Long non-coding RNA XIST protects chondrocytes ATDC5 and CHON-001 from IL-1β-induced injury via regulating miR-653-5p/SIRT1 axis.

Chondrocyte apoptosis is linked to cartilage degeneration, and considered as a crucial event during the development of osteoarthritis (OA). X inactive specific transcript (XIST) is an oncogenic long non-coding RNA (lncRNA). However, its role in the pathophysiological process of OA remains largely unknown. In this work, quantitative real-time reverse transcriptase PCR (qRT-PCR) was employed to measure the expression of XIST, miR-653-5p and sirtuin1 (SIRT1) mRNA in OA and normal cartilage tissues. Chondrocyte cell lines, CHON-001 and ATDC5, were treated with different doses of interleukin- 1β (IL-1β) to mimic the inflammatory environment of OA in vitro. Overexpression plasmids, microRNA (miRNA) mimics, miRNA inhibitors and small interfering RNAs (siRNAs) were constructed and transfected into CHON-001 and ATDC5 cells. 3-(4,5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2H-tetrazolium bromide (MTT) assay was adopted to determine the cell viability. Western blot was used to detect the expression of apoptosis-related proteins. Enzyme-linked immunosorbent assay (ELISA) was employed to probe the expression levels of inflammatory factors. Flow cytometry was used to analyze the cell apoptosis. StarBase and TargetScan databases were used to predict the binding sites between XIST and miR-653-5p, miR-653-5p and 3'UTR of SIRT1, respectively, which were then verified by dual luciferase reporter assay. The data in the present study demonstrated that XIST and SIRT1 were down-regulated while miR-653-5p was up-regulated in OA tissues and cell models. The up-regulation of XIST increased the viability of CHON-001 and ATDC5 cells, while it impeded their apoptosis and inflammatory response induced by IL-1β. Conversely, miR-653-5p had opposite effects. It was proved that miR-653-5p could be sponged and suppressed by XIST. Additionally, SIRT1 was identified as a target of miR-653-5p, and SIRT1 could be suppressed by XIST indirectly. In conclusion, down-regulated XIST was involved in the injury of chondrocytes during the pathophysiological process of OA, and XIST up-regulation protected chondrocytes from inflammatory injury via regulating miR-653-5p/SIRT1 axis. Copyright 2020 Biolife Sas. www.biolifesas.org.