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Zhu D, Huang MF, Xu A, Gao X, Huang YW, Phan TTT, Lu L, Chi TY, Dai Y, Pang LK, Gingold JA, Tu J, Huo Z, Bazer DA, Shoemaker R, Wang J, Ambrose CG, Shen J, Kameoka J, Zhao Z, Wang LL, Zhang Y, Zhao R, Lee DF. Systematic transcriptome profiling of hPSC-derived osteoblasts unveils CORIN's mastery in governing osteogenesis through CEBPD modulation. J Biol Chem 2024; 300:107494. [PMID: 38925326 PMCID: PMC11301355 DOI: 10.1016/j.jbc.2024.107494] [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: 01/23/2024] [Revised: 05/21/2024] [Accepted: 06/09/2024] [Indexed: 06/28/2024] Open
Abstract
The commitment of stem cells to differentiate into osteoblasts is a highly regulated and complex process that involves the coordination of extrinsic signals and intrinsic transcriptional machinery. While rodent osteoblastic differentiation has been extensively studied, research on human osteogenesis has been limited by cell sources and existing models. Here, we systematically dissect human pluripotent stem cell-derived osteoblasts to identify functional membrane proteins and their downstream transcriptional networks involved in human osteogenesis. Our results reveal an enrichment of type II transmembrane serine protease CORIN in humans but not rodent osteoblasts. Functional analyses demonstrated that CORIN depletion significantly impairs osteogenesis. Genome-wide chromatin immunoprecipitation enrichment and mechanistic studies show that p38 MAPK-mediated CCAAT enhancer binding protein delta (CEBPD) upregulation is required for CORIN-modulated osteogenesis. Contrastingly, the type I transmembrane heparan sulfate proteoglycan SDC1 enriched in mesenchymal stem cells exerts a negative regulatory effect on osteogenesis through a similar mechanism. Chromatin immunoprecipitation-seq, bulk and single-cell transcriptomes, and functional validations indicated that CEBPD plays a critical role in controlling osteogenesis. In summary, our findings uncover previously unrecognized CORIN-mediated CEBPD transcriptomic networks in driving human osteoblast lineage commitment.
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Affiliation(s)
- Dandan Zhu
- Department of Integrative Biology and Pharmacology, McGovern Medical School, The University of Texas Health Science Center at Houston, Houston, Texas, USA
| | - Mo-Fan Huang
- Department of Integrative Biology and Pharmacology, McGovern Medical School, The University of Texas Health Science Center at Houston, Houston, Texas, USA; The University of Texas MD Anderson Cancer Center UTHealth Houston Graduate School of Biomedical Sciences, Houston, Texas, USA
| | - An Xu
- Department of Integrative Biology and Pharmacology, McGovern Medical School, The University of Texas Health Science Center at Houston, Houston, Texas, USA
| | - Xueqin Gao
- Department of Integrative Biology and Pharmacology, McGovern Medical School, The University of Texas Health Science Center at Houston, Houston, Texas, USA; Linda and Mitch Hart Center for Regenerative and Personalized Medicine, Steadman Philippon Research Institute, Vail, Colorado, USA
| | - Yu-Wen Huang
- Department of Integrative Biology and Pharmacology, McGovern Medical School, The University of Texas Health Science Center at Houston, Houston, Texas, USA
| | - Trinh T T Phan
- Department of Integrative Biology and Pharmacology, McGovern Medical School, The University of Texas Health Science Center at Houston, Houston, Texas, USA
| | - Linchao Lu
- Department of Pediatrics, Baylor College of Medicine, Texas Children's Hospital, Houston, Texas, USA
| | - Ting-Yen Chi
- Department of Materials Science and Engineering, Texas A&M University, College Station, Texas, USA
| | - Yulin Dai
- Center for Precision Health, School of Biomedical Informatics, The University of Texas Health Science Center at Houston, Houston, Texas, USA
| | - Lon Kai Pang
- Department of Pediatrics, Baylor College of Medicine, Texas Children's Hospital, Houston, Texas, USA
| | - Julian A Gingold
- Department of Obstetrics & Gynecology and Women's Health, Einstein/Montefiore Medical Center, Bronx, New York, USA
| | - Jian Tu
- Department of Integrative Biology and Pharmacology, McGovern Medical School, The University of Texas Health Science Center at Houston, Houston, Texas, USA
| | - Zijun Huo
- Department of Integrative Biology and Pharmacology, McGovern Medical School, The University of Texas Health Science Center at Houston, Houston, Texas, USA
| | - Danielle A Bazer
- Department of Neurology, Renaissance School of Medicine at Stony Brook University, Stony Brook, New York, USA
| | - Rachel Shoemaker
- Department of Integrative Biology and Pharmacology, McGovern Medical School, The University of Texas Health Science Center at Houston, Houston, Texas, USA; The University of Texas MD Anderson Cancer Center UTHealth Houston Graduate School of Biomedical Sciences, Houston, Texas, USA
| | - Jun Wang
- The University of Texas MD Anderson Cancer Center UTHealth Houston Graduate School of Biomedical Sciences, Houston, Texas, USA; Department of Pediatrics, McGovern Medical School, The University of Texas Health Science Center at Houston, Houston, Texas, USA
| | - Catherine G Ambrose
- Department of Orthopedic Surgery, McGovern Medical School, The University of Texas Health Science Center at Houston, Houston, Texas, USA
| | - Jingnan Shen
- Department of Musculoskeletal Oncology, The First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, PR China
| | - Jun Kameoka
- Department of Materials Science and Engineering, Texas A&M University, College Station, Texas, USA; Department of Electrical and Computer Engineering, Texas A&M University, College Station, College Station, Texas, USA
| | - Zhongming Zhao
- The University of Texas MD Anderson Cancer Center UTHealth Houston Graduate School of Biomedical Sciences, Houston, Texas, USA; Center for Precision Health, School of Biomedical Informatics, The University of Texas Health Science Center at Houston, Houston, Texas, USA
| | - Lisa L Wang
- Department of Pediatrics, Baylor College of Medicine, Texas Children's Hospital, Houston, Texas, USA
| | - Yang Zhang
- College of Science, Harbin Institute of Technology (Shenzhen), Shenzhen, Guangdong, China.
| | - Ruiying Zhao
- Department of Integrative Biology and Pharmacology, McGovern Medical School, The University of Texas Health Science Center at Houston, Houston, Texas, USA.
| | - Dung-Fang Lee
- Department of Integrative Biology and Pharmacology, McGovern Medical School, The University of Texas Health Science Center at Houston, Houston, Texas, USA; The University of Texas MD Anderson Cancer Center UTHealth Houston Graduate School of Biomedical Sciences, Houston, Texas, USA; Center for Precision Health, School of Biomedical Informatics, The University of Texas Health Science Center at Houston, Houston, Texas, USA; Center for Stem Cell and Regenerative Medicine, The Brown Foundation Institute of Molecular Medicine for the Prevention of Human Diseases, The University of Texas Health Science Center at Houston, Houston, Texas, USA.
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Duan G, Lu YF, Chen HL, Zhu ZQ, Yang S, Wang YQ, Wang JQ, Jia XH. Smurf1-targeting microRNA-136-5p-modified bone marrow mesenchymal stem cells combined with 3D-printed β-tricalcium phosphate scaffolds strengthen osteogenic activity and alleviate bone defects. Kaohsiung J Med Sci 2024; 40:621-630. [PMID: 38820598 DOI: 10.1002/kjm2.12847] [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: 10/18/2023] [Revised: 04/12/2024] [Accepted: 04/25/2024] [Indexed: 06/02/2024] Open
Abstract
Suitable biomaterials with seed cells have promising potential to repair bone defects. However, bone marrow mesenchymal stem cells (BMSCs), one of the most common seed cells used in tissue engineering, cannot differentiate efficiently and accurately into functional osteoblasts. In view of this, a new tissue engineering technique combined with BMSCs and scaffolds is a major task for bone defect repair. Lentiviruses interfering with miR-136-5p or Smurf1 expression were transfected into BMSCs. The effects of miR-136-5p or Smurf1 on the osteogenic differentiation (OD) of BMSCs were evaluated by measuring alkaline phosphatase activity and calcium deposition. Then, the targeting relationship between miR-136-5p and Smurf1 was verified by bioinformatics website analysis and dual luciferase reporter assay. Then, a rabbit femoral condyle bone defect model was established. miR-136-5p/BMSCs/β-TCP scaffold was implanted into the defect, and the repair of the bone defect was detected by Micro-CT and HE staining. Elevating miR-136-5p-3p or suppressing Smurf1 could stimulate OD of BMSCs. miR-136-5p negatively regulated Smurf1 expression. Overexpressing Smurf1 reduced the promoting effect of miR-136-5p on the OD of BMSCs. miR-136-5p/BMSCs/β-TCP could strengthen bone density in the defected area and accelerate bone repair. SmurF1-targeting miR-136-5p-modified BMSCs combined with 3D-printed β-TCP scaffolds can strengthen osteogenic activity and alleviate bone defects.
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Affiliation(s)
- Gang Duan
- Department of Orthopedics, The Second Affiliated Hospital of Xuzhou Medical University, Xuzhou, Jiangsu, China
| | - Ya-Fei Lu
- Graduate School of Xuzhou Medical University, Xuzhou, Jiangsu, China
| | - Hong-Liang Chen
- Department of Orthopedics, The Affiliated Hospital of Xuzhou Medical University, Xuzhou, Jiangsu, China
| | - Zi-Qiang Zhu
- Department of Orthopedics, The Second Affiliated Hospital of Xuzhou Medical University, Xuzhou, Jiangsu, China
| | - Shuo Yang
- Department of Orthopedics, The Second Affiliated Hospital of Xuzhou Medical University, Xuzhou, Jiangsu, China
| | - Yun-Qing Wang
- Department of Orthopedics, The Second Affiliated Hospital of Xuzhou Medical University, Xuzhou, Jiangsu, China
| | - Jian-Qiang Wang
- Department of Orthopedics, The Second Affiliated Hospital of Xuzhou Medical University, Xuzhou, Jiangsu, China
| | - Xing-Hai Jia
- Department of Orthopedics, The Second Affiliated Hospital of Xuzhou Medical University, Xuzhou, Jiangsu, China
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Wu M, Wu S, Chen W, Li YP. The roles and regulatory mechanisms of TGF-β and BMP signaling in bone and cartilage development, homeostasis and disease. Cell Res 2024; 34:101-123. [PMID: 38267638 PMCID: PMC10837209 DOI: 10.1038/s41422-023-00918-9] [Citation(s) in RCA: 98] [Impact Index Per Article: 98.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/26/2023] [Accepted: 12/15/2023] [Indexed: 01/26/2024] Open
Abstract
Transforming growth factor-βs (TGF-βs) and bone morphometric proteins (BMPs) belong to the TGF-β superfamily and perform essential functions during osteoblast and chondrocyte lineage commitment and differentiation, skeletal development, and homeostasis. TGF-βs and BMPs transduce signals through SMAD-dependent and -independent pathways; specifically, they recruit different receptor heterotetramers and R-Smad complexes, resulting in unique biological readouts. BMPs promote osteogenesis, osteoclastogenesis, and chondrogenesis at all differentiation stages, while TGF-βs play different roles in a stage-dependent manner. BMPs and TGF-β have opposite functions in articular cartilage homeostasis. Moreover, TGF-β has a specific role in maintaining the osteocyte network. The precise activation of BMP and TGF-β signaling requires regulatory machinery at multiple levels, including latency control in the matrix, extracellular antagonists, ubiquitination and phosphorylation in the cytoplasm, nucleus-cytoplasm transportation, and transcriptional co-regulation in the nuclei. This review weaves the background information with the latest advances in the signaling facilitated by TGF-βs and BMPs, and the advanced understanding of their diverse physiological functions and regulations. This review also summarizes the human diseases and mouse models associated with disordered TGF-β and BMP signaling. A more precise understanding of the BMP and TGF-β signaling could facilitate the development of bona fide clinical applications in treating bone and cartilage disorders.
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Affiliation(s)
- Mengrui Wu
- Department of Cell and Developmental Biology, College of Life Sciences, Zhejiang University, Hangzhou, Zhejiang, China.
| | - Shali Wu
- Department of Cell and Developmental Biology, College of Life Sciences, Zhejiang University, Hangzhou, Zhejiang, China
| | - Wei Chen
- Division in Cellular and Molecular Medicine, Department of Pathology and Laboratory Medicine, Tulane University School of Medicine, Tulane University, New Orleans, LA, USA
| | - Yi-Ping Li
- Division in Cellular and Molecular Medicine, Department of Pathology and Laboratory Medicine, Tulane University School of Medicine, Tulane University, New Orleans, LA, USA.
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Xiao Y, Shen Q, Li W, Zhang Y, Yin K, Xu Y. 280 mT static magnetic field promotes the growth of postpartum condylar cartilage. Connect Tissue Res 2022; 64:248-261. [PMID: 36469671 DOI: 10.1080/03008207.2022.2148527] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
PURPOSE Functional appliances made of permanent magnets have been used in jaw orthopedic treatment. However, whether the static magnetic field (SMF) generated by permanent magnets promotes the developmental sequence of condylar cartilage and thus promotes the growth of the mandible remains to be studied. The aim of this study was to investigate the effects of 280 mT SMF on postnatal condylar chondrogenesis and endochondral ossification and the roles of FLRT3, FGF2 and BMP2 signaling in this chondrodevelopmental sequences. METHODS Forty-eight rats were assigned to two groups (control and SMF). The condyles were collected at the specified time points. The histomorphological changes in the condyle were observed by histological staining. The expression of proteins related to the proliferation and differentiation of the condylar cartilage and the changes in subchondral bone microstructure were analyzed by immunohistochemical staining and micro-CT scanning. FLRT3, FGF2, and BMP2 expression was detected by immunofluorescence staining. RESULTS Under SMF stimulation, the cartilage of young rats grew longitudinally and laterally, and the thickness of the cartilage became thinner as it grew. The SMF promoted the proliferation and differentiation of condylar chondrocytes and endochondral ossification and increased subchondral bone mineral density, and BMP2 signaling was involved. Moreover, under SMF loading, the increased expression of FGF2 and FLRT3 were involved in regulating cartilage morphogenesis and growth. In late development, the decreased expression of FGF2/FLRT3 and the increased expression of BMP2 promoted endochondral ossification. The SMF accelerated this opposite expression trend. CONCLUSION FGF2/FLRT3 and BMP2 signals are involved in the regulatory effect of SMF exposure on chondrogenesis and endochondral ossification, which provides a theoretical basis for the clinical use of magnetic appliances to promote condylar growth.
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Affiliation(s)
- Yiwen Xiao
- Department of Orthodontics, Kunming Medical University School and Hospital of Stomatology, Kunming, China.,Department of Stomatology, Hubei NO. 3 People's Hospital of Jianghan University, Wuhan, China.,Yunnan Key Laboratory of Stomatology, Kunming, China
| | - Qinhao Shen
- Yunnan Key Laboratory of Stomatology, Kunming, China.,Department of the first dental clinic, Kunming Medical University School and Hospital of Stomatology, Kunming, China
| | - Weihao Li
- Yunnan Key Laboratory of Stomatology, Kunming, China
| | - Yibo Zhang
- Yunnan Key Laboratory of Stomatology, Kunming, China
| | - Kang Yin
- Department of Orthodontics, Kunming Medical University School and Hospital of Stomatology, Kunming, China
| | - Yanhua Xu
- Department of Orthodontics, Kunming Medical University School and Hospital of Stomatology, Kunming, China
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Xu K, Chu Y, Liu Q, Fan W, He H, Huang F. NEDD4 E3 Ligases: Functions and Mechanisms in Bone and Tooth. Int J Mol Sci 2022; 23:ijms23179937. [PMID: 36077334 PMCID: PMC9455957 DOI: 10.3390/ijms23179937] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/28/2022] [Revised: 08/26/2022] [Accepted: 08/29/2022] [Indexed: 11/29/2022] Open
Abstract
Protein ubiquitination is a precisely controlled enzymatic cascade reaction belonging to the post-translational modification of proteins. In this process, E3 ligases catalyze the binding of ubiquitin (Ub) to protein substrates and define specificity. The neuronally expressed developmentally down-regulated 4 (NEDD4) subfamily, belonging to the homology to E6APC terminus (HECT) class of E3 ligases, has recently emerged as an essential determinant of multiple cellular processes in different tissues, including bone and tooth. Here, we place special emphasis on the regulatory role of the NEDD4 subfamily in the molecular and cell biology of osteogenesis. We elucidate in detail the specific roles, downstream substrates, and upstream regulatory mechanisms of the NEDD4 subfamily. Further, we provide an overview of the involvement of E3 ligases and deubiquitinases in the development, repair, and regeneration of another mineralized tissue—tooth.
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Affiliation(s)
- Ke Xu
- Hospital of Stomatology, Guanghua School of Stomatology, Sun Yat-sen University, Guangzhou 510008, China
- Guangdong Provincial Key Laboratory of Stomatology, Guangzhou 510008, China
| | - Yanhao Chu
- Hospital of Stomatology, Guanghua School of Stomatology, Sun Yat-sen University, Guangzhou 510008, China
- Guangdong Provincial Key Laboratory of Stomatology, Guangzhou 510008, China
| | - Qin Liu
- Hospital of Stomatology, Guanghua School of Stomatology, Sun Yat-sen University, Guangzhou 510008, China
- Guangdong Provincial Key Laboratory of Stomatology, Guangzhou 510008, China
| | - Wenguo Fan
- Hospital of Stomatology, Guanghua School of Stomatology, Sun Yat-sen University, Guangzhou 510008, China
- Guangdong Provincial Key Laboratory of Stomatology, Guangzhou 510008, China
| | - Hongwen He
- Guangdong Provincial Key Laboratory of Stomatology, Guangzhou 510008, China
- Correspondence: (H.H.); (F.H.)
| | - Fang Huang
- Hospital of Stomatology, Guanghua School of Stomatology, Sun Yat-sen University, Guangzhou 510008, China
- Guangdong Provincial Key Laboratory of Stomatology, Guangzhou 510008, China
- Correspondence: (H.H.); (F.H.)
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Gan Q, Pan H, Zhang W, Yuan Y, Qian J, Liu C. Fabrication and evaluation of a BMP-2/dexamethasone co-loaded gelatin sponge scaffold for rapid bone regeneration. Regen Biomater 2022; 9:rbac008. [PMID: 35592142 PMCID: PMC9113239 DOI: 10.1093/rb/rbac008] [Citation(s) in RCA: 6] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/20/2021] [Revised: 12/31/2021] [Accepted: 01/10/2022] [Indexed: 11/26/2022] Open
Abstract
Improving the osteogenic activity of BMP-2 in vivo has significant clinical application value. In this research, we use a clinical gelatin sponge scaffold loaded with BMP-2 and dexamethasone (Dex) to evaluate the osteogenic activity of dual drugs via ectopic osteogenesis in vivo. We also investigate the mechanism of osteogenesis induced by BMP-2 and Dex with C2C12, a multipotent muscle-derived progenitor cell. The results show that the gelatin scaffold with Dex and BMP-2 can significantly accelerate osteogenesis in vivo. It is indicated that compared with the BMP-2 or Dex alone, 100 nM of Dex can dramatically enhance the BMP-2-induced alkaline phosphatase activity (ALP), ALP mRNA expression and mineralization. Further studies show that 100 nM of Dex can maintain the secondary structure of BMP-2 and facilitate recognition of BMP-2 with its receptors on the surface of C2C12 cells. We also find that in C2C12, Dex has no obvious effect on the BMP-2-induced Smad1/5/8 protein expression and the STAT3-dependent pathway, but Runx2-dependent pathway is involved in the Dex-stimulated osteoblast differentiation of BMP-2 both in vitro and in vivo. Based on these results, a potential mechanism model about the synergistic osteoinductive effect of Dex and BMP-2 in C2C12 cells via Runx2 activation is proposed. This may provide a theoretical basis for the pre-clinical application of Dex and BMP-2 for bone regeneration.
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Affiliation(s)
- Qi Gan
- Key Laboratory for Ultrafine Materials of Ministry of Education, East China University of Science and Technology, Shanghai, 200237, PR China
- The State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, 200237, PR China
| | - Hao Pan
- Engineering Research Center for Biomedical Materials of the Ministry of Education, East China University of Science and Technology, Shanghai, 200237, PR China
| | - Wenjing Zhang
- Engineering Research Center for Biomedical Materials of the Ministry of Education, East China University of Science and Technology, Shanghai, 200237, PR China
| | - Yuan Yuan
- Key Laboratory for Ultrafine Materials of Ministry of Education, East China University of Science and Technology, Shanghai, 200237, PR China
| | - Jiangchao Qian
- The State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, 200237, PR China
| | - Changsheng Liu
- Key Laboratory for Ultrafine Materials of Ministry of Education, East China University of Science and Technology, Shanghai, 200237, PR China
- Engineering Research Center for Biomedical Materials of the Ministry of Education, East China University of Science and Technology, Shanghai, 200237, PR China
- Frontiers Science Center for Materiobiology and Dynamic Chemistry, East China University of Science and Technology, Shanghai, 200237, PR China
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Huang Z. Simplifying cell fate map by determining lineage history of core pathway activation during fate specification. TRENDS IN DEVELOPMENTAL BIOLOGY 2022; 15:53-62. [PMID: 37396969 PMCID: PMC10312135] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Subscribe] [Scholar Register] [Indexed: 07/04/2023]
Abstract
A fundamental question in developmental biology is how a single genome gives rise to the diversity of cell fates. In essence, each cell fate in the human body is a unique but stable output state of the genome, maintained by positive and negative feedbacks from both inside and outside the cell (a stable cell state). Traditionally, defining a cell fate means identifying a unique combination of transcriptional factors expressed by the specific cell type. The hundreds of transcriptional factors in the genome, however, have complicated the task of simplifying cell fate representation and obtaining insights into its regulation. Moreover, results from this approach provides only a mostly static picture, with each cell fate/state disconnected from one another. An alternative approach instead defines cell fates by determining their relationship to each other, through identifying the signaling pathways that control each step of their lineage transition from a common progenitor during development. Decades of studies have shown only a handful of signaling pathways are sufficient to specify all cell fates in the body, simplifying the execution of such a strategy. In this review, I will argue this alternative approach is not only feasible but also has the potential of simplifying the cell fate landscape as well as facilitating the engineering of different cell fates for regenerative medicine.
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Affiliation(s)
- Zhen Huang
- Departments of Neuroscience and Neurology, University of Wisconsin-Madison, Madison, WI 53705 USA
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8
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Krstić J, Mojsilović S, Mojsilović SS, Santibanez JF. Regulation of the mesenchymal stem cell fate by interleukin-17: Implications in osteogenic differentiation. World J Stem Cells 2021; 13:1696-1713. [PMID: 34909118 PMCID: PMC8641017 DOI: 10.4252/wjsc.v13.i11.1696] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/05/2021] [Revised: 05/14/2021] [Accepted: 10/18/2021] [Indexed: 02/06/2023] Open
Abstract
Bone regeneration is a tightly regulated process that ensures proper repair and functionality after injury. The delicate balance between bone formation and resorption is governed by cytokines and signaling molecules released during the inflammatory response. Interleukin (IL)-17A, produced in the early phase of inflammation, influences the fate of osteoprogenitors. Due to their inherent capacity to differentiate into osteoblasts, mesenchymal stem/stromal cells (MSCs) contribute to bone healing and regeneration. This review presents an overview of IL-17A signaling and the leading cellular and molecular mechanisms by which it regulates the osteogenic differentiation of MSCs. The main findings demonstrating IL-17A’s influence on osteoblastogenesis are described. To this end, divergent information exists about the capacity of IL-17A to regulate MSCs’ osteogenic fate, depending on the tissue context and target cell type, along with contradictory findings in the same cell types. Therefore, we summarize the data showing both the pro-osteogenic and anti-osteogenic roles of IL-17, which may help in the understanding of IL-17A function in bone repair and regeneration.
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Affiliation(s)
- Jelena Krstić
- Gottfried Schatz Research Center, Medical University of Graz, Graz 8010, Austria
| | - Slavko Mojsilović
- Group for Hematology and Stem Cells, Institute for Medical Research, National Institute of Republic of Serbia, University of Belgrade, Belgrade 11129, Serbia
| | - Sonja S Mojsilović
- Group for Immunology, Institute for Medical Research, National Institute of Republic of Serbia, Belgrade 11129, Serbia
| | - Juan F Santibanez
- Group for Molecular Oncology, Institute for Medical Research, National Institute of Republic of Serbia, University of Belgrade, Belgrade 11000, Serbia
- Centro Integrativo de Biología y Química Aplicada, Universidad Bernardo O’Higgins, Chile 8370993, Chile
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Park JH, Ameri AH, Dempsey KE, Conrad DN, Kem M, Mino-Kenudson M, Demehri S. Nuclear IL-33/SMAD signaling axis promotes cancer development in chronic inflammation. EMBO J 2021; 40:e106151. [PMID: 33616251 DOI: 10.15252/embj.2020106151] [Citation(s) in RCA: 37] [Impact Index Per Article: 9.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/05/2020] [Revised: 12/27/2020] [Accepted: 01/11/2021] [Indexed: 12/16/2022] Open
Abstract
Interleukin (IL)-33 cytokine plays a critical role in allergic diseases and cancer. IL-33 also has a nuclear localization signal. However, the nuclear function of IL-33 and its impact on cancer is unknown. Here, we demonstrate that nuclear IL-33-mediated activation of SMAD signaling pathway in epithelial cells is essential for cancer development in chronic inflammation. Using RNA and ChIP sequencing, we found that nuclear IL-33 repressed the expression of an inhibitory SMAD, Smad6, by interacting with its transcription factor, RUNX2. IL-33 was highly expressed in the skin and pancreatic epithelial cells in chronic inflammation, leading to a markedly repressed Smad6 expression as well as dramatically upregulated p-SMAD2/3 and p-SMAD1/5 in the epithelial cells. Blocking TGF-β/SMAD signaling attenuated the IL-33-induced cell proliferation in vitro and inhibited IL-33-dependent epidermal hyperplasia and skin cancer development in vivo. IL-33 and SMAD signaling were upregulated in human skin cancer, pancreatitis, and pancreatitis-associated pancreatic cancer. Collectively, our findings reveal that nuclear IL-33/SMAD signaling is a cell-autonomous tumor-promoting axis in chronic inflammation, which can be targeted by small-molecule inhibitors for cancer treatment and prevention.
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Affiliation(s)
- Jong Ho Park
- Center for Cancer Immunology and Cutaneous Biology Research Center, Department of Dermatology, Center for Cancer Research, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA
| | - Amir H Ameri
- Center for Cancer Immunology and Cutaneous Biology Research Center, Department of Dermatology, Center for Cancer Research, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA
| | - Kaitlin E Dempsey
- Center for Cancer Immunology and Cutaneous Biology Research Center, Department of Dermatology, Center for Cancer Research, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA
| | - Danielle N Conrad
- Center for Cancer Immunology and Cutaneous Biology Research Center, Department of Dermatology, Center for Cancer Research, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA
| | - Marina Kem
- Department of Pathology, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA
| | - Mari Mino-Kenudson
- Department of Pathology, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA
| | - Shadmehr Demehri
- Center for Cancer Immunology and Cutaneous Biology Research Center, Department of Dermatology, Center for Cancer Research, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA
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10
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Human kidney clonal proliferation disclose lineage-restricted precursor characteristics. Sci Rep 2020; 10:22097. [PMID: 33328501 PMCID: PMC7745030 DOI: 10.1038/s41598-020-78366-3] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/26/2020] [Accepted: 11/02/2020] [Indexed: 01/10/2023] Open
Abstract
In-vivo single cell clonal analysis in the adult mouse kidney has previously shown lineage-restricted clonal proliferation within varying nephron segments as a mechanism responsible for cell replacement and local regeneration. To analyze ex-vivo clonal growth, we now preformed limiting dilution to generate genuine clonal cultures from one single human renal epithelial cell, which can give rise to up to 3.4 * 106 cells, and analyzed their characteristics using transcriptomics. A comparison between clonal cultures revealed restriction to either proximal or distal kidney sub-lineages with distinct cellular and molecular characteristics; rapidly amplifying de-differentiated clones and a stably proliferating cuboidal epithelial-appearing clones, respectively. Furthermore, each showed distinct molecular features including cell-cycle, epithelial-mesenchymal transition, oxidative phosphorylation, BMP signaling pathway and cell surface markers. In addition, analysis of clonal versus bulk cultures show early clones to be more quiescent, with elevated expression of renal developmental genes and overall reduction in renal identity markers, but with an overlapping expression of nephron segment identifiers and multiple identity. Thus, ex-vivo clonal growth mimics the in-vivo situation displaying lineage-restricted precursor characteristics of mature renal cells. These data suggest that for reconstruction of varying renal lineages with human adult kidney based organoid technology and kidney regeneration ex-vivo, use of multiple heterogeneous precursors is warranted.
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Xiong A, He Y, Gao L, Li G, Weng J, Kang B, Wang D, Zeng H. Smurf1-targeting miR-19b-3p-modified BMSCs combined PLLA composite scaffold to enhance osteogenic activity and treat critical-sized bone defects. Biomater Sci 2020; 8:6069-6081. [PMID: 33000773 DOI: 10.1039/d0bm01251c] [Citation(s) in RCA: 18] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/15/2022]
Abstract
Over the past few years, tissue-engineering technology provided a new direction for bone defects therapy, which involved developing applicable biological materials composite with seed cells to repair bone defects tissue. However, as one of the commonest seed cells for tissue engineering, BMSCs (bone marrow mesenchymal stem cells), are still lacking an efficient and accurate differentiation ability into functional osteoblast. Given these facts, the development of a novel tissue engineering technology integrated BMSCs and scaffold materials have become an urgent need for bone defects repair. In this work, we found that miR-19b-3p could suppress the expression of Smurf1 which is a negative regulator of osteogenesis. By employing lentivirus pLVTHM-miR-19b-3p transfected BMSCs, we verified that miR-19b-3p could promote BMSCs osteogenic differentiation via suppressing Smurf1 expression. Furthermore, we designed a new porous PLLA/POSS scaffold combined with BMSCs for tissue engineering. In vitro experiment showed that miR-19b-3p modified BMSCs facilitated the expansion and proliferation of BMSCs when culturing with the PLLA/POSS scaffold. We established rats calvarial critical-sized defect model, after transplanting the BMSCs/PLLA/POSS for 3 month, the pathology, immunohistochemical and Micro-CT results showed that miR-19b-BMSCs/PLLA/POSS significantly facilitated the osteogenesis differentiation, enhanced the bone density of defect area and accelerated the repair of bone defect. We elucidated the mechanism that miR-19b-3p suppressed the expression of Smurf1 and provided a novel tissue engineering strategy for using microRNA gene-modified BMSCs combined with PLLA/POSS scaffold in bone tissue engineering.
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Affiliation(s)
- Ao Xiong
- Department of Bone & Joint Surgery, Peking University Shenzhen Hospital, Shenzhen, 518036, PR China.
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12
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Van Gils M, Nollet L, Verly E, Deianova N, Vanakker OM. Cellular signaling in pseudoxanthoma elasticum: an update. Cell Signal 2019; 55:119-129. [PMID: 30615970 DOI: 10.1016/j.cellsig.2018.12.009] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/23/2018] [Revised: 12/20/2018] [Accepted: 12/20/2018] [Indexed: 12/27/2022]
Abstract
Pseudoxanthoma elasticum is an autosomal recessive genodermatosis with variable expression, due to mutations in the ABCC6 or ENPP1 gene. It is characterized by elastic fiber mineralization and fragmentation, resulting in skin, eye and cardiovascular symptoms. Significant advances have been made in the last 20 years with respect to the phenotypic characterization and pathophysiological mechanisms leading to elastic fiber mineralization. Nonetheless, the substrates of the ABCC6 transporter - the main cause of PXE - remain currently unknown. Though the precise mechanisms linking the ABCC6 transporter to mineralization of the extracellular matrix are unclear, several studies have looked into the cellular consequences of ABCC6 deficiency in PXE patients and/or animal models. In this paper, we compile the evidence on cellular signaling in PXE, which seems to revolve mainly around TGF-βs, BMPs and inorganic pyrophosphate signaling cascades. Where conflicting results or fragmented data are present, we address these with novel signaling data. This way, we aim to better understand the up- and down-stream signaling of TGF-βs and BMPs in PXE and we demonstrate that ANKH deficiency can be an additional mechanism contributing to decreased serum PPi levels in PXE patients.
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Affiliation(s)
- M Van Gils
- Center for Medical Genetics, Ghent University Hospital, Ghent, Belgium; Department of Biomolecular Medicine, Ghent University, Belgium
| | - L Nollet
- Center for Medical Genetics, Ghent University Hospital, Ghent, Belgium
| | - E Verly
- Center for Medical Genetics, Ghent University Hospital, Ghent, Belgium
| | - N Deianova
- Center for Medical Genetics, Ghent University Hospital, Ghent, Belgium
| | - O M Vanakker
- Center for Medical Genetics, Ghent University Hospital, Ghent, Belgium; Department of Biomolecular Medicine, Ghent University, Belgium.
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13
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Abstract
Bone morphogenetic proteins (BMPs) constitute the largest subdivision of the transforming growth factor-β family of ligands. BMPs exhibit widespread utility and pleiotropic, context-dependent effects, and the strength and duration of BMP pathway signaling is tightly regulated at numerous levels via mechanisms operating both inside and outside the cell. Defects in the BMP pathway or its regulation underlie multiple human diseases of different organ systems. Yet much remains to be discovered about the BMP pathway in its original context, i.e., the skeleton. In this review, we provide a comprehensive overview of the intricacies of the BMP pathway and its inhibitors in bone development, homeostasis, and disease. We frame the content of the review around major unanswered questions for which incomplete evidence is available. First, we consider the gene regulatory network downstream of BMP signaling in osteoblastogenesis. Next, we examine why some BMP ligands are more osteogenic than others and what factors limit BMP signaling during osteoblastogenesis. Then we consider whether specific BMP pathway components are required for normal skeletal development, and if the pathway exerts endogenous effects in the aging skeleton. Finally, we propose two major areas of need of future study by the field: greater resolution of the gene regulatory network downstream of BMP signaling in the skeleton, and an expanded repertoire of reagents to reliably and specifically inhibit individual BMP pathway components.
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Affiliation(s)
- Jonathan W Lowery
- Division of Biomedical Science, Marian University College of Osteopathic Medicine , Indianapolis, Indiana ; and Department of Developmental Biology, Harvard School of Dental Medicine , Boston, Massachusetts
| | - Vicki Rosen
- Division of Biomedical Science, Marian University College of Osteopathic Medicine , Indianapolis, Indiana ; and Department of Developmental Biology, Harvard School of Dental Medicine , Boston, Massachusetts
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14
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Abstract
The transforming growth factor-β (TGF-β) family of ligands elicit their biological effects by initiating new programs of gene expression. The best understood signal transducers for these ligands are the SMADs, which essentially act as transcription factors that are activated in the cytoplasm and then accumulate in the nucleus in response to ligand induction where they bind to enhancer/promoter sequences in the regulatory regions of target genes to either activate or repress transcription. This review focuses on the mechanisms whereby the SMADs achieve this and the functional implications. The SMAD complexes have weak affinity for DNA and limited specificity and, thus, they cooperate with other site-specific transcription factors that act either to actively recruit the SMAD complexes or to stabilize their DNA binding. In some situations, these cooperating transcription factors function to integrate the signals from TGF-β family ligands with environmental cues or with information about cell lineage. Activated SMAD complexes regulate transcription via remodeling of the chromatin template. Consistent with this, they recruit a variety of coactivators and corepressors to the chromatin, which either directly or indirectly modify histones and/or modulate chromatin structure.
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Affiliation(s)
- Caroline S Hill
- The Francis Crick Institute, Lincoln's Inn Fields Laboratory, London WC2A 3LY, United Kingdom
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15
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Senarath-Yapa K, Li S, Walmsley GG, Zielins E, Paik K, Britto JA, Grigoriadis AE, Wan DC, Liu KJ, Longaker MT, Quarto N. Small Molecule Inhibition of Transforming Growth Factor Beta Signaling Enables the Endogenous Regenerative Potential of the Mammalian Calvarium. Tissue Eng Part A 2016; 22:707-20. [PMID: 27036931 DOI: 10.1089/ten.tea.2015.0527] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/12/2022] Open
Abstract
Current approaches for the treatment of skeletal defects are suboptimal, principally because the ability of bone to repair and regenerate is poor. Although the promise of effective cellular therapies for skeletal repair is encouraging, these approaches are limited by the risks of infection, cellular contamination, and tumorigenicity. Development of a pharmacological approach would therefore help avoid some of these potential risks. This study identifies transforming growth factor beta (TGFβ) signaling as a potential pathway for pharmacological modulation in vivo. We demonstrate that inhibition of TGFβ signaling by the small molecule SB431542 potentiates calvarial skeletal repair through activation of bone morphogenetic protein (BMP) signaling on osteoblasts and dura mater cells participating in healing of calvarial defects. Cells respond to inhibition of TGFβ signaling by producing higher levels of BMP2 that upregulates inhibitory Smad6 expression, thus providing a negative feedback loop to contain excessive BMP signaling. Importantly, study on human osteoblasts indicates that molecular mechanism(s) triggered by SB431542 are conserved. Collectively, these data provide insights into the use of small molecules to modulate key signaling pathways for repairing skeletal defects.
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Affiliation(s)
- Kshemendra Senarath-Yapa
- 1 Hagey Laboratory for Pediatric Regenerative Medicine, Department of Surgery, Stanford University School of Medicine , Stanford, California.,2 Department of Craniofacial Development and Stem Cell Biology, Dental Institute , King's College London, London, United Kingdom .,3 Department of Plastic and Reconstructive Surgery, North Western Deanery , Manchester, United Kingdom
| | - Shuli Li
- 1 Hagey Laboratory for Pediatric Regenerative Medicine, Department of Surgery, Stanford University School of Medicine , Stanford, California
| | - Graham G Walmsley
- 1 Hagey Laboratory for Pediatric Regenerative Medicine, Department of Surgery, Stanford University School of Medicine , Stanford, California.,4 Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine , Stanford, California
| | - Elizabeth Zielins
- 1 Hagey Laboratory for Pediatric Regenerative Medicine, Department of Surgery, Stanford University School of Medicine , Stanford, California
| | - Kevin Paik
- 1 Hagey Laboratory for Pediatric Regenerative Medicine, Department of Surgery, Stanford University School of Medicine , Stanford, California
| | - Jonathan A Britto
- 5 Department of Craniofacial Surgery, Great Ormond Street Hospital , London, United Kingdom
| | - Agamemnon E Grigoriadis
- 2 Department of Craniofacial Development and Stem Cell Biology, Dental Institute , King's College London, London, United Kingdom
| | - Derrick C Wan
- 1 Hagey Laboratory for Pediatric Regenerative Medicine, Department of Surgery, Stanford University School of Medicine , Stanford, California
| | - Karen J Liu
- 2 Department of Craniofacial Development and Stem Cell Biology, Dental Institute , King's College London, London, United Kingdom
| | - Michael T Longaker
- 1 Hagey Laboratory for Pediatric Regenerative Medicine, Department of Surgery, Stanford University School of Medicine , Stanford, California.,4 Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine , Stanford, California
| | - Natalina Quarto
- 1 Hagey Laboratory for Pediatric Regenerative Medicine, Department of Surgery, Stanford University School of Medicine , Stanford, California.,6 Dipartimento di Scienze Biomediche Avanzate, Universita' degli Studi di Napoli Federico II , Napoli, Italy
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16
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Wu M, Chen G, Li YP. TGF-β and BMP signaling in osteoblast, skeletal development, and bone formation, homeostasis and disease. Bone Res 2016; 4:16009. [PMID: 27563484 PMCID: PMC4985055 DOI: 10.1038/boneres.2016.9] [Citation(s) in RCA: 1146] [Impact Index Per Article: 127.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/29/2015] [Revised: 03/04/2016] [Accepted: 03/07/2016] [Indexed: 12/11/2022] Open
Abstract
Transforming growth factor-beta (TGF-β) and bone morphogenic protein (BMP) signaling has fundamental roles in both embryonic skeletal development and postnatal bone homeostasis. TGF-βs and BMPs, acting on a tetrameric receptor complex, transduce signals to both the canonical Smad-dependent signaling pathway (that is, TGF-β/BMP ligands, receptors, and Smads) and the non-canonical-Smad-independent signaling pathway (that is, p38 mitogen-activated protein kinase/p38 MAPK) to regulate mesenchymal stem cell differentiation during skeletal development, bone formation and bone homeostasis. Both the Smad and p38 MAPK signaling pathways converge at transcription factors, for example, Runx2 to promote osteoblast differentiation and chondrocyte differentiation from mesenchymal precursor cells. TGF-β and BMP signaling is controlled by multiple factors, including the ubiquitin–proteasome system, epigenetic factors, and microRNA. Dysregulated TGF-β and BMP signaling result in a number of bone disorders in humans. Knockout or mutation of TGF-β and BMP signaling-related genes in mice leads to bone abnormalities of varying severity, which enable a better understanding of TGF-β/BMP signaling in bone and the signaling networks underlying osteoblast differentiation and bone formation. There is also crosstalk between TGF-β/BMP signaling and several critical cytokines’ signaling pathways (for example, Wnt, Hedgehog, Notch, PTHrP, and FGF) to coordinate osteogenesis, skeletal development, and bone homeostasis. This review summarizes the recent advances in our understanding of TGF-β/BMP signaling in osteoblast differentiation, chondrocyte differentiation, skeletal development, cartilage formation, bone formation, bone homeostasis, and related human bone diseases caused by the disruption of TGF-β/BMP signaling.
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Affiliation(s)
- Mengrui Wu
- Department of Pathology, University of Alabama at Birmingham , Birmingham, USA
| | - Guiqian Chen
- Department of Pathology, University of Alabama at Birmingham, Birmingham, USA; Department of neurology, Bruke Medical Research Institute, Weil Cornell Medicine of Cornell University, White Plains, USA
| | - Yi-Ping Li
- Department of Pathology, University of Alabama at Birmingham , Birmingham, USA
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17
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Mafi Golchin M, Heidari L, Ghaderian SMH, Akhavan-Niaki H. Osteoporosis: A Silent Disease with Complex Genetic Contribution. J Genet Genomics 2016; 43:49-61. [PMID: 26924688 DOI: 10.1016/j.jgg.2015.12.001] [Citation(s) in RCA: 35] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/13/2015] [Revised: 10/30/2015] [Accepted: 12/26/2015] [Indexed: 12/17/2022]
Abstract
Osteoporosis is the most common multifactorial metabolic bone disorder worldwide with a strong genetic component. In this review, the evidence for a genetic contribution to osteoporosis and related phenotypes is summarized alongside with methods used to identify osteoporosis susceptibility genes. The key biological pathways involved in the skeleton and bone development are discussed with a particular focus on master genes clustered in these pathways and their mode of action. Furthermore, the most studied single nucleotide polymorphisms (SNPs) analyzed for their importance as genetic markers of the disease are presented. New data generated by next-generation sequencing in conjunction with extensive meta-analyses should contribute to a better understanding of the genetic basis of osteoporosis and related phenotype variability. These data could be ultimately used for identifying at-risk patients for disease prevention by both controlling environmental factors and providing possible therapeutic targets.
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Affiliation(s)
- Maryam Mafi Golchin
- Department of Genetics, Faculty of Medicine, Babol University of Medical Sciences, Babol 4717647745, Iran
| | - Laleh Heidari
- Department of Medical Genetics, Faculty of Medicine, Shahid Beheshti University of Medical Sciences & Health Services, Tehran 1985717443, Iran
| | - Seyyed Mohammad Hossein Ghaderian
- Department of Medical Genetics, Faculty of Medicine, Shahid Beheshti University of Medical Sciences & Health Services, Tehran 1985717443, Iran
| | - Haleh Akhavan-Niaki
- Department of Genetics, Faculty of Medicine, Babol University of Medical Sciences, Babol 4717647745, Iran.
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18
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Yu W, Qiao Y, Tang X, Ma L, Wang Y, Zhang X, Weng W, Pan Q, Yu Y, Sun F, Wang J. Tumor suppressor long non-coding RNA, MT1DP is negatively regulated by YAP and Runx2 to inhibit FoxA1 in liver cancer cells. Cell Signal 2014; 26:2961-8. [PMID: 25261601 DOI: 10.1016/j.cellsig.2014.09.011] [Citation(s) in RCA: 88] [Impact Index Per Article: 8.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/21/2014] [Accepted: 09/05/2014] [Indexed: 02/08/2023]
Abstract
Recent studies are indicative for strong carcinogenetic roles of Runt related transcription factor 2 (Runx2) and Yes associated protein (YAP) in several cancer types. However, whether and how the interaction between Runx2 and YAP plays a role in liver tumorigenesis still remain illusive. Here, we identified a close relationship between Runx2 and YAP in liver cancer cells. Runx2 had a positive role on YAP expression and vice versa. We also found that Rux2 and YAP were capable of inhibiting long non-coding RNA (lncRNA), Metallothionein 1D, Pseudogene (MT1DP) expression through direct promoter binding. Overexpression of MT1DP resulted in reduced cell proliferation and colony formation in soft agar, but increased apoptosis in liver cancer cells, whereas knockdown of this lncRNA had the opposite effect, indicating that MT1DP acts as a tumor suppressor. Furthermore, MT1DP was revealed as a negative regulator of Alfa-fetoprotein (AFP), a classic liver cancer tumor marker, through inhibiting protein synthesis of Forkhead box A1 (FoxA1), an important transcription factor in liver development and cancer progression. Furthermore, we found that FoxA1 plays a positive role on YAP and Runx2 expression. Specially, opening the compacted chromatin by FoxA1 around CREB binding site within the YAP promoter facilitates CREB-mediated YAP transcription. Finally, MT1DP-inhibited in vivo liver cancer cell growth could be rescued by a combination of overexpression of FoxA1, Runx2 and YAP. Taken together, the close relationship between Rnux2 and YAP plays a pro-carcinogenetic role in liver cancer cells through inhibiting tumor suppressor lncRNA, MT1DP in a FoxA1 dependent manner.
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Affiliation(s)
- Wenjun Yu
- Department of Clinical Laboratory Medicine, Shanghai Tenth People's Hospital of Tongji University, Shanghai 200072, China
| | - Yongxia Qiao
- School of Public Health, Shanghai Jiaotong University School of Medicine, Shanghai 200025, China
| | - Xun Tang
- Department of Clinical Laboratory Medicine, Shanghai Tenth People's Hospital of Tongji University, Shanghai 200072, China
| | - Lifang Ma
- Department of Clinical Laboratory Medicine, Shanghai Tenth People's Hospital of Tongji University, Shanghai 200072, China
| | - Yulan Wang
- Department of Clinical Laboratory Medicine, Shanghai Tenth People's Hospital of Tongji University, Shanghai 200072, China
| | - Xiao Zhang
- Department of Clinical Laboratory Medicine, Shanghai Tenth People's Hospital of Tongji University, Shanghai 200072, China
| | - Wenhao Weng
- Department of Clinical Laboratory Medicine, Shanghai Tenth People's Hospital of Tongji University, Shanghai 200072, China
| | - Qiuhui Pan
- Department of Central Laboratory, Shanghai Tenth People's Hospital of Tongji University, Shanghai 200072, China
| | - Yongchun Yu
- Shanghai Municipal Hospital of Traditional Chinese Medicine affiliated to Shanghai TCM University, Shanghai 200071, China
| | - Fenyong Sun
- Department of Clinical Laboratory Medicine, Shanghai Tenth People's Hospital of Tongji University, Shanghai 200072, China.
| | - Jiayi Wang
- Department of Clinical Laboratory Medicine, Shanghai Tenth People's Hospital of Tongji University, Shanghai 200072, China.
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19
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Li S, Shu B, Zhang Y, Li J, Guo J, Wang Y, Ren F, Xiao G, Chang Z, Chen D. Carboxyl terminus of Hsp70-interacting protein regulation of osteoclast formation in mice through promotion of tumor necrosis factor receptor-associated factor 6 protein degradation. Arthritis Rheumatol 2014; 66:1854-63. [PMID: 24578159 DOI: 10.1002/art.38521] [Citation(s) in RCA: 18] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/25/2013] [Accepted: 02/18/2014] [Indexed: 12/20/2022]
Abstract
OBJECTIVE Carboxyl terminus of Hsp70-interacting protein (CHIP or STUB1) is an E3 ligase that regulates the stability of several proteins involved in tumor growth and metastasis. However, the role of CHIP in bone growth and bone remodeling in vivo has not been reported. This study was undertaken to investigate the role and mechanism of CHIP in regulation of bone mass and bone remodeling. METHODS The bone phenotype of Chip(-/-) mice was assessed by histologic, histomorphometric, and micro-computed tomographic analyses. The mechanism by which CHIP regulates the degradation of tumor necrosis factor receptor-associated factor 6 (TRAF6) and the inhibition of NF-κB signaling was examined by immunoprecipitation, Western blot, and luciferase reporter assays. RESULTS Deletion of the Chip gene led to an osteopenic phenotype and increased osteoclast formation. TRAF6, an adaptor protein that is a key regulator of NF-κB signaling and is critical for RANKL-induced osteoclastogenesis, was up-regulated in osteoclasts from Chip(-/-) mice. CHIP interacted with TRAF6 to promote TRAF6 ubiquitination and proteasome degradation. Further, CHIP inhibited p65 nuclear translocation, leading to the repression of TRAF6-mediated NF-κB transcription. CONCLUSION CHIP inhibits NF-κB signaling by promoting TRAF6 degradation and plays an important role in osteoclastogenesis and bone remodeling. These findings suggest that CHIP may be a novel therapeutic target in bone loss-associated disorders.
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Affiliation(s)
- Shan Li
- Tsinghua University School of Medicine, Beijing, China, and Rush University Medical Center, Chicago, Illinois
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20
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Kim S, Lee JC, Cho ES, Kwon J. COMP-Ang1 accelerates chondrocyte maturation by decreasing HO-1 expression. J Cell Biochem 2014; 114:2513-21. [PMID: 24030957 DOI: 10.1002/jcb.24596] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/12/2012] [Accepted: 05/14/2013] [Indexed: 01/08/2023]
Abstract
Endochondral ossification is essential for new bone formation and remodeling during the distraction stage. Endochondral ossification is attributed to chondrocyte maturation, which is induced by various factors, such as the cellular environment, gene transcription, and growth factor expression. Cartilage oligomeric matrix protein (COMP)-angiopoietin 1 (Ang1) is more soluble, stable, and potent than endogenous Ang1, and COMP-Ang1 treatment has osteogenic and angiogenic effects in an in vivo model of bone fracture healing. Although the osteogenic effects of COMP-Ang1 have been demonstrated, the precise mechanism by which COMP-Ang1 induces chondrocyte maturation and triggers endochondral ossification is not understood. Here, we investigated the possible mechanism by which COMP-Ang1 induces chondrocyte maturation. First, using a WST assay, we found that COMP-Ang1 is nontoxic in rat chondrocytes. Then, we isolated total RNA from COMP-Ang1-treated rat chondrocytes, and analyzed the decrease in chondrogenic gene expression and the increase in osteogenic gene expression using real-time RT-PCR. Gene and protein expression of heme oxygenase-1 (HO-1), which maintains chondrocytes in an immature stage, decreased in a dose-dependent manner upon COMP-Ang1 treatment. To clarify the relationship between HO-1 and COMP-Ang1 in chondrocyte maturation, we used cobalt protoporphyrin IX (CoPP IX), an HO-1 inducer, and tin protoporphyrin IX (SnPP-IX), an HO-1 inhibitor. Treatment with various combinations of CoPP IX, SnPP IX, and COMP-Ang1 confirmed that COMP-Ang1 accelerates chondrocyte maturation by reducing HO-1. In conclusion, our results suggest that COMP-Ang1 accelerates chondrocyte maturation by interacting with HO-1.
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Affiliation(s)
- Sokho Kim
- Department of Laboratory Animal Medicine, College of Veterinary Medicine, Institute of Oral Biosciences and BK21 Program, Chonbuk National University, Jeonju, 561-156, Republic of Korea
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21
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Shen J, Li J, Wang B, Jin H, Wang M, Zhang Y, Yang Y, Im HJ, O'Keefe R, Chen D. Deletion of the transforming growth factor β receptor type II gene in articular chondrocytes leads to a progressive osteoarthritis-like phenotype in mice. ACTA ACUST UNITED AC 2014; 65:3107-19. [PMID: 23982761 DOI: 10.1002/art.38122] [Citation(s) in RCA: 154] [Impact Index Per Article: 14.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/24/2012] [Accepted: 08/01/2013] [Indexed: 12/11/2022]
Abstract
OBJECTIVE While transforming growth factor β (TGFβ) signaling plays a critical role in chondrocyte metabolism, the TGFβ signaling pathways and target genes involved in cartilage homeostasis and the development of osteoarthritis (OA) remain unclear. Using an in vitro cell culture method and an in vivo mouse genetic approach, we undertook this study to investigate TGFβ signaling in chondrocytes and to determine whether Mmp13 and Adamts5 are critical downstream target genes of TGFβ signaling. METHODS TGFβ receptor type II (TGFβRII)-conditional knockout (KO) (TGFβRII(Col2ER)) mice were generated by breeding TGFβRII(flox/flox) mice with Col2-CreER-transgenic mice. Histologic, histomorphometric, and gene expression analyses were performed. In vitro TGFβ signaling studies were performed using chondrogenic rat chondrosarcoma cells. To determine whether Mmp13 and Adamts5 are critical downstream target genes of TGFβ signaling, TGFβRII/matrix metalloproteinase 13 (MMP-13)- and TGFβRII/ADAMTS-5-double-KO mice were generated and analyzed. RESULTS Inhibition of TGFβ signaling (deletion of the Tgfbr2 gene in chondrocytes) resulted in up-regulation of Runx2, Mmp13, and Adamts5 expression in articular cartilage tissue and progressive OA development in TGFβRII(Col2ER) mice. Deletion of the Mmp13 or Adamts5 gene significantly ameliorated the OA-like phenotype induced by the loss of TGFβ signaling. Treatment of TGFβRII(Col2ER) mice with an MMP-13 inhibitor also slowed OA progression. CONCLUSION Mmp13 and Adamts5 are critical downstream target genes involved in the TGFβ signaling pathway during the development of OA.
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Affiliation(s)
- Jie Shen
- Rush University Medical Center, Chicago, Illinois; University of Rochester School of Medicine, Rochester, New York
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22
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Kaneda A, Fujita T, Anai M, Yamamoto S, Nagae G, Morikawa M, Tsuji S, Oshima M, Miyazono K, Aburatani H. Activation of Bmp2-Smad1 signal and its regulation by coordinated alteration of H3K27 trimethylation in Ras-induced senescence. PLoS Genet 2011; 7:e1002359. [PMID: 22072987 PMCID: PMC3207904 DOI: 10.1371/journal.pgen.1002359] [Citation(s) in RCA: 57] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/02/2011] [Accepted: 09/11/2011] [Indexed: 02/06/2023] Open
Abstract
Cellular senescence involves epigenetic alteration, e.g. loss of H3K27me3 in Ink4a-Arf locus. Using mouse embryonic fibroblast (MEF), we here analyzed transcription and epigenetic alteration during Ras-induced senescence on genome-wide scale by chromatin immunoprecipitation (ChIP)-sequencing and microarray. Bmp2 was the most activated secreted factor with H3K4me3 gain and H3K27me3 loss, whereas H3K4me3 loss and de novo formation of H3K27me3 occurred inversely in repression of nine genes, including two BMP-SMAD inhibitors Smad6 and Noggin. DNA methylation alteration unlikely occurred. Ras-activated cells senesced with nuclear accumulation of phosphorylated SMAD1/5/8. Senescence was bypassed in Ras-activated cells when Bmp2/Smad1 signal was blocked by Bmp2 knockdown, Smad6 induction, or Noggin induction. Senescence was induced when recombinant BMP2 protein was added to Bmp2-knocked-down Ras-activated cells. Downstream Bmp2-Smad1 target genes were then analyzed genome-wide by ChIP-sequencing using anti-Smad1 antibody in MEF that was exposed to BMP2. Smad1 target sites were enriched nearby transcription start sites of genes, which significantly correlated to upregulation by BMP2 stimulation. While Smad6 was one of Smad1 target genes to be upregulated by BMP2 exposure, Smad6 repression in Ras-activated cells with increased enrichment of Ezh2 and gain of H3K27me3 suggested epigenetic disruption of negative feedback by Polycomb. Among Smad1 target genes that were upregulated in Ras-activated cells without increased repressive mark, Parvb was found to contribute to growth inhibition as Parvb knockdown lead to escape from senescence. It was revealed through genome-wide analyses in this study that Bmp2-Smad1 signal and its regulation by harmonized epigenomic alteration play an important role in Ras-induced senescence. To avoid becoming cancer cells, cells have a barrier system to block cellular proliferation by falling into irreversible growth arrest, so-called cellular senescence. For future strategy of cancer treatment, it is important to understand how cancer occurs, and investigation of underlying mechanism in senescence can lead to clarification of carcinogenesis mechanism. Epigenetic mechanism including DNA methylation and histone modification may be important to regulate gene expressions properly in senescence. Here, taking advantage of recent technical and methodological advance of genome-wide analyses, we examine epigenome and gene expression alteration in senescence induced by Ras oncogene. We identify that Bmp2-Smad1 signal is critical. We further examine downstream target genes of this critical signal on a genome-wide scale. We show dynamic and coordinated H3K27me3 alteration, e.g. activation of Bmp2 by loss of H3K27me3, repression of the signal inhibitors and the negative feedback loop by gain of H3K27me3, and selective activation of downstream target genes that may contribute to growth arrest. Our findings are helpful in understanding the importance of epigenetic regulation and a critical signal in the physiological barrier system against oncogenic transformation and the importance of disruption of BMP-SMAD signal in cancer, and they may provide an idea how cancer with Ras mutation occurs.
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Affiliation(s)
- Atsushi Kaneda
- Genome Science Division, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan.
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23
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Shu B, Zhang M, Xie R, Wang M, Jin H, Hou W, Tang D, Harris SE, Mishina Y, O'Keefe RJ, Hilton MJ, Wang Y, Chen D. BMP2, but not BMP4, is crucial for chondrocyte proliferation and maturation during endochondral bone development. J Cell Sci 2011; 124:3428-40. [PMID: 21984813 DOI: 10.1242/jcs.083659] [Citation(s) in RCA: 188] [Impact Index Per Article: 13.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022] Open
Abstract
The BMP signaling pathway has a crucial role in chondrocyte proliferation and maturation during endochondral bone development. To investigate the specific function of the Bmp2 and Bmp4 genes in growth plate chondrocytes during cartilage development, we generated chondrocyte-specific Bmp2 and Bmp4 conditional knockout (cKO) mice and Bmp2,Bmp4 double knockout (dKO) mice. We found that deletion of Bmp2 and Bmp4 genes or the Bmp2 gene alone results in a severe chondrodysplasia phenotype, whereas deletion of the Bmp4 gene alone produces a minor cartilage phenotype. Both dKO and Bmp2 cKO mice exhibit severe disorganization of chondrocytes within the growth plate region and display profound defects in chondrocyte proliferation, differentiation and apoptosis. To understand the mechanism by which BMP2 regulates these processes, we explored the specific relationship between BMP2 and Runx2, a key regulator of chondrocyte differentiation. We found that BMP2 induces Runx2 expression at both the transcriptional and post-transcriptional levels. BMP2 enhances Runx2 protein levels through inhibition of CDK4 and subsequent prevention of Runx2 ubiquitylation and proteasomal degradation. Our studies provide novel insights into the genetic control and molecular mechanism of BMP signaling during cartilage development.
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Affiliation(s)
- Bing Shu
- Department of Orthopaedics and Rehabilitation, Center for Musculoskeletal Research, University of Rochester School of Medicine, Rochester, NY 14642, USA
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Wang YK, Yu X, Cohen DM, Wozniak MA, Yang MT, Gao L, Eyckmans J, Chen CS. Bone morphogenetic protein-2-induced signaling and osteogenesis is regulated by cell shape, RhoA/ROCK, and cytoskeletal tension. Stem Cells Dev 2011; 21:1176-86. [PMID: 21967638 DOI: 10.1089/scd.2011.0293] [Citation(s) in RCA: 192] [Impact Index Per Article: 13.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/02/2023] Open
Abstract
Osteogenic differentiation of human mesenchymal stem cells (hMSCs) is classically thought to be mediated by different cytokines such as the bone morphogenetic proteins (BMPs). Here, we report that cell adhesion to extracellular matrix (ECM), and its effects on cell shape and cytoskeletal mechanics, regulates BMP-induced signaling and osteogenic differentiation of hMSCs. Using micropatterned substrates to progressively restrict cell spreading and flattening against ECM, we demonstrated that BMP-induced osteogenesis is progressively antagonized with decreased cell spreading. BMP triggered rapid and sustained RhoA/Rho-associated protein kinase (ROCK) activity and contractile tension only in spread cells, and this signaling was required for BMP-induced osteogenesis. Exploring the molecular basis for this effect, we found that restricting cell spreading, reducing ROCK signaling, or inhibiting cytoskeletal tension prevented BMP-induced SMA/mothers against decapentaplegic (SMAD)1 c-terminal phosphorylation, SMAD1 dimerization with SMAD4, and SMAD1 translocation into the nucleus. Together, these findings demonstrate the direct involvement of cell spreading and RhoA/ROCK-mediated cytoskeletal tension generation in BMP-induced signaling and early stages of in vitro osteogenesis, and highlight the essential interplay between biochemical and mechanical cues in stem cell differentiation.
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Affiliation(s)
- Yang-Kao Wang
- Department of Bioengineering, University of Pennsylvania, Philadelphia, PA 19104, USA
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25
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Lin M, Stewart DJ, Spitz MR, Hildebrandt MA, Lu C, Lin J, Gu J, Huang M, Lippman SM, Wu X. Genetic variations in the transforming growth factor-beta pathway as predictors of survival in advanced non-small cell lung cancer. Carcinogenesis 2011; 32:1050-6. [PMID: 21515830 PMCID: PMC3128559 DOI: 10.1093/carcin/bgr067] [Citation(s) in RCA: 27] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/16/2010] [Revised: 03/01/2011] [Accepted: 03/25/2011] [Indexed: 12/26/2022] Open
Abstract
The magnitude of benefit is variable for advanced non-small cell lung cancer (NSCLC) patients receiving platinum-based chemotherapy. The purpose of this study is to determine whether genetic variations in the transforming growth factor-beta (TGF-β) pathway are associated with clinical outcomes in NSCLC patients receiving first-line platinum-based chemotherapy. Five hundred and ninety-eight advanced-stage NSCLC patients who received first-line platinum-based chemotherapy with or without radiotherapy were recruited at the MD Anderson Cancer Center between 1995 and 2007. DNA from blood was genotyped for 227 single nucleotide polymorphisms (SNPs) in 23 TGF-β pathway-related genes to evaluate their associations with overall survival. In individual SNP analysis, 22 variants were significantly associated with overall survival, of which the strongest associations were found for BMP2:rs235756 [hazard ratio (HR) = 1.45; 95% confidence interval (CI), 1.11-1.90] and SMAD3:rs4776342 (HR = 1.25; 95% CI, 1.06-1.47). Fifteen and 18 genetic loci displayed treatment-specific associations for chemotherapy and chemoradiation, respectively, identifying a majority of the cases who would be predicted to respond favorably to a specific treatment regimen. BMP2:rs235753 and a haplotype in SMAD3 were associated with overall survival for both treatment modalities. Cumulative effect analysis showed that multiple risk genotypes had a significant dose-dependent effect on overall survival (P(trend) = 2.44 x 10(-15)). Survival tree analysis identified subgroups of patients with dramatically different median survival times of 45.39 versus 13.55 months and 18.02 versus 5.89 months for high- and low- risk populations when treated with chemoradiation and chemotherapy, respectively. These results suggest that genetic variations in the TGF-β pathway are potential predictors of overall survival in NSCLC patients treated with platinum-based chemotherapy with or without radiation.
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Affiliation(s)
| | - David J. Stewart
- Thoracic/Head and Neck Medical Oncology, University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA
| | | | | | - Charles Lu
- Thoracic/Head and Neck Medical Oncology, University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA
| | | | | | | | - Scott M. Lippman
- Thoracic/Head and Neck Medical Oncology, University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA
| | - Xifeng Wu
- To whom correspondence should be addressed. Tel: +1 713 745 2485; Fax: +1 713 792 4657;
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26
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Knezevic K, Bee T, Wilson NK, Janes ME, Kinston S, Polderdijk S, Kolb-Kokocinski A, Ottersbach K, Pencovich N, Groner Y, de Bruijn M, Göttgens B, Pimanda JE. A Runx1-Smad6 rheostat controls Runx1 activity during embryonic hematopoiesis. Mol Cell Biol 2011; 31:2817-26. [PMID: 21576367 PMCID: PMC3133398 DOI: 10.1128/mcb.01305-10] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/15/2010] [Revised: 01/05/2011] [Accepted: 04/22/2011] [Indexed: 12/12/2022] Open
Abstract
The oncogenic transcription factor Runx1 is required for the specification of definitive hematopoietic stem cells (HSC) in the developing embryo. The activity of this master regulator is tightly controlled during development. The transcription factors that upregulate the expression of Runx1 also upregulate the expression of Smad6, the inhibitory Smad, which controls Runx1 activity by targeting it to the proteasome. Here we show that Runx1, in conjunction with Fli1, Gata2, and Scl, directly regulates the expression of Smad6 in the aorta-gonad-mesonephros (AGM) region in the developing embryo, where HSCs originate. Runx1 regulates Smad6 activity via a novel upstream enhancer, and Runx1 null embryos show reduced Smad6 transcripts in the yolk-sac and c-Kit-positive fetal liver cells. By directly regulating the expression of Smad6, Runx1 sets up a functional rheostat to control its own activity. The perturbation of this rheostat, using a proteasomal inhibitor, results in an increase in Runx1 and Smad6 levels that can be directly attributed to increased Runx1 binding to tissue-specific regulatory elements of these genes. Taken together, we describe a scenario in which a key hematopoietic transcription factor controls its own expression levels by transcriptionally controlling its controller.
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Affiliation(s)
- Kathy Knezevic
- Lowy Cancer Research Centre and the Prince of Wales Clinical School, University of New South Wales, Sydney, New South Wales 2052, Australia
| | - Thomas Bee
- Lowy Cancer Research Centre and the Prince of Wales Clinical School, University of New South Wales, Sydney, New South Wales 2052, Australia
| | - Nicola K. Wilson
- Department of Haematology, Cambridge Institute for Medical Research, University of Cambridge, Cambridge CB2 0XY, United Kingdom
| | - Mary E. Janes
- Lowy Cancer Research Centre and the Prince of Wales Clinical School, University of New South Wales, Sydney, New South Wales 2052, Australia
| | - Sarah Kinston
- Department of Haematology, Cambridge Institute for Medical Research, University of Cambridge, Cambridge CB2 0XY, United Kingdom
| | - Stéphanie Polderdijk
- Department of Haematology, Cambridge Institute for Medical Research, University of Cambridge, Cambridge CB2 0XY, United Kingdom
| | | | - Katrin Ottersbach
- Department of Haematology, Cambridge Institute for Medical Research, University of Cambridge, Cambridge CB2 0XY, United Kingdom
| | - Niv Pencovich
- Department of Molecular Genetics, The Weizmann Institute of Science, Rehovot 76100, Israel
| | - Yoram Groner
- Department of Molecular Genetics, The Weizmann Institute of Science, Rehovot 76100, Israel
| | - Marella de Bruijn
- Weatherall Institute of Molecular Medicine, University of Oxford, Oxford OX3 9DS, United Kingdom
| | - Berthold Göttgens
- Department of Haematology, Cambridge Institute for Medical Research, University of Cambridge, Cambridge CB2 0XY, United Kingdom
| | - John E. Pimanda
- Lowy Cancer Research Centre and the Prince of Wales Clinical School, University of New South Wales, Sydney, New South Wales 2052, Australia
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27
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Fustino N, Rakheja D, Ateek CS, Neumann JC, Amatruda JF. Bone morphogenetic protein signalling activity distinguishes histological subsets of paediatric germ cell tumours. ACTA ACUST UNITED AC 2011; 34:e218-33. [PMID: 21696393 DOI: 10.1111/j.1365-2605.2011.01186.x] [Citation(s) in RCA: 42] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/04/2023]
Abstract
Germ cell tumours (GCTs) are cancers of the testis, ovary or extragonadal sites that occur in infants, children and adults. Testicular GCT is the most common cancer in young men aged 15-40 years. Abnormalities in developmental signalling pathways such as wnt/β-catenin, TGF-β/BMP and Hedgehog have been described in many childhood tumours. To date, however, the status of BMP signalling in GCTs has not been described. Herein, we examine BMP-SMAD signalling in a set of clinically-annotated paediatric GCTs. We find that BMP signalling activity is absent in undifferentiated tumours such as seminomas and dysgerminomas, but robustly present in most yolk sac tumours, a differentiated tumour type. Gene expression profiling of TGF-β/BMP pathway genes in germinomas and yolk sac tumours reveals a set of genes that distinguish the two tumour types. There is significant intertumoural heterogeneity between tumours of the same histological subclass, implying that the BMP pathway can be differentially regulated in individual tumours. Finally, through miRNA expression profiling, we identify differential regulation of a set of miRNAs predicted to target the TGF-β/BMP pathway at multiple sites. Taken together, these results suggest that the BMP signalling pathway may represent a new therapeutical target for childhood GCTs.
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Affiliation(s)
- N Fustino
- Division of Hematology-Oncology, Department of Pediatrics, University of Texas Southwestern Medical Center, Dallas, Texas 75390-8534, USA
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28
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Lu K, Li P, Zhang M, Xing G, Li X, Zhou W, Bartlam M, Zhang L, Rao Z, He F. Pivotal role of the C2 domain of the Smurf1 ubiquitin ligase in substrate selection. J Biol Chem 2011; 286:16861-70. [PMID: 21402695 PMCID: PMC3089529 DOI: 10.1074/jbc.m110.211979] [Citation(s) in RCA: 36] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/17/2010] [Revised: 03/04/2011] [Indexed: 01/17/2023] Open
Abstract
The C2-WW-HECT-type ubiquitin ligases Smurf1 and Smurf2 play a critical role in embryogenesis and adult bone homeostasis via regulation of bone morphogenetic protein, Wnt, and RhoA signaling pathways. The intramolecular interaction between C2 and HECT domains autoinhibits the ligase activity of Smurf2. However, the role of the Smurf1 C2 domain remains elusive. Here, we show that the C2-HECT autoinhibition mechanism is not observed in Smurf1, and instead its C2 domain functions in substrate selection. The Smurf1 C2 domain exerts a key role in localization to the plasma membrane and endows Smurf1 with differential activity toward RhoA versus Smad5 and Runx2. Crystal structure analysis reveals that the Smurf1 C2 domain possesses a typical anti-parallel β-sandwich fold. Examination of the sulfate-binding site analysis reveals two key lysine residues, Lys-28 and Lys-85, within the C2 domain that are important for Smurf1 localization at the plasma membrane, regulation on cell migration, and robust ligase activity toward RhoA, which further supports a Ca(2+)-independent localization mechanism for Smurf1. These findings demonstrate a previously unidentified role of the Smurf1 C2 domain in substrate selection and cellular localization.
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Affiliation(s)
- Kefeng Lu
- From the State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, Beijing 100850 and
| | - Ping Li
- the Tianjin Key Laboratory of Protein Sciences, College of Life Sciences, Nankai University, Tianjin 300071, China
| | - Minghua Zhang
- From the State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, Beijing 100850 and
| | - Guichun Xing
- From the State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, Beijing 100850 and
| | - Xin Li
- the Tianjin Key Laboratory of Protein Sciences, College of Life Sciences, Nankai University, Tianjin 300071, China
| | - Weihong Zhou
- the Tianjin Key Laboratory of Protein Sciences, College of Life Sciences, Nankai University, Tianjin 300071, China
| | - Mark Bartlam
- the Tianjin Key Laboratory of Protein Sciences, College of Life Sciences, Nankai University, Tianjin 300071, China
| | - Lingqiang Zhang
- From the State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, Beijing 100850 and
| | - Zihe Rao
- the Tianjin Key Laboratory of Protein Sciences, College of Life Sciences, Nankai University, Tianjin 300071, China
| | - Fuchu He
- From the State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, Beijing 100850 and
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29
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Laux DW, Febbo JA, Roman BL. Dynamic analysis of BMP-responsive smad activity in live zebrafish embryos. Dev Dyn 2011; 240:682-94. [PMID: 21337466 PMCID: PMC4287217 DOI: 10.1002/dvdy.22558] [Citation(s) in RCA: 43] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 12/23/2010] [Indexed: 11/06/2022] Open
Abstract
Bone morphogenetic proteins (BMPs) are critical players in development and disease, regulating such diverse processes as dorsoventral patterning, palate formation, and ossification. These ligands are classically considered to signal via BMP receptor-specific Smad proteins 1, 5, and 8. To determine the spatiotemporal pattern of Smad1/5/8 activity and thus canonical BMP signaling in the developing zebrafish embryo, we generated a transgenic line expressing EGFP under the control of a BMP-responsive element. EGFP is expressed in many established BMP signaling domains and is responsive to alterations in BMP type I receptor activity and smad1 and smad5 expression. This transgenic Smad1/5/8 reporter line will be useful for determining ligand and receptor requirements for specific domains of BMP activity, as well as for genetic and pharmacological screens aimed at identifying enhancers or suppressors of canonical BMP signaling.
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Affiliation(s)
- Derek W. Laux
- Department of Biological Sciences, University of Pittsburgh, Pittsburgh, PA 15260
| | - Jennifer A. Febbo
- Department of Biological Sciences, University of Pittsburgh, Pittsburgh, PA 15260
| | - Beth L. Roman
- Department of Biological Sciences, University of Pittsburgh, Pittsburgh, PA 15260
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30
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Developmental mechanisms in articular cartilage degradation in osteoarthritis. ARTHRITIS 2010; 2011:683970. [PMID: 22046522 PMCID: PMC3199933 DOI: 10.1155/2011/683970] [Citation(s) in RCA: 66] [Impact Index Per Article: 4.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Subscribe] [Scholar Register] [Received: 08/05/2010] [Accepted: 12/09/2010] [Indexed: 01/16/2023]
Abstract
Osteoarthritis is the most common arthritic condition, which involves progressive degeneration of articular cartilage. The most recent accomplishments have significantly advanced our understanding on the mechanisms of the disease development and progression. The most intriguing is the growing evidence indicating that extracellular matrix destruction in osteoarthritic articular cartilage resembles that in the hypertrophic zone of fetal growth plate during endochondral ossification. This suggests common regulatory mechanisms of matrix degradation in OA and in the development and can provide new approaches for the treatment of the disease by targeting reparation of chondrocyte phenotype.
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31
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Grusch M, Petz M, Metzner T, Öztürk D, Schneller D, Mikulits W. The crosstalk of RAS with the TGF-β family during carcinoma progression and its implications for targeted cancer therapy. Curr Cancer Drug Targets 2010; 10:849-57. [PMID: 20718708 PMCID: PMC3044462 DOI: 10.2174/156800910793357943] [Citation(s) in RCA: 44] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/28/2009] [Accepted: 08/18/2010] [Indexed: 11/22/2022]
Abstract
Both RAS and transforming growth factor (TGF)-β signaling cascades are central in tumorigenesis and show synergisms depending on tumor stage and tissue context. In this review we focus on the interaction of RAS subeffector proteins with signaling components of the TGF-β family including those of TGF-βs, activins and bone morphogenic proteins. Compelling evidence indicates that RAS signaling is essentially involved in the switch from tumor-suppressive to tumor-promoting functions of the TGF-β family leading to enhanced cancer growth and metastatic dissemination of primary tumors. Thus, the interface of these signaling cascades is considered as a promising target for the development of novel cancer therapeutics. The current pharmacological anti-cancer concepts combating the molecular cooperation between RAS and TGF-β family signaling during carcinoma progression are critically discussed.
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Affiliation(s)
- M. Grusch
- Department of Medicine I, Division, Institute of Cancer Research, Medical University of Vienna, Borschke-Gasse 8a, A-1090 Vienna, Austria
| | - M. Petz
- Department of Medicine I, Division, Institute of Cancer Research, Medical University of Vienna, Borschke-Gasse 8a, A-1090 Vienna, Austria
| | - T. Metzner
- Department of Medicine I, Division, Institute of Cancer Research, Medical University of Vienna, Borschke-Gasse 8a, A-1090 Vienna, Austria
| | - D. Öztürk
- Department of Medicine I, Division, Institute of Cancer Research, Medical University of Vienna, Borschke-Gasse 8a, A-1090 Vienna, Austria
| | - D. Schneller
- Department of Medicine I, Division, Institute of Cancer Research, Medical University of Vienna, Borschke-Gasse 8a, A-1090 Vienna, Austria
| | - W. Mikulits
- Department of Medicine I, Division, Institute of Cancer Research, Medical University of Vienna, Borschke-Gasse 8a, A-1090 Vienna, Austria
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32
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Ayrault O, Zhao H, Zindy F, Qu C, Sherr CJ, Roussel MF. Atoh1 inhibits neuronal differentiation and collaborates with Gli1 to generate medulloblastoma-initiating cells. Cancer Res 2010; 70:5618-27. [PMID: 20516124 PMCID: PMC2896438 DOI: 10.1158/0008-5472.can-09-3740] [Citation(s) in RCA: 86] [Impact Index Per Article: 5.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/04/2023]
Abstract
The morphogen and mitogen Sonic Hedgehog (Shh) activates a Gli1-dependent transcription program that drives proliferation of granule neuron progenitors (GNP) within the external germinal layer of the postnatally developing cerebellum. Medulloblastomas with mutations activating the Shh signaling pathway preferentially arise within the external germinal layer, and the tumor cells closely resemble GNPs. Atoh1/Math1, a basic helix-loop-helix transcription factor essential for GNP histogenesis, does not induce medulloblastomas when expressed in primary mouse GNPs that are explanted from the early postnatal cerebellum and transplanted back into the brains of naïve mice. However, enforced expression of Atoh1 in primary GNPs enhances the oncogenicity of cells overexpressing Gli1 by almost three orders of magnitude. Unlike Gli1, Atoh1 cannot support GNP proliferation in the absence of Shh signaling and does not govern expression of canonical cell cycle genes. Instead, Atoh1 maintains GNPs in a Shh-responsive state by regulating genes that trigger neuronal differentiation, including many expressed in response to bone morphogenic protein-4. Therefore, by targeting multiple genes regulating the differentiation state of GNPs, Atoh1 collaborates with the pro-proliferative Gli1-dependent transcriptional program to influence medulloblastoma development.
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Affiliation(s)
- Olivier Ayrault
- Department of Genetics and Tumor Cell Biology, 262 Danny Thomas Place, Memphis, Tennessee, 38105
| | - Haotian Zhao
- Department of Genetics and Tumor Cell Biology, 262 Danny Thomas Place, Memphis, Tennessee, 38105
| | - Frederique Zindy
- Department of Genetics and Tumor Cell Biology, 262 Danny Thomas Place, Memphis, Tennessee, 38105
| | - Chunxu Qu
- Department of Information Sciences, 262 Danny Thomas Place, Memphis, Tennessee, 38105
| | - Charles J. Sherr
- Department of Genetics and Tumor Cell Biology, 262 Danny Thomas Place, Memphis, Tennessee, 38105
- Howard Hughes Medical Institute at St. Jude Children’s Research Hospital, 262 Danny Thomas Place, Memphis, Tennessee, 38105
| | - Martine F. Roussel
- Department of Genetics and Tumor Cell Biology, 262 Danny Thomas Place, Memphis, Tennessee, 38105
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Abstract
The homologous to the E6-associated protein carboxyl terminus (HECT) domain E3 ubiquitin ligase Smurf1 is the first E3 ligase to be implicated in regulating bone cell function. The involvement of Smurf1 in multiple signaling pathways and pathological conditions is presently an area of extensive scientific interest. This review highlights recent works exploring Smurf-regulated biological processes in bone cells and highlights recent discoveries surrounding the regulatory mechanisms modulating its catalytic activity and substrate recognition capability. Moreover, we discuss the relevance of targeting the HECT E3s through the development of small-molecule inhibitors as an anticancer therapeutic strategy.
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Affiliation(s)
- Lianping Xing
- Department of Pathology, University of Rochester School of Medicine, Rochester, New York 14642
- Center for Musculoskeletal Research, University of Rochester School of Medicine, Rochester, New York 14642
| | - Ming Zhang
- Center for Musculoskeletal Research, University of Rochester School of Medicine, Rochester, New York 14642
- Department of Orthopaedics and Rehabilitation, University of Rochester School of Medicine, Rochester, New York 14642
| | - Di Chen
- Center for Musculoskeletal Research, University of Rochester School of Medicine, Rochester, New York 14642
- Department of Orthopaedics and Rehabilitation, University of Rochester School of Medicine, Rochester, New York 14642
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34
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Jiang Y, Chen LK, Zhu DC, Zhang GR, Guo C, Qi YY, Ouyang HW. The Inductive Effect of Bone Morphogenetic Protein-4 on Chondral-Lineage Differentiation and In Situ Cartilage Repair. Tissue Eng Part A 2010; 16:1621-32. [DOI: 10.1089/ten.tea.2009.0681] [Citation(s) in RCA: 51] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/13/2022] Open
Affiliation(s)
- YangZi Jiang
- Center for Stem Cell and Tissue Engineering, Zhejiang University, Hangzhou, China
| | - Long Kun Chen
- Center for Stem Cell and Tissue Engineering, Zhejiang University, Hangzhou, China
| | - Ding Cheng Zhu
- Center for Stem Cell and Tissue Engineering, Zhejiang University, Hangzhou, China
| | - Guo Rong Zhang
- Center for Stem Cell and Tissue Engineering, Zhejiang University, Hangzhou, China
| | - Chao Guo
- Center for Stem Cell and Tissue Engineering, Zhejiang University, Hangzhou, China
| | - Yi Ying Qi
- Center for Stem Cell and Tissue Engineering, Zhejiang University, Hangzhou, China
| | - Hong Wei Ouyang
- Center for Stem Cell and Tissue Engineering, Zhejiang University, Hangzhou, China
- Institute of Cell Biology, School of Medicine, Zhejiang University, Hangzhou, China
- Department of Orthopedic Surgery, Sir Run Run Shaw Hospital, Zhejiang University, Hangzhou, China
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35
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Park D, Estevez A, Riddle DL. Antagonistic Smad transcription factors control the dauer/non-dauer switch in C. elegans. Development 2010; 137:477-85. [PMID: 20081192 DOI: 10.1242/dev.043752] [Citation(s) in RCA: 34] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022]
Abstract
The C. elegans daf-8 gene encodes an R-Smad that is expressed in a subset of head neurons, the intestine, gonadal distal tip cells and the excretory cell. We found that DAF-8, which inhibits the DAF-3 Co-Smad, is associated with DAF-3 and the DAF-14 Smad in vivo and in vitro. Overexpression of daf-8 conferred a dauer-defective phenotype and suppressed constitutive dauer formation in daf-8 and daf-14 mutants. In contrast to mammalian systems described thus far, active DAF-3 drives a feedback regulatory loop that represses transcription of daf-7 (a TGFbeta ligand) and daf-8 by directly binding to their regulatory regions. Hence, DAF-8 and DAF-3 are mutually antagonistic. The feedback repression may reinforce the developmental switch by allowing DAF-3 to freely activate dauer transcription in target tissues, unless sufficiently inhibited by DAF-8 and DAF-14. In the adult, DAF-8 downregulates lag-2 expression in the distal tip cells, thus promoting germ line meiosis. This function does not involve DAF-3, thereby avoiding the feedback loop that functions in the dauer switch.
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Affiliation(s)
- Donha Park
- Michael Smith Laboratories, University of British Columbia, Vancouver, BC V6T1Z4, Canada
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36
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Wu WKK, Sung JJY, To KF, Yu L, Li HT, Li ZJ, Chu KM, Yu J, Cho CH. The host defense peptide LL-37 activates the tumor-suppressing bone morphogenetic protein signaling via inhibition of proteasome in gastric cancer cells. J Cell Physiol 2010; 223:178-86. [PMID: 20054823 DOI: 10.1002/jcp.22026] [Citation(s) in RCA: 46] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/13/2023]
Abstract
The human cathelicidin LL-37, a pleiotropic host defense peptide, is down-regulated in gastric adenocarcinomas. We therefore investigated whether this peptide suppresses gastric cancer growth. LL-37 lowered gastric cancer cell proliferation and delayed G(1)-S transition in vitro and inhibits the growth of gastric cancer xenograft in vivo. In this connection, LL-37 increased the tumor-suppressing bone morphogenetic protein (BMP) signaling, manifested as an increase in BMP4 expression and the subsequent Smad1/5 phosphorylation and the induction of p21(Waf1/Cip1). The anti-mitogenic effect, Smad1/5 phosphorylation, and p21(Waf1/Cip1) up-regulation induced by LL-37 were reversed by the knockdown of BMP receptor II. The activation of BMP signaling was paralleled by the inhibition of chymotrypsin-like and caspase-like activity of proteasome. In this regard, proteasome inhibitor MG-132 mimicked the effect of LL-37 by up-regulating BMP4 expression and Smad1/5 phosphorylation. Further analysis of clinical samples revealed that LL-37 and p21(Waf1/Cip1) mRNA expressions were both down-regulated in gastric cancer tissues and their expressions were positively correlated. Collectively, we describe for the first time that LL-37 inhibits gastric cancer cell proliferation through activation of BMP signaling via a proteasome-dependent mechanism. This unique biological activity may open up novel therapeutic avenue for the treatment of gastric cancer.
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Affiliation(s)
- William Ka Kei Wu
- Department of Medicine & Therapeutics, Faculty of Medicine, The Chinese University of Hong Kong, Hong Kong, China.
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Walker EC, McGregor NE, Poulton IJ, Solano M, Pompolo S, Fernandes TJ, Constable MJ, Nicholson GC, Zhang JG, Nicola NA, Gillespie MT, Martin TJ, Sims NA. Oncostatin M promotes bone formation independently of resorption when signaling through leukemia inhibitory factor receptor in mice. J Clin Invest 2010; 120:582-92. [PMID: 20051625 DOI: 10.1172/jci40568] [Citation(s) in RCA: 229] [Impact Index Per Article: 15.3] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/20/2009] [Accepted: 11/11/2009] [Indexed: 11/17/2022] Open
Abstract
Effective osteoporosis therapy requires agents that increase the amount and/or quality of bone. Any modification of osteoclast-mediated bone resorption by disease or drug treatment, however, elicits a parallel change in osteoblast-mediated bone formation because the processes are tightly coupled. Anabolic approaches now focus on uncoupling osteoblast action from osteoclast formation, for example, by inhibiting sclerostin, an inhibitor of bone formation that does not influence osteoclast differentiation. Here, we report that oncostatin M (OSM) is produced by osteoblasts and osteocytes in mouse bone and that it has distinct effects when acting through 2 different receptors, OSM receptor (OSMR) and leukemia inhibitory factor receptor (LIFR). Specifically, mouse OSM (mOSM) inhibited sclerostin production in a stromal cell line and in primary murine osteoblast cultures by acting through LIFR. In contrast, when acting through OSMR, mOSM stimulated RANKL production and osteoclast formation. A key role for OSMR in bone turnover was confirmed by the osteopetrotic phenotype of mice lacking OSMR. Furthermore, in contrast to the accepted model, in which mOSM acts only through OSMR, mOSM inhibited sclerostin expression in Osmr-/- osteoblasts and enhanced bone formation in vivo. These data reveal what we believe to be a novel pathway by which bone formation can be stimulated independently of bone resorption and provide new insights into OSMR and LIFR signaling that are relevant to other medical conditions, including cardiovascular and neurodegenerative diseases and cancer.
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Abstract
In recent years, much progress has been made in understanding the factors that regulate the gene expression program that underlies the induction, proliferation, differentiation, and maturation of osteoblasts. A large and growing number of transcription factors make important contributions to the precise control of osteoblast formation and function. It has become increasingly clear that these diverse transcription factors and the signals that regulate their activity cannot be viewed as discrete, separate signaling pathways. Rather, they form a highly interconnected, cooperative network that permits gene expression to be closely regulated. There has also been a substantial increase in our understanding of the mechanistic control of gene expression by cofactors such as acetyltransferases and histone deacetylases. The purpose of this review is to highlight recent progress in understanding the major transcription factors and epigenetic coregulators, including histone deacetylases and microRNAs, involved in osteoblastogenesis and the mechanisms that determine their functions as regulators of gene expression.
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Affiliation(s)
- Eric D Jensen
- Department of Diagnostic and Biological Sciences, School of Dentistry, University of Minnesota, Minneapolis, MN 55455
| | - Rajaram Gopalakrishnan
- Department of Diagnostic and Biological Sciences, School of Dentistry, University of Minnesota, Minneapolis, MN 55455
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Grcevic D, Jajic Z, Kovacic N, Lukic IK, Velagic V, Grubisic F, Ivcevic S, Marusic A. Peripheral blood expression profiles of bone morphogenetic proteins, tumor necrosis factor-superfamily molecules, and transcription factor Runx2 could be used as markers of the form of arthritis, disease activity, and therapeutic responsiveness. J Rheumatol 2009; 37:246-56. [PMID: 20008919 DOI: 10.3899/jrheum.090167] [Citation(s) in RCA: 42] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/25/2022]
Abstract
OBJECTIVE To assess whether different forms of arthritis and disease activity could be distinguished by peripheral blood expression profiles of bone-regulatory factors including tumor necrosis factor (TNF)-superfamily [TNF-related apoptosis-inducing ligand (TRAIL), the Fas ligand (FasL), and the ligand for herpesvirus entry mediator (LIGHT)] and bone morphogenetic protein (BMP)-family members (BMP-2, BMP-4, BMP-6) as well as osteoblast differentiation gene Runx2. METHODS Blood cells from healthy controls (n = 25) and patients at different disease stages with rheumatoid arthritis (RA; n = 49), osteoarthritis (OA; n = 17), or spondyloarthritis, including ankylosing spondylitis (AS; n = 27) or psoriatic arthritis (PsA; n = 23), were processed for quantitative polymerase chain reaction. Gene expression was assessed in comparison with control samples, correlated with clinical data of different forms of arthritis, and analyzed for discriminative efficacy between groups by receiver-operation characteristic (ROC) curves. Results were confirmed on diagnostic RA (n = 5) and AS (n = 8) samples. RESULTS BMP-4, BMP-6, and Runx2 expressions were significantly decreased in patients with RA and OA versus controls. Patients with RA also had decreased FasL and LIGHT expression, while patients with AS had increased Runx2 expression. Negative correlation with disease activity was found for BMP-4, FasL, and Runx2 in RA and for Runx2 in PsA, while positive correlation was found for BMP-4 in PsA. Gene expression was higher in the therapy-resistant form of AS (for BMP-4, LIGHT, and Runx2) and in methotrexate-treated patients in RA (for BMP-2 and LIGHT). ROC curve analysis confirmed discrimination between groups, particularly decreased LIGHT and Runx2 for RA and increased Runx2 for AS. CONCLUSION Our study identified BMP and Runx2 as possible biomarkers of bone metabolism in several forms of arthritis, while lower FasL and LIGHT were associated with RA. Correlation between gene expression and disease activity may be clinically useful in assessing therapeutic effectiveness and disease monitoring.
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Affiliation(s)
- Danka Grcevic
- Department of Physiology and Immunology, University of Zagreb School of Medicine, Zagreb, Croatia.
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40
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Abstract
Transforming growth factor beta (TGFbeta) pathways are implicated in metazoan development, adult homeostasis and disease. TGFbeta ligands signal via receptor serine/threonine kinases that phosphorylate, and activate, intracellular Smad effectors as well as other signaling proteins. Oligomeric Smad complexes associate with chromatin and regulate transcription, defining the biological response of a cell to TGFbeta family members. Signaling is modulated by negative-feedback regulation via inhibitory Smads. We review here the mechanisms of TGFbeta signal transduction in metazoans and emphasize events crucial for embryonic development.
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41
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Jonason JH, Xiao G, Zhang M, Xing L, Chen D. Post-translational Regulation of Runx2 in Bone and Cartilage. J Dent Res 2009; 88:693-703. [PMID: 19734454 DOI: 10.1177/0022034509341629] [Citation(s) in RCA: 124] [Impact Index Per Article: 7.8] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/02/2023] Open
Abstract
The Runx2 gene product is essential for mammalian bone development. In humans, Runx2 haploinsufficiency results in cleidocranial dysplasia, a skeletal disorder characterized by bone and dental abnormalities. At the molecular level, Runx2 acts as a transcription factor for genes expressed in hypertrophic chondrocytes and osteoblasts. Runx2 gene expression and protein function are regulated on multiple levels, including transcription, translation, and post-translational modification. Furthermore, Runx2 is involved in numerous protein-protein interactions, most of which either activate or repress transcription of target genes. In this review, we discuss expression of Runx2 during development as well as the post-translational regulation of Runx2 through modification by phosphorylation, ubiquitination, and acetylation.
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Affiliation(s)
- J H Jonason
- Department of Orthopaedics, Center for Musculoskeletal Research, University of Rochester School of Medicine, 601 Elmwood Avenue, Box 665, Rochester, NY 14642, USA
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Blitz IL, Cho KWY. Finding partners: how BMPs select their targets. Dev Dyn 2009; 238:1321-31. [PMID: 19441058 DOI: 10.1002/dvdy.21984] [Citation(s) in RCA: 38] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/03/2023] Open
Abstract
The bone morphogenetic protein (BMP) signaling pathway is a conserved and evolutionarily ancient regulatory module affecting a large variety of cellular behaviors. The evolutionary flexibility in using BMP responses presumably arose by co-option of a canonical BMP signaling cascade to regulate the transcription of diverse batteries of target genes. This begs the question of how seemingly interchangeable BMP signaling components elicit widely different outputs in different cell types, an important issue in the context of understanding how BMP signaling integrates with gene regulatory networks to control development. Because a molecular understanding of how BMP signaling activates different batteries of target genes is an essential prerequisite to comprehending the roles of BMPs in regulating cellular responses, here we review the current knowledge of how BMP-regulated target genes are selected by the signal transduction machinery. We highlight recent studies suggesting the evolutionary conservation of BMP target gene regulation signaling by Schnurri family zinc finger proteins. Developmental Dynamics 238:1321-1331, 2009. (c) 2009 Wiley-Liss, Inc.
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Affiliation(s)
- Ira L Blitz
- Department of Developmental and Cell Biology and the Developmental Biology Center, University of California, Irvine, California, USA.
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Cruzat F, Henriquez B, Villagra A, Hepp M, Lian JB, van Wijnen AJ, Stein JL, Imbalzano AN, Stein GS, Montecino M. SWI/SNF-independent nuclease hypersensitivity and an increased level of histone acetylation at the P1 promoter accompany active transcription of the bone master gene Runx2. Biochemistry 2009; 48:7287-95. [PMID: 19545172 PMCID: PMC2760825 DOI: 10.1021/bi9004792] [Citation(s) in RCA: 23] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/26/2023]
Abstract
The Runx2 transcription factor is essential for skeletal development as it regulates expression of several key bone-related genes. Multiple lines of evidence indicate that expression of the Runx2/p57 isoform in osteoblasts is controlled by the distal P1 promoter. Alterations of chromatin structure are often associated with transcription and can be mediated by members of the SWI/SNF family of chromatin remodeling complexes, or by transcriptional coactivators that possess enzymatic activities that covalently modify structural components of the chromatin. Here, we report that a specific chromatin remodeling process at the proximal region (residues -400 to 35) of the Runx2 gene P1 promoter accompanies transcriptional activity in osteoblasts. This altered chromatin organization is reflected by the presence of two DNase I hypersensitive sites that span key regulatory elements for Runx2/p57 transcription. Chromatin remodeling and transcription of the Runx2 gene are associated with elevated levels of histone acetylation at the P1 promoter region and binding of active RNA polymerase II and are independent of the activity of the SWI/SNF chromatin remodeling complex. Changes in chromatin organization at the P1 promoter are stimulated during differentiation of C2C12 mesenchymal cells to the osteoblastic lineage by treatment with BMP2. Together, our results support a model in which changes in chromatin organization occur at very early stages of mesenchymal differentiation to facilitate subsequent expression of the Runx2/p57 isoform in osteoblastic cells.
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Affiliation(s)
- Fernando Cruzat
- Departamento de Bioquimica y Biologia Molecular, Facultad de Ciencias Biologicas, Universidad de Concepcion, Concepcion, Chile
| | - Berta Henriquez
- Departamento de Bioquimica y Biologia Molecular, Facultad de Ciencias Biologicas, Universidad de Concepcion, Concepcion, Chile
| | - Alejandro Villagra
- Departamento de Bioquimica y Biologia Molecular, Facultad de Ciencias Biologicas, Universidad de Concepcion, Concepcion, Chile
| | - Matias Hepp
- Departamento de Bioquimica y Biologia Molecular, Facultad de Ciencias Biologicas, Universidad de Concepcion, Concepcion, Chile
| | - Jane B. Lian
- Department of Cell Biology, University of Massachusetts Medical School, Worcester, MA
| | - Andre J. van Wijnen
- Department of Cell Biology, University of Massachusetts Medical School, Worcester, MA
| | - Janet L. Stein
- Department of Cell Biology, University of Massachusetts Medical School, Worcester, MA
| | - Anthony N. Imbalzano
- Department of Cell Biology, University of Massachusetts Medical School, Worcester, MA
| | - Gary S. Stein
- Department of Cell Biology, University of Massachusetts Medical School, Worcester, MA
| | - Martin Montecino
- Departamento de Bioquimica y Biologia Molecular, Facultad de Ciencias Biologicas, Universidad de Concepcion, Concepcion, Chile
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Chen YL, Liu B, Zhou ZN, Hu RY, Fei C, Xie ZH, Ding X. Smad6 inhibits the transcriptional activity of Tbx6 by mediating its degradation. J Biol Chem 2009; 284:23481-90. [PMID: 19561075 DOI: 10.1074/jbc.m109.007864] [Citation(s) in RCA: 23] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/10/2023] Open
Abstract
Members of the bone morphogenetic protein (BMP) and T-box gene families play several critical roles in the early embryonic development and tissue homeostasis. Although BMP proteins are the upstream regulators of T-box genes, few studies have investigated the molecular mechanisms between these two protein families. Here, we report that Tbx6 interacts directly with Smad6, an inhibitory Smad that antagonizes the BMP signal. This interaction is mediated through the Mad homology 2 (MH2) domain of Smad6 and residues 90-180 of Tbx6. We demonstrate that Smad6 facilitates the degradation of Tbx6 protein through recruitment of Smurf1, a ubiquitin E3 ligase. Consequently, Smad6 reduces Tbx6-mediated Myf-5 gene activation. Furthermore, specific knockdown of endogenous Smad6 and Smurf1 by small interfering RNA increases the protein levels of Tbx6 and enhance the expression of Tbx6 target genes. Collectively, these findings reveal that Smad6 serves as a critical mediator of BMP signal via a functional interaction with Tbx6, thus regulating the activation of Tbx6 downstream genes during cell differentiation.
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Affiliation(s)
- Yue-Lei Chen
- Laboratory of Molecular Cell Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Graduate School of the Chinese Academy of Sciences, Shanghai 200031, China
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Abstract
Transforming growth factor (TGF)-β is a pleiotropic cytokine regulating a variety of cellular processes such as cell growth, differentiation, apoptosis, migration, cell adhesion, and immune response. In the well-understood classical TGF-β signaling pathway, TGF-β activates Smad signalling via its two cell surface receptors such as TβRII and ALK5/TβRI, leading to Smad-mediated transcriptional regulation. In addition, TGF-β may also activate other signaling pathways like mitogen-activated protein kinase, PI3K, etc. The signaling of TGF-β is finely regulated at different levels. Inhibitory Smads, including Smad6 and Smad7, are key regulators of TGF-β/bone morphogenetic protein (BMP) signaling by negative feedback loops. They can form stable complexes with activated type I receptors and thereby blocking the phosphorylation of R-Smads, or recruit ubiquitin E3 ligases, such as Smurf1/2, resulting in the ubiquitination and degradation of the activated type I receptors. Besides, these inhibitory Smad proteins also inhibit TGF-β/BMP signaling in the nucleus by interacting with transcriptional repressors, such as histone deacetylases, Hoxc-8, and CtBP, or disrupting the formation of the TGF-β-induced functional Smad-DNA complexes. Smad7 is in turn regulated by different stimuli, including TGF-β, IFN-γ, TNF-α as well as ultraviolet and TPA, and mediates the crosstalk between TGF-β and other signaling pathways. Deregulation of Smad7 expression has been associated with various human diseases, such as tissue fibrosis, inflammatory disease as well as carcinogenesis. Overexpression of Smad7 has been shown to antagonize TGF-β-mediated fibrosis, carcinogenesis, and inflammation, suggesting a therapeutic potential of Smad7 to treat these diseases.
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Affiliation(s)
- Xiaohua Yan
- State Key Laboratory of Biomembrane and Membrane Biotechnology, Department of Biological Sciences and Biotechnology, Tsinghua University, Beijing 100084, China
| | - Ziying Liu
- State Key Laboratory of Biomembrane and Membrane Biotechnology, Department of Biological Sciences and Biotechnology, Tsinghua University, Beijing 100084, China
| | - Yeguang Chen
- State Key Laboratory of Biomembrane and Membrane Biotechnology, Department of Biological Sciences and Biotechnology, Tsinghua University, Beijing 100084, China
- Correspondence address. Tel: +86-10-62795184; Fax: +86-10-62794376; E-mail:
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Cardiotrophin-1 is an osteoclast-derived stimulus of bone formation required for normal bone remodeling. J Bone Miner Res 2008; 23:2025-32. [PMID: 18665789 DOI: 10.1359/jbmr.080706] [Citation(s) in RCA: 144] [Impact Index Per Article: 8.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 11/18/2022]
Abstract
Cardiotrophin (CT-1) signals through gp130 and the LIF receptor (LIFR) and plays a major role in cardiac, neurological, and liver biology. We report here that CT-1 is also expressed within bone in osteoclasts and that CT-1 is capable of increasing osteoblast activity and mineralization both in vitro and in vivo. Furthermore, CT-1 stimulated CAAT/enhancer-binding protein-delta (C/EBP delta) expression and runt-related transcription factor 2 (runx2) activation. In neonate CT-1(-/-) mice, we detected low bone mass associated with reduced osteoblasts and many large osteoclasts, but increased cartilage remnants within the bone, suggesting impaired resorption. Cultured bone marrow (BM) from CT-1(-/-) mice generated many oversized osteoclasts and mineralized poorly compared with wildtype BM. As the CT-1(-/-) mice aged, the reduced osteoblast surface (ObS/BS) was no longer detected, but impaired bone resorption continued resulting in an osteopetrotic phenotype in adult bone. CT-1 may now be classed as an essential osteoclast-derived stimulus of both bone formation and resorption.
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47
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Jäger M, Fischer J, Dohrn W, Li X, Ayers DC, Czibere A, Prall WC, Lensing-Höhn S, Krauspe R. Dexamethasone modulates BMP-2 effects on mesenchymal stem cells in vitro. J Orthop Res 2008; 26:1440-8. [PMID: 18404732 DOI: 10.1002/jor.20565] [Citation(s) in RCA: 62] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/04/2023]
Abstract
Dexamethasone/ascorbic acid/glycerolphosphate (DAG) and bone morphogenic protein (BMP)-2 are potent agents in cell proliferation and differentiation pathways. This study investigates the in vitro interactions between dexamethasone and BMP-2 for an osteoblastic differentiation of mesenchymal stem cells (MSCs). Bone marrow-derived human MSCs were cultured with DAG (group A), BMP-2 + DAG (group B), and DAG + BMP-2 combined with a porous collagen I/III scaffold (group C). RT-PCR, ELISA, immuncytochemical stainings and flow cytometry analysis served to evaluate the osteogenic-promoting potency of each of the above conditions in terms of cell morphology/viability, antigen presentation, and gene expression. DAG induced collagen I secretion from MSCs, which was further increased by the combination of DAG + BMP-2. In comparison, the collagen scaffold and the control samples showed no significant influence on collagen I secretion of MSCs. DAG stimulation of MSCs led also to a steady but not significant increase of BMP-2 level. A DAG and more, a DAG + BMP-2, stimulation increased the number of mesenchymal cells (CD105+/CD73+). All samples showed mRNA of ALP, osteopontin, Runx2, Twist 1 and 2, Notch-1/2, osteonectin, osteocalcin, BSP, and collagen-A1 after 28 days of in vitro culture. Culture media of all samples showed a decrease in Ca(2+) and PO(4) (2-) concentration, whereas a collagen-I-peak only occurred at day 28 in DAG- and DAG + BMP-2-stimulated bone marrow cells. In conclusion, BMP-2 enhances DAG-induced osteogenic differentiation in mesenchymal bone marrow cells. Both agents interact in various ways and can modify osteoblastic bone formation.
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Affiliation(s)
- Marcus Jäger
- Research Laboratory for Regenerative Medicine and Biomaterials, Department of Orthopaedics, Heinrich-Heine University Medical School, Moorenstr. 5, D-40225 Düsseldorf, Germany.
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Pan Q, Yu Y, Chen Q, Li C, Wu H, Wan Y, Ma J, Sun F. Sox9, a key transcription factor of bone morphogenetic protein-2-induced chondrogenesis, is activated through BMP pathway and a CCAAT box in the proximal promoter. J Cell Physiol 2008; 217:228-41. [PMID: 18506848 DOI: 10.1002/jcp.21496] [Citation(s) in RCA: 115] [Impact Index Per Article: 6.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/09/2022]
Abstract
Mouse embryonic fibroblasts (MEFs) can be differentiated into fully functional chondrocytes in response to bone morphogenetic protein-2 (BMP-2). The expression of Sox9, a critical transcription factor for the multiple steps of chondrogenesis, has been reported to be upregulated during this process. But the molecular mechanisms by which BMP-2 promotes chondrogenesis still remain largely unknown. The aim of the present study was therefore to investigate the underlying mechanism. In the MEFs, BMP-2 efficiently induced Sox9 expression along with chondrogenic differentiation in a time- and dose-dependent manner. SB203580, a specific inhibitor for p38 pathway, blocked BMP-2-induced chondrogenic differentiation as well as Sox9 expression and its transactivation of downstream genes. Forced expression of Smad6, a natural antagonist for BMP/Smad pathway, only inhibited Sox9 protein function without rendering any effects on its mRNA expression. A CCAAT box was identified in Sox9 promoter as the cis-elements responsible for BMP-2 stimulation. This study provides insight into the mechanisms underlying BMP-2-regulated Sox9 expression and activity in MEFs, and suggests differential roles of BMP-2/p38 and BMP-2/Smad pathways in modulating the function of Sox9 during chondrogenesis.
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Affiliation(s)
- Qiuhui Pan
- Medical Research Center, the Second Affiliated Hospital, Sun Yat-sen University, Guangzhou City, Guangdong Province, PR China
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49
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Liu F, Bloch N, Bhushan KR, De Grand AM, Tanaka E, Solazzo S, Mertyna PM, Goldberg N, Frangioni JV, Lenkinski RE. Humoral Bone Morphogenetic Protein 2 Is Sufficient for Inducing Breast Cancer Microcalcification. Mol Imaging 2008. [DOI: 10.2310/7290.2008.00018a] [Citation(s) in RCA: 24] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/18/2022] Open
Affiliation(s)
- Fangbing Liu
- From the Division of Hematology/Oncology and Department of Radiology, Beth Israel Deaconess Medical Center/Harvard Medical School, Boston, MA
| | - Nathalie Bloch
- From the Division of Hematology/Oncology and Department of Radiology, Beth Israel Deaconess Medical Center/Harvard Medical School, Boston, MA
| | - Kumar R. Bhushan
- From the Division of Hematology/Oncology and Department of Radiology, Beth Israel Deaconess Medical Center/Harvard Medical School, Boston, MA
| | - Alec M. De Grand
- From the Division of Hematology/Oncology and Department of Radiology, Beth Israel Deaconess Medical Center/Harvard Medical School, Boston, MA
| | - Eiichi Tanaka
- From the Division of Hematology/Oncology and Department of Radiology, Beth Israel Deaconess Medical Center/Harvard Medical School, Boston, MA
| | - Stephanie Solazzo
- From the Division of Hematology/Oncology and Department of Radiology, Beth Israel Deaconess Medical Center/Harvard Medical School, Boston, MA
| | - Pawel M. Mertyna
- From the Division of Hematology/Oncology and Department of Radiology, Beth Israel Deaconess Medical Center/Harvard Medical School, Boston, MA
| | - Nahum Goldberg
- From the Division of Hematology/Oncology and Department of Radiology, Beth Israel Deaconess Medical Center/Harvard Medical School, Boston, MA
| | - John V. Frangioni
- From the Division of Hematology/Oncology and Department of Radiology, Beth Israel Deaconess Medical Center/Harvard Medical School, Boston, MA
| | - Robert E. Lenkinski
- From the Division of Hematology/Oncology and Department of Radiology, Beth Israel Deaconess Medical Center/Harvard Medical School, Boston, MA
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50
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Liu F, Bloch N, Bhushan KR, De Grand AM, Tanaka E, Solazzo S, Mertyna PM, Goldberg N, Frangioni JV, Lenkinski RE. Humoral bone morphogenetic protein 2 is sufficient for inducing breast cancer microcalcification. Mol Imaging 2008; 7:175-86. [PMID: 19123988 PMCID: PMC2768041] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 05/27/2023] Open
Abstract
Microcalcifications are an important diagnostic marker for breast cancer on mammograms, yet the mechanism of their formation is poorly understood. Indeed, there is presently no short-latency, high-yield, syngeneic rodent model of the process. Bone morphogenetic protein 2 (BMP-2) is a key mediator of physiologic bone formation and pathologic vasculature calcification, but its role in breast cancer microcalcification is unknown. In this study, R3230 rat breast tumors were adapted to cell culture, transduced with adenoviral BMP-2, and inoculated into a syngeneic host. Tumor growth and calcium salt deposition were quantified in living animals over time using micro-computed tomography and probed chemically using near-infrared fluorescence. Plasma BMP-2 levels were quantified over time by enzyme-linked immunosorbent assay. Within 3 weeks, 100% of the breast tumors developed microcalcifications, which were absent from all normal tissues. Importantly, when two tumors were initiated in a single host, the ipsilateral tumor expressing BMP-2 was able to induce microcalcification in the contralateral tumor that was not expressing BMP-2, suggesting that BMP-2 can act humorally. Taken together, we describe the first reproducible rodent model of breast cancer microcalcification, prove that BMP-2 expression is sufficient for initiating the process, and lay the foundation for a new generation of targeted diagnostic agents.
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Affiliation(s)
- Fangbing Liu
- Division of Hematology/Oncology, Beth Israel Deaconess Medical Center/Harvard Medical School, Boston, MA 02215
| | - Nathalie Bloch
- Department of Radiology, Beth Israel Deaconess Medical Center/Harvard Medical School, Boston, MA 02215
| | - Kumar R. Bhushan
- Division of Hematology/Oncology, Beth Israel Deaconess Medical Center/Harvard Medical School, Boston, MA 02215
| | - Alec M. De Grand
- Department of Radiology, Beth Israel Deaconess Medical Center/Harvard Medical School, Boston, MA 02215
| | - Eiichi Tanaka
- Division of Hematology/Oncology, Beth Israel Deaconess Medical Center/Harvard Medical School, Boston, MA 02215
| | - Stephanie Solazzo
- Department of Radiology, Beth Israel Deaconess Medical Center/Harvard Medical School, Boston, MA 02215
| | - Pawel M. Mertyna
- Department of Radiology, Beth Israel Deaconess Medical Center/Harvard Medical School, Boston, MA 02215
| | - Nahum Goldberg
- Department of Radiology, Beth Israel Deaconess Medical Center/Harvard Medical School, Boston, MA 02215
| | - John V. Frangioni
- Division of Hematology/Oncology, Beth Israel Deaconess Medical Center/Harvard Medical School, Boston, MA 02215
- Department of Radiology, Beth Israel Deaconess Medical Center/Harvard Medical School, Boston, MA 02215
| | - Robert E. Lenkinski
- Department of Radiology, Beth Israel Deaconess Medical Center/Harvard Medical School, Boston, MA 02215
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