Abstract
BackgroundChondrogenic differentiation of human adipose-derived stem cells (hADSCs) is important for cartilage generation and degradation. LncRNAs play an essential role in stem cell differentiation. However, the role and mechanism of lncRNA in hADSCs remain unclear. Our previous study showed that miR-490-5p was downregulated during chondrogenic differentiation of hADSCs. In this study, we investigated the effect and mechanism of lncRNA NONHSAT030515 interacting with miR-490-5p on chondrogenic differentiation of hADSCs.MethodsAlcian blue staining was used to assess the deposition of chondromatrix proteins following chondrogenic differentiation of human adipose stem cells. Immunohistochemistry was used to evaluate the expression of collagenII. TargetScan, miRTarBase and miRDB database analyses were applied to find the miRNA and target genes of lncRNA NONHSAT030515. A dual luciferase experiment was conducted to identify the direct target of NONHSAT030515. pcDNA3.1- NONHSAT030515 transfection and sh- NONHSAT030515 treatment were conducted to verify the role of lncRNA NONHSAT030515 in chondrogenic differentiation. The levels of Aggrecan, SOX9 and COL2A1 were analyzed by qRT-PCR and Western blot assay.ResultsAlcian blue staining, immunocytochemical, qRT-PCR, and Western blot have determined that lncRNA NONHSAT030515 can promote the chondrogenic differentiation of hADSCs. MiR-490- 5p was the direct target of NONHSAT030515, while BMPR2 was the target gene. This result was confirmed by luciferase reporter assay. Up-regulation of NONHSAT030515 promoted BMPR2 protein expression and promoted chondrogenic differentiation, whereas down-regulation of NONHSAT030515 caused completely opposite results.ConclusionLncRNA NONHSAT030515 promotes the chondrogenic differentiation of hADSCs through increasing BMPR2 expression by regulating miR-490- 5p.
Highlights
Osteoarthritis (OA) is a chronic degenerative disease due to the growth of age, overwork, and other factors caused by articular cartilage damage, called osteoarthrosis [1]
Results lncRNA NONHSAT030515 is upregulated during chondrogenic differentiation of human adipose-derived stem cells (hADSCs) To determine the expression level of lncRNA NONHSAT030515 during the chondrogenic differentiation of hADSCs, hADSCs were induced by microsphere culture for chondrogenic differentiation for 0, 7, 14 and 21 days
Chondrogenic differentiation of hADSCs was assessed by Alcian blue staining, and the deposition of cartilage matrix protein was most obvious after 21 days of induction
Summary
Osteoarthritis (OA) is a chronic degenerative disease due to the growth of age, overwork, and other factors caused by articular cartilage damage, called osteoarthrosis [1]. Studies on the mechanism of differentiation of mesenchymal stem cells into chondrocyte phenotypes and their regulation may help to develop a new therapeutic strategy OA [22]. Human adipose-derived stem cells (hADSCs) are considered to be a more ideal chondrogenic inducer than bone marrow mesenchymal stem cells (BMSCs) because of their easy access to a large number of cells and strong proliferative ability [3]. SOX9 and COL2A1 have been reported to be significantly up-regulated after chondrogenic differentiation of human adipose-derived stem cells, and they are markers of chondrogenic differentiation [9]. Chondrogenic differentiation of human adipose-derived stem cells (hADSCs) is important for cartilage generation and degradation. Our previous study showed that miR-490-5p was downregulated during chondrogenic differentiation of hADSCs. In this study, we investigated the effect and mechanism of lncRNA NONHSAT030515 interacting with miR-490-5p on chondrogenic differentiation of hADSCs
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