RESUMO
Osteoarthritis is a degenerative joint disease and a leading cause of adult disability. Our previous study has reported that mesenchymal stem cell-derived exosomes (MSC-Exo) mediated long non-coding RNA KLF3-AS1 improves osteoarthritis. This study aims to investigate the molecular mechanism of KLF3-AS1 in osteoarthritis. Chondrocytes were treated with IL-1ß to induce chondrocyte injury, followed by MSC-Exo treatment. We found that MSC-Exo enhanced KLF3-AS1 expression in IL-1ß-treated chondrocytes. IL-1ß treatment reduced cell viability and enhanced apoptosis in chondrocytes. MSC-Exo-mediated KLF3-AS1 promoted cell viability and repressed apoptosis of IL-1ß-treated chondrocytes. Rapamycin (autophagy activator) promoted cell viability and suppressed apoptosis of chondrocytes by activating autophagy. Moreover, KLF3-AS1 interacted with YBX1 in chondrocytes. MSC-Exo-mediated KLF3-AS1 activated PI3K/Akt/mTOR signaling pathway, which was abrogated by YBX1 silencing. MSC-Exo-mediated KLF3-AS1 repressed autophagy and apoptosis of chondrocytes by activating PI3K/Akt/mTOR signaling pathway. In conclusion, our data demonstrate that MSC-Exo-mediated KLF3-AS1 inhibits autophagy and apoptosis of IL-1ß-treated chondrocyte through PI3K/Akt/mTOR signaling pathway. KLF3-AS1 activates PI3K/Akt/mTOR signaling pathway by targeting YBX1 to improve the progression of osteoarthritis. Thus, this work suggests that MSC-Exo-mediated KLF3-AS1 may be a potential therapeutic target for osteoarthritis.
Assuntos
Exossomos , Células-Tronco Mesenquimais , Osteoartrite , RNA Longo não Codificante , Apoptose/genética , Autofagia/genética , Condrócitos/metabolismo , Exossomos/metabolismo , Humanos , Fatores de Transcrição Kruppel-Like/genética , Fatores de Transcrição Kruppel-Like/metabolismo , Células-Tronco Mesenquimais/metabolismo , Osteoartrite/genética , Osteoartrite/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Serina-Treonina Quinases TOR/metabolismoRESUMO
BACKGROUND: Exosomes secreted by human mesenchymal stem cells (hMSCs) have been shown to promote cartilage regeneration. This study aimed to explore whether exosomal lncRNA-KLF3-AS1 derived from hMSCs can promote chondrocyte proliferation via miR-206/GIT1 axis in osteoarthritis (OA). METHODS: hMSCs and MSC-derived exosomes (MSC-exo) were prepared for morphological observation and identification by transmission electron microscopy (TEM) and flow cytometry. IL-1ß-induced OA chondrocytes and collagenase-induced mouse OA model were established for the further experiments. Luciferase activity assay was performed to test whether miR-206 could bind to KLF3-AS1 or GIT1. Cell proliferation and apoptosis were evaluated by CCK-8 assay and flow cytometry, respectively. RESULTS: MSC-Exos increased chondrogenic genes Col2a1 (type II collagen alpha 1) and aggrecan, decreased hondrocyte hypertrophy markers MMP-13 (matrix metalloproteinase-13) and Runx2 (runt-related transcription factor 2) in chondrocytes isolated from OA model mice. Furthermore, MSC-Exos attenuated IL-1ß-induced chondrocyte proliferation inhibition and apoptosis induction. Moreover, MSCKLF3-AS1-Exos (exosomes derived from KLF3-AS1-overexpressing-MSCs) ameliorated IL-1ß-induced chondrocyte injury. Results also demonstrated that KLF3-AS1 acted as a competitive endogenous RNA (ceRNA) by sponging miR-206 to facilitate GIT1 expression. In addition, miR-206 overexpression and GIT1 knockdown reversed MSCKLF3-AS1-Exos-mediated attenuation of chondrocyte injury. CONCLUSION: Exosomal KLF3-AS1 derived from MSCs involved in MSC-Exos-mediated chondrocyte proliferation induction and chondrocyte apoptosis inhibition via miR-206/GIT1 axis. Abbreviation: G-protein-coupled receptor kinase interacting protein-1 (GIT1).
Assuntos
Apoptose , Artrite Experimental/cirurgia , Proteínas de Ciclo Celular/metabolismo , Proliferação de Células , Condrócitos/metabolismo , Exossomos/transplante , Proteínas Ativadoras de GTPase/metabolismo , Articulações/cirurgia , Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais , MicroRNAs/metabolismo , RNA Longo não Codificante/metabolismo , Animais , Artrite Experimental/genética , Artrite Experimental/metabolismo , Artrite Experimental/patologia , Proteínas de Ciclo Celular/genética , Células Cultivadas , Condrócitos/patologia , Condrogênese , Técnicas de Cocultura , Exossomos/genética , Exossomos/metabolismo , Exossomos/ultraestrutura , Proteínas Ativadoras de GTPase/genética , Humanos , Articulações/metabolismo , Articulações/patologia , Células-Tronco Mesenquimais/metabolismo , Células-Tronco Mesenquimais/ultraestrutura , Camundongos Endogâmicos C57BL , MicroRNAs/genética , RNA Longo não Codificante/genética , Transdução de SinaisRESUMO
The present study was designed to explore whether exosomal lncRNA-KLF3-AS1 derived from human mesenchymal stem cells (hMSCs) can serve as a positive treatment for osteoarthritis (OA). hMSCs and MSC-derived exosomes (MSC-exo) were prepared for morphological observation and identification by transmission electron microscopy and flow cytometry. IL-1ß-induced OA chondrocytes and collagenase-induced rat model of OA were established for the further experiments. Lentivirus-mediated siRNA targeting KLF3-AS1 was transfected into MSCs for silencing KLF3-AS1. The real-time quantitative PCR and western blotting analysis were performed to examine the mRNA and protein levels of type II collagen alpha 1 (Col2a1), aggrecan, matrix metalloproteinase 13 and runt-related transcription factor 2. Cell proliferation, apoptosis and migration were evaluated by CCK-8 assay, flow cytometry and transwell assay. HE (hematoxylin and eosin) staining and immunohistochemistry were used for histopathological studies. MSC-exo ameliorated IL-1ß-induced cartilage injury. Furthermore, lncRNA KLF3-AS1 was markedly enriched in MSC-exo, and exosomal KLF3-AS1 suppressed IL-1ß-induced apoptosis of chondrocytes. Further in vivo investigation indicated that exosomal KLF3-AS1 promoted cartilage repair in a rat model of OA. Exosomal KLF3-AS1 promoted cartilage repair and chondrocyte proliferation in a rat model of OA, which might be an underlying therapeutic target for OA.