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Hong M, Hong S, Song JM. 3D Bioprinted Multidrug Resistance (MDR)-Dependent Tumor Spheroids. ACS APPLIED MATERIALS & INTERFACES 2025; 17:7377-7394. [PMID: 39853257 DOI: 10.1021/acsami.4c19291] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/26/2025]
Abstract
Multidrug resistance (MDR) refers to the ability of cancer cells to resist various anticancer drugs and release them from the cells. This phenomenon is widely recognized as a significant barrier that must be overcome in chemotherapy. MDR varies depending on the number and expression level of the ATP-binding cassette transporter (ABC transporter), which is expressed differently in various cancer cells. Therefore, the dose of anticancer drugs should be adjusted according to the extent of MDR. The demand for drug screening that considers the differences in MDR is increasing in the process of drug discovery. In this study, three types of tumor spheroids were fabricated from HeLa (MRP1-/BCRP-), HepG2 (MRP1+/BCRP-), and A549 cells (MRP1+/BCRP+) using three-dimensional (3D) bioprinting. The fabricated tumor spheroids maintained their own MDR phenotypes. The EC50 values of doxorubicin (DOX) against the three tumor spheroids were more than 2-fold higher than those against the 2D cells. In addition, the EC50 value of DOX against tumor spheroids was proportional to the number of ABC transporters. The EC50 value of DOX against A549 tumor spheroids had the largest value of 9.5 μM among the three spheroids. In addition, the EC50 values of DOX against HepG2 and A549 tumor spheroids were remarkably reduced when they were treated with ABC transporter inhibitors, such as MK-571 against MRP1 and/or NOV against BCRP. These results demonstrate the successful construction of a 3D bioprinting-based screening platform to quantitatively evaluate the anticancer efficacy of chemodrugs, considering the MDR of cancer cells.
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Affiliation(s)
- Minki Hong
- College of Pharmacy, Seoul National University, Seoul 08826, South Korea
| | - Sera Hong
- College of Pharmacy, Seoul National University, Seoul 08826, South Korea
| | - Joon Myong Song
- College of Pharmacy, Seoul National University, Seoul 08826, South Korea
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Gonzalez GC, Li CM, Pasolini I, Pete SI, Verheyen C, Vignolo SM, De Toni T, Stock AA, Tomei AA. High-Yield Generation of Glucose-Responsive Pseudoislets From Murine Insulinoma Cells for In Vitro Studies and Longitudinal Monitoring of Graft Survival In Vivo. Cell Transplant 2025; 34:9636897251315123. [PMID: 39881520 PMCID: PMC11780636 DOI: 10.1177/09636897251315123] [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: 11/01/2024] [Revised: 12/14/2024] [Accepted: 01/06/2025] [Indexed: 01/31/2025] Open
Abstract
Compared to primary pancreatic islets, insulinoma cell-derived 3D pseudoislets offer a more accessible, consistent, renewable, and widely applicable model system for optimization and mechanistic studies in type 1 diabetes (T1D). Here, we report a simple and efficient method for generating 3D pseudoislets from MIN6 and NIT-1 murine insulinoma cells. These pseudoislets are homogeneous in size and morphology (~150 µm), exhibit functional glucose-stimulated insulin secretion (GSIS) up to 18 days (NIT-1) enabling long-term studies, are produced in high yield [>35,000 Islet Equivalence from 30 ml culture], and are suitable for both in vitro and in vivo studies, including for encapsulation studies. To enable non-invasive longitudinal monitoring of graft survival in vivo, we transduced NIT-1 cells with green fluorescent protein-luciferase and confirmed comparable morphology, viability, and GSIS to untransduced cells in vitro. After subcutaneous implantation, we show capability to monitor graft survival in immunodeficient mice, recurrence of autoimmunity in non-obese diabetic mice, and allorejection in C57BL/6 mice. Overall, this platform provides an accessible protocol for generating high yields of 3D pseudoislets and non-invasive longitudinal monitoring of graft survival in different models offer advantages over primary islets for optimization and mechanistic studies of β cell biology, drug discovery, T1D pathogenesis and prevention, and β cell transplantation.
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Affiliation(s)
- Grisell C. Gonzalez
- Diabetes Research Institute, University of Miami Miller School of Medicine, Miami, FL, USA
| | - Chris M. Li
- Diabetes Research Institute, University of Miami Miller School of Medicine, Miami, FL, USA
- Department of Microbiology and Immunology, University of Miami Miller School of Medicine, Miami, FL, USA
| | - Ilaria Pasolini
- Diabetes Research Institute, University of Miami Miller School of Medicine, Miami, FL, USA
- Department of Biomedical Engineering, University of Miami, Miami, FL, USA
| | - Sophia I. Pete
- Diabetes Research Institute, University of Miami Miller School of Medicine, Miami, FL, USA
- Department of Biomedical Engineering, University of Miami, Miami, FL, USA
| | - Connor Verheyen
- Diabetes Research Institute, University of Miami Miller School of Medicine, Miami, FL, USA
- Department of Biomedical Engineering, University of Miami, Miami, FL, USA
| | - Sofia M. Vignolo
- Diabetes Research Institute, University of Miami Miller School of Medicine, Miami, FL, USA
- Department of Biomedical Engineering, University of Miami, Miami, FL, USA
| | - Teresa De Toni
- Diabetes Research Institute, University of Miami Miller School of Medicine, Miami, FL, USA
- Department of Biomedical Engineering, University of Miami, Miami, FL, USA
| | - Aaron A. Stock
- Diabetes Research Institute, University of Miami Miller School of Medicine, Miami, FL, USA
- Department of Biomedical Engineering, University of Miami, Miami, FL, USA
| | - Alice A. Tomei
- Diabetes Research Institute, University of Miami Miller School of Medicine, Miami, FL, USA
- Department of Microbiology and Immunology, University of Miami Miller School of Medicine, Miami, FL, USA
- Department of Biomedical Engineering, University of Miami, Miami, FL, USA
- Department of Surgery, University of Miami Miller School of Medicine, Miami, FL, USA
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Yadav P, Singh S, Jaiswal S, Kumar R. Synthetic and natural polymer hydrogels: A review of 3D spheroids and drug delivery. Int J Biol Macromol 2024; 280:136126. [PMID: 39349080 DOI: 10.1016/j.ijbiomac.2024.136126] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/22/2024] [Revised: 09/25/2024] [Accepted: 09/27/2024] [Indexed: 10/02/2024]
Abstract
This review centers on the synthesis and characterization of both natural and synthetic hydrogels, highlighting their diverse applications across various fields. We will delve into the evolution of hydrogels, focusing on the importance of polysaccharide-based and synthetic variants, which have been particularly chosen for 3D spheroid development in cancer research and drug delivery. A detailed background on the research and specific methodologies, including the in-situ free radical polymerization used for synthesizing these hydrogels, will be extensively discussed. Additionally, the review will explore various applications of these hydrogels, such as their self-healing properties, swelling ratios, pH responsiveness, and cell viability. A comprehensive literature review will support this investigation. Ultimately, this review aims to clearly outline the objectives and significance of hydrogel synthesis and their applications.
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Affiliation(s)
- Paramjeet Yadav
- Department of Chemistry, Institute of Science, Banaras Hindu University, Varanasi 221005, UP, India
| | - Shiwani Singh
- Department of Chemistry, Institute of Science, Banaras Hindu University, Varanasi 221005, UP, India
| | - Sheetal Jaiswal
- Department of Chemistry, Institute of Science, Banaras Hindu University, Varanasi 221005, UP, India
| | - Rajesh Kumar
- Department of Chemistry, Institute of Science, Banaras Hindu University, Varanasi 221005, UP, India.
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Ohguro H, Watanabe M, Sato T, Nishikiori N, Umetsu A, Higashide M, Yano T, Suzuki H, Miyazaki A, Takada K, Uhara H, Furuhashi M, Hikage F. Application of Single Cell Type-Derived Spheroids Generated by Using a Hanging Drop Culture Technique in Various In Vitro Disease Models: A Narrow Review. Cells 2024; 13:1549. [PMID: 39329734 PMCID: PMC11430518 DOI: 10.3390/cells13181549] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/28/2024] [Revised: 08/21/2024] [Accepted: 08/23/2024] [Indexed: 09/28/2024] Open
Abstract
Cell culture methods are indispensable strategies for studies in biological sciences and for drug discovery and testing. Most cell cultures have been developed using two-dimensional (2D) culture methods, but three-dimensional (3D) culture techniques enable the establishment of in vitro models that replicate various pathogenic conditions and they provide valuable insights into the pathophysiology of various diseases as well as more precise results in tests for drug efficacy. However, one difficulty in the use of 3D cultures is selection of the appropriate 3D cell culture technique for the study purpose among the various techniques ranging from the simplest single cell type-derived spheroid culture to the more sophisticated organoid cultures. In the simplest single cell type-derived spheroid cultures, there are also various scaffold-assisted methods such as hydrogel-assisted cultures, biofilm-assisted cultures, particle-assisted cultures, and magnet particle-assisted cultures, as well as non-assisted methods, such as static suspension cultures, floating cultures, and hanging drop cultures. Since each method can be differently influenced by various factors such as gravity force, buoyant force, centrifugal force, and magnetic force, in addition to non-physiological scaffolds, each method has its own advantages and disadvantages, and the methods have different suitable applications. We have been focusing on the use of a hanging drop culture method for modeling various non-cancerous and cancerous diseases because this technique is affected only by gravity force and buoyant force and is thus the simplest method among the various single cell type-derived spheroid culture methods. We have found that the biological natures of spheroids generated even by the simplest method of hanging drop cultures are completely different from those of 2D cultured cells. In this review, we focus on the biological aspects of single cell type-derived spheroid culture and its applications in in vitro models for various diseases.
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Affiliation(s)
- Hiroshi Ohguro
- Departments of Ophthalmology, Sapporo Medical University, S1W17, Chuo-ku, Sapporo 060-8556, Japan; (M.W.); (N.N.); (A.U.); (M.H.)
| | - Megumi Watanabe
- Departments of Ophthalmology, Sapporo Medical University, S1W17, Chuo-ku, Sapporo 060-8556, Japan; (M.W.); (N.N.); (A.U.); (M.H.)
| | - Tatsuya Sato
- Departments of Cardiovascular, Renal and Metabolic Medicine, Sapporo Medical University, S1W17, Chuo-ku, Sapporo 060-8556, Japan; (T.S.); (T.Y.); (M.F.)
- Departments of Cellular Physiology and Signal Transduction, Sapporo Medical University, S1W17, Chuo-ku, Sapporo 060-8556, Japan
| | - Nami Nishikiori
- Departments of Ophthalmology, Sapporo Medical University, S1W17, Chuo-ku, Sapporo 060-8556, Japan; (M.W.); (N.N.); (A.U.); (M.H.)
| | - Araya Umetsu
- Departments of Ophthalmology, Sapporo Medical University, S1W17, Chuo-ku, Sapporo 060-8556, Japan; (M.W.); (N.N.); (A.U.); (M.H.)
| | - Megumi Higashide
- Departments of Ophthalmology, Sapporo Medical University, S1W17, Chuo-ku, Sapporo 060-8556, Japan; (M.W.); (N.N.); (A.U.); (M.H.)
| | - Toshiyuki Yano
- Departments of Cardiovascular, Renal and Metabolic Medicine, Sapporo Medical University, S1W17, Chuo-ku, Sapporo 060-8556, Japan; (T.S.); (T.Y.); (M.F.)
| | - Hiromu Suzuki
- Departments of Molecular Biology, Sapporo Medical University, S1W17, Chuo-ku, Sapporo 060-8556, Japan;
| | - Akihiro Miyazaki
- Departments of Oral Surgery, Sapporo Medical University, S1W17, Chuo-ku, Sapporo 060-8556, Japan;
| | - Kohichi Takada
- Departments of Medical Oncology, Sapporo Medical University, S1W17, Chuo-ku, Sapporo 060-8556, Japan;
| | - Hisashi Uhara
- Departments of Dermatology, Sapporo Medical University, S1W17, Chuo-ku, Sapporo 060-8556, Japan;
| | - Masato Furuhashi
- Departments of Cardiovascular, Renal and Metabolic Medicine, Sapporo Medical University, S1W17, Chuo-ku, Sapporo 060-8556, Japan; (T.S.); (T.Y.); (M.F.)
| | - Fumihito Hikage
- Departments of Ophthalmology, Sapporo Medical University, S1W17, Chuo-ku, Sapporo 060-8556, Japan; (M.W.); (N.N.); (A.U.); (M.H.)
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Česnik AB, Švajger U. The issue of heterogeneity of MSC-based advanced therapy medicinal products-a review. Front Cell Dev Biol 2024; 12:1400347. [PMID: 39129786 PMCID: PMC11310176 DOI: 10.3389/fcell.2024.1400347] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/13/2024] [Accepted: 07/15/2024] [Indexed: 08/13/2024] Open
Abstract
Mesenchymal stromal stem cells (MSCs) possess a remarkable potential for numerous clinical applications due to their unique properties including self-renewal, immunomodulation, paracrine actions and multilineage differentiation. However, the translation of MSC-based Advanced Therapy Medicinal Products (ATMPs) into the clinic has frequently met with inconsistent outcomes. One of the suspected reasons for this issue is the inherent and extensive variability that exists among such ATMPs, which makes the interpretation of their clinical efficacy difficult to assess, as well as to compare the results of various studies. This variability stems from numerous reasons including differences in tissue sources, donor attributes, variances in manufacturing protocols, as well as modes of administration. MSCs can be isolated from various tissues including bone marrow, umbilical cord, adipose tissue and others, each with its unique phenotypic and functional characteristics. While MSCs from different sources do share common features, they also exhibit distinct gene expression profiles and functional properites. Donor-specific factors such as age, sex, body mass index, and underlying health conditions can influence MSC phenotype, morphology, differentiation potential and function. Moreover, variations in preparation of MSC products introduces additional heterogeneity as a result of cell culture media composition, presence or absence of added growth factors, use of different serum supplements and culturing techniques. Once MSC products are formulated, storage protocols play a pivotal role in its efficacy. Factors that affect cell viability include cell concentration, delivery solution and importantly, post-thawing protocols where applicable. Ensuing, differences in administration protocols can critically affect the distribution and functionallity of administered cells. As MSC-based therapies continue to advance through numerous clinical trials, implication of strategies to reduce product heterogeneity is imperative. Central to addressing these challenges is the need for precise prediction of clinical responses, which require well-defined MSC populations and harmonized assessment of their specific functions. By addressing these issues by meaningful approaches, such as, e.g., MSC pooling, the field can overcome barriers to advance towards more consistent and effective MSC-based therapies.
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Affiliation(s)
- Ana Bajc Česnik
- Slovenian Institute for Transfusion Medicine, Department for Therapeutic Services, Ljubljana, Slovenia
| | - Urban Švajger
- Slovenian Institute for Transfusion Medicine, Department for Therapeutic Services, Ljubljana, Slovenia
- Faculty of Pharmacy, University of Ljubljana, Ljubljana, Slovenia
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Sahu RK, Tandon S, Singh S, Das BC, Hedau ST. Methyl CpG binding protein MBD2 has a regulatory role on the BRCA1 gene expression and its modulation by resveratrol in ER+, PR+ & triple-negative breast cancer cells. BMC Cancer 2024; 24:566. [PMID: 38711004 PMCID: PMC11071212 DOI: 10.1186/s12885-024-12274-x] [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: 11/08/2023] [Accepted: 04/16/2024] [Indexed: 05/08/2024] Open
Abstract
BACKGROUND Resveratrol has demonstrated its ability to regulate BRCA1 gene expression in breast cancer cells, and previous studies have established the binding of MBD proteins to BRCA1 gene promoter regions. However, the molecular mechanism underlying these interactions remains to be elucidated. The aimed to evaluate the impact of MBD proteins on the regulation of BRCA1, BRCA2, and p16 genes and their consequential effects on breast cancer cells. METHODS Efficacy of resveratrol was assessed using the MTT assay. Binding interactions were investigated through EMSA, ChIP, & MeIP assay. Expression analyses of MBD genes and proteins were conducted using qRT-PCR and western blotting, respectively. Functional assays, including clonogenic, migratory, and sphere formation assays were used to assess cancer cells' colony-forming, metastatic, and tumor-forming abilities. The cytotoxicity of resveratrol on cancer cells was also tested using an apoptosis assay. RESULTS The study determined an IC50 of 30µM for resveratrol. MBD proteins were found to bind to the BRCA1 gene promoter. Resveratrol exhibited regulatory effects on MBD gene expression, subsequently impacting BRCA1 gene expression and protein levels. Higher concentrations of resveratrol resulted in reduced colony and sphere formation, decreases migration of cancer cells, and an increases number of apoptotic cells in breast cancer cells. Impact Identification of MBD2-BRCA1 axis indicates their significant role in the induction of apoptosis and reduction of metastasis and proliferation in breast cancer cells. Further therapy can be designed to target these MBD proteins and resveratrol could be used along with other anticancer drugs to target breast cancer. CONCLUSIONS In conclusion MBD2 protein interact to the BRCA1 gene promoter, and resveratrol modulates MBD2 gene expression, which in turn regulates BRCA1 gene expression, and inhibits cell proliferation, migration, and induces apoptosis in ER+, PR+ & Triple negative breast cancer cells.
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Affiliation(s)
- Ram Krishna Sahu
- Division of Molecular Oncology, ICMR-National Institute of Cancer Prevention and Research, I -7, Sector - 39, Noida, Uttar Pradesh, 201301, India
- Amity Institute of Molecular Medicine & Stem Cell Research, Amity University, Noida, Uttar Pradesh, 201313, India
| | | | - Shalini Singh
- Division of Clinical Oncology, ICMR-National Institute of Cancer Prevention and Research, I -7, Sector - 39, Noida, Uttar Pradesh, 201301, India
| | - Bhudev Chandra Das
- Amity Institute of Molecular Medicine & Stem Cell Research, Amity University, Noida, Uttar Pradesh, 201313, India
| | - Suresh T Hedau
- Division of Molecular Oncology, ICMR-National Institute of Cancer Prevention and Research, I -7, Sector - 39, Noida, Uttar Pradesh, 201301, India.
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Kamprom W, Tangporncharoen R, Vongthaiwan N, Tragoonlugkana P, Phetfong J, Pruksapong C, Supokawej A. Enhanced potent immunosuppression of intracellular adipose tissue-derived stem cell extract by priming with three-dimensional spheroid formation. Sci Rep 2024; 14:9084. [PMID: 38643332 PMCID: PMC11032398 DOI: 10.1038/s41598-024-59910-x] [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: 12/29/2023] [Accepted: 04/16/2024] [Indexed: 04/22/2024] Open
Abstract
Immunomodulatory properties of mesenchymal stem cells are widely studied, supporting the use of MSCs as cell-based therapy in immunological diseases. This study aims to generate cell-free MSC extract and improves their immunomodulatory potential. Intracellular extracts were prepared from adipose-derived stem cells (ADSC) spheroid via a freeze-thawing method. The immunomodulatory capacities of ADSC spheroid extracts were investigated in vitro, including lymphocyte proliferation, T regulatory cell expansion, and macrophage assays. A comparative study was conducted with ADSC monolayer extract. The key immunomodulatory mediators presented in ADSC extract were identified. The results revealed that ADSC spheroid extract could suppress lymphocyte activation while enhancing T regulatory cell expansion. Immunomodulatory molecules such as COX-2, TSG-6, and TGF-β1 were upregulated in ADSC priming via spheroid culture. Selective inhibition of COX-2 abrogates the effect of ADSC extract on inducing T regulatory cell expansion. Thus, ADSC spheroid extract gains high efficacy in regulating the immune responses which are associated in part by COX-2 generation. Furthermore, ADSC spheroid extract possessed a potent anti-inflammation by manipulation of TNF-α production from LPS-activated macrophage. Our current study has highlighted the opportunity of using cell-free extracts from adipose tissue-derived mesenchymal stem cells spheroid as novel immunomodulators for the treatment of immunological-associated diseases.
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Affiliation(s)
- Witchayapon Kamprom
- Department of Clinical Microbiology and Applied Technology, Faculty of Medical Technology, Mahidol University, Nakhon Pathom, Thailand
| | - Rattanawan Tangporncharoen
- Department of Clinical Microscopy, Faculty of Medical Technology, Mahidol University, 999 Phutthamonthon Sai 4, Salaya, Phutthamonthon, Nakhon Pathom, 73170, Thailand
| | - Nuttapoom Vongthaiwan
- Department of Clinical Microscopy, Faculty of Medical Technology, Mahidol University, 999 Phutthamonthon Sai 4, Salaya, Phutthamonthon, Nakhon Pathom, 73170, Thailand
| | - Patcharapa Tragoonlugkana
- Department of Clinical Microscopy, Faculty of Medical Technology, Mahidol University, 999 Phutthamonthon Sai 4, Salaya, Phutthamonthon, Nakhon Pathom, 73170, Thailand
| | - Jitrada Phetfong
- Department of Clinical Microscopy, Faculty of Medical Technology, Mahidol University, 999 Phutthamonthon Sai 4, Salaya, Phutthamonthon, Nakhon Pathom, 73170, Thailand
| | - Chatchai Pruksapong
- Department of Surgery, Phramongkutklao Hospital and College of Medicine, Bangkok, Thailand
| | - Aungkura Supokawej
- Department of Clinical Microscopy, Faculty of Medical Technology, Mahidol University, 999 Phutthamonthon Sai 4, Salaya, Phutthamonthon, Nakhon Pathom, 73170, Thailand.
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Moriwaki T, Tani H, Haga K, Morita-Umei Y, Soma Y, Umei TC, Sekine O, Takatsuna K, Kishino Y, Kanazawa H, Fujita J, Fukuda K, Tohyama S, Ieda M. Scalable production of homogeneous cardiac organoids derived from human pluripotent stem cells. CELL REPORTS METHODS 2023; 3:100666. [PMID: 38113855 PMCID: PMC10753388 DOI: 10.1016/j.crmeth.2023.100666] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 01/06/2023] [Revised: 08/24/2023] [Accepted: 11/16/2023] [Indexed: 12/21/2023]
Abstract
Three-dimensional (3D) cultures are known to more closely mimic in vivo conditions compared with 2D cultures. Cardiac spheroids (CSs) and organoids (COs) are useful for 3D tissue engineering and are advantageous for their simplicity and mass production for regenerative therapy and drug discovery. Herein, we describe a large-scale method for producing homogeneous human induced pluripotent stem cell (hiPSC)-derived CSs (hiPSC-CSs) and COs without scaffolds using a porous 3D microwell substratum with a suction system. Our method has many advantages, such as increased efficiency and improved functionality, homogeneity, and sphericity of hiPSC-CSs. Moreover, we have developed a substratum on a clinically relevant large scale for regenerative therapy and have succeeded in producing approximately 40,000 hiPSC-CSs with high sphericity at once. Furthermore, we efficiently produced a fused CO model consisting of hiPSC-derived atrial and ventricular cardiomyocytes localized on opposite sides of one organoid. This method will facilitate progress toward hiPSC-based clinical applications.
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Affiliation(s)
- Taijun Moriwaki
- Department of Cardiology, Keio University School of Medicine, Shinjuku, Tokyo, Japan
| | - Hidenori Tani
- Department of Cardiology, Keio University School of Medicine, Shinjuku, Tokyo, Japan; Joint Research Laboratory for Medical Innovation in Heart Disease, Keio University School of Medicine, Shinjuku, Tokyo, Japan
| | - Kotaro Haga
- Department of Cardiology, Keio University School of Medicine, Shinjuku, Tokyo, Japan
| | - Yuika Morita-Umei
- Department of Cardiology, Keio University School of Medicine, Shinjuku, Tokyo, Japan; Kanagawa Institute of Industrial Science and Technology (KISTEC), Kawasaki, Kanagawa, Japan
| | - Yusuke Soma
- Department of Cardiology, Keio University School of Medicine, Shinjuku, Tokyo, Japan
| | - Tomohiko C Umei
- Department of Cardiology, Keio University School of Medicine, Shinjuku, Tokyo, Japan
| | - Otoya Sekine
- Department of Cardiology, Keio University School of Medicine, Shinjuku, Tokyo, Japan
| | - Kaworu Takatsuna
- Department of Cardiology, Keio University School of Medicine, Shinjuku, Tokyo, Japan
| | - Yoshikazu Kishino
- Department of Cardiology, Keio University School of Medicine, Shinjuku, Tokyo, Japan
| | - Hideaki Kanazawa
- Department of Cardiology, Keio University School of Medicine, Shinjuku, Tokyo, Japan
| | - Jun Fujita
- Department of Cardiology, Keio University School of Medicine, Shinjuku, Tokyo, Japan; Department of Pathology & Immunology, Baylor College of Medicine, Houston, TX, USA
| | - Keiichi Fukuda
- Department of Cardiology, Keio University School of Medicine, Shinjuku, Tokyo, Japan
| | - Shugo Tohyama
- Department of Cardiology, Keio University School of Medicine, Shinjuku, Tokyo, Japan.
| | - Masaki Ieda
- Department of Cardiology, Keio University School of Medicine, Shinjuku, Tokyo, Japan
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Senrung A, Lalwani S, Janjua D, Tripathi T, Kaur J, Ghuratia N, Aggarwal N, Chhokar A, Yadav J, Chaudhary A, Joshi U, Bharti AC. 3D tumor spheroids: morphological alterations a yardstick to anti-cancer drug response. IN VITRO MODELS 2023; 2:219-248. [PMID: 39872501 PMCID: PMC11756486 DOI: 10.1007/s44164-023-00059-8] [Citation(s) in RCA: 7] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 06/02/2023] [Revised: 08/13/2023] [Accepted: 08/21/2023] [Indexed: 01/30/2025]
Abstract
Tumor spheroids are one of the well-characterized 3D culture systems bearing close resemblance to the physiological tissue organization and complexity of avascular solid tumor stage with hypoxic core. They hold a wide-spread application in the field of pharmaceutical science and anti-cancer drug research. However, the difficulty in determining optimal technique for the generation of spheroids with uniform size and shape, evaluation of experimental outputs, or mass production often limits their usage in anti-cancer research and in high-throughput drug screening. In recent times, several studies have demonstrated various simple techniques for generating uniform-size 3D spheroids, including the hanging drop (HD), liquid overlay technique (LOT), and microfluidic approaches. Morphological alterations apart from biochemical assays, and staining techniques are suitably employed for the evaluation of experimental outcomes within 3D spheroid models. Morphological alterations in response to effective anti-cancer drug treatment in 3D tumor spheroids such as reduced spheroid size, loss of spheroid compactness and integrity or smooth surface, are highly reliable. These alterations can significantly reduce the need for biochemical assays and staining techniques, resulting in both time and cost savings. The present article specifically covers a variety of available procedures in spheroid generation. For practical applicability, we have supplemented our review study with the generation of glioblastoma U87 spheroids using HD and LOT methods. Additionally, we have also incorporated the outcome of U87 spheroid treatment with doxorubicin on spheroid morphology.
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Affiliation(s)
- Anna Senrung
- Molecular Oncology Laboratory, Department of Zoology, University of Delhi (North Campus), Delhi, 110007 India
- Neuropharmacology & Drug Delivery Laboratory, Zoology Department, Daulat Ram College, University of Delhi, Delhi, 110007 India
| | - Sakshi Lalwani
- Neuropharmacology & Drug Delivery Laboratory, Zoology Department, Daulat Ram College, University of Delhi, Delhi, 110007 India
| | - Divya Janjua
- Molecular Oncology Laboratory, Department of Zoology, University of Delhi (North Campus), Delhi, 110007 India
| | - Tanya Tripathi
- Molecular Oncology Laboratory, Department of Zoology, University of Delhi (North Campus), Delhi, 110007 India
| | - Jasleen Kaur
- Neuropharmacology & Drug Delivery Laboratory, Zoology Department, Daulat Ram College, University of Delhi, Delhi, 110007 India
| | - Netra Ghuratia
- Neuropharmacology & Drug Delivery Laboratory, Zoology Department, Daulat Ram College, University of Delhi, Delhi, 110007 India
| | - Nikita Aggarwal
- Molecular Oncology Laboratory, Department of Zoology, University of Delhi (North Campus), Delhi, 110007 India
| | - Arun Chhokar
- Molecular Oncology Laboratory, Department of Zoology, University of Delhi (North Campus), Delhi, 110007 India
- Department of Zoology, Deshbandhu College, University of Delhi, Delhi, India
| | - Joni Yadav
- Molecular Oncology Laboratory, Department of Zoology, University of Delhi (North Campus), Delhi, 110007 India
| | - Apoorva Chaudhary
- Molecular Oncology Laboratory, Department of Zoology, University of Delhi (North Campus), Delhi, 110007 India
| | - Udit Joshi
- Molecular Oncology Laboratory, Department of Zoology, University of Delhi (North Campus), Delhi, 110007 India
| | - Alok Chandra Bharti
- Molecular Oncology Laboratory, Department of Zoology, University of Delhi (North Campus), Delhi, 110007 India
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Rakhmatullina AR, Mingaleeva RN, Gafurbaeva DU, Glazunova ON, Sagdeeva AR, Bulatov ER, Rizvanov AA, Miftakhova RR. Adipose-Derived Mesenchymal Stem Cell (MSC) Immortalization by Modulation of hTERT and TP53 Expression Levels. J Pers Med 2023; 13:1621. [PMID: 38003936 PMCID: PMC10672200 DOI: 10.3390/jpm13111621] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/11/2023] [Revised: 11/03/2023] [Accepted: 11/13/2023] [Indexed: 11/26/2023] Open
Abstract
Mesenchymal stem cells (MSCs) are pivotal players in tissue repair and hold great promise as cell therapeutic agents for regenerative medicine. Additionally, they play a significant role in the development of various human diseases. Studies on MSC biology have encountered a limiting property of these cells, which includes a low number of passages and a decrease in differentiation potential during in vitro culture. Although common methods of immortalization through gene manipulations of cells are well established, the resulting MSCs vary in differentiation potential compared to primary cells and eventually undergo senescence. This study aimed to immortalize primary adipose-derived MSCs by overexpressing human telomerase reverse transcriptase (hTERT) gene combined with a knockdown of TP53. The research demonstrated that immortalized MSCs maintained a stable level of differentiation into osteogenic and chondrogenic lineages during 30 passages, while also exhibiting an increase in cell proliferation rate and differentiation potential towards the adipogenic lineage. Long-term culture of immortalized cells did not alter cell morphology and self-renewal potential. Consequently, a genetically stable line of immortalized adipose-derived MSCs (iMSCs) was established.
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Affiliation(s)
- Aigul R. Rakhmatullina
- Institute of Fundamental Medicine and Biology, Kazan Federal University, 420008 Kazan, Russia
| | - Rimma N. Mingaleeva
- Institute of Fundamental Medicine and Biology, Kazan Federal University, 420008 Kazan, Russia
| | - Dina U. Gafurbaeva
- Institute of Fundamental Medicine and Biology, Kazan Federal University, 420008 Kazan, Russia
| | - Olesya N. Glazunova
- Institute of Fundamental Medicine and Biology, Kazan Federal University, 420008 Kazan, Russia
| | - Aisylu R. Sagdeeva
- Institute of Fundamental Medicine and Biology, Kazan Federal University, 420008 Kazan, Russia
| | - Emil R. Bulatov
- Institute of Fundamental Medicine and Biology, Kazan Federal University, 420008 Kazan, Russia
- Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russia
| | - Albert A. Rizvanov
- Institute of Fundamental Medicine and Biology, Kazan Federal University, 420008 Kazan, Russia
| | - Regina R. Miftakhova
- Institute of Fundamental Medicine and Biology, Kazan Federal University, 420008 Kazan, Russia
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11
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Bumroongthai K, Kavanagh DPJ, Genever P, Kalia N. Improving vasculoprotective effects of MSCs in coronary microvessels - benefits of 3D culture, sub-populations and heparin. Front Immunol 2023; 14:1257497. [PMID: 37954606 PMCID: PMC10635425 DOI: 10.3389/fimmu.2023.1257497] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/12/2023] [Accepted: 10/02/2023] [Indexed: 11/14/2023] Open
Abstract
Introduction Opening occluded coronary arteries in patients with myocardial infarction (MI) damages the delicate coronary microvessels through a process called myocardial ischaemia-reperfusion injury. Although mesenchymal stromal cells (MSCs) have the potential to limit this injury, clinical success remains limited. This may be due to (i) poor MSC homing to the heart (ii) infused MSCs, even if derived from the same site, being a heterogeneous population with varying therapeutic efficacy and (iii) conventional 2D culture of MSCs decreasing their homing and beneficial properties. This study investigated whether 3D culture of two distinctly different bone marrow (BM)-derived MSC sub-populations could improve their homing and coronary vasculoprotective efficacy. Methods Intravital imaging of the anaesthetised mouse beating heart was used to investigate the trafficking and microvascular protective effects of two clonally-derived BM-derived MSC lines, namely CD317neg MSCs-Y201 and CD317pos MSCs-Y202, cultured using conventional monolayer and 3D hanging drop methods. Results 3D culture consistently improved the adhesive behaviour of MSCs-Y201 to various substrates in vitro. However, it was their differential ability to reduce neutrophil events within the coronary capillaries and improve ventricular perfusion in vivo that was most remarkable. Moreover, dual therapy combined with heparin further improved the vasculoprotection afforded by 3D cultured MSCs-Y201 by also modifying platelet as well as neutrophil recruitment, which subsequently led to the greatest salvage of viable myocardium. Therapeutic benefit could mechanistically be explained by reductions in coronary endothelial oxidative stress and intercellular adhesion molecule-1 (ICAM-1)/vascular cell adhesion molecule-1 (VCAM-1) expression. However, since this was noted by both 2D and 3D cultured MSCs-Y201, therapeutic benefit is likely explained by the fact that 3D cultured MSCs-Y201 were the most potent sub-population at reducing serum levels of several pro-inflammatory cytokines. Conclusion This novel study highlights the importance of not only 3D culture, but also of a specific CD317neg MSC sub-population, as being critical to realising their full coronary vasculoprotective potential in the injured heart. Since the smallest coronary blood vessels are increasingly recognised as a primary target of reperfusion injury, therapeutic interventions must be able to protect these delicate structures from inflammatory cells and maintain perfusion in the heart. We propose that relatively feasible technical modifications in a specific BM-derived MSC sub-population could achieve this.
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Affiliation(s)
- Kobkaew Bumroongthai
- Microcirculation Research Group, Institute of Cardiovascular Sciences, College of Medical and Dental Sciences, University of Birmingham, Birmingham, United Kingdom
| | - Dean P. J. Kavanagh
- Microcirculation Research Group, Institute of Cardiovascular Sciences, College of Medical and Dental Sciences, University of Birmingham, Birmingham, United Kingdom
| | - Paul Genever
- Department of Biology, University of York, York, United Kingdom
| | - Neena Kalia
- Microcirculation Research Group, Institute of Cardiovascular Sciences, College of Medical and Dental Sciences, University of Birmingham, Birmingham, United Kingdom
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12
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Zingales V, Esposito MR, Torriero N, Taroncher M, Cimetta E, Ruiz MJ. The Growing Importance of Three-Dimensional Models and Microphysiological Systems in the Assessment of Mycotoxin Toxicity. Toxins (Basel) 2023; 15:422. [PMID: 37505691 PMCID: PMC10467068 DOI: 10.3390/toxins15070422] [Citation(s) in RCA: 5] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/22/2023] [Revised: 06/21/2023] [Accepted: 06/25/2023] [Indexed: 07/29/2023] Open
Abstract
Current investigations in the field of toxicology mostly rely on 2D cell cultures and animal models. Although well-accepted, the traditional 2D cell-culture approach has evident drawbacks and is distant from the in vivo microenvironment. To overcome these limitations, increasing efforts have been made in the development of alternative models that can better recapitulate the in vivo architecture of tissues and organs. Even though the use of 3D cultures is gaining popularity, there are still open questions on their robustness and standardization. In this review, we discuss the current spheroid culture and organ-on-a-chip techniques as well as the main conceptual and technical considerations for the correct establishment of such models. For each system, the toxicological functional assays are then discussed, highlighting their major advantages, disadvantages, and limitations. Finally, a focus on the applications of 3D cell culture for mycotoxin toxicity assessments is provided. Given the known difficulties in defining the safety ranges of exposure for regulatory agency policies, we are confident that the application of alternative methods may greatly improve the overall risk assessment.
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Affiliation(s)
- Veronica Zingales
- Laboratory of Toxicology, Faculty of Pharmacy, University of Valencia, Av. Vicent Andrés Estellés s/n, 46100 Valencia, Spain;
- Department of Industrial Engineering (DII), University of Padua, Via Marzolo 9, 35131 Padova, Italy; (M.R.E.); (N.T.); (E.C.)
- Fondazione Istituto di Ricerca Pediatrica Cittá Della Speranza (IRP)—Lab BIAMET, Corso Stati Uniti 4, 35127 Padova, Italy
| | - Maria Rosaria Esposito
- Department of Industrial Engineering (DII), University of Padua, Via Marzolo 9, 35131 Padova, Italy; (M.R.E.); (N.T.); (E.C.)
- Fondazione Istituto di Ricerca Pediatrica Cittá Della Speranza (IRP)—Lab BIAMET, Corso Stati Uniti 4, 35127 Padova, Italy
| | - Noemi Torriero
- Department of Industrial Engineering (DII), University of Padua, Via Marzolo 9, 35131 Padova, Italy; (M.R.E.); (N.T.); (E.C.)
- Fondazione Istituto di Ricerca Pediatrica Cittá Della Speranza (IRP)—Lab BIAMET, Corso Stati Uniti 4, 35127 Padova, Italy
| | - Mercedes Taroncher
- Laboratory of Toxicology, Faculty of Pharmacy, University of Valencia, Av. Vicent Andrés Estellés s/n, 46100 Valencia, Spain;
| | - Elisa Cimetta
- Department of Industrial Engineering (DII), University of Padua, Via Marzolo 9, 35131 Padova, Italy; (M.R.E.); (N.T.); (E.C.)
- Fondazione Istituto di Ricerca Pediatrica Cittá Della Speranza (IRP)—Lab BIAMET, Corso Stati Uniti 4, 35127 Padova, Italy
| | - María-José Ruiz
- Laboratory of Toxicology, Faculty of Pharmacy, University of Valencia, Av. Vicent Andrés Estellés s/n, 46100 Valencia, Spain;
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13
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Henrique RBL, Lima RRM, Monteiro CAP, Oliveira WF, Pereira G, Cabral Filho PE, Fontes A. Advances in the study of spheroids as versatile models to evaluate biological interactions of inorganic nanoparticles. Life Sci 2022; 302:120657. [PMID: 35609631 DOI: 10.1016/j.lfs.2022.120657] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/18/2022] [Revised: 05/10/2022] [Accepted: 05/18/2022] [Indexed: 12/26/2022]
Abstract
Spheroids are in vitro three-dimensional multicellular microstructures able to mimic the biological microenvironment, including the complexity of tumor architecture. Therefore, results closer to those expected for in vivo organisms can be reached using spheroids compared to the cell culture monolayer model. Inorganic nanoparticles (NPs) have also been playing relevant roles in the comprehension of biological processes. Moreover, they have been probed as novel diagnostic and therapeutical nanosystems. In this context, in this review, we present applications, published in the last five years, which show that spheroids can be versatile models to study and evaluate biological interactions involving inorganic NPs. Applications of spheroids associated with (i) basic studies to assess the penetration profile of nanostructures, (ii) the evaluation of NP toxicity, and (iii) NP-based therapeutical approaches are described. Fundamentals of spheroids and their formation methods are also included. We hope that this review can be a reference and guide future investigations related to this interesting three-dimensional biological model, favoring advances to Nanobiotechnology.
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Affiliation(s)
- Rafaella B L Henrique
- Departamento de Biofísica e Radiobiologia, Centro de Biociências, Universidade Federal de Pernambuco, Recife, PE, Brazil
| | - Rennan R M Lima
- Departamento de Biofísica e Radiobiologia, Centro de Biociências, Universidade Federal de Pernambuco, Recife, PE, Brazil
| | - Camila A P Monteiro
- Departamento de Biofísica e Radiobiologia, Centro de Biociências, Universidade Federal de Pernambuco, Recife, PE, Brazil
| | - Weslley F Oliveira
- Departamento de Bioquímica, Centro de Biociências, Universidade Federal de Pernambuco, Recife, PE, Brazil
| | - Goreti Pereira
- Departamento de Química Fundamental, Centro de Ciências Exatas e da Natureza, Universidade Federal de Pernambuco, Recife, PE, Brazil
| | - Paulo E Cabral Filho
- Departamento de Biofísica e Radiobiologia, Centro de Biociências, Universidade Federal de Pernambuco, Recife, PE, Brazil.
| | - Adriana Fontes
- Departamento de Biofísica e Radiobiologia, Centro de Biociências, Universidade Federal de Pernambuco, Recife, PE, Brazil.
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Genetically engineered and enucleated human mesenchymal stromal cells for the targeted delivery of therapeutics to diseased tissue. Nat Biomed Eng 2022; 6:882-897. [PMID: 34931077 PMCID: PMC9207157 DOI: 10.1038/s41551-021-00815-9] [Citation(s) in RCA: 70] [Impact Index Per Article: 23.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/03/2020] [Accepted: 07/07/2021] [Indexed: 02/05/2023]
Abstract
Targeting the delivery of therapeutics specifically to diseased tissue enhances their efficacy and decreases their side effects. Here we show that mesenchymal stromal cells with their nuclei removed by density-gradient centrifugation following the genetic modification of the cells for their display of chemoattractant receptors and endothelial-cell-binding molecules are effective vehicles for the targeted delivery of therapeutics. The enucleated cells neither proliferate nor permanently engraft in the host, yet retain the organelles for energy and protein production, undergo integrin-regulated adhesion to inflamed endothelial cells, and actively home to chemokine gradients established by diseased tissues. In mouse models of acute inflammation and of pancreatitis, systemically administered enucleated cells expressing two types of chemokine receptor and an endothelial adhesion molecule enhanced the delivery of an anti-inflammatory cytokine to diseased tissue (with respect to unmodified stromal cells and to exosomes derived from bone-marrow-derived stromal cells), attenuating inflammation and ameliorating disease pathology. Enucleated cells retain most of the cells' functionality, yet acquire the cargo-carrying characteristics of cell-free delivery systems, and hence represent a versatile delivery vehicle and therapeutic system.
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15
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Lee SY, Lee JW. 3D Spheroid Cultures of Stem Cells and Exosome Applications for Cartilage Repair. LIFE (BASEL, SWITZERLAND) 2022; 12:life12070939. [PMID: 35888029 PMCID: PMC9317836 DOI: 10.3390/life12070939] [Citation(s) in RCA: 20] [Impact Index Per Article: 6.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 05/26/2022] [Revised: 06/20/2022] [Accepted: 06/21/2022] [Indexed: 11/16/2022]
Abstract
Cartilage is a connective tissue that constitutes the structure of the body and consists of chondrocytes that produce considerable collagenous extracellular matrix and plentiful ground substances, such as proteoglycan and elastin fibers. Self-repair is difficult when the cartilage is damaged because of insufficient blood supply, low cellularity, and limited progenitor cell numbers. Therefore, three-dimensional (3D) culture systems, including pellet culture, hanging droplets, liquid overlays, self-injury, and spinner culture, have attracted attention. In particular, 3D spheroid culture strategies can enhance the yield of exosome production of mesenchymal stem cells (MSCs) when compared to two-dimensional culture, and can improve cellular restorative function by enhancing the paracrine effects of MSCs. Exosomes are membrane-bound extracellular vesicles, which are intercellular communication systems that carry RNAs and proteins. Information transfer affects the phenotype of recipient cells. MSC-derived exosomes can facilitate cartilage repair by promoting chondrogenic differentiation and proliferation. In this article, we reviewed recent major advances in the application of 3D culture techniques, cartilage regeneration with stem cells using 3D spheroid culture system, the effect of exosomes on chondrogenic differentiation, and chondrogenic-specific markers related to stem cell derived exosomes. Furthermore, the utilization of MSC-derived exosomes to enhance chondrogenic differentiation for osteoarthritis is discussed. If more mechanistic studies at the molecular level are conducted, MSC-spheroid-derived exosomes will supply a better therapeutic option to improve osteoarthritis.
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Affiliation(s)
- Seung Yeon Lee
- Department of Molecular Medicine, College of Medicine, Gachon University, 155, Gaetbeol-ro, Yeonsu-ku, Incheon 21999, Korea;
| | - Jin Woo Lee
- Department of Molecular Medicine, College of Medicine, Gachon University, 155, Gaetbeol-ro, Yeonsu-ku, Incheon 21999, Korea;
- Department of Health Sciences and Technology, GAIHST, Gachon University, 155, Gaetbeol-ro, Yeonsu-ku, Incheon 21999, Korea
- Correspondence: ; Tel.: +82-32-899-6516; Fax: +82-32-899-6039
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16
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Kim MJ, Moon W, Heo J, Lim S, Lee SH, Jeong JY, Lee SJ. Optimization of adipose tissue-derived mesenchymal stromal cells transplantation for bone marrow repopulation following irradiation. World J Stem Cells 2022; 14:245-263. [PMID: 35432736 PMCID: PMC8968216 DOI: 10.4252/wjsc.v14.i3.245] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/19/2021] [Revised: 01/12/2022] [Accepted: 02/27/2022] [Indexed: 02/06/2023] Open
Abstract
BACKGROUND Bone marrow (BM) suppression is one of the most common side effects of radiotherapy and the primary cause of death following exposure to irradiation. Despite concerted efforts, there is no definitive treatment method available. Recent studies have reported using mesenchymal stromal cells (MSCs), but their therapeutic effects are contested. AIM We administered and examined the effects of various amounts of adipose-derived MSCs (ADSCs) in mice with radiation-induced BM suppression. METHODS Mice were divided into three groups: Normal control group, irradiated (RT) group, and stem cell-treated group following whole-body irradiation (WBI). Mouse ADSCs (mADSCs) were transplanted into the peritoneal cavity either once or three times at 5 × 105 cells/200 μL. The white blood cell count and the levels of, plasma cytokines, BM mRNA, and BM surface markers were compared between the three groups. Human BM-derived CD34+ hematopoietic progenitor cells were co-cultured with human ADSCs (hADSCs) or incubated in the presence of hADSCs conditioned media to investigate the effect on human cells in vitro. RESULTS The survival rate of mice that received one transplant of mADSCs was higher than that of mice that received three transplants. Multiple transplantations of ADSCs delayed the repopulation of BM hematopoietic stem cells. Anti-inflammatory effects and M2 polarization by intraperitoneal ADSCs might suppress erythropoiesis and induce myelopoiesis in sub-lethally RT mice. CONCLUSION The results suggested that an optimal amount of MSCs could improve survival rates post-WBI.
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Affiliation(s)
- Min-Jung Kim
- Department of Biochemistry, Cancer Research Institute Kosin University College of Medicine, Seo-gu 49267, Busan, South Korea
| | - Won Moon
- Department of Internal Medicine, Kosin University College of Medicine, Seo-gu 49267, Busan, South Korea
| | - Jeonghoon Heo
- Department of Molecular Biology and Immunology, Kosin University College of Medicine, Seo-gu 49267, Busan, South Korea
| | - Sangwook Lim
- Department of Radiation Oncology, Kosin University College of Medicine, Seo-gu 49267, Busan, South Korea
| | - Seung-Hyun Lee
- Department of General Surgery, Kosin University College of Medicine, Seo-gu 49267, Busan, South Korea
| | - Jee-Yeong Jeong
- Department of Biochemistry, Cancer Research Institute Kosin University College of Medicine, Seo-gu 49267, Busan, South Korea
| | - Sang Joon Lee
- Department of Ophthalmology, Gospel Hospital, Kosin University College of Medicine, Seo-gu 49267, Busan, South Korea.
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Ali R, Huwaizi S, Alhallaj A, Al Subait A, Barhoumi T, Al Zahrani H, Al Anazi A, Latif Khan A, Boudjelal M. New Born Calf Serum Can Induce Spheroid Formation in Breast Cancer KAIMRC1 Cell Line. Front Mol Biosci 2022; 8:769030. [PMID: 35004846 PMCID: PMC8740237 DOI: 10.3389/fmolb.2021.769030] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/01/2021] [Accepted: 11/02/2021] [Indexed: 11/29/2022] Open
Abstract
Three-dimensional (3D) cell culture systems have become very popular in the field of drug screening and discovery. There is an immense demand for highly efficient and easy methods to produce 3D spheroids in any cell format. We have developed a novel and easy method to produce spheroids from the newly isolated KAIMRC1 cell line in vitro. It can be used as a 3D model to study proliferation, differentiation, cell death, and drug response of cancer cells. Our procedure requires growth media supplemented with 10% new born calf serum (NBCS) and regular cell culture plates to generate KAIMRC1 spheroids without the need for any specialized 3D cell culture system. This procedure generates multiple spheroids within a 12–24-h culture. KAIMRC1 spheroids are compact, homogeneous in size and morphology with a mean size of 55.8 µm (±3.5). High content imaging (HCI) of KAIMRC1 spheroids treated with a panel of 240 compounds resulted in the identification of several highly specific compounds towards spheroids. Immunophenotyping of KAIMRC1 spheroids revealed phosphorylation of FAK, cJUN, and E-cadherin, which suggests the involvement of JNK/JUN pathway in the KAIMRC1 spheroids formation. Gene expression analysis showed upregulation of cell junction genes, GJB3, DSC1, CLDN5, CLDN8, and PLAU. Furthermore, co-culture of KAIMRC1 cells with primary cancer-associated-fibroblasts (CAFs) showcased the potential of these cells in drug discovery application.
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Affiliation(s)
- Rizwan Ali
- Medical Research Core Facility and Platforms, King Abdullah International Medical Research Center (KAIMRC), King Saud bin Abdulaziz University for Health Sciences (KSAU-HS), MNGHA, Riyadh, Saudi Arabia
| | - Sarah Huwaizi
- Medical Research Core Facility and Platforms, King Abdullah International Medical Research Center (KAIMRC), King Saud bin Abdulaziz University for Health Sciences (KSAU-HS), MNGHA, Riyadh, Saudi Arabia
| | - Alshaimaa Alhallaj
- Medical Research Core Facility and Platforms, King Abdullah International Medical Research Center (KAIMRC), King Saud bin Abdulaziz University for Health Sciences (KSAU-HS), MNGHA, Riyadh, Saudi Arabia
| | - Arwa Al Subait
- Medical Research Core Facility and Platforms, King Abdullah International Medical Research Center (KAIMRC), King Saud bin Abdulaziz University for Health Sciences (KSAU-HS), MNGHA, Riyadh, Saudi Arabia
| | - Tlili Barhoumi
- Medical Research Core Facility and Platforms, King Abdullah International Medical Research Center (KAIMRC), King Saud bin Abdulaziz University for Health Sciences (KSAU-HS), MNGHA, Riyadh, Saudi Arabia
| | - Hajar Al Zahrani
- Medical Research Core Facility and Platforms, King Abdullah International Medical Research Center (KAIMRC), King Saud bin Abdulaziz University for Health Sciences (KSAU-HS), MNGHA, Riyadh, Saudi Arabia
| | - Abdullah Al Anazi
- Department of Pathology and Laboratory Medicine, King Abdulaziz Medical City (KAMC), MNGHA, Riyadh, Saudi Arabia
| | - Abdul Latif Khan
- Department of Pathology and Laboratory Medicine, King Abdulaziz Medical City (KAMC), MNGHA, Riyadh, Saudi Arabia
| | - Mohamed Boudjelal
- Medical Research Core Facility and Platforms, King Abdullah International Medical Research Center (KAIMRC), King Saud bin Abdulaziz University for Health Sciences (KSAU-HS), MNGHA, Riyadh, Saudi Arabia
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Shestovskaya MV, Bozhkova SA, Sopova JV, Khotin MG, Bozhokin MS. Methods of Modification of Mesenchymal Stem Cells and Conditions of Their Culturing for Hyaline Cartilage Tissue Engineering. Biomedicines 2021; 9:biomedicines9111666. [PMID: 34829895 PMCID: PMC8615732 DOI: 10.3390/biomedicines9111666] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/01/2021] [Revised: 11/04/2021] [Accepted: 11/05/2021] [Indexed: 12/24/2022] Open
Abstract
The use of mesenchymal stromal cells (MSCs) for tissue engineering of hyaline cartilage is a topical area of regenerative medicine that has already entered clinical practice. The key stage of this procedure is to create conditions for chondrogenic differentiation of MSCs, increase the synthesis of hyaline cartilage extracellular matrix proteins by these cells and activate their proliferation. The first such works consisted in the indirect modification of cells, namely, in changing the conditions in which they are located, including microfracturing of the subchondral bone and the use of 3D biodegradable scaffolds. The most effective methods for modifying the cell culture of MSCs are protein and physical, which have already been partially introduced into clinical practice. Genetic methods for modifying MSCs, despite their effectiveness, have significant limitations. Techniques have not yet been developed that allow studying the effectiveness of their application even in limited groups of patients. The use of MSC modification methods allows precise regulation of cell culture proliferation, and in combination with the use of a 3D biodegradable scaffold, it allows obtaining a hyaline-like regenerate in the damaged area. This review is devoted to the consideration and comparison of various methods used to modify the cell culture of MSCs for their use in regenerative medicine of cartilage tissue.
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Affiliation(s)
- Maria V. Shestovskaya
- Institute of Cytology of the Russian Academy of Sciences, Tikhoretsky Ave. 4, 194064 St. Petersburg, Russia; (M.V.S.); (J.V.S.); (M.G.K.)
| | - Svetlana A. Bozhkova
- Vreden National Medical Research Center of Traumatology and Orthopedics, Academica Baykova Str., 8, 195427 St. Petersburg, Russia;
| | - Julia V. Sopova
- Institute of Cytology of the Russian Academy of Sciences, Tikhoretsky Ave. 4, 194064 St. Petersburg, Russia; (M.V.S.); (J.V.S.); (M.G.K.)
- Center of Transgenesis and Genome Editing, St. Petersburg State University, Universitetskaja Emb., 7/9, 199034 St. Petersburg, Russia
| | - Mikhail G. Khotin
- Institute of Cytology of the Russian Academy of Sciences, Tikhoretsky Ave. 4, 194064 St. Petersburg, Russia; (M.V.S.); (J.V.S.); (M.G.K.)
| | - Mikhail S. Bozhokin
- Institute of Cytology of the Russian Academy of Sciences, Tikhoretsky Ave. 4, 194064 St. Petersburg, Russia; (M.V.S.); (J.V.S.); (M.G.K.)
- Vreden National Medical Research Center of Traumatology and Orthopedics, Academica Baykova Str., 8, 195427 St. Petersburg, Russia;
- Correspondence:
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Habanjar O, Diab-Assaf M, Caldefie-Chezet F, Delort L. 3D Cell Culture Systems: Tumor Application, Advantages, and Disadvantages. Int J Mol Sci 2021; 22:12200. [PMID: 34830082 PMCID: PMC8618305 DOI: 10.3390/ijms222212200] [Citation(s) in RCA: 251] [Impact Index Per Article: 62.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/27/2021] [Revised: 11/05/2021] [Accepted: 11/07/2021] [Indexed: 01/09/2023] Open
Abstract
The traditional two-dimensional (2D) in vitro cell culture system (on a flat support) has long been used in cancer research. However, this system cannot be fully translated into clinical trials to ideally represent physiological conditions. This culture cannot mimic the natural tumor microenvironment due to the lack of cellular communication (cell-cell) and interaction (cell-cell and cell-matrix). To overcome these limitations, three-dimensional (3D) culture systems are increasingly developed in research and have become essential for tumor research, tissue engineering, and basic biology research. 3D culture has received much attention in the field of biomedicine due to its ability to mimic tissue structure and function. The 3D matrix presents a highly dynamic framework where its components are deposited, degraded, or modified to delineate functions and provide a platform where cells attach to perform their specific functions, including adhesion, proliferation, communication, and apoptosis. So far, various types of models belong to this culture: either the culture based on natural or synthetic adherent matrices used to design 3D scaffolds as biomaterials to form a 3D matrix or based on non-adherent and/or matrix-free matrices to form the spheroids. In this review, we first summarize a comparison between 2D and 3D cultures. Then, we focus on the different components of the natural extracellular matrix that can be used as supports in 3D culture. Then we detail different types of natural supports such as matrigel, hydrogels, hard supports, and different synthetic strategies of 3D matrices such as lyophilization, electrospiding, stereolithography, microfluid by citing the advantages and disadvantages of each of them. Finally, we summarize the different methods of generating normal and tumor spheroids, citing their respective advantages and disadvantages in order to obtain an ideal 3D model (matrix) that retains the following characteristics: better biocompatibility, good mechanical properties corresponding to the tumor tissue, degradability, controllable microstructure and chemical components like the tumor tissue, favorable nutrient exchange and easy separation of the cells from the matrix.
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Affiliation(s)
- Ola Habanjar
- Université Clermont-Auvergne, INRAE, UNH, Unité de Nutrition Humaine, CRNH-Auvergne, 63000 Clermont-Ferrand, France; (O.H.); (F.C.-C.)
| | - Mona Diab-Assaf
- Equipe Tumorigénèse Pharmacologie Moléculaire et Anticancéreuse, Faculté des Sciences II, Université Libanaise Fanar, Beyrouth 1500, Liban;
| | - Florence Caldefie-Chezet
- Université Clermont-Auvergne, INRAE, UNH, Unité de Nutrition Humaine, CRNH-Auvergne, 63000 Clermont-Ferrand, France; (O.H.); (F.C.-C.)
| | - Laetitia Delort
- Université Clermont-Auvergne, INRAE, UNH, Unité de Nutrition Humaine, CRNH-Auvergne, 63000 Clermont-Ferrand, France; (O.H.); (F.C.-C.)
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20
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Kim H, Han Y, Suhito IR, Choi Y, Kwon M, Son H, Kim HR, Kim TH. Raman Spectroscopy-Based 3D Analysis of Odontogenic Differentiation of Human Dental Pulp Stem Cell Spheroids. Anal Chem 2021; 93:9995-10004. [PMID: 34241992 DOI: 10.1021/acs.analchem.0c05165] [Citation(s) in RCA: 13] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/25/2023]
Abstract
Numerous efforts have been made to establish three-dimensional (3D) cell cultures that mimic the structure, cell composition, and functions of actual tissues and organs in vitro; however, owing to its intrinsic complexity, precise characterization of 3D differentiation remains challenging and often results in high variations in the resulting differentiated spheroids. This study reports a 3D Raman mapping-based analytical method that helps us to identify the crucial factors responsible for inducing variability in differentiated stem cell spheroids. Human dental pulp stem cell spheroids were generated at various cell densities using the hanging drop (HD) and molded parafilm-based (MP) methods and were then further subjected to odontogenic differentiation. Thereafter, the 3D cellular differentiation in these spheroids was analyzed based on three different Raman peaks, namely, 960, 1156/1528, and 2935 cm-1, which correspond to hydroxyapatite (HA, odontogenic differentiation marker), β-carotene (precursor of HA), and proteins/cellular components (cell reference). By correlating such cell differentiation-related peaks and water/medium peaks (3400 cm-1), we discovered that the diffusion of the medium containing various nutrients and differentiation factors was crucial in determining the variations in 3D differentiation of stem cell spheroids. Odontogenic differentiation was majorly induced at the outer shell of HD spheroids (up to ∼20 μm), whereas odontogenic differentiation was markedly induced in MP spheroids (up to 40-50 μm). Considering the challenges associated with high variations in spheroid and organoid differentiation, we conclude that the proposed Raman-based 3D analysis plays a pivotal role in stem cell-based regenerative therapy and drug screening.
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Affiliation(s)
- Huijung Kim
- School of Integrative Engineering, Chung-Ang University, 84 Heukseok-ro, Dongjak-gu, Seoul 06974, Republic of Korea
| | - Yoojoong Han
- R&D Division, Nanobase, Inc., Seoul 08502, Republic of Korea
| | - Intan Rosalina Suhito
- School of Integrative Engineering, Chung-Ang University, 84 Heukseok-ro, Dongjak-gu, Seoul 06974, Republic of Korea
| | - Yoon Choi
- School of Integrative Engineering, Chung-Ang University, 84 Heukseok-ro, Dongjak-gu, Seoul 06974, Republic of Korea
| | - Minkyeong Kwon
- School of Integrative Engineering, Chung-Ang University, 84 Heukseok-ro, Dongjak-gu, Seoul 06974, Republic of Korea
| | - Hyungbin Son
- School of Integrative Engineering, Chung-Ang University, 84 Heukseok-ro, Dongjak-gu, Seoul 06974, Republic of Korea
| | - Hyung-Ryong Kim
- College of Dentistry, Dankook University, Cheonan 31116, Republic of Korea
| | - Tae-Hyung Kim
- School of Integrative Engineering, Chung-Ang University, 84 Heukseok-ro, Dongjak-gu, Seoul 06974, Republic of Korea.,Integrative Research Center for Two-Dimensional Functional Materials, Institute of Interdisciplinary Convergence Research, Chung-Ang University, Seoul 06974, Republic of Korea
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21
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Lobo DA, Ginestra P, Ceretti E, Miquel TP, Ciurana J. Cancer Cell Direct Bioprinting: A Focused Review. MICROMACHINES 2021; 12:764. [PMID: 34203530 PMCID: PMC8305105 DOI: 10.3390/mi12070764] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 05/26/2021] [Revised: 06/23/2021] [Accepted: 06/25/2021] [Indexed: 12/24/2022]
Abstract
Three-dimensional printing technologies allow for the fabrication of complex parts with accurate geometry and less production time. When applied to biomedical applications, two different approaches, known as direct or indirect bioprinting, may be performed. The classical way is to print a support structure, the scaffold, and then culture the cells. Due to the low efficiency of this method, direct bioprinting has been proposed, with or without the use of scaffolds. Scaffolds are the most common technology to culture cells, but bioassembly of cells may be an interesting methodology to mimic the native microenvironment, the extracellular matrix, where the cells interact between themselves. The purpose of this review is to give an updated report about the materials, the bioprinting technologies, and the cells used in cancer research for breast, brain, lung, liver, reproductive, gastric, skin, and bladder associated cancers, to help the development of possible treatments to lower the mortality rates, increasing the effectiveness of guided therapies. This work introduces direct bioprinting to be considered as a key factor above the main tissue engineering technologies.
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Affiliation(s)
- David Angelats Lobo
- Department of Mechanical and Industrial Engineering, University of Brescia, V. Branze 38, 25123 Brescia, Italy; (D.A.L.); (E.C.)
- New Therapeutic Targets Laboratory (TargetsLab), Oncology Unit, Department of Medical Sciences, Girona Institute for Biomedical Research, University of Girona, Emili Grahit 77, 17003 Girona, Spain;
| | - Paola Ginestra
- Department of Mechanical and Industrial Engineering, University of Brescia, V. Branze 38, 25123 Brescia, Italy; (D.A.L.); (E.C.)
| | - Elisabetta Ceretti
- Department of Mechanical and Industrial Engineering, University of Brescia, V. Branze 38, 25123 Brescia, Italy; (D.A.L.); (E.C.)
| | - Teresa Puig Miquel
- New Therapeutic Targets Laboratory (TargetsLab), Oncology Unit, Department of Medical Sciences, Girona Institute for Biomedical Research, University of Girona, Emili Grahit 77, 17003 Girona, Spain;
| | - Joaquim Ciurana
- Product, Process and Production Engineering Research Group (GREP), Department of Mechanical Engineering and Industrial Construction, University of Girona, Maria Aurèlia Capmany 61, 17003 Girona, Spain;
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22
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Selim OA, Lakhani S, Midha S, Mosahebi A, Kalaskar DM. Three-Dimensional Engineered Peripheral Nerve: Toward a New Era of Patient-Specific Nerve Repair Solutions. TISSUE ENGINEERING PART B-REVIEWS 2021; 28:295-335. [PMID: 33593147 DOI: 10.1089/ten.teb.2020.0355] [Citation(s) in RCA: 15] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/22/2022]
Abstract
Reconstruction of peripheral nerve injuries (PNIs) with substance loss remains challenging because of limited treatment solutions and unsatisfactory patient outcomes. Currently, nerve autografting is the first-line management choice for bridging critical-sized nerve defects. The procedure, however, is often complicated by donor site morbidity and paucity of nerve tissue, raising a quest for better alternatives. The application of other treatment surrogates, such as nerve guides, remains questionable, and it is inefficient in irreducible nerve gaps. More importantly, these strategies lack customization for personalized patient therapy, which is a significant drawback of these nerve repair options. This negatively impacts the fascicle-to-fascicle regeneration process, critical to restoring the physiological axonal pathway of the disrupted nerve. Recently, the use of additive manufacturing (AM) technologies has offered major advancements to the bioengineering solutions for PNI therapy. These techniques aim at reinstating the native nerve fascicle pathway using biomimetic approaches, thereby augmenting end-organ innervation. AM-based approaches, such as three-dimensional (3D) bioprinting, are capable of biofabricating 3D-engineered nerve graft scaffolds in a patient-specific manner with high precision. Moreover, realistic in vitro models of peripheral nerve tissues that represent the physiologically and functionally relevant environment of human organs could also be developed. However, the technology is still nascent and faces major translational hurdles. In this review, we spotlighted the clinical burden of PNIs and most up-to-date treatment to address nerve gaps. Next, a summarized illustration of the nerve ultrastructure that guides research solutions is discussed. This is followed by a contrast of the existing bioengineering strategies used to repair peripheral nerve discontinuities. In addition, we elaborated on the most recent advances in 3D printing and biofabrication applications in peripheral nerve modeling and engineering. Finally, the major challenges that limit the evolution of the field along with their possible solutions are also critically analyzed.
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Affiliation(s)
- Omar A Selim
- Department of Surgical Biotechnology, Division of Surgery and Interventional Sciences, Royal Free Hospital, University College London (UCL), London, United Kingdom
| | - Saad Lakhani
- Department of Surgical Biotechnology, Division of Surgery and Interventional Sciences, Royal Free Hospital, University College London (UCL), London, United Kingdom
| | - Swati Midha
- Department of Surgical Biotechnology, Division of Surgery and Interventional Sciences, Royal Free Hospital, University College London (UCL), London, United Kingdom.,Department of Surgical Biotechnology, Special Centre for Nanoscience, Jawaharlal Nehru University, New Delhi, India
| | - Afshin Mosahebi
- Department of Plastic Surgery, Royal Free Hospital, University College London (UCL), London, United Kingdom
| | - Deepak M Kalaskar
- Department of Surgical Biotechnology, Division of Surgery and Interventional Sciences, Royal Free Hospital, University College London (UCL), London, United Kingdom.,Department of Surgical Biotechnology, Institute of Orthopaedics and Musculoskeletal Science, Royal National Orthopaedic Hospital, University College London (UCL), Stanmore, United Kingdom
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23
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Functional Properties of Human-Derived Mesenchymal Stem Cell Spheroids: A Meta-Analysis and Systematic Review. Stem Cells Int 2021; 2021:8825332. [PMID: 33884001 PMCID: PMC8041538 DOI: 10.1155/2021/8825332] [Citation(s) in RCA: 18] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/23/2020] [Revised: 01/31/2021] [Accepted: 02/12/2021] [Indexed: 02/06/2023] Open
Abstract
Mesenchymal stem cells (MSC) are adult multi-potent cells that can be isolated from many types of tissues including adipose tissue, bone marrow, and umbilical cord. They show great potential for cell therapy-based treatments, which is why they are being used in numerous clinical trials for a wide range of diseases. However, the success of placebo-controlled clinical trials has been limited, so new ways of improving the therapeutic effects of MSC are being developed, such as their assembly in a 3D conformation. In this meta-analysis, we review aggregate formation, in vitro functional properties and in vivo therapeutic potential displayed by adipose tissue, bone marrow, and umbilical cord-derived MSC, assembled as spheroids. The databases PubMed and SciELO were used to find eligible articles, using free-words and MeSH terms related to the subject, finding 28 published articles meeting all inclusion and exclusion criteria. Of the articles selected 15 corresponded to studies using MSC derived from bone marrow, 10 from adipose tissue and 3 from umbilical cord blood or tissue. The MSC spheroids properties analyzed that displayed enhancement in comparison with monolayer 2D culture, are stemness, angiogenesis, differentiation potential, cytokine secretion, paracrine and immunomodulatory effects. Overall studies reveal that the application of MSC spheroids in vivo enhanced therapeutic effects. For instance, research exhibited reduced inflammation, faster wound healing, and closure, functional recovery and tissue repair due to immunomodulatory effects, better MSC engraftment in damaged tissue, higher MSC survival and less apoptosis at the injury. Still, further research and clinical studies with controlled and consistent results are needed to see the real therapeutic efficacy of MSC spheroids.
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24
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Nasser MI, Qi X, Zhu S, He Y, Zhao M, Guo H, Zhu P. Current situation and future of stem cells in cardiovascular medicine. Biomed Pharmacother 2020; 132:110813. [PMID: 33068940 DOI: 10.1016/j.biopha.2020.110813] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/02/2020] [Revised: 09/22/2020] [Accepted: 09/25/2020] [Indexed: 12/21/2022] Open
Abstract
Cardiovascular disease (CVD) is one of the leading causes of death worldwide. Currently, many methods have been proposed by researchers for the prevention and treatment of CVD; among them, stem cell-based therapies are the most promising. As the cells of origin for various mature cells, stem cells have the ability to self-renew and differentiate. Stem cells have a powerful ability to regenerate biologically, self-repair, and enhance damaged functional tissues or organs. Allogeneic stem cells and somatic stem cells are two types of cells that can be used for cardiac repair. Theoretically, dilated cardiomyopathy and acute myocardial infarction can be treated with such cells. In addition, stem cell transplantation procedures, including intravenous, epicardial, cardiac, and endocardial injections, have been reported to provide significant benefits in clinical practice; however, there are still a number of issues that need further study and consideration, such as the form and quantity of transplanted cells and post-transplantation health. The goal of this analysis was to summarize the recent advances in stem cell-based therapies and their efficacy in cardiovascular regenerative medicine.
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Affiliation(s)
- M I Nasser
- Guangdong Cardiovascular Institute, Guangdong Provincial People's Hospital, Guangdong Academy of Medical Sciences, Guangzhou, Guangdong, 510100, China. Address: 106 Zhongshan Er Road, Guangzhou, 510080, PR China
| | - Xiao Qi
- Guangdong Cardiovascular Institute, Guangdong Provincial People's Hospital, Guangdong Academy of Medical Sciences, Guangzhou, Guangdong, 510100, China. Address: 106 Zhongshan Er Road, Guangzhou, 510080, PR China
| | - Shuoji Zhu
- Guangdong Cardiovascular Institute, Guangdong Provincial People's Hospital, Guangdong Academy of Medical Sciences, Guangzhou, Guangdong, 510100, China. Address: 106 Zhongshan Er Road, Guangzhou, 510080, PR China
| | - Yin He
- Guangdong Cardiovascular Institute, Guangdong Provincial People's Hospital, Guangdong Academy of Medical Sciences, Guangzhou, Guangdong, 510100, China. Address: 106 Zhongshan Er Road, Guangzhou, 510080, PR China
| | - Mingyi Zhao
- Guangdong Cardiovascular Institute, Guangdong Provincial People's Hospital, Guangdong Academy of Medical Sciences, Guangzhou, Guangdong, 510100, China. Address: 106 Zhongshan Er Road, Guangzhou, 510080, PR China
| | - Huiming Guo
- Guangdong Cardiovascular Institute, Guangdong Provincial People's Hospital, Guangdong Academy of Medical Sciences, Guangzhou, Guangdong, 510100, China. Address: 106 Zhongshan Er Road, Guangzhou, 510080, PR China
| | - Ping Zhu
- Guangdong Cardiovascular Institute, Guangdong Provincial People's Hospital, Guangdong Academy of Medical Sciences, Guangzhou, Guangdong, 510100, China. Address: 106 Zhongshan Er Road, Guangzhou, 510080, PR China.
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25
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Soder RP, Dawn B, Weiss ML, Dunavin N, Weir S, Mitchell J, Li M, Shune L, Singh AK, Ganguly S, Morrison M, Abdelhakim H, Godwin AK, Abhyankar S, McGuirk J. A Phase I Study to Evaluate Two Doses of Wharton's Jelly-Derived Mesenchymal Stromal Cells for the Treatment of De Novo High-Risk or Steroid-Refractory Acute Graft Versus Host Disease. Stem Cell Rev Rep 2020; 16:979-991. [PMID: 32740891 PMCID: PMC9289888 DOI: 10.1007/s12015-020-10015-8] [Citation(s) in RCA: 23] [Impact Index Per Article: 4.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
Abstract
BACKGROUND Because of their well-described immunosuppressive properties, allogeneic adult human mesenchymal stromal cells (MSC) derived from bone marrow have demonstrated safety and efficacy in steroid refractory acute graft versus host disease (SR aGVHD). Clinical trials have resulted in variable success and an optimal source of MSC has yet to be defined. Based on the importance of maternal-fetal interface immune tolerance, extraembryonic fetal tissues, such as the umbilical cord, may provide an superior tissue source of MSC to mediate immunomodulation in aGVHD. METHODS A two-dose cohort trial allogeneic Wharton's Jelly-derived mesenchymal stromal cells (WJMSC, referred to as MSCTC-0010, here) were tested in 10 patients with de novo high risk (HR) or SR aGVHD post allogeneic hematopoietic stem cell transplantation (allo-HCT). Following Good Manufacturing Practices isolation, expansion and cryostorage, WJMSC were thawed and administered via intravenous infusions on days 0 and 7 at one of two doses (low dose cohort, 2 × 106/kg, n = 5; high dose cohort, 10 × 106/kg, n = 5). To evaluate safety, patients were monitored for infusion related toxicity, Treatment Related Adverse Events (TRAE) til day 42, or ectopic tissue formation at day 90. Clinical responses were monitored at time points up to 180 days post infusion. Serum biomarkers ST2 and REG3α were acquired 1 day prior to first MSCTC-0010 infusion and on day 14. RESULTS Safety was indicated, e.g., no infusion-related toxicity, no development of TRAE, nor ectopic tissue formation in either low or high dose cohort was observed. Clinical response was suggested at day 28: the overall response rate (ORR) was 70%, 4 of 10 patients had a complete response (CR) and 3 had a partial response (PR). By study day 90, the addition of escalated immunosuppressive therapy was necessary in 2 of 9 surviving patients. Day 100 and 180 post infusion survival was 90% and 60%, respectively. Serum biomarker REG3α decreased, particularly in the high dose cohort, and with REG3α decrease correlated with clinical response. CONCLUSIONS Treatment of patients with de novo HR or SR aGVHD with low or high dose MSCTC-0010 was safe: the infusion was well-tolerated, and no TRAEs or ectopic tissue formation was observed. A clinical improvement was seen in about 70% patients, with 4 of 10 showing a complete response that may have been attributable to MSCTC-0010 infusions. These observations indicate safety of two different doses of MSCTC-0010, and suggest that the 10 × 106 cells/ kg dose be tested in an expanded randomized, controlled Phase 2 trial. Graphical abstract.
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Affiliation(s)
- Rupal P Soder
- Midwest Stem Cell Therapy Center, University of Kansas Medical Center, Kansas City, KS, USA
| | - Buddhadeb Dawn
- University of Nevada, Las Vegas School of Medicine, Las Vegas, NV, USA
| | - Mark L Weiss
- Midwest Institute of Comparative Stem Cell Biotechnology and Department of Anatomy and Physiology, Kansas State University, Manhattan, KS, USA
| | - Neil Dunavin
- University of California, San Francisco, CA, USA
| | - Scott Weir
- Institute for Advancing Medical Innovation Medical Center, University of Kansas, Kansas City, USA
| | - James Mitchell
- Midwest Stem Cell Therapy Center, University of Kansas Medical Center, Kansas City, KS, USA
| | - Meizhang Li
- Pathology & Laboratory Medicine, Univeristy of Kansas Medical Center, Kansas City, USA
| | - Leyla Shune
- Blood and Marrow Transplant Program, Division of Hematologic Malignancies and Cellular Therapeutics, University of Kansas Medical Center, 2330 Shawnee Mission Parkway, Suite 210, Westwood, KS, 66205, USA
| | - Anurag K Singh
- Blood and Marrow Transplant Program, Division of Hematologic Malignancies and Cellular Therapeutics, University of Kansas Medical Center, 2330 Shawnee Mission Parkway, Suite 210, Westwood, KS, 66205, USA
| | - Siddhartha Ganguly
- Blood and Marrow Transplant Program, Division of Hematologic Malignancies and Cellular Therapeutics, University of Kansas Medical Center, 2330 Shawnee Mission Parkway, Suite 210, Westwood, KS, 66205, USA
| | - Marc Morrison
- Blood and Marrow Transplant Program, Division of Hematologic Malignancies and Cellular Therapeutics, University of Kansas Medical Center, 2330 Shawnee Mission Parkway, Suite 210, Westwood, KS, 66205, USA
| | - Haitham Abdelhakim
- Blood and Marrow Transplant Program, Division of Hematologic Malignancies and Cellular Therapeutics, University of Kansas Medical Center, 2330 Shawnee Mission Parkway, Suite 210, Westwood, KS, 66205, USA
| | - Andrew K Godwin
- Pathology & Laboratory Medicine, Univeristy of Kansas Medical Center, Kansas City, USA
| | - Sunil Abhyankar
- Midwest Stem Cell Therapy Center, University of Kansas Medical Center, Kansas City, KS, USA
- Blood and Marrow Transplant Program, Division of Hematologic Malignancies and Cellular Therapeutics, University of Kansas Medical Center, 2330 Shawnee Mission Parkway, Suite 210, Westwood, KS, 66205, USA
| | - Joseph McGuirk
- Blood and Marrow Transplant Program, Division of Hematologic Malignancies and Cellular Therapeutics, University of Kansas Medical Center, 2330 Shawnee Mission Parkway, Suite 210, Westwood, KS, 66205, USA.
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26
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Soheilmoghaddam M, Padmanabhan H, Cooper-White JJ. Biomimetic cues from poly(lactic-co-glycolic acid)/hydroxyapatite nano-fibrous scaffolds drive osteogenic commitment in human mesenchymal stem cells in the absence of osteogenic factor supplements. Biomater Sci 2020; 8:5677-5689. [PMID: 32915185 DOI: 10.1039/d0bm00946f] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/21/2022]
Abstract
Mimicking the complex hierarchical architecture of the 'osteon', the functional unit of cortical bone, from the bottom-up offers the possibility of generating mature bone tissue in tissue engineered bone substitutes. In this work, a modular 'bottom-up' approach has been developed to assemble bone niche-mimicking nanocomposite scaffolds composed of aligned electrospun nanofibers of poly(lactic-co-glycolic acid) (PLGA) encapsulating aligned rod-shape nano-sized hydroxyapatite (nHA). By encoding axial orientation of the nHA within these aligned nanocomposite fibers, significant improvements in mechanical properties, surface roughness, hydrophilicity and in vitro simulated body fluid (SBF) mineral deposition were achieved. Moreover, these hierarchical scaffolds induced robust formation of bone hydroxyapatite and osteoblastic maturation of human bone marrow-derived mesenchymal stem cells (hBMSCs) in growth media that was absent of any soluble osteogenic differentiation factors. The results of this investigation confirm that these tailored, aligned nanocomposite fibers, in the absence of media-bone inductive factors, offer the requisite biophysical and biochemical cues to hBMSCs to promote and support their differentiation into mature osteoblast cells and form early bone-like tissue in vitro.
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Affiliation(s)
- Mohammad Soheilmoghaddam
- Tissue Engineering and Microfluidics Laboratory (TE&M), Australian Institute for Bioengineering and Nanotechnology (AIBN), University Of Queensland, St Lucia, QLD, Australia.
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27
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Shanbhag S, Suliman S, Bolstad AI, Stavropoulos A, Mustafa K. Xeno-Free Spheroids of Human Gingiva-Derived Progenitor Cells for Bone Tissue Engineering. Front Bioeng Biotechnol 2020; 8:968. [PMID: 32974308 PMCID: PMC7466771 DOI: 10.3389/fbioe.2020.00968] [Citation(s) in RCA: 12] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/22/2020] [Accepted: 07/27/2020] [Indexed: 12/19/2022] Open
Abstract
Gingiva has been identified as a minimally invasive source of multipotent progenitor cells (GPCs) for use in bone tissue engineering (BTE). To facilitate clinical translation, it is important to characterize GPCs in xeno-free cultures. Recent evidence indicates several advantages of three-dimensional (3D) spheroid cultures of mesenchymal stromal cells (MSCs) over conventional 2D monolayers. The present study aimed to characterize human GPCs in xeno-free 2D cultures, and to test their osteogenic potential in 3D cultures, in comparison to bone marrow MSCs (BMSCs). Primary GPCs and BMSCs were expanded in human platelet lysate (HPL) or fetal bovine serum (FBS) and characterized based on in vitro proliferation, immunophenotype and multi-lineage differentiation. Next, 3D spheroids of GPCs and BMSCs were formed via self-assembly and cultured in HPL. Expression of stemness- (SOX2, OCT4, NANOG) and osteogenesis-related markers (BMP2, RUNX2, OPN, OCN) was assessed at gene and protein levels in 3D and 2D cultures. The cytokine profile of 3D and 2D GPCs and BMSCs was assessed via a multiplex immunoassay. Monolayer GPCs in both HPL and FBS demonstrated a characteristic MSC-like immunophenotype and multi-lineage differentiation; osteogenic differentiation of GPCs was enhanced in HPL vs. FBS. CD271+ GPCs in HPL spontaneously acquired a neuronal phenotype and strongly expressed neuronal/glial markers. 3D spheroids of GPCs and BMSCs with high cell viability were formed in HPL media. Expression of stemness- and osteogenesis-related genes was significantly upregulated in 3D vs. 2D GPCs/BMSCs; the latter was independent of osteogenic induction. Synthesis of SOX2, BMP2 and OCN was confirmed via immunostaining, and in vitro mineralization via Alizarin red staining. Finally, secretion of several growth factors and chemokines was enhanced in GPC/BMSC spheroids, while that of pro-inflammatory cytokines was reduced, compared to monolayers. In summary, monolayer GPCs expanded in HPL demonstrate enhanced osteogenic differentiation potential, comparable to that of BMSCs. Xeno-free spheroid culture further enhances stemness- and osteogenesis-related gene expression, and cytokine secretion in GPCs, comparable to that of BMSCs.
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Affiliation(s)
- Siddharth Shanbhag
- Department of Clinical Dentistry, Faculty of Medicine, University of Bergen, Bergen, Norway
| | - Salwa Suliman
- Department of Clinical Dentistry, Faculty of Medicine, University of Bergen, Bergen, Norway
| | - Anne Isine Bolstad
- Department of Clinical Dentistry, Faculty of Medicine, University of Bergen, Bergen, Norway
| | - Andreas Stavropoulos
- Department of Periodontology, Faculty of Odontology, Malmö University, Malmö, Sweden.,Division of Regenerative Medicine and Periodontology, University Clinics of Dental Medicine, University of Geneva, Geneva, Switzerland
| | - Kamal Mustafa
- Department of Clinical Dentistry, Faculty of Medicine, University of Bergen, Bergen, Norway
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28
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Burand AJ, Di L, Boland LK, Boyt DT, Schrodt MV, Santillan DA, Ankrum JA. Aggregation of Human Mesenchymal Stromal Cells Eliminates Their Ability to Suppress Human T Cells. Front Immunol 2020; 11:143. [PMID: 32158443 PMCID: PMC7052295 DOI: 10.3389/fimmu.2020.00143] [Citation(s) in RCA: 18] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/05/2019] [Accepted: 01/20/2020] [Indexed: 12/15/2022] Open
Abstract
Mesenchymal stromal cells (MSCs) are administered locally to treat sites of inflammation. Local delivery is known to cause MSCs to aggregate into “spheroids,” which alters gene expression and phenotype. While adherent MSCs are highly efficient in their inhibition of T cells, whether or not this property is altered upon MSC aggregation has not been thoroughly determined. In this study, we discovered that aggregation of MSCs into spheroids causes them to lose their T cell-suppressive abilities. Interestingly, adding budesonide, a topical glucocorticoid steroid, alongside spheroids partially restored MSC suppression of T cell proliferation. Through a series of inhibition and add-back studies, we determined budesonide acts synergistically with spheroid MSC-produced PGE2 to suppress T cell proliferation through the PGE2 receptors EP2 and EP4. These findings highlight critical differences between adherent and spheroid MSC interactions with human immune cells that have significant translational consequences. In addition, we uncovered a mechanism through which spheroid MSC suppression of T cells can be partly restored. By understanding the phenotypic changes that occur upon MSC aggregation and the impact of MSC drug interactions, improved immunosuppressive MSC therapies for localized delivery can be designed.
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Affiliation(s)
- Anthony J Burand
- Roy J. Carver Department of Biomedical Engineering, College of Engineering, University of Iowa, Iowa City, IA, United States.,Fraternal Order of Eagles Diabetes Research Center, University of Iowa, Iowa City, IA, United States
| | - Lin Di
- Roy J. Carver Department of Biomedical Engineering, College of Engineering, University of Iowa, Iowa City, IA, United States.,Fraternal Order of Eagles Diabetes Research Center, University of Iowa, Iowa City, IA, United States
| | - Lauren K Boland
- Roy J. Carver Department of Biomedical Engineering, College of Engineering, University of Iowa, Iowa City, IA, United States.,Fraternal Order of Eagles Diabetes Research Center, University of Iowa, Iowa City, IA, United States
| | - Devlin T Boyt
- Roy J. Carver Department of Biomedical Engineering, College of Engineering, University of Iowa, Iowa City, IA, United States.,Fraternal Order of Eagles Diabetes Research Center, University of Iowa, Iowa City, IA, United States
| | - Michael V Schrodt
- Roy J. Carver Department of Biomedical Engineering, College of Engineering, University of Iowa, Iowa City, IA, United States.,Fraternal Order of Eagles Diabetes Research Center, University of Iowa, Iowa City, IA, United States
| | - Donna A Santillan
- Department of Obstetrics and Gynecology, Carver College of Medicine, University of Iowa, Iowa City, IA, United States.,Center for Immunology and Immune Based Diseases, Roy J. and Lucille A. Carver College of Medicine, University of Iowa, Iowa City, IA, United States.,Center for Hypertension Research, University of Iowa, Iowa City, IA, United States
| | - James A Ankrum
- Roy J. Carver Department of Biomedical Engineering, College of Engineering, University of Iowa, Iowa City, IA, United States.,Fraternal Order of Eagles Diabetes Research Center, University of Iowa, Iowa City, IA, United States
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29
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Spheroid Culture System Methods and Applications for Mesenchymal Stem Cells. Cells 2019; 8:cells8121620. [PMID: 31842346 PMCID: PMC6953111 DOI: 10.3390/cells8121620] [Citation(s) in RCA: 306] [Impact Index Per Article: 51.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/26/2019] [Revised: 12/09/2019] [Accepted: 12/09/2019] [Indexed: 12/16/2022] Open
Abstract
Owing to the importance of stem cell culture systems in clinical applications, researchers have extensively studied them to optimize the culture conditions and increase efficiency of cell culture. A spheroid culture system provides a similar physicochemical environment in vivo by facilitating cell–cell and cell–matrix interaction to overcome the limitations of traditional monolayer cell culture. In suspension culture, aggregates of adjacent cells form a spheroid shape having wide utility in tumor and cancer research, therapeutic transplantation, drug screening, and clinical study, as well as organic culture. There are various spheroid culture methods such as hanging drop, gel embedding, magnetic levitation, and spinner culture. Lately, efforts are being made to apply the spheroid culture system to the study of drug delivery platforms and co-cultures, and to regulate differentiation and pluripotency. To study spheroid cell culture, various kinds of biomaterials are used as building forms of hydrogel, film, particle, and bead, depending upon the requirement. However, spheroid cell culture system has limitations such as hypoxia and necrosis in the spheroid core. In addition, studies should focus on methods to dissociate cells from spheroid into single cells.
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Dong G, Wang S, Ge Y, Deng Q, Cao Q, Wang Q, Shang Z, OuYang W, Li J, Liu C, Tang J, Zhao W, Gu Y. Serum-Free Culture System for Spontaneous Human Mesenchymal Stem Cell Spheroid Formation. Stem Cells Int 2019; 2019:6041816. [PMID: 31737076 PMCID: PMC6815607 DOI: 10.1155/2019/6041816] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/15/2019] [Revised: 08/12/2019] [Accepted: 09/03/2019] [Indexed: 12/19/2022] Open
Abstract
Human mesenchymal stem cells (hMSCs) are widely used in clinical research because of their multipotential, immunomodulatory, and reparative properties. Previous studies determined that hMSC spheroids from a three-dimensional (3D) culture possess higher therapeutic efficacy than conventional hMSCs from a monolayer (2D) culture. To date, various 3D culture methods have been developed to form hMSC spheroids but most of them used culture medium containing fetal bovine serum (FBS), which is not suitable for further clinical use. Here, we demonstrate that dissociated single MSCs seeded in induced pluripotent stem medium (MiPS) adhere loosely to the dish and spontaneously migrate to form spheroids during day 3 to day 6. Through component deletion screening and complementation experiments, the knockout serum replacement (KSR) was identified as necessary and sufficient for hMSC spheroid formation. Transcriptome analysis showed that the overall expression profiles were highly similar between 2D culture with FBS and KSR-derived spheroids. Interestingly, genes related to inflammatory response, immune response, and angiogenesis were upregulated in spheroids at day 6 and qPCR results further validated the increased expression level of related genes, including STC1, CCL7, HGF, IL24, and TGFB3. When spheroids were replated in normal FBS medium, cells formed a typical spindle-shaped morphology and FACS results showed that the recovered cells retained MSC-specific surface markers, such as CD73, CD90, and CD105. In summary, we developed a practical and convenient method to generate hMSC spheroids for clinical research and therapy.
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Affiliation(s)
- Guoyi Dong
- BGI-Shenzhen, Shenzhen 518083, China
- China National GeneBank, BGI-Shenzhen, Shenzhen 518120, China
| | - Shengpeng Wang
- BGI-Shenzhen, Shenzhen 518083, China
- China National GeneBank, BGI-Shenzhen, Shenzhen 518120, China
| | - Yuping Ge
- BGI-Shenzhen, Shenzhen 518083, China
- China National GeneBank, BGI-Shenzhen, Shenzhen 518120, China
| | - Qiuting Deng
- BGI-Shenzhen, Shenzhen 518083, China
- China National GeneBank, BGI-Shenzhen, Shenzhen 518120, China
| | - Qi Cao
- BGI-Shenzhen, Shenzhen 518083, China
| | - Quanlei Wang
- BGI-Shenzhen, Shenzhen 518083, China
- China National GeneBank, BGI-Shenzhen, Shenzhen 518120, China
| | - Zhouchun Shang
- BGI-Shenzhen, Shenzhen 518083, China
- China National GeneBank, BGI-Shenzhen, Shenzhen 518120, China
| | - Wenjie OuYang
- BGI-Shenzhen, Shenzhen 518083, China
- China National GeneBank, BGI-Shenzhen, Shenzhen 518120, China
| | - Jing Li
- BGI-Shenzhen, Shenzhen 518083, China
- China National GeneBank, BGI-Shenzhen, Shenzhen 518120, China
| | - Chao Liu
- BGI-Shenzhen, Shenzhen 518083, China
- China National GeneBank, BGI-Shenzhen, Shenzhen 518120, China
| | - Jie Tang
- Shenzhen Second People's Hospital, First Affiliated Hospital of Shenzhen University, Shenzhen 518035, Guangdong, China
| | - Weihua Zhao
- Shenzhen Second People's Hospital, First Affiliated Hospital of Shenzhen University, Shenzhen 518035, Guangdong, China
| | - Ying Gu
- BGI-Shenzhen, Shenzhen 518083, China
- China National GeneBank, BGI-Shenzhen, Shenzhen 518120, China
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Tan HX, Cao ZB, He TT, Huang T, Xiang CL, Liu Y. TGFβ1 is essential for MSCs-CAFs differentiation and promotes HCT116 cells migration and invasion via JAK/STAT3 signaling. Onco Targets Ther 2019; 12:5323-5334. [PMID: 31308702 PMCID: PMC6615717 DOI: 10.2147/ott.s178618] [Citation(s) in RCA: 40] [Impact Index Per Article: 6.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/28/2018] [Accepted: 04/17/2019] [Indexed: 12/12/2022] Open
Abstract
Background and purpose Colorectal cancer (CRC) frequently metastasizes to the liver, which involves the participation of multiple cytokines. Tumor microenvironment (TME) composed of cancer-associated fibroblasts (CAFs) and tumor cells acts as an essential factor in cancer metastasis. Transforming growth factor β1 (TGFβ1) is a vital cytokine involved in migration and invasion of cancer cells. However, the underlying mechanisms remain unclear. In the present study, we aimed to investigate the role and molecular mechanisms of TGFβ1 in TME. Methods The conditioned medium prepared from colorectal cancer HCT116 and HT29 cells was used to culture mesenchymal stem cells (MSCs). The differentiation of MSCs to CAFs was detected by flow cytometry. The role of TGFβ1 in colorectal cancer cells metastasis was examined by wound-healing assay and transwell assay. And the activation of the Janus kinase/signal transducer and activator of transcription 3 (JAK/STAT3) signaling pathway was measured by Western blot assay. Results TGFβ1 induced the differentiation of MSCs to CAFs and improved HCT116 and HT29 cells migration and invasion. Meanwhile, TGFβ1 also upregulated the phosphorylation of STAT3 and enhanced the nuclear localization of p-STAT3, which activated JAK/STAT3 signaling pathway. Conclusion TGFβ1 induced the differentiation of MSCs into CAFs and promoted the migration and invasion of HCT116 and HT29 cells, which depended on the activation of JAK/STAT3 signaling pathway.
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Affiliation(s)
- Hao-Xiang Tan
- Department of Gastrointestinal Surgery, Hunan Provincial People's Hospital, Changsha 410002, People's Republic of China.,Department of Gastrointestinal Surgery, Ruikang Hospital Affiliated to Guangxi University of Chinese Medicine, Nanning 530011, People's Republic of China.,Department of Gastrointestinal Surgery, First Affiliated Hospital of Hunan Normal University, Changsha 410002, People's Republic of China
| | - Zhen-Bin Cao
- Department of Radiology, Ruikang Hospital Affiliated to Guangxi University of Chinese Medicine, Nanning 530011, People's Republic of China
| | - Ting-Ting He
- Department of Pathology, Ruikang Hospital Affiliated to Guangxi University of Chinese Medicine, Nanning 530011, People's Republic of China
| | - Tao Huang
- Department of Gastrointestinal Surgery, Hunan Provincial People's Hospital, Changsha 410002, People's Republic of China.,Department of Gastrointestinal Surgery, First Affiliated Hospital of Hunan Normal University, Changsha 410002, People's Republic of China
| | - Cai-Ling Xiang
- Department of Gastrointestinal Surgery, Hunan Provincial People's Hospital, Changsha 410002, People's Republic of China.,Department of Gastrointestinal Surgery, First Affiliated Hospital of Hunan Normal University, Changsha 410002, People's Republic of China
| | - Yu Liu
- Department of Gastrointestinal Surgery, Hunan Provincial People's Hospital, Changsha 410002, People's Republic of China.,Department of Gastrointestinal Surgery, First Affiliated Hospital of Hunan Normal University, Changsha 410002, People's Republic of China
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Spontaneously Formed Spheroids from Mouse Compact Bone-Derived Cells Retain Highly Potent Stem Cells with Enhanced Differentiation Capability. Stem Cells Int 2019; 2019:8469012. [PMID: 31191686 PMCID: PMC6525826 DOI: 10.1155/2019/8469012] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/19/2018] [Revised: 02/26/2019] [Accepted: 03/10/2019] [Indexed: 02/07/2023] Open
Abstract
The results from our recent study showed the presence of two distinct spheroid-forming mechanisms, i.e., spontaneous and mechanical. In this study, we focused on the spontaneously formed spheroids, and the character of spontaneously formed spheroids from mouse compact bone-derived cells (CBDCs) was explored. Cells from (C57BL/6J) mouse leg bones were isolated, and compact bone-derived cells were cultured after enzymatic digestion. Spontaneous spheroid formation was achieved on a culture plate with specific water contact angle as reported. The expression levels of embryonic stem cell markers were analyzed using immunofluorescence and quantitative reverse transcription polymerase chain reaction. Then, the cells from spheroids were induced into osteogenic and neurogenic lineages. The spontaneously formed spheroids from CBDCs were positive for ES cell markers such as SSEA1, Sox2, Oct4, and Nanog. Additionally, the expressions of fucosyltransferase 4/FUT4 (SSEA1), Sox2, and Nanog were significantly higher than those in monolayer cultured cells. The gene expression of mesenchymal stem cell markers was almost identical in both spheroids and monolayer-cultured cells, but the expression of Sca-1 was higher in spheroids. Spheroid-derived cells showed significantly higher osteogenic and neurogenic marker expression than monolayer-cultured cells after induction. Spontaneously formed spheroids expressed stem cell markers and showed enhanced osteogenic and neurogenic differentiation capabilities than cells from the conventional monolayer culture, which supports the superior stemness.
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Noronha NDC, Mizukami A, Caliári-Oliveira C, Cominal JG, Rocha JLM, Covas DT, Swiech K, Malmegrim KCR. Priming approaches to improve the efficacy of mesenchymal stromal cell-based therapies. Stem Cell Res Ther 2019; 10:131. [PMID: 31046833 PMCID: PMC6498654 DOI: 10.1186/s13287-019-1224-y] [Citation(s) in RCA: 383] [Impact Index Per Article: 63.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022] Open
Abstract
Multipotent mesenchymal stromal cells (MSC) have been widely explored for cell-based therapy of immune-mediated, inflammatory, and degenerative diseases, due to their immunosuppressive, immunomodulatory, and regenerative potentials. Preclinical studies and clinical trials have demonstrated promising therapeutic results although these have been somewhat limited. Aspects such as low in vivo MSC survival in inhospitable disease microenvironments, requirements for ex vivo cell overexpansion prior to infusions, intrinsic differences between MSC and different sources and donors, variability of culturing protocols, and potency assays to evaluate MSC products have been described as limitations in the field. In recent years, priming approaches to empower MSC have been investigated, thereby generating cellular products with improved potential for different clinical applications. Herein, we review the current priming approaches that aim to increase MSC therapeutic efficacy. Priming with cytokines and growth factors, hypoxia, pharmacological drugs, biomaterials, and different culture conditions, as well as other diverse molecules, are revised from current and future perspectives.
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Affiliation(s)
- Nádia de Cássia Noronha
- Center for Cell-based Therapy, Regional Blood Center of Ribeirão Preto, Ribeirão Preto Medical School, University of São Paulo, Ribeirão Preto, Brazil.,Graduate Program on Bioscience and Biotechnology, School of Pharmaceutical Sciences of Ribeirão Preto, University of São Paulo, Ribeirão Preto, Brazil
| | - Amanda Mizukami
- Center for Cell-based Therapy, Regional Blood Center of Ribeirão Preto, Ribeirão Preto Medical School, University of São Paulo, Ribeirão Preto, Brazil
| | | | - Juçara Gastaldi Cominal
- Center for Cell-based Therapy, Regional Blood Center of Ribeirão Preto, Ribeirão Preto Medical School, University of São Paulo, Ribeirão Preto, Brazil.,Graduate Program on Bioscience and Biotechnology, School of Pharmaceutical Sciences of Ribeirão Preto, University of São Paulo, Ribeirão Preto, Brazil
| | - José Lucas M Rocha
- Center for Cell-based Therapy, Regional Blood Center of Ribeirão Preto, Ribeirão Preto Medical School, University of São Paulo, Ribeirão Preto, Brazil.,Graduate Program on Basic and Applied Immunology, Ribeirão Preto Medical School, University of São Paulo, Ribeirão Preto, Brazil
| | - Dimas Tadeu Covas
- Center for Cell-based Therapy, Regional Blood Center of Ribeirão Preto, Ribeirão Preto Medical School, University of São Paulo, Ribeirão Preto, Brazil
| | - Kamilla Swiech
- Center for Cell-based Therapy, Regional Blood Center of Ribeirão Preto, Ribeirão Preto Medical School, University of São Paulo, Ribeirão Preto, Brazil.,Department of Pharmaceutical Sciences, School of Pharmaceutical Sciences of Ribeirão Preto, University of São Paulo, Ribeirão Preto, Brazil
| | - Kelen C R Malmegrim
- Center for Cell-based Therapy, Regional Blood Center of Ribeirão Preto, Ribeirão Preto Medical School, University of São Paulo, Ribeirão Preto, Brazil. .,Department of Clinical, Toxicological and Bromatological Analysis, School of Pharmaceutical Sciences of Ribeirão Preto, University of São Paulo, Avenida do Café, s/n°, Ribeirão Preto, SP, 14010-903, Brazil.
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Shin HS, Lee S, Hong HJ, Lim YC, Koh WG, Lim JY. Stem cell properties of human clonal salivary gland stem cells are enhanced by three-dimensional priming culture in nanofibrous microwells. Stem Cell Res Ther 2018; 9:74. [PMID: 29566770 PMCID: PMC5863805 DOI: 10.1186/s13287-018-0829-x] [Citation(s) in RCA: 17] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/25/2018] [Revised: 03/05/2018] [Accepted: 03/07/2018] [Indexed: 01/05/2023] Open
Abstract
BACKGROUND Three-dimensional (3D) cultures recapitulate the microenvironment of tissue-resident stem cells and enable them to modulate their properties. We determined whether salivary gland-resident stem cells (SGSCs) are primed by a 3D spheroid culture prior to treating irradiation-induced salivary hypofunction using in-vitro coculture and in-vivo transplant models. METHODS 3D spheroid-derived SGSCs (SGSCs3D) were obtained from 3D culture in microwells consisting of a nanofiber bottom and cell-repellent hydrogel walls, and were examined for salivary stem or epithelial gene/protein expression, differentiation potential, and paracrine secretory function compared with monolayer-cultured SGSCs (SGSCs2D) in vitro and in vivo. RESULTS SGSCs3D expressed increased salivary stem cell markers (LGR5 and THY1) and pluripotency markers (POU5F1 and NANOG) compared with SGSCs2D. Also, SGSCs3D exhibited enhanced potential to differentiate into salivary epithelial cells upon differentiation induction and increased paracrine secretion as compared to SGSCs2D. Wnt signaling was activated by 3D spheroid formation in the microwells and suppression of the Wnt/β-catenin pathway led to reduced stemness of SGSCs3D. Enhanced radioprotective properties of SGSCs3D against radiation-induced salivary hypofunction was confirmed by an organotypic 3D coculture and in-vivo transplantation experiments. CONCLUSION The 3D spheroid culture of SGSCs in nanofibrous microwells promotes stem cell properties via activation of Wnt signaling. This may contribute to SGSC priming prior to regenerative therapy to restore salivary hypofunction after radiotherapy.
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Affiliation(s)
- Hyun-Soo Shin
- Department of Otorhinolaryngology, Gangnam Severance Hospital, Yonsei University College of Medicine, 211 Eonju-ro, Gangnam-gu, Seoul, 06273, Republic of Korea
| | - Songyi Lee
- Department of Otorhinolaryngology, Gangnam Severance Hospital, Yonsei University College of Medicine, 211 Eonju-ro, Gangnam-gu, Seoul, 06273, Republic of Korea
| | - Hye Jin Hong
- Department of Chemical and Biomolecular Engineering, Yonsei University, Seoul, Republic of Korea
| | - Young Chang Lim
- Department of Otorhinolaryngology, Head and Neck Surgery, Konkuk University School of Medicine, Seoul, Republic of Korea
| | - Won-Gun Koh
- Department of Chemical and Biomolecular Engineering, Yonsei University, Seoul, Republic of Korea
| | - Jae-Yol Lim
- Department of Otorhinolaryngology, Gangnam Severance Hospital, Yonsei University College of Medicine, 211 Eonju-ro, Gangnam-gu, Seoul, 06273, Republic of Korea.
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Balikov DA, Crowder SW, Lee JB, Lee Y, Ko UH, Kang ML, Kim WS, Shin JH, Sung HJ. Aging Donor-Derived Human Mesenchymal Stem Cells Exhibit Reduced Reactive Oxygen Species Loads and Increased Differentiation Potential Following Serial Expansion on a PEG-PCL Copolymer Substrate. Int J Mol Sci 2018; 19:ijms19020359. [PMID: 29370101 PMCID: PMC5855581 DOI: 10.3390/ijms19020359] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/09/2018] [Revised: 01/22/2018] [Accepted: 01/23/2018] [Indexed: 12/13/2022] Open
Abstract
Human mesenchymal stem cells (hMSCs) have been widely studied for therapeutic development in tissue engineering and regenerative medicine. They can be harvested from human donors via tissue biopsies, such as bone marrow aspiration, and cultured to reach clinically relevant cell numbers. However, an unmet issue lies in the fact that the hMSC donors for regenerative therapies are more likely to be of advanced age. Their stem cells are not as potent compared to those of young donors, and continue to lose healthy, stemness-related activities when the hMSCs are serially passaged in tissue culture plates. Here, we have developed a cheap, scalable, and effective copolymer film to culture hMSCs obtained from aged human donors over several passages without loss of reactive oxygen species (ROS) handling or differentiation capacity. Assays of cell morphology, reactive oxygen species load, and differentiation potential demonstrate the effectiveness of copolymer culture on reduction in senescence-related activities of aging donor-derived hMSCs that could hinder the therapeutic potential of autologous stem cell therapies.
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Affiliation(s)
- Daniel A Balikov
- Department of Biomedical Engineering, Vanderbilt University, Nashville, TN 37235, USA.
| | - Spencer W Crowder
- Department of Materials and Department of Bioengineering, Imperial College London, London SW7 2AZ, UK.
| | - Jung Bok Lee
- Department of Biomedical Engineering, Vanderbilt University, Nashville, TN 37235, USA.
- Severance Biomedical Science Institute, College of Medicine, Yonsei University, Seoul 03722, Korea.
| | - Yunki Lee
- Department of Biomedical Engineering, Vanderbilt University, Nashville, TN 37235, USA.
- Department of Mechanical Engineering, Georgia Institute of Technology, Atlanta, GA 30332, USA.
| | - Ung Hyun Ko
- Department of Mechanical Engineering, Korea Advanced Institute of Science and Technology, Daejeon 34141, Korea.
| | - Mi-Lan Kang
- Severance Biomedical Science Institute, College of Medicine, Yonsei University, Seoul 03722, Korea.
| | - Won Shik Kim
- Department of Otorhinolaryngology, College of Medicine, Yonsei University, Seoul 03722, Korea.
| | - Jennifer H Shin
- Department of Mechanical Engineering, Korea Advanced Institute of Science and Technology, Daejeon 34141, Korea.
| | - Hak-Joon Sung
- Severance Biomedical Science Institute, College of Medicine, Yonsei University, Seoul 03722, Korea.
- Department of Mechanical Engineering, Georgia Institute of Technology, Atlanta, GA 30332, USA.
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Generating Chondromimetic Mesenchymal Stem Cell Spheroids by Regulating Media Composition and Surface Coating. Cell Mol Bioeng 2017; 11:99-115. [PMID: 29623134 DOI: 10.1007/s12195-017-0517-4] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/22/2022] Open
Abstract
INTRODUCTION Spheroids of mesenchymal stem cells (MSCs) in cartilage tissue engineering have been shown to enhance regenerative potential owing to their 3D structure. In this study, we explored the possibility of priming spheroids under different media to replace the use of inductive surface coatings for chondrogenic differentiation. METHODS Rat bone marrow-derived MSCs were organized into cell spheroids by the hanging drop technique and subsequently cultured on hyaluronic acid (HA) coated or non-coated well plates under different cell media conditions. Endpoint analysis included cell viability, DNA and Glycosaminoglycan (GAG) and collagen content, gene expression and immunohistochemistry. RESULTS For chondrogenic applications, MSC spheroids derived on non-coated surfaces outperformed the spheroids derived from HA-coated surfaces in matrix synthesis and collagen II gene expression. Spheroids on non-coated surfaces gave rise to the highest collagen and GAG when primed with medium containing insulin-like growth factor (IGF) for 1 week during spheroid formation. Spheroids that were grown in chondroinductive raw material-inclusive media such as aggrecan or chondroitin sulfate exhibited the highest Collagen II gene expression in the non-coated surface at 1 week. CONCLUSION Media priming by growth factors and raw materials might be a more predictive influencer of chondrogenesis compared to inductive-surfaces. Such tailored bioactivity of the stem cell spheroids in the stage of the spheroid formation may give rise to a platform technology that may eventually produce spheroids capable of chondrogenesis achieved by mere media manipulation, skipping the need for additional culture on a modified surface, that paves the way for cost-effective technologies.
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Nandi S, Brown AC. Characterizing Cell Migration Within Three-dimensional In Vitro Wound Environments. J Vis Exp 2017. [PMID: 28872146 DOI: 10.3791/56099] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/26/2023] Open
Abstract
Currently, most in vitro models of wound healing, such as well-established scratch assays, involve studying cell migration and wound closure on two-dimensional surfaces. However, the physiological environment in which in vivo wound healing takes place is three-dimensional rather than two-dimensional. It is becoming increasingly clear that cell behavior differs greatly in two-dimensional vs. three-dimensional environments; therefore, there is a need for more physiologically relevant in vitro models for studying cell migration behaviors in wound closure. The method described herein allows for the study of cell migration in a three-dimensional model that better reflects physiological conditions than previously established two-dimensional scratch assays. The purpose of this model is to evaluate cell outgrowth via the examination of cell migration away from a spheroid body embedded within a fibrin matrix in the presence of pro- or anti-migratory factors. Using this method, cell outgrowth from the spheroid body in a three-dimensional matrix can be observed and is easily quantifiable over time via brightfield microscopy and analysis of spheroid body area. The effect of pro-migratory and/or inhibitory factors on cell migration can also be evaluated in this system. This method provides researchers with a simple method of analyzing cell migration in three-dimensional wound associated matrices in vitro, thus increasing the relevance of in vitro cell studies prior to the use of in vivo animal models.
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Affiliation(s)
- Seema Nandi
- Joint Department of Biomedical Engineering, North Carolina State University and The University of North Carolina - Chapel Hill; Comparative Medicine Institute, North Carolina State University
| | - Ashley C Brown
- Joint Department of Biomedical Engineering, North Carolina State University and The University of North Carolina - Chapel Hill; Comparative Medicine Institute, North Carolina State University;
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Tang H, Zhang Y, Jansen JA, van den Beucken JJJP. Effect of monocytes/macrophages on the osteogenic differentiation of adipose-derived mesenchymal stromal cells in 3D co-culture spheroids. Tissue Cell 2017; 49:461-469. [PMID: 28684045 DOI: 10.1016/j.tice.2017.06.002] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/11/2017] [Revised: 06/02/2017] [Accepted: 06/06/2017] [Indexed: 02/08/2023]
Abstract
This study aimed to investigate the distinctive roles of the monocytes and macrophages on osteogenic differentiation of adipose-derived mesenchymal stromal cells (ADMSCs) in 3D spheroid co-cultures. We hypothesized that monocytes or macrophages (subtypes pro-inflammatory M1 and pro-wound healing M2) would affect the osteogenic differentiation of ADMSCs in 3D spheroids and that cell-cell interactions between monocytes/macrophages and ADMSCs play an important role in the osteogenic differentiation process of ADMSCs. The obtained results indicated that the osteogenic differentiation of ADMSCs was inhibited by monocytes and both macrophage subtypes in 3D spheroids. Monocytes and M2 macrophages had a stronger inhibiting effect than M1 macrophages. Cell-cell interactions mediated by N-cadherin likely played a role in the inhibiting effect of monocytes/macrophages on the osteogenic differentiation of ADMSCs.
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Affiliation(s)
- Hongbo Tang
- Department of Biomaterials, Radboudumc, Nijmegen, the Netherlands; Department of Plastic Surgery, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China
| | - Yang Zhang
- Department of Biomaterials, Radboudumc, Nijmegen, the Netherlands
| | - John A Jansen
- Department of Biomaterials, Radboudumc, Nijmegen, the Netherlands
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Petrenko Y, Syková E, Kubinová Š. The therapeutic potential of three-dimensional multipotent mesenchymal stromal cell spheroids. Stem Cell Res Ther 2017; 8:94. [PMID: 28446248 PMCID: PMC5406927 DOI: 10.1186/s13287-017-0558-6] [Citation(s) in RCA: 163] [Impact Index Per Article: 20.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
The efficiency of clinical trials involving transplantation of multipotent mesenchymal stromal cells (MSCs) is often insufficient due to harsh conditions present within the target tissue including hypoxia, low nutrient supply as well as inflammatory reactions. This indicates the necessity for optimization of cell-based therapy approaches which might include either modification of the cell manufacturing process or specific cell pretreatment procedures prior to transplantation. Recent reports confirm evidence that the aggregation of MSCs into three-dimensional (3D) multicellular spheroids results in enhancement of the overall therapeutic potential of cells, by improving the anti-inflammatory and angiogenic properties, stemness and survival of MSCs after transplantation. Such an MSCs spheroid generation approach may open new opportunities for the enlargement of MSCs applications in clinical research and therapy. However, the unification and optimization of 3D spheroid generation techniques, including the selection of appropriate clinical-grade culture conditions and methods for their large-scale production, are still of great importance. The current review addresses questions regarding therapeutic-associated properties of 3D multicellular MSCs spheroids in vitro and during preclinical animal studies, with special attention to the possibilities of translating these research achievements toward further clinical manufacturing and applications.
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Affiliation(s)
- Yuriy Petrenko
- Department of Biomaterials and Biophysical Methods, Institute of Experimental Medicine AS CR v. v. i, Vídeňská 1083, 14220, Prague 4-Krč, Czech Republic.
| | - Eva Syková
- Department of Neuroscience, Charles University, Second Faculty of Medicine, V Uvalu 84, 15006, Prague, Czech Republic
| | - Šárka Kubinová
- Department of Biomaterials and Biophysical Methods, Institute of Experimental Medicine AS CR v. v. i, Vídeňská 1083, 14220, Prague 4-Krč, Czech Republic
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Bae YJ, Kwon YR, Kim HJ, Lee S, Kim YJ. Enhanced differentiation of mesenchymal stromal cells by three-dimensional culture and azacitidine. Blood Res 2017; 52:18-24. [PMID: 28401097 PMCID: PMC5383582 DOI: 10.5045/br.2017.52.1.18] [Citation(s) in RCA: 18] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/10/2016] [Revised: 12/16/2016] [Accepted: 01/06/2017] [Indexed: 12/27/2022] Open
Abstract
BACKGROUND Mesenchymal stromal cells (MSCs) are useful for cell therapy because of their potential for multilineage differentiation. However, MSCs that are expanded in traditional two-dimensional (2D) culture systems eventually lose their differentiation abilities. Therefore, we investigated whether azacitidine (AZA) supplementation and three-dimensional culture (3D) could improve the differentiation properties of MSCs. METHODS 2D- or 3D-cultured MSCs which were prepared according to the conventional or hanging-drop culture method respectively, were treated with or without AZA (1 µM for 72 h), and their osteogenic and adipogenic differentiation potential were determined and compared. RESULTS AZA treatment did not affect the cell apoptosis or viability in both 2D- and 3D-cultured MSCs. However, compared to conventionally cultured 2D-MSCs, AZA-treated 2D-MSCs showed marginally increased differentiation abilities. In contrast, 3D-MSCs showed significantly increased osteogenic and adipogenic differentiation ability. When 3D culture was performed in the presence of AZA, the osteogenic differentiation ability was further increased, whereas adipogenic differentiation was not affected. CONCLUSION 3D culture efficiently promoted the multilineage differentiation of MSCs, and in combination with AZA, it could help MSCs to acquire greater osteogenic differentiation ability. This optimized culture method can enhance the therapeutic potential of MSCs.
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Affiliation(s)
- Yoo-Jin Bae
- Laboratory of Hematological Disease and Immunology, Seoul St. Mary's Hospital, College of Medicine, The Catholic University of Korea, Seoul, Korea
| | - Yong-Rim Kwon
- Laboratory of Hematological Disease and Immunology, Seoul St. Mary's Hospital, College of Medicine, The Catholic University of Korea, Seoul, Korea
| | - Hye Joung Kim
- Laboratory of Hematological Disease and Immunology, Seoul St. Mary's Hospital, College of Medicine, The Catholic University of Korea, Seoul, Korea
| | - Seok Lee
- Leukemia Research Institute, Seoul St. Mary's Hospital, College of Medicine, The Catholic University of Korea, Seoul, Korea.; Department of Hematology, Catholic Blood and Marrow Transplantation Center, Seoul St. Mary's Hospital, College of Medicine, The Catholic University of Korea, Seoul, Korea
| | - Yoo-Jin Kim
- Laboratory of Hematological Disease and Immunology, Seoul St. Mary's Hospital, College of Medicine, The Catholic University of Korea, Seoul, Korea.; Leukemia Research Institute, Seoul St. Mary's Hospital, College of Medicine, The Catholic University of Korea, Seoul, Korea.; Department of Hematology, Catholic Blood and Marrow Transplantation Center, Seoul St. Mary's Hospital, College of Medicine, The Catholic University of Korea, Seoul, Korea
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41
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Zhou F, Hui Y, Xin H, Xu YD, Lei HE, Yang BC, Guan RL, Li M, Hou JQ, Xin ZC. Therapeutic effects of adipose-derived stem cells-based microtissues on erectile dysfunction in streptozotocin-induced diabetic rats. Asian J Androl 2017; 19:91-97. [PMID: 27345005 PMCID: PMC5227681 DOI: 10.4103/1008-682x.182817] [Citation(s) in RCA: 18] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
This study aimed to explore the therapeutic effects of adipose-derived stem cells (ADSCs)-based microtissues (MTs) on erectile dysfunction (ED) in streptozotocin (STZ)-induced diabetic rats. Fifty-six 8-week-old Sprague-Dawley rats received intraperitoneal injection of STZ (60 mg kg−1), and 8 weeks later, the determined diabetic rats randomly received intracavernous (IC) injection of phosphate buffer solution (PBS), ADSCs, or MTs. Another eight normal rats equally got IC injection of PBS. MTs were generated with a hanging drop method, and the injected cells were tracked in ADSC- and MT-injected rats. Four weeks after the treatments, intracavernous pressure (ICP), histopathological changes in corpus cavernosum (CC), and functional proteins were measured. Rat cytokine antibody array was used to detect ADSCs or MTs lysate. The results showed that MTs expressed vascular endothelial growth factor (VEGF), nerve growth factor (NGF), and tumor necrosis factor-stimulated gene-6 (TSG-6). MTs injection had a higher retention than ADSCs injection and MTs treatment improved ICP, neuronal nitric oxide synthase (nNOS) expression, smooth muscle, and endothelial contents in diabetic rats, ameliorated local inflammation in CC better. Thus, our findings demonstrate that IC injection of MTs improves erectile function and histopathological changes in STZ-induced diabetic rats and appears to be more promising than traditional ADSCs. The underlying mechanisms involve increased cell retention accompanied with neuroprotection and anti-inflammatory behaviors of the paracrine factors.
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Affiliation(s)
- Feng Zhou
- Molecular Biology Laboratory of Andrology Center, Peking University First Hospital, Peking University, Beijing 100034, China.,Department of Urology, First Affiliated Hospital of Soochow University, Soochow University, Suzhou 215006, China
| | - Yu Hui
- Molecular Biology Laboratory of Andrology Center, Peking University First Hospital, Peking University, Beijing 100034, China.,Department of Urology, First Affiliated Hospital of Soochow University, Soochow University, Suzhou 215006, China
| | - Hua Xin
- Department of Ophthalmology, Beijing Chao-Yang Hospital, Capital Medical University, Beijing 100043, China
| | - Yong-De Xu
- Molecular Biology Laboratory of Andrology Center, Peking University First Hospital, Peking University, Beijing 100034, China
| | - Hong-En Lei
- Molecular Biology Laboratory of Andrology Center, Peking University First Hospital, Peking University, Beijing 100034, China
| | - Bi-Cheng Yang
- Molecular Biology Laboratory of Andrology Center, Peking University First Hospital, Peking University, Beijing 100034, China
| | - Rui-Li Guan
- Molecular Biology Laboratory of Andrology Center, Peking University First Hospital, Peking University, Beijing 100034, China
| | - Meng Li
- Department of Urology, General Hospital of Ningxia Medical University, Ningxia Medical University, Ningxia 750021, China
| | - Jian-Quan Hou
- Department of Urology, First Affiliated Hospital of Soochow University, Soochow University, Suzhou 215006, China
| | - Zhong-Cheng Xin
- Molecular Biology Laboratory of Andrology Center, Peking University First Hospital, Peking University, Beijing 100034, China
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42
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Aktas E, Chamberlain CS, Saether EE, Duenwald-Kuehl SE, Kondratko-Mittnacht J, Stitgen M, Lee JS, Clements AE, Murphy WL, Vanderby R. Immune modulation with primed mesenchymal stem cells delivered via biodegradable scaffold to repair an Achilles tendon segmental defect. J Orthop Res 2017; 35:269-280. [PMID: 27061844 DOI: 10.1002/jor.23258] [Citation(s) in RCA: 58] [Impact Index Per Article: 7.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/25/2015] [Accepted: 04/04/2016] [Indexed: 02/06/2023]
Abstract
Tendon healing is a complex coordinated series of events resulting in protracted recovery, limited regeneration, and scar formation. Mesenchymal stem cell (MSC) therapy has shown promise as a new technology to enhance soft tissue and bone healing. A challenge with MSC therapy involves the ability to consistently control the inflammatory response and subsequent healing. Previous studies suggest that preconditioning MSCs with inflammatory cytokines, such as IFN-γ, TNF-α, and IL-1β may accelerate cutaneous wound closure. The objective of this study was to therefore elucidate these effects in tendon. That is, the in vivo healing effects of TNF-α primed MSCs were studied using a rat Achilles segmental defect model. Rat Achilles tendons were subjected to a unilateral 3 mm segmental defect and repaired with either a PLG scaffold alone, MSC-seeded PLG scaffold, or TNF-α-primed MSC-seeded PLG scaffold. Achilles tendons were analyzed at 2 and 4 weeks post-injury. In vivo, MSCs, regardless of priming, increased IL-10 production and reduced the inflammatory factor, IL-1α. Primed MSCs reduced IL-12 production and the number of M1 macrophages, as well as increased the percent of M2 macrophages, and synthesis of the anti-inflammatory factor IL-4. Primed MSC treatment also increased the concentration of type I procollagen in the healing tissue and increased failure stress of the tendon 4 weeks post-injury. Taken together delivery of TNF-α primed MSCs via 3D PLG scaffold modulated macrophage polarization and cytokine production to further accentuate the more regenerative MSC-induced healing response. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:269-280, 2017.
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Affiliation(s)
- Erdem Aktas
- Department of Orthopedics, Ankara Oncology Research and Training Hospital, Ankara, Turkey
| | - Connie S Chamberlain
- Department of Orthopedics and Rehabilitation, University of Wisconsin, Madison, Wisconsin, 53705
| | - Erin E Saether
- Department of Biomedical Engineering, University of Wisconsin, Madison, Wisconsin, 53705
| | - Sarah E Duenwald-Kuehl
- Department of Biomedical Engineering, University of Wisconsin, Madison, Wisconsin, 53705
| | | | - Michael Stitgen
- Department of Biomedical Engineering, University of Wisconsin, Madison, Wisconsin, 53705
| | - Jae Sung Lee
- Department of Biomedical Engineering, University of Wisconsin, Madison, Wisconsin, 53705
| | - Anna E Clements
- Department of Orthopedics and Rehabilitation, University of Wisconsin, Madison, Wisconsin, 53705
| | - William L Murphy
- Department of Orthopedics and Rehabilitation, University of Wisconsin, Madison, Wisconsin, 53705.,Department of Biomedical Engineering, University of Wisconsin, Madison, Wisconsin, 53705
| | - Ray Vanderby
- Department of Orthopedics and Rehabilitation, University of Wisconsin, Madison, Wisconsin, 53705.,Department of Biomedical Engineering, University of Wisconsin, Madison, Wisconsin, 53705
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43
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Crowder SW, Balikov DA, Boire TC, McCormack D, Lee JB, Gupta MK, Skala MC, Sung HJ. Copolymer-Mediated Cell Aggregation Promotes a Proangiogenic Stem Cell Phenotype In Vitro and In Vivo. Adv Healthc Mater 2016; 5:2866-2871. [PMID: 27717208 PMCID: PMC5152909 DOI: 10.1002/adhm.201600819] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/28/2016] [Revised: 08/19/2016] [Indexed: 12/31/2022]
Abstract
Material-induced cell aggregation drives a proangiogenic expression profile. Copolymer substrates containing cell-repellent and cell-adhesive domains force the aggregation of human mesenchymal stem cells, which results in enhanced tubulogenesis in vitro and stabilization of vasculature in vivo. These findings can be used to design instructive biomaterial scaffolds for clinical use.
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Affiliation(s)
- Spencer W. Crowder
- Department of Biomedical Engineering, Vanderbilt University, Nashville, TN 37235, USA
| | - Daniel A. Balikov
- Department of Biomedical Engineering, Vanderbilt University, Nashville, TN 37235, USA
| | - Timothy C. Boire
- Department of Biomedical Engineering, Vanderbilt University, Nashville, TN 37235, USA
| | - Devin McCormack
- Department of Biomedical Engineering, Vanderbilt University, Nashville, TN 37235, USA
| | - Jung Bok Lee
- Department of Biomedical Engineering, Vanderbilt University, Nashville, TN 37235, USA
| | - Mukesh K. Gupta
- Department of Biomedical Engineering, Vanderbilt University, Nashville, TN 37235, USA
| | - Melissa C. Skala
- Department of Biomedical Engineering, Vanderbilt University, Nashville, TN 37235, USA
| | - Hak-Joon Sung
- Department of Biomedical Engineering, Vanderbilt University, Nashville, TN 37235, USA
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44
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Bartosh TJ, Ullah M, Zeitouni S, Beaver J, Prockop DJ. Cancer cells enter dormancy after cannibalizing mesenchymal stem/stromal cells (MSCs). Proc Natl Acad Sci U S A 2016; 113:E6447-E6456. [PMID: 27698134 PMCID: PMC5081643 DOI: 10.1073/pnas.1612290113] [Citation(s) in RCA: 132] [Impact Index Per Article: 14.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022] Open
Abstract
Patients with breast cancer often develop malignant regrowth of residual drug-resistant dormant tumor cells years after primary treatment, a process defined as cancer relapse. Deciphering the causal basis of tumor dormancy therefore has obvious therapeutic significance. Because cancer cell behavior is strongly influenced by stromal cells, particularly the mesenchymal stem/stromal cells (MSCs) that are actively recruited into tumor-associated stroma, we assessed the impact of MSCs on breast cancer cell (BCC) dormancy. Using 3D cocultures to mimic the cellular interactions of an emerging tumor niche, we observed that MSCs sequentially surrounded the BCCs, promoted formation of cancer spheroids, and then were internalized/degraded through a process resembling the well-documented yet ill-defined clinical phenomenon of cancer cell cannibalism. This suspected feeding behavior was less appreciable in the presence of a rho kinase inhibitor and in 2D monolayer cocultures. Notably, cannibalism of MSCs enhanced survival of BCCs deprived of nutrients but suppressed their tumorigenicity, together suggesting the cancer cells entered dormancy. Transcriptome profiles revealed that the resulting BCCs acquired a unique molecular signature enriched in prosurvival factors and tumor suppressors, as well as inflammatory mediators that demarcate the secretome of senescent cells, also referred to as the senescence-associated secretory phenotype. Overall, our results provide intriguing evidence that cancer cells under duress enter dormancy after cannibalizing MSCs. Importantly, our practical 3D coculture model could provide a valuable tool to understand the antitumor activity of MSCs and cell cannibalism further, and therefore open new therapeutic avenues for the prevention of cancer recurrence.
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Affiliation(s)
- Thomas J Bartosh
- Institute for Regenerative Medicine, College of Medicine, Texas A&M University System Health Science Center, Temple, TX 76502; Medical Physiology, College of Medicine, Texas A&M University System Health Science Center, Temple, TX 76504
| | - Mujib Ullah
- Institute for Regenerative Medicine, College of Medicine, Texas A&M University System Health Science Center, Temple, TX 76502
| | - Suzanne Zeitouni
- Institute for Regenerative Medicine, College of Medicine, Texas A&M University System Health Science Center, Temple, TX 76502
| | - Joshua Beaver
- Institute for Regenerative Medicine, College of Medicine, Texas A&M University System Health Science Center, Temple, TX 76502; Medical Physiology, College of Medicine, Texas A&M University System Health Science Center, Temple, TX 76504
| | - Darwin J Prockop
- Institute for Regenerative Medicine, College of Medicine, Texas A&M University System Health Science Center, Temple, TX 76502;
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45
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de Soure AM, Fernandes-Platzgummer A, da Silva CL, Cabral JMS. Scalable microcarrier-based manufacturing of mesenchymal stem/stromal cells. J Biotechnol 2016; 236:88-109. [PMID: 27527397 DOI: 10.1016/j.jbiotec.2016.08.007] [Citation(s) in RCA: 57] [Impact Index Per Article: 6.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/30/2016] [Revised: 08/02/2016] [Accepted: 08/09/2016] [Indexed: 12/17/2022]
Abstract
Due to their unique features, mesenchymal stem/stromal cells (MSC) have been exploited in clinical settings as therapeutic candidates for the treatment of a variety of diseases. However, the success in obtaining clinically-relevant MSC numbers for cell-based therapies is dependent on efficient isolation and ex vivo expansion protocols, able to comply with good manufacturing practices (GMP). In this context, the 2-dimensional static culture systems typically used for the expansion of these cells present several limitations that may lead to reduced cell numbers and compromise cell functions. Furthermore, many studies in the literature report the expansion of MSC using fetal bovine serum (FBS)-supplemented medium, which has been critically rated by regulatory agencies. Alternative platforms for the scalable manufacturing of MSC have been developed, namely using microcarriers in bioreactors, with also a considerable number of studies now reporting the production of MSC using xenogeneic/serum-free medium formulations. In this review we provide a comprehensive overview on the scalable manufacturing of human mesenchymal stem/stromal cells, depicting the various steps involved in the process from cell isolation to ex vivo expansion, using different cell tissue sources and culture medium formulations and exploiting bioprocess engineering tools namely microcarrier technology and bioreactors.
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Affiliation(s)
- António M de Soure
- Department of Bioengineering and iBB-Institute for Bioengineering and Biosciences, Instituto Superior Técnico, Universidade de Lisboa, Av. Rovisco Pais, Lisboa, Portugal
| | - Ana Fernandes-Platzgummer
- Department of Bioengineering and iBB-Institute for Bioengineering and Biosciences, Instituto Superior Técnico, Universidade de Lisboa, Av. Rovisco Pais, Lisboa, Portugal
| | - Cláudia L da Silva
- Department of Bioengineering and iBB-Institute for Bioengineering and Biosciences, Instituto Superior Técnico, Universidade de Lisboa, Av. Rovisco Pais, Lisboa, Portugal
| | - Joaquim M S Cabral
- Department of Bioengineering and iBB-Institute for Bioengineering and Biosciences, Instituto Superior Técnico, Universidade de Lisboa, Av. Rovisco Pais, Lisboa, Portugal.
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46
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Berg J, Roch M, Altschüler J, Winter C, Schwerk A, Kurtz A, Steiner B. Human adipose-derived mesenchymal stem cells improve motor functions and are neuroprotective in the 6-hydroxydopamine-rat model for Parkinson's disease when cultured in monolayer cultures but suppress hippocampal neurogenesis and hippocampal memory function when cultured in spheroids. Stem Cell Rev Rep 2015; 11:133-49. [PMID: 25120226 DOI: 10.1007/s12015-014-9551-y] [Citation(s) in RCA: 41] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/30/2023]
Abstract
Adult human adipose-derived mesenchymal stem cells (MSC) have been reported to induce neuroprotective effects in models for Parkinson's disease (PD). However, these effects strongly depend on the most optimal application of the transplant. In the present study we compared monolayer-cultured (aMSC) and spheroid (sMSC) MSC following transplantation into the substantia nigra (SN) of 6-OHDA lesioned rats regarding effects on the local microenvironment, degeneration of dopaminergic neurons, neurogenesis in the hippocampal DG as well as motor and memory function in the 6-OHDA-rat model for PD. aMSC transplantation significantly increased tyrosine hydroxylase (TH) and brain-derived neurotrophic factor (BDNF) levels in the SN, increased the levels of the glial fibrillary acidic protein (GFAP) and improved motor functions compared to untreated and sMSC treated animals. In contrast, sMSC grafting induced an increased local microgliosis, decreased TH levels in the SN and reduced numbers of newly generated cells in the dentate gyrus (DG) without yet affecting hippocampal learning and memory function. We conclude that the neuroprotective potential of adipose-derived MSC in the rat model of PD crucially depends on the applied cellular phenotype.
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Affiliation(s)
- Jürgen Berg
- Department of Neurology, Charité University Medicine Berlin, CCM, Charitéplatz 1, D-10117, Berlin, Germany
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47
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Gleeson BM, Martin K, Ali MT, Kumar AHS, Pillai MGK, Kumar SPG, O'Sullivan JF, Whelan D, Stocca A, Khider W, Barry FP, O'Brien T, Caplice NM. Bone Marrow-Derived Mesenchymal Stem Cells Have Innate Procoagulant Activity and Cause Microvascular Obstruction Following Intracoronary Delivery: Amelioration by Antithrombin Therapy. Stem Cells 2015; 33:2726-37. [PMID: 25969127 DOI: 10.1002/stem.2050] [Citation(s) in RCA: 88] [Impact Index Per Article: 8.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/03/2014] [Revised: 04/02/2015] [Accepted: 04/07/2015] [Indexed: 12/17/2022]
Abstract
Mesenchymal stem cells (MSCs) are currently under investigation as tools to preserve cardiac structure and function following acute myocardial infarction (AMI). However, concerns have emerged regarding safety of acute intracoronary (IC) MSC delivery. This study aimed to characterize innate prothrombotic activity of MSC and identify means of its mitigation toward safe and efficacious therapeutic IC MSC delivery post-AMI. Expression of the initiator of the coagulation cascade tissue factor (TF) on MSC was detected and quantified by immunofluorescence, FACS, and immunoblotting. MSC-derived TF antigen was catalytically active and capable of supporting thrombin generation in vitro. Addition of MSCs to whole citrated blood enhanced platelet thrombus deposition on collagen at arterial shear, an effect abolished by heparin coadministration. In a porcine AMI model, IC infusion of 25 × 10(6) MSC during reperfusion was associated with a decrease in coronary flow reserve but not when coadministered with an antithrombin agent (heparin). Heparin reduced MSC-associated thrombosis incorporating platelets and VWF within the microvasculature. Heparin-assisted therapeutic MSC delivery also reduced apoptosis in the infarct border zone at 24 hours, significantly improved infarct size, left ventricular (LV) ejection fraction, LV volumes, wall motion, and attenuated histologic evidence of scar formation at 6 weeks post-AMI. Heparin alone or heparin-assisted fibroblast control cell delivery had no such effect. Procoagulant TF activity of therapeutic MSCs is associated with reductions in myocardial perfusion when delivered IC may be successfully managed by heparin coadministration. This study highlights an important mechanistic insight into safety concerns associated with therapeutic IC MSC delivery for AMI.
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Affiliation(s)
- Birgitta M Gleeson
- Centre for Research in Vascular Biology (CRVB), Biosciences Institute, University College Cork, Cork, Ireland
| | - Kenneth Martin
- Centre for Research in Vascular Biology (CRVB), Biosciences Institute, University College Cork, Cork, Ireland
| | - Mohammed T Ali
- Centre for Research in Vascular Biology (CRVB), Biosciences Institute, University College Cork, Cork, Ireland
| | - Arun H S Kumar
- Centre for Research in Vascular Biology (CRVB), Biosciences Institute, University College Cork, Cork, Ireland
| | - M Gopala-Krishnan Pillai
- Centre for Research in Vascular Biology (CRVB), Biosciences Institute, University College Cork, Cork, Ireland
| | - Sujith P G Kumar
- Centre for Research in Vascular Biology (CRVB), Biosciences Institute, University College Cork, Cork, Ireland
| | - John F O'Sullivan
- Centre for Research in Vascular Biology (CRVB), Biosciences Institute, University College Cork, Cork, Ireland
| | - Derek Whelan
- Centre for Research in Vascular Biology (CRVB), Biosciences Institute, University College Cork, Cork, Ireland
| | - Alessia Stocca
- Centre for Research in Vascular Biology (CRVB), Biosciences Institute, University College Cork, Cork, Ireland
| | - Wisam Khider
- Centre for Research in Vascular Biology (CRVB), Biosciences Institute, University College Cork, Cork, Ireland
| | - Frank P Barry
- Regenerative Medicine Institute (REMEDI), National University of Ireland, Galway, Ireland
| | - Timothy O'Brien
- Regenerative Medicine Institute (REMEDI), National University of Ireland, Galway, Ireland
| | - Noel M Caplice
- Centre for Research in Vascular Biology (CRVB), Biosciences Institute, University College Cork, Cork, Ireland
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48
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McGuirk JP, Smith JR, Divine CL, Zuniga M, Weiss ML. Wharton's Jelly-Derived Mesenchymal Stromal Cells as a Promising Cellular Therapeutic Strategy for the Management of Graft-versus-Host Disease. Pharmaceuticals (Basel) 2015; 8:196-220. [PMID: 25894816 PMCID: PMC4491656 DOI: 10.3390/ph8020196] [Citation(s) in RCA: 22] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/10/2014] [Revised: 03/13/2015] [Accepted: 04/08/2015] [Indexed: 02/06/2023] Open
Abstract
Allogeneic hematopoietic cell transplantation (allo-HCT), a treatment option in hematologic malignancies and bone marrow failure syndromes, is frequently complicated by Graft-versus-host disease (GVHD). The primary treatment for GVHD involves immune suppression by glucocorticoids. However, patients are often refractory to the steroid therapy, and this results in a poor prognosis. Therefore alternative therapies are needed to treat GVHD. Here, we review data supporting the clinical investigation of a novel cellular therapy using Wharton’s jelly (WJ)-derived mesenchymal stromal cells (MSCs) as a potentially safe and effective therapeutic strategy in the management of GVHD. Adult-derived sources of MSCs have demonstrated signals of efficacy in the management of GVHD. However, there are limitations, including: limited proliferation capacity; heterogeneity of cell sources; lengthy expansion time to clinical dose; expansion failure in vitro; and a painful, invasive, isolation procedure for the donor. Therefore, alternative MSC sources for cellular therapy are sought. The reviewed data suggests MSCs derived from WJ may be a safe and effective cellular therapy for GVHD. Laboratories investigated and defined the immune properties of WJ-MSCs for potential use in cellular therapy. These cells represent a more uniform cell population than bone marrow-derived MSCs, displaying robust immunosuppressive properties and lacking significant immunogenicity. They can be collected safely and painlessly from individuals at birth, rapidly expanded and stored cryogenically for later clinical use. Additionally, data we reviewed suggested licensing MSCs (activating MSCs by exposure to cytokines) to enhance effectiveness in treating GVHD. Therefore, WJCs should be tested as a second generation, relatively homogeneous allogeneic cell therapy for the treatment of GVHD.
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Affiliation(s)
- Joseph P McGuirk
- Blood and Marrow Transplant Program, The University of Kansas Medical Center, 2330 Shawnee Mission Pkwy., Suite 210 Mailstop 5003, Westwood, KS 66205, USA.
| | - J Robert Smith
- Department of Anatomy and Physiology, Kansas State University, 1600 Denison Ave., Coles Hall 228, Manhattan, KS 66506-5802, USA.
| | - Clint L Divine
- Blood and Marrow Transplant Program, The University of Kansas Medical Center, 2330 Shawnee Mission Pkwy., Suite 210 Mailstop 5003, Westwood, KS 66205, USA
| | - Micheal Zuniga
- Department of Anatomy and Physiology, Kansas State University, 1600 Denison Ave., Coles Hall 228, Manhattan, KS 66506-5802, USA.
| | - Mark L Weiss
- Department of Anatomy and Physiology, Kansas State University, 1600 Denison Ave., Coles Hall 228, Manhattan, KS 66506-5802, USA.
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49
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Schwerk A, Altschüler J, Roch M, Gossen M, Winter C, Berg J, Kurtz A, Steiner B. Human adipose-derived mesenchymal stromal cells increase endogenous neurogenesis in the rat subventricular zone acutely after 6-hydroxydopamine lesioning. Cytotherapy 2015; 17:199-214. [DOI: 10.1016/j.jcyt.2014.09.005] [Citation(s) in RCA: 31] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/11/2014] [Revised: 09/05/2014] [Accepted: 09/20/2014] [Indexed: 01/07/2023]
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50
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Magruder HT, Quinn JA, Schwartzbauer JE, Reichner J, Huang A, Filardo EJ. The G protein-coupled estrogen receptor-1, GPER-1, promotes fibrillogenesis via a Shc-dependent pathway resulting in anchorage-independent growth. Discov Oncol 2014; 5:390-404. [PMID: 25096985 DOI: 10.1007/s12672-014-0195-9] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/30/2014] [Accepted: 07/27/2014] [Indexed: 02/02/2023] Open
Abstract
The G protein-coupled estrogen receptor-1, GPER-1, coordinates fibronectin (FN) matrix assembly and release of heparan-bound epidermal growth factor (HB-EGF). This mechanism of action results in the recruitment of FN-engaged integrin α5β1 to fibrillar adhesions and the formation of integrin α5β1-Shc adaptor protein complexes. Here, we show that GPER-1 stimulation of murine 4 T1 or human SKBR3 breast cancer cells with 17β-estradiol (E2β) promotes the formation of focal adhesions and actin stress fibers and results in increased cellular adhesion and haptotaxis on FN, but not collagen. These actions are also induced by the xenoestrogen, bisphenol A, and the estrogen receptor (ER) antagonist, ICI 182, 780, but not the inactive stereoisomer, 17α-estradiol (E2α). In addition, we show that GPER-1 stimulation of breast cancer cells allows for FN-dependent, anchorage-independent growth and FN fibril formation in "hanging drop" assays, indicating that these GPER-1-mediated actions occur independently of adhesion to solid substrata. Stable expression of Shc mutant Y317F lacking its primary tyrosyl phosphorylation site disrupts E2β-induced focal adhesion and actin stress fiber formation and abolishes E2β-enhanced haptotaxis on FN and anchorage-dependent growth. Collectively, these data demonstrate that E2β action via GPER-1 enhances cellular adhesivity and FN matrix assembly and allows for anchorage-independent growth, cellular events that may allow for cellular survival, and tumor progression.
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Affiliation(s)
- Hilary T Magruder
- Division of Hematology and Oncology, Rhode Island Hospital and the Warren Alpert Medical School of Brown University, Providence, RI, 02903, USA
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