Enhancement of human mesenchymal stem cell differentiation by combination treatment with 5-azacytidine and trichostatin A

Kim, Hye Joung; Kwon, Yong-Rim; Bae, Yu-Jin; Kim, Yoo-Jin

doi:10.1007/s10529-015-1949-3

Cited by 11 publications

(11 citation statements)

References 15 publications

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“…found that TSA enhanced hMSCs chondrogenesis in pellets cultured in chondrogenic media 46 . Kim et al found TSA treatment increased type II collagen expression over control levels seen during hMSC chondrogenesis 47 . The mechanism by which TSA affects cartilage development and degradation may be attributable to histone modification by HDACs.…”

Section: Discussionmentioning

confidence: 97%

MicroRNA-193b-3p regulates chondrogenesis and chondrocyte metabolism by targeting HDAC3

Meng¹,

Li²,

Zhang³

et al. 2018

Theranostics

119

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Histone deacetylase 3 (HDAC3) plays a pivotal role in the repression of cartilage-specific gene expression in human chondrocytes. The aim of this study was to determine whether microRNA-193b-3p (miR-193b-3p) regulates the expression of HDAC3 during chondrogenesis and chondrocyte metabolism.Methods: miR-193b-3p expression was assessed in a human mesenchymal stem cell (hMSC) model of chondrogenesis, in interleukin-1β (IL-1β)-treated primary human chondrocytes (PHCs), and in non-degraded and degraded cartilage. hMSCs and PHCs were transfected with miR-193b-3p or its antisense inhibitor. A direct interaction between miR-193b-3p and its putative binding site in the 3′-untranslated region (3′-UTR) of HDAC3 mRNA was confirmed by performing luciferase reporter assays. Chondrocytes were transfected with miR-193b-3p before performing a chromatin immunoprecipitation assay with an anti-acetylated histone H3 antibody. To investigate miR-193b-3p-transfected PHCs in vivo, they were seeded in tricalcium phosphate-collagen-hyaluronate (TCP-COL-HA) scaffolds, which were then implanted in nude mice. In addition, plasma exosomal miR-193b-3p in samples from normal controls and patients with osteoarthritis (OA) were measured.Results: miR-193b-3p expression was elevated in chondrogenic and hypertrophic hMSCs, while expression was significantly reduced in degraded cartilage compared to non-degraded cartilage. In addition, miR-193b-3p suppressed the activity of reporter constructs containing the 3′-UTR of HDAC3, inhibited HDAC3 expression, and promoted histone H3 acetylation in the COL2A1, AGGRECAN, COMP, and SOX9 promoters. Treatment with the HDAC inhibitor trichostatin A (TSA) increased cartilage-specific gene expression and enhanced hMSCs chondrogenesis. TSA also increased AGGRECAN expression and decreased MMP13 expression in IL-1β-treated PHCs. Further, 8 weeks after implanting PHC-seeded TCP-COL-HA scaffolds subcutaneously in nude mice, we found that miR-193b overexpression strongly enhanced in vivo cartilage formation compared to that found under control conditions. We also found that patients with OA had lower plasma exosomal miR-193b levels than control subjects.Conclusions: These findings indicate that miR-193b-3p directly targets HDAC3, promotes H3 acetylation, and regulates hMSC chondrogenesis and metabolism in PHCs.

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Section: Discussionmentioning

confidence: 97%

MicroRNA-193b-3p regulates chondrogenesis and chondrocyte metabolism by targeting HDAC3

Meng¹,

Li²,

Zhang³

et al. 2018

Theranostics

119

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“…For example, chromatin modification by epigenetic drugs has been proven to enhance multilineage differentiation of MSCs to osteocytes, adipocytes chondrocytes, and cardiomyocytes [192021]; however, the data regarding the influence of AZA on MSCs remain controversial [22]. We have previously observed that a combination of AZA and trichostatin A treatment enhanced multilineage differentiation, with AZA-induced hypomethylation being the main contributing mechanism [13]. However, in the present study, we observed no significant changes in the expression levels of key regulatory molecules in 2D-MSCs following AZA treatment.…”

Section: Discussionmentioning

confidence: 99%

“…For adipogenesis, cells plated in 24-well plates and grown to confluence were incubated in adipogenic differentiation medium for 2 weeks (Thermo Fisher Scientific, Waltham, MA, USA), and were either used for RNA isolation for genetic analysis or stained with Oil Red O for marker detection (Sigma). For osteogenesis, cells grown on 24-well plates to confluence were incubated with osteogenic differentiation medium (DMEM with 10% MSC-FBS, 10 −7 M dexamethasone, 10 mM β-glycerophosphate, and 0.3 mM ascorbic acid) [13] for 2 weeks, and were either processed for RNA isolation or stained with alkaline phosphatase (ALP) to highlight calcium deposits.…”

Section: Methodsmentioning

confidence: 99%

“…In addition to direct genetic modification of such regulators [9], control of gene expression as well as other genomic functions could be achieved by epigenetic modifications, for example by changing the local chromatin configuration or nuclear architecture [1011]. In line with these epigenetic mechanisms, our team previously reported that epigenetic agents such as hypomethylating agents or a histone deacetylation inhibitor could change the gene expression pattern and function of cells [1213], which suggests that such epigenetic modification could also be used to change the characteristics of MSCs.…”

Section: Introductionmentioning

confidence: 99%

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Enhanced differentiation of mesenchymal stromal cells by three-dimensional culture and azacitidine

et al. 2017

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BackgroundMesenchymal 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.Methods2D- 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.ResultsAZA 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.Conclusion3D 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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“…In our previous study, epigenetic modification of human MSCs (hMSCs) with HMAs plus HDACi resulted in higher expression of Runx-2, BDNF, and Sox-9 than the control treatment. HMAs and HDACi enhance the in-vitro differentiation of MSCs, suggesting that epigenetic modification could alter the function of hMSCs [ 22 ].…”

Section: Introductionmentioning

confidence: 99%

Epigenetic modification of mesenchymal stromal cells enhances their suppressive effects on the Th17 responses of cells from rheumatoid arthritis patients

Kim

et al. 2018

Stem Cell Res Ther

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BackgroundThe aim of this study was to investigate if epigenetically modified human mesenchymal stromal cells (hMSCs) can regulate the Th17-related immune responses.MethodsWe tested epigenetic drug combinations at various doses and selected the four combinations that resulted in maximal interleukin (IL)-10 and indoleamine 2,3-dioxygenase gene expression in hMSCs. We examined the effects of epigenetically modified hMSCs (epi-hMSCs) on CD4+ T-cell proliferation and inflammatory cytokine secretion under Th0- and Th17-polarizing conditions using mixed lymphocyte reactions and enzyme-linked immunosorbent assays (ELISAs). We determined Th17 cytokine levels and the percentage of Th17 cells among synovial fluid mononuclear cells (SFMCs) from rheumatoid arthritis (RA) patients by ELISA and flow cytometry.ResultsEpi-hMSCs inhibited the development of IL-17-producing cells in culture. The percentages of IL-17+ and interferon (IFN)-γ+ cells among peripheral blood mononuclear cells from healthy donors were lower under both the Th0 and Th17 conditions in the presence of epi-hMSCs than in the presence of no or untreated hMSCs. Epi-hMSC-treated RA patient SFMCs secreted lower levels of IL-17 and IFN-γ than RA patient SFMCs cultured without hMSCs or with untreated hMSCs.ConclusionsAn optimal combination of hypomethylating agents and histone deacetylase inhibitors can enhance the immunomodulatory potential of hMSCs, which may be useful for RA treatment.

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Enhancement of human mesenchymal stem cell differentiation by combination treatment with 5-azacytidine and trichostatin A

Abstract: HMAs and HDACs enhanced in vitro differentiation of MSCs, which was maximized when the two drugs were combined, with HMA having the dominant effect.

Cited by 11 publications

References 15 publications

MicroRNA-193b-3p regulates chondrogenesis and chondrocyte metabolism by targeting HDAC3

MicroRNA-193b-3p regulates chondrogenesis and chondrocyte metabolism by targeting HDAC3

Enhanced differentiation of mesenchymal stromal cells by three-dimensional culture and azacitidine

Epigenetic modification of mesenchymal stromal cells enhances their suppressive effects on the Th17 responses of cells from rheumatoid arthritis patients

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