue Regeneration, Vienna, Austria
| | - M Danzer
- Austrian Red Cross Blood Transfusion Service of Upper Austria, Linz, Austria
| | - B Schädl
- Ludwig Boltzmann Institute for Experimental and Clinical Traumatology, AUVA Trauma Research Center, Vienna, Austria; Austrian Cluster for Tissue Regeneration, Vienna, Austria; University Clinic of Dentistry, Medical University of Vienna, Vienna, Austria
| | - S Nürnberger
- Ludwig Boltzmann Institute for Experimental and Clinical Traumatology, AUVA Trauma Research Center, Vienna, Austria; Austrian Cluster for Tissue Regeneration, Vienna, Austria; Department of Orthopedics and Trauma-Surgery, Division of Trauma-Surgery, Medical University of Vienna, Vienna, Austria
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Hu H, Liu W, Sun C, Wang Q, Yang W, Zhang Z, Xia Z, Shao Z, Wang B. Endogenous Repair and Regeneration of Injured Articular Cartilage: A Challenging but Promising Therapeutic Strategy. Aging Dis 2021; 12:886-901. [PMID: 34094649 PMCID: PMC8139200 DOI: 10.14336/ad.2020.0902] [Citation(s) in RCA: 21] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/31/2020] [Accepted: 09/02/2020] [Indexed: 12/12/2022] Open
Abstract
Articular cartilage (AC) has a very limited intrinsic repair capacity after injury or disease. Although exogenous cell-based regenerative approaches have obtained acceptable outcomes, they are usually associated with complicated procedures, donor-site morbidities and cell differentiation during ex vivo expansion. In recent years, endogenous regenerative strategy by recruiting resident mesenchymal stem/progenitor cells (MSPCs) into the injured sites, as a promising alternative, has gained considerable attention. It takes full advantage of body's own regenerative potential to repair and regenerate injured tissue while avoiding exogenous regenerative approach-associated limitations. Like most tissues, there are also multiple stem-cell niches in AC and its surrounding tissues. These MSPCs have the potential to migrate into injured sites to produce replacement cells under appropriate stimuli. Traditional microfracture procedure employs the concept of MSPCs recruitment usually fails to regenerate normal hyaline cartilage. The reasons for this failure might be attributed to an inadequate number of recruiting cells and adverse local tissue microenvironment after cartilage injury. A strategy that effectively improves local matrix microenvironment and recruits resident MSPCs may enhance the success of endogenous AC regeneration (EACR). In this review, we focused on the reasons why AC cannot regenerate itself in spite of potential self-repair capacity and summarized the latest developments of the three key components in the field of EACR. In addition, we discussed the challenges facing in the present EACR strategy. This review will provide an increasing understanding of EACR and attract more researchers to participate in this promising research arena.
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Affiliation(s)
- Hongzhi Hu
- 1Department of Orthopaedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China
| | - Weijian Liu
- 1Department of Orthopaedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China
| | - Caixia Sun
- 2Department of Gynecology, General Hospital of the Yangtze River Shipping, Wuhan 430022, China
| | - Qiuyuan Wang
- 3Department of Nephrology, Xiangyang Central Hospital, Affiliated Hospital of Hubei University of Arts and Science, Xiangyang 441100, China
| | - Wenbo Yang
- 1Department of Orthopaedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China
| | - ZhiCai Zhang
- 1Department of Orthopaedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China
| | - Zhidao Xia
- 4Centre for Nanohealth, ILS2, Swansea university Medical school, Swansea, SA2 8PP, UK
| | - Zengwu Shao
- 1Department of Orthopaedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China
| | - Baichuan Wang
- 1Department of Orthopaedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China.,4Centre for Nanohealth, ILS2, Swansea university Medical school, Swansea, SA2 8PP, UK
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43
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Hou M, Zhang Y, Zhou X, Liu T, Yang H, Chen X, He F, Zhu X. Kartogenin prevents cartilage degradation and alleviates osteoarthritis progression in mice via the miR-146a/NRF2 axis. Cell Death Dis 2021; 12:483. [PMID: 33986262 PMCID: PMC8119954 DOI: 10.1038/s41419-021-03765-x] [Citation(s) in RCA: 41] [Impact Index Per Article: 10.3] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/28/2020] [Revised: 03/29/2021] [Accepted: 03/30/2021] [Indexed: 02/06/2023]
Abstract
Osteoarthritis (OA) is a common articular degenerative disease characterized by loss of cartilage matrix and subchondral bone sclerosis. Kartogenin (KGN) has been reported to improve chondrogenic differentiation of mesenchymal stem cells. However, the therapeutic effect of KGN on OA-induced cartilage degeneration was still unclear. This study aimed to explore the protective effects and underlying mechanisms of KGN on articular cartilage degradation using mice with post-traumatic OA. To mimic the in vivo arthritic environment, in vitro cultured chondrocytes were exposed to interleukin-1β (IL-1β). We found that KGN barely affected the cell proliferation of chondrocytes; however, KGN significantly enhanced the synthesis of cartilage matrix components such as type II collagen and aggrecan in a dose-dependent manner. Meanwhile, KGN markedly suppressed the expression of matrix degradation enzymes such as MMP13 and ADAMTS5. In vivo experiments showed that intra-articular administration of KGN ameliorated cartilage degeneration and inhibited subchondral bone sclerosis in an experimental OA mouse model. Molecular biology experiments revealed that KGN modulated intracellular reactive oxygen species in IL-1β-stimulated chondrocytes by up-regulating nuclear factor erythroid 2-related factor 2 (NRF2), while barely affecting its mRNA expression. Microarray analysis further revealed that IL-1β significantly up-regulated miR-146a that played a critical role in regulating the protein levels of NRF2. KGN treatment showed a strong inhibitory effect on the expression of miR-146a in IL-1β-stimulated chondrocytes. Over-expression of miR-146a abolished the anti-arthritic effects of KGN not only by down-regulating the protein levels of NRF2 but also by up-regulating the expression of matrix degradation enzymes. Our findings demonstrate, for the first time, that KGN exerts anti-arthritic effects via activation of the miR-146a-NRF2 axis and KGN is a promising heterocyclic molecule to prevent OA-induced cartilage degeneration.
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Affiliation(s)
- Mingzhuang Hou
- Department of Orthopaedics, The First Affiliated Hospital of Soochow University, Soochow University, Suzhou, China.,Orthopaedic Institute, Medical College, Soochow University, Suzhou, China
| | - Yijian Zhang
- Department of Orthopaedics, The First Affiliated Hospital of Soochow University, Soochow University, Suzhou, China.,Orthopaedic Institute, Medical College, Soochow University, Suzhou, China
| | - Xinfeng Zhou
- Department of Orthopaedics, The First Affiliated Hospital of Soochow University, Soochow University, Suzhou, China.,Orthopaedic Institute, Medical College, Soochow University, Suzhou, China
| | - Tao Liu
- Department of Orthopaedics, The First Affiliated Hospital of Soochow University, Soochow University, Suzhou, China
| | - Huilin Yang
- Department of Orthopaedics, The First Affiliated Hospital of Soochow University, Soochow University, Suzhou, China.,Orthopaedic Institute, Medical College, Soochow University, Suzhou, China
| | - Xi Chen
- Department of Pathology, The Third Affiliated Hospital of Soochow University, Changzhou, China.
| | - Fan He
- Department of Orthopaedics, The First Affiliated Hospital of Soochow University, Soochow University, Suzhou, China. .,Orthopaedic Institute, Medical College, Soochow University, Suzhou, China.
| | - Xuesong Zhu
- Department of Orthopaedics, The First Affiliated Hospital of Soochow University, Soochow University, Suzhou, China. .,Orthopaedic Institute, Medical College, Soochow University, Suzhou, China.
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van Eegher S, Perez-Lozano ML, Toillon I, Valour D, Pigenet A, Citadelle D, Bourrier C, Courtade-Gaïani S, Grégoire L, Cléret D, Malbos S, Nourissat G, Sautet A, Lafage-Proust MH, Pastoureau P, Rolland-Valognes G, De Ceuninck F, Berenbaum F, Houard X. The differentiation of prehypertrophic into hypertrophic chondrocytes drives an OA-remodeling program and IL-34 expression. Osteoarthritis Cartilage 2021; 29:257-268. [PMID: 33301945 DOI: 10.1016/j.joca.2020.10.013] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/11/2019] [Revised: 09/16/2020] [Accepted: 10/08/2020] [Indexed: 02/02/2023]
Abstract
OBJECTIVES We hypothesize that chondrocytes from the deepest articular cartilage layer are pivotal in maintaining cartilage integrity and that the modification of their prehypertrophic phenotype to a hypertrophic phenotype will drive cartilage degradation in osteoarthritis. DESIGN Murine immature articular chondrocytes (iMACs) were successively cultured into three different culture media to induce a progressive hypertrophic differentiation. Chondrocyte were phenotypically characterized by whole-genome microarray analysis. The expression of IL-34 and its receptors PTPRZ1 and CSF1R in chondrocytes and in human osteoarthritis tissues was assessed by RT-qPCR, ELISA and immunohistochemistry. The expression of bone remodeling and angiogenesis factors and the cell response to IL-1β and IL-34 were investigated by RT-qPCR and ELISA. RESULTS Whole-genome microarray analysis showed that iMACs, prehypertrophic and hypertrophic chondrocytes each displayed a specific phenotype. IL-1β induced a stronger catabolic effect in prehypertrophic chondrocytes than in iMACs. Hypertrophic differentiation of prehypertrophic chondrocytes increased Bmp-2 (95%CI [0.78; 1.98]), Bmp-4 (95%CI [0.89; 1.59]), Cxcl12 (95%CI [2.19; 5.41]), CCL2 (95%CI [3.59; 11.86]), Mmp 3 (95%CI [10.29; 32.14]) and Vegf mRNA expression (95%CI [0.20; 1.74]). Microarray analysis identified IL-34, PTPRZ1 and CSFR1 as being strongly overexpressed in hypertrophic chondrocytes. IL-34 was released by human osteoarthritis cartilage; its receptors were expressed in human osteoarthritis tissues. IL-34 stimulated CCL2 and MMP13 in osteoblasts and hypertrophic chondrocytes but not in iMACs or prehypertrophic chondrocytes. CONCLUSION Our results identify prehypertrophic chondrocytes as being potentially pivotal in the control of cartilage and subchondral bone integrity. Their differentiation into hypertrophic chondrocytes initiates a remodeling program in which IL-34 may be involved.
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Affiliation(s)
- S van Eegher
- Sorbonne Université, INSERM, Centre de Recherche Saint-Antoine (CRSA), F-75012, Paris, France
| | - M-L Perez-Lozano
- Sorbonne Université, INSERM, Centre de Recherche Saint-Antoine (CRSA), F-75012, Paris, France
| | - I Toillon
- Sorbonne Université, INSERM, Centre de Recherche Saint-Antoine (CRSA), F-75012, Paris, France
| | - D Valour
- Servier Research Institute, F-78290, Croissy-sur-Seine, France
| | - A Pigenet
- Sorbonne Université, INSERM, Centre de Recherche Saint-Antoine (CRSA), F-75012, Paris, France
| | - D Citadelle
- Sorbonne Université, INSERM, Centre de Recherche Saint-Antoine (CRSA), F-75012, Paris, France
| | - C Bourrier
- Servier Research Institute, F-78290, Croissy-sur-Seine, France
| | | | - L Grégoire
- Soladis, 94 Rue Saint-Lazare, F-75009, Paris, France
| | - D Cléret
- Université de Lyon - Université Jean Monnet, INSERM U1059, Faculté de Médecine, F-42270, Saint-Priest en Jarez, France
| | - S Malbos
- Sorbonne Université, INSERM, Centre de Recherche Saint-Antoine (CRSA), F-75012, Paris, France
| | - G Nourissat
- Sorbonne Université, INSERM, Centre de Recherche Saint-Antoine (CRSA), F-75012, Paris, France; Clinique Maussins-Nollet, Ramsay Générale de Santé, F-75019, Paris, France
| | - A Sautet
- Department of Orthopaedic Surgery and Traumatology, APHP Saint-Antoine Hospital, F-75012, Paris, France
| | - M-H Lafage-Proust
- Université de Lyon - Université Jean Monnet, INSERM U1059, Faculté de Médecine, F-42270, Saint-Priest en Jarez, France
| | - P Pastoureau
- Servier Research Institute, F-78290, Croissy-sur-Seine, France
| | | | - F De Ceuninck
- Servier Research Institute, F-78290, Croissy-sur-Seine, France
| | - F Berenbaum
- Sorbonne Université, INSERM, Centre de Recherche Saint-Antoine (CRSA), F-75012, Paris, France; Sorbonne Université, INSERM CRSA, AP-HP Hopital Saint Antoine, Paris.
| | - X Houard
- Sorbonne Université, INSERM, Centre de Recherche Saint-Antoine (CRSA), F-75012, Paris, France
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Sun AR, Udduttula A, Li J, Liu Y, Ren PG, Zhang P. Cartilage tissue engineering for obesity-induced osteoarthritis: Physiology, challenges, and future prospects. J Orthop Translat 2021; 26:3-15. [PMID: 33437618 PMCID: PMC7773977 DOI: 10.1016/j.jot.2020.07.004] [Citation(s) in RCA: 29] [Impact Index Per Article: 7.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/10/2020] [Revised: 06/25/2020] [Accepted: 07/21/2020] [Indexed: 12/14/2022] Open
Abstract
UNLABELLED Osteoarthritis (OA) is a multifactorial joint disease with pathological changes that affect whole joint tissue. Obesity is acknowledged as the most influential risk factor for both the initiation and progression of OA in weight-bearing and non-weight-bearing joints. Obesity-induced OA is a newly defined phenotypic group in which chronic low-grade inflammation has a central role. Aside from persistent chronic inflammation, abnormal mechanical loading due to increased body weight on weight-bearing joints is accountable for the initiation and progression of obesity-induced OA. The current therapeutic approaches for OA are still evolving. Tissue-engineering-based strategy for cartilage regeneration is one of the most promising treatment breakthroughs in recent years. However, patients with obesity-induced OA are often excluded from cartilage repair attempts due to the abnormal mechanical demands, altered biomechanical and biochemical activities of cells, persistent chronic inflammation, and other obesity-associated factors. With the alarming increase in the number of obese populations globally, the need for an innovative therapeutic approach that could effectively repair and restore the damaged synovial joints is of significant importance for this sub-population of patients. In this review, we discuss the involvement of the systemic and localized inflammatory response in obesity-induced OA and the impact of altered mechanical loading on pathological changes in the synovial joint. Moreover, we examine the current strategies in cartilage tissue engineering and address the critical challenges of cell-based therapies for OA. Besides, we provide examples of innovative ways and potential strategies to overcome the obstacles in the treatment of obesity-induced OA. THE TRANSLATIONAL POTENTIAL OF THIS ARTICLE Altogether, this review delivers insight into obesity-induced OA and offers future research direction on the creation of tissue engineering-based therapies for obesity-induced OA.
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Affiliation(s)
- Antonia RuJia Sun
- Center for Translational Medicine Research and Development, Shenzhen Institutes of Advanced Technology, Chinese Academy of Science, Shenzhen, Guangdong, 518055, China
| | - Anjaneyulu Udduttula
- Center for Energy Metabolism and Reproduction, Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences, Shenzhen, Guangdong, 518055, China
| | - Jian Li
- Center for Translational Medicine Research and Development, Shenzhen Institutes of Advanced Technology, Chinese Academy of Science, Shenzhen, Guangdong, 518055, China
| | - Yanzhi Liu
- Center for Translational Medicine Research and Development, Shenzhen Institutes of Advanced Technology, Chinese Academy of Science, Shenzhen, Guangdong, 518055, China
- Guangdong Key Laboratory for Research and Development of Natural Drugs, Department of Pharmacology, Guangdong Medical University, Zhanjiang, Guangdong, 524023, China
| | - Pei-Gen Ren
- Center for Energy Metabolism and Reproduction, Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences, Shenzhen, Guangdong, 518055, China
| | - Peng Zhang
- Center for Translational Medicine Research and Development, Shenzhen Institutes of Advanced Technology, Chinese Academy of Science, Shenzhen, Guangdong, 518055, China
- Shenzhen Engineering Research Center for Medical Bioactive Materials, Shenzhen, Guangdong, 518055, China
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Zhang C, Gullbrand SE, Schaer TP, Lau YK, Jiang Z, Dodge GR, Elliott DM, Mauck RL, Malhotra NR, Smith LJ. Inflammatory cytokine and catabolic enzyme expression in a goat model of intervertebral disc degeneration. J Orthop Res 2020; 38:2521-2531. [PMID: 32091156 PMCID: PMC7483272 DOI: 10.1002/jor.24639] [Citation(s) in RCA: 32] [Impact Index Per Article: 6.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/11/2019] [Revised: 01/28/2020] [Accepted: 02/19/2020] [Indexed: 02/04/2023]
Abstract
Intervertebral disc degeneration is implicated as a leading cause of low back pain. Persistent, local inflammation within the disc nucleus pulposus (NP) and annulus fibrosus (AF) is an important mediator of disc degeneration and negatively impacts the performance of therapeutic stem cells. There is a lack of validated large animal models of disc degeneration that recapitulate clinically relevant local inflammation. We recently described a goat model of disc degeneration in which increasing doses of chondroitinase ABC (ChABC) were used to reproducibly induce a spectrum of degenerative changes. The objective of this study was to extend the clinical relevance of this model by establishing whether these degenerative changes are associated with the local expression of inflammatory cytokines and catabolic enzymes. Degeneration was induced in goat lumbar discs using ChABC at different doses. After 12 weeks, degeneration severity was determined histologically and using quantitative magnetic resonance imaging (MRI). Expression levels of inflammatory cytokines (tumor necrosis factor-α [TNF-α], interleukin-1β [IL-1β], and IL-6) and catabolic enzymes (matrix metalloproteinases-1 [MMPs-1] and 13, and a disintegrin and metalloproteinase with thrombospondin type-1 motifs-4 [ADAMTS-4]) were assessed as the percentage of immunopositive cells in the NP and AF. With the exception of MMP-1, cytokine, and enzyme expression levels were significantly elevated in ChABC-treated discs in the NP and AF. Expression levels of TNF-α, IL1-β, and ADAMTS-4 were positively correlated with histological grade, while all cytokines and ADAMTS-4 were negatively correlated with MRI T2 and T1ρ scores. These results demonstrate that degenerate goat discs exhibit elevated expression of clinically relevant inflammatory mediators, and further validate this animal model as a platform for evaluating new therapeutic approaches for disc degeneration.
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Affiliation(s)
- Chenghao Zhang
- Translational Musculoskeletal Research Center, Corporal Michael J. Crescenz Philadelphia VA Medical Center, 3900 Woodland Avenue, Philadelphia, PA, USA
- Department of Neurosurgery, Perelman School of Medicine, University of Pennsylvania, 3450 Hamilton Walk, Philadelphia, PA, USA
- Department of Orthopaedic Surgery, Perelman School of Medicine, University of Pennsylvania, 3450 Hamilton Walk, Philadelphia, PA, USA, Philadelphia, PA, USA
| | - Sarah E. Gullbrand
- Translational Musculoskeletal Research Center, Corporal Michael J. Crescenz Philadelphia VA Medical Center, 3900 Woodland Avenue, Philadelphia, PA, USA
- Department of Orthopaedic Surgery, Perelman School of Medicine, University of Pennsylvania, 3450 Hamilton Walk, Philadelphia, PA, USA, Philadelphia, PA, USA
| | - Thomas P. Schaer
- Comparative Orthopaedic Research Laboratory, New Bolton Center, School of Veterinary Medicine, University of Pennsylvania, 382 W Street Rd, Kennett Square, PA, USA
| | - Yian Khai Lau
- Translational Musculoskeletal Research Center, Corporal Michael J. Crescenz Philadelphia VA Medical Center, 3900 Woodland Avenue, Philadelphia, PA, USA
- Department of Neurosurgery, Perelman School of Medicine, University of Pennsylvania, 3450 Hamilton Walk, Philadelphia, PA, USA
- Department of Orthopaedic Surgery, Perelman School of Medicine, University of Pennsylvania, 3450 Hamilton Walk, Philadelphia, PA, USA, Philadelphia, PA, USA
| | - Zhirui Jiang
- Translational Musculoskeletal Research Center, Corporal Michael J. Crescenz Philadelphia VA Medical Center, 3900 Woodland Avenue, Philadelphia, PA, USA
- Department of Neurosurgery, Perelman School of Medicine, University of Pennsylvania, 3450 Hamilton Walk, Philadelphia, PA, USA
- Department of Orthopaedic Surgery, Perelman School of Medicine, University of Pennsylvania, 3450 Hamilton Walk, Philadelphia, PA, USA, Philadelphia, PA, USA
| | - George R. Dodge
- Translational Musculoskeletal Research Center, Corporal Michael J. Crescenz Philadelphia VA Medical Center, 3900 Woodland Avenue, Philadelphia, PA, USA
- Department of Orthopaedic Surgery, Perelman School of Medicine, University of Pennsylvania, 3450 Hamilton Walk, Philadelphia, PA, USA, Philadelphia, PA, USA
| | - Dawn M. Elliott
- Department of Biomedical Engineering, University of Delaware, Newark, DE, USA
| | - Robert L. Mauck
- Translational Musculoskeletal Research Center, Corporal Michael J. Crescenz Philadelphia VA Medical Center, 3900 Woodland Avenue, Philadelphia, PA, USA
- Department of Orthopaedic Surgery, Perelman School of Medicine, University of Pennsylvania, 3450 Hamilton Walk, Philadelphia, PA, USA, Philadelphia, PA, USA
| | - Neil R. Malhotra
- Translational Musculoskeletal Research Center, Corporal Michael J. Crescenz Philadelphia VA Medical Center, 3900 Woodland Avenue, Philadelphia, PA, USA
| | - Lachlan J. Smith
- Translational Musculoskeletal Research Center, Corporal Michael J. Crescenz Philadelphia VA Medical Center, 3900 Woodland Avenue, Philadelphia, PA, USA
- Department of Neurosurgery, Perelman School of Medicine, University of Pennsylvania, 3450 Hamilton Walk, Philadelphia, PA, USA
- Department of Orthopaedic Surgery, Perelman School of Medicine, University of Pennsylvania, 3450 Hamilton Walk, Philadelphia, PA, USA, Philadelphia, PA, USA
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Pleiotropic actions of Vitamin D in composite musculoskeletal trauma. Injury 2020; 51:2099-2109. [PMID: 32624209 DOI: 10.1016/j.injury.2020.06.023] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/06/2020] [Revised: 05/21/2020] [Accepted: 06/13/2020] [Indexed: 02/02/2023]
Abstract
Composite tissue injuries are the result of high energy impacts caused by motor vehicle accidents, gunshot wounds or blasts. These are highly traumatic injuries characterized by wide-spread, penetrating wounds affecting the entire musculoskeletal system, and are generally defined by frank volumetric muscle loss with concomitant segmental bone defects. At the tissue level, the breadth of damage to multiple tissue systems, and potential for infection from penetration, have been shown to lead to an exaggerated, often chronic inflammatory response with subsequent dysregulation of normal musculoskeletal healing mechanisms. Aside from the direct effects of inflammation on myogenesis and osteogenesis, frank muscle loss has been shown to directly impair fracture union and ultimately contribute to failed wound regeneration. Care for these injuries requires extensive surgical intervention and acute care strategies. However, often these interventions do not adequately mitigate inflammation or promote proper musculoskeletal injury repair and force amputation of the limb. Therefore, identification of factors that can promote tissue regeneration and mitigate inflammation could be key to restoring wound healing after composite tissue injury. One such factor that may directly affect both inflammation and tissue regeneration in response to these multi-tissue injuries may be Vitamin D. Beyond traditional roles, the pleiotropic and localized actions of Vitamin D are increasingly being recognized in most aspects of wound healing in complex tissue injuries - e.g., regulation of inflammation, myogenesis, fracture callus mineralization and remodeling. Conversely, pre-existing Vitamin D deficiency leads to musculoskeletal dysfunction, increased fracture risk or fracture non-unions, decreased strength/function and reduced capacity to heal wounds through increased inflammation. This Vitamin D deficient state requires acute supplementation in order to quickly restore circulating levels to an optimal level, thereby facilitating a robust wound healing response. Herein, the purpose of this review is to address the roles and critical functions of Vitamin D throughout the wound healing process. Findings from this review suggest that careful monitoring and/or supplementation of Vitamin D may be critical for wound regeneration in composite tissue injuries.
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Jauković A, Kukolj T, Obradović H, Okić-Đorđević I, Mojsilović S, Bugarski D. Inflammatory niche: Mesenchymal stromal cell priming by soluble mediators. World J Stem Cells 2020; 12:922-937. [PMID: 33033555 PMCID: PMC7524701 DOI: 10.4252/wjsc.v12.i9.922] [Citation(s) in RCA: 14] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/14/2020] [Revised: 05/13/2020] [Accepted: 09/02/2020] [Indexed: 02/06/2023] Open
Abstract
Mesenchymal stromal/stem cells (MSCs) are adult stem cells of stromal origin that possess self-renewal capacity and the ability to differentiate into multiple mesodermal cell lineages. They play a critical role in tissue homeostasis and wound healing, as well as in regulating the inflammatory microenvironment through interactions with immune cells. Hence, MSCs have garnered great attention as promising candidates for tissue regeneration and cell therapy. Because the inflammatory niche plays a key role in triggering the reparative and immunomodulatory functions of MSCs, priming of MSCs with bioactive molecules has been proposed as a way to foster the therapeutic potential of these cells. In this paper, we review how soluble mediators of the inflammatory niche (cytokines and alarmins) influence the regenerative and immunomodulatory capacity of MSCs, highlighting the major advantages and concerns regarding the therapeutic potential of these inflammatory primed MSCs. The data summarized in this review may provide a significant starting point for future research on priming MSCs and establishing standardized methods for the application of preconditioned MSCs in cell therapy.
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Affiliation(s)
- Aleksandra Jauković
- Laboratory for Experimental Hematology and Stem Cells, Institute for Medical Research, University of Belgrade, Belgrade 11129, Serbia
| | - Tamara Kukolj
- Laboratory for Experimental Hematology and Stem Cells, Institute for Medical Research, University of Belgrade, Belgrade 11129, Serbia
| | - Hristina Obradović
- Laboratory for Experimental Hematology and Stem Cells, Institute for Medical Research, University of Belgrade, Belgrade 11129, Serbia
| | - Ivana Okić-Đorđević
- Laboratory for Experimental Hematology and Stem Cells, Institute for Medical Research, University of Belgrade, Belgrade 11129, Serbia
| | - Slavko Mojsilović
- Laboratory for Experimental Hematology and Stem Cells, Institute for Medical Research, University of Belgrade, Belgrade 11129, Serbia
| | - Diana Bugarski
- Laboratory for Experimental Hematology and Stem Cells, Institute for Medical Research, University of Belgrade, Belgrade 11129, Serbia
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Otahal A, Kramer K, Kuten-Pella O, Weiss R, Stotter C, Lacza Z, Weber V, Nehrer S, De Luna A. Characterization and Chondroprotective Effects of Extracellular Vesicles From Plasma- and Serum-Based Autologous Blood-Derived Products for Osteoarthritis Therapy. Front Bioeng Biotechnol 2020; 8:584050. [PMID: 33102466 PMCID: PMC7546339 DOI: 10.3389/fbioe.2020.584050] [Citation(s) in RCA: 34] [Impact Index Per Article: 6.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/16/2020] [Accepted: 08/31/2020] [Indexed: 12/11/2022] Open
Abstract
Autologous blood products gain increasing interest in the field of regenerative medicine as well as in orthopedics, aesthetic surgery, and cosmetics. Currently, citrate-anticoagulated platelet-rich plasma (CPRP) preparations are often applied in osteoarthritis (OA), but more physiological and cell-free alternatives such as hyperacute serum (hypACT) are under development. Besides growth factors, blood products also bring along extracellular vesicles (EVs) packed with signal molecules, which open up a new level of complexity at evaluating the functional spectrum of blood products. Large proportions of EVs originated from platelets in CPRP and hypACT, whereas very low erythrocyte and monocyte-derived EVs were detected via flow cytometry. EV treatment of chondrocytes enhanced the expression of anabolic markers type II collagen, SRY-box transcription factor 9 (SOX9), and aggrecan compared to full blood products, but also the catabolic marker and tissue remodeling factor matrix metalloproteinase 3, whereas hypACT EVs prevented type I collagen expression. CPRP blood product increased SOX9 protein expression, in contrast to hypACT blood product. However, hypACT EVs induced SOX9 protein expression while preventing interleukin-6 secretion. The results indicate that blood EVs are sufficient to induce chondrogenic gene expression changes in OA chondrocytes, while preventing proinflammatory cytokine release compared to full blood product. This highlights the potential of autologous blood-derived EVs as regulators of cartilage extracellular matrix metabolism and inflammation, as well as candidates for new cell-free therapeutic approaches for OA.
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Affiliation(s)
- Alexander Otahal
- Center for Regenerative Medicine, Department for Health Sciences, Medicine and Research, Danube University Krems, Krems an der Donau, Austria
| | - Karina Kramer
- Center for Regenerative Medicine, Department for Health Sciences, Medicine and Research, Danube University Krems, Krems an der Donau, Austria
| | - Olga Kuten-Pella
- Center for Regenerative Medicine, Department for Health Sciences, Medicine and Research, Danube University Krems, Krems an der Donau, Austria.,OrthoSera GmbH, Krems an der Donau, Austria
| | - René Weiss
- Center for Biomedical Technology, Department for Biomedical Research, Danube University Krems, Krems an der Donau, Austria
| | - Christoph Stotter
- Center for Regenerative Medicine, Department for Health Sciences, Medicine and Research, Danube University Krems, Krems an der Donau, Austria
| | - Zsombor Lacza
- Deptartment Sports Physiology, University of Physical Education, Budapest, Hungary
| | - Viktoria Weber
- Center for Biomedical Technology, Department for Biomedical Research, Danube University Krems, Krems an der Donau, Austria
| | - Stefan Nehrer
- Center for Regenerative Medicine, Department for Health Sciences, Medicine and Research, Danube University Krems, Krems an der Donau, Austria
| | - Andrea De Luna
- Center for Regenerative Medicine, Department for Health Sciences, Medicine and Research, Danube University Krems, Krems an der Donau, Austria
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50
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Miyatake K, Kumagai K, Imai S, Yamaguchi Y, Inaba Y. Sclerostin inhibits interleukin-1β-induced late stage chondrogenic differentiation through downregulation of Wnt/β-catenin signaling pathway. PLoS One 2020; 15:e0239651. [PMID: 32976505 PMCID: PMC7518574 DOI: 10.1371/journal.pone.0239651] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/30/2019] [Accepted: 09/11/2020] [Indexed: 11/19/2022] Open
Abstract
It is known that Wnt/β-catenin signaling induces endochondral ossification and plays a significant role in the pathophysiology of osteoarthritis (OA). Sclerostin is a potent inhibitor of the Wnt/β-catenin signaling pathway. This study investigated the role of sclerostin in the endochondral differentiation under an OA-like condition induced by proinflammatory cytokines. ATDC5 cells were used to investigate chondrogenic differentiation and terminal calcification, and 10 ng/ml IL-1β and/or 200 ng/ml sclerostin were added to the culture medium. IL-1β impaired early chondrogenesis from undifferentiated state into proliferative chondrocytes, and it was not altered by sclerostin. IL-1β induced progression of chondrogenic differentiation in the late stage and promoted terminal calcification. These processes were inhibited by sclerostin and chondrogenic phenotype was restored. In addition, sclerostin restored IL-1β-induced upregulation of Wnt/β-catenin signaling in the late stage. This study provides insights into the possible role of sclerostin in the chondrogenic differentiation under the IL-1β-induced OA-like environment. Suppression of Wnt signaling by an antagonist may play a key role in the maintenance of articular homeostasis and has a potential to prevent the progression of OA. Thus, sclerostin is a candidate treatment option for OA.
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Affiliation(s)
- Kazuma Miyatake
- Department of Orthopaedic Surgery and Musculoskeletal Science, Graduate School of Medicine, Yokohama City University, Yokohama, Japan
| | - Ken Kumagai
- Department of Orthopaedic Surgery and Musculoskeletal Science, Graduate School of Medicine, Yokohama City University, Yokohama, Japan
- * E-mail:
| | - Sosuke Imai
- Department of Orthopaedic Surgery and Musculoskeletal Science, Graduate School of Medicine, Yokohama City University, Yokohama, Japan
| | - Yasuteru Yamaguchi
- Department of Orthopaedic Surgery and Musculoskeletal Science, Graduate School of Medicine, Yokohama City University, Yokohama, Japan
| | - Yutaka Inaba
- Department of Orthopaedic Surgery and Musculoskeletal Science, Graduate School of Medicine, Yokohama City University, Yokohama, Japan
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