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.
Lian LP
，Xi XY
《JOURNAL OF BIOLOGICAL REGULATORS AND HOMEOSTATIC AGENTS》
Down-regulation of MiR-138-5p Protects Chondrocytes ATDC5 and CHON-001 from IL-1 β-induced Inflammation Via Up-regulating SOX9.
Osteoarthritis (OA) pertains to a chronic disease of degenerative joints distinguished by articular cartilage destruction, subchondral bone remodeling, osteophyte formation, and inflammatory changes. Chondrocyte apoptosis is inextricably linked to cartilage degeneration. SRY-related high-mobility-group-box 9 (SOX9) is a well-acknowledged transcription factor in the chondrogenesis. Nevertheless, the detailed function of miR-138-5p/SOX9 in OA remains to be fully clarified.
qRT-PCR was performed to measure the expressions of miR-138-5p and SOX9 mRNA in OA and normal cartilage tissues and cells. Human chondrocyte cell lines, CHON-001 and ATDC5, were treated with different doses of interleukin-1β (IL-1β) to simulate the inflammatory response environment of OA. miR-138-5p mimics, miR-138-5p inhibitors, and SOX9 small interfering RNA (siRNA) were constructed and transfected into CHON-001 and ATDC5 cells. CCK-8 was conducted to determine the cell viability and transwell assay was used to monitor the migration of cells. Western blot was carried out to detect the expressions of apoptosis- related factors. Enzyme-linked immunosorbent assay (ELISA) was adopted to measure the contents of inflammatory factors. TargetScan predicted SOX9 was a target gene of miR-138-5p, which was then verified by luciferase assay.
miR-138-5p expression was down-regulated in OA and regulated SOX9 expression. The downregulation of miR-138-5p facilitated the proliferation and migration of CHON-001 and ATDC5 cells, while impeded their apoptosis and inflammatory response. Besides, down-regulated SOX9 can counteract the promoting effect of down-regulated miR-138-5p on the proliferation and migration of chondrocytes.
miR-138-5p can arrest the proliferation and migration of CHON-001 and ATDC5 via restraining SOX9, and facilitate the apoptosis and inflammation. This study revealed the protective effect of down-regulated miR-138-5p on the inflammatory injury of chondrocytes caused by IL-1β.
Chunlei H
，Chang Z
，Sheng L
，Yanchun Z
，Lulin L
，Daozhang C
... -
《-》
lncRNA OIP5-AS1 attenuates the osteoarthritis progression in IL-1β-stimulated chondrocytes.
In view of the association between long noncoding RNA OIP5-AS1 and osteoarthritis (OA) pathology, the corresponding potential mechanism is worthy of exploration. Primary chondrocytes were identified by morphological observation and immunohistochemical staining of collagen II. The association between OIP5-AS1 and miR-338-3p was analyzed by StarBase and dual-luciferase reporter assay. After the expression of OIP5-AS1 or miR-338-3p in interleukin (IL)-1β-stimulated primary chondrocytes and CHON-001 cells was manipulated, cell viability, proliferation, apoptosis rate, apoptosis-related protein (cleaved caspase-9, Bax) expressions, extracellular matrix (ECM) (matrix metalloproteinase (MMP)-3, MMP-13, aggrecan, and collagen II), PI3K/AKT pathway, and mRNA expressions of inflammatory factors (IL-6 and IL-8), OIP5-AS1, and miR-338-3p were determined by cell counting kit-8, EdU, flow cytometry, Western blot, and quantitative reverse transcription-polymerase chain reaction. As a result, the expression of OIP5-AS1 was downregulated in IL-1β-activated chondrocytes, while miR-338-3p was overexpressed. OIP5-AS1 overexpression reversed the effects of IL-1β on viability, proliferation, apoptosis, ECM degradation, and inflammation in chondrocytes. However, OIP5-AS1 knockdown exhibited opposite effects. Interestingly, the effects of OIP5-AS1 overexpression were partially offset by miR-338-3p overexpression. Furthermore, OIP5-AS1 overexpression blocked the PI3K/AKT pathway by modulating miR-338-3p expression. In sum, OIP5-AS1 promotes viability and proliferation, and inhibits apoptosis and ECM degradation in IL-1β-activated chondrocytes by targeting miR-338-3p through blocking the PI3K/AKT pathway, indicating an attractive strategy for OA treatment.
Zhang X
，Wang Z
，Wang B
，Li J
，Yuan H
... -
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