BackgroundAseptic loosening of artificial hip joint is a major complication affecting the long-term use of the artificial hip joint, and is the main cause of joint replacement failure. However, the mechanism of aseptic loosening of THR has not yet cleared. The aim of this study was to investigate the underlying mechanism of DANCR in osteoblast differentiation (OD).MethodsWe detected the expressions of DANCR and FOXO1 in clinical samples and mesenchymal stem cells (MSCs) by qRT-PCR and western blotting. The effects of polymethylmethacrylate (PMMA) on OD of MSCs were examined by alkaline phosphatase (ALP) activity and Alizarin Red S (ARS) staining. The expressions of OD markers were measured by qRT-PCR and western blotting. The mechanism of DANCR in OD was detected by RNA pull-down, RNA immunoprecipitation (RIP) assay and ubiquitination assays.ResultsCompared with the surrounding normal tissues, DANCR expression was up-regulated and FOXO1 expression was down-regulated in periprosthetic tissues. PMMA suppressed ALP activity, increased DANCR expression, and decreased the expressions of FOXO1, Runx2, Osterix (Ostx) and osteocalcin (OCN). ARS staining showed that PMMA inhibited the OD of MSCs. Knockdown of DANCR attenuated the inhibitory effect of PMMA on OD. Knockdown of FOXO1 could reverse the effect of si-DANC. RNA pull-down and RIP assay implicated that DANCR bound to FOXO1. Ubiquitination assay indicated that si-DANCR could repress Skp2-mediated ubiquitination of FOXO1.ConclusionLncRNA DANCR could inhibit OD by regulating FOXO1 expression.
Increasing evidence indicates that long non-coding RNAs (lncRNAs) play critical roles in osteoarthritis (OA). The present study aimed to investigate the underlying molecular mechanism of lncRNA musculin antisense RNA 1 (MSC-AS1) in OA. RT-qPCR was used to detect MSC-AS1 levels in cartilage tissues from patients with OA. The effects of MSC-AS1 knockdown on the viability and apoptosis in OA were evaluated via CCK-8 and TUNEL assays. The StarBase database was used to predict the binding sites between microRNA (miR)-369-3p and MSC-AS1 or JAK2, which were confirmed via the dual-luciferase reporter assay. The results demonstrated that MSC-AS1 expression was downregulated in OA. Functional analysis indicated that the addition of MSC-AS1 promoted viability and inhibited inflammation and the apoptosis of chondrocytes. In addition, MSC-AS1 regulated the survival of OA chondrocytes by sponging miR-369-3p. JAK2 was confirmed as a direct target of miR-369-3p, and MSC-AS1 regulated JAK2/STAT3 signaling via miR-369-3p in OA chondrocytes. Taken together, our results suggest that MSC-AS1 may regulate the miR-369-3p/JAK2/STAT3 signaling pathway to accelerate the viability, and inhibit inflammation and cell apoptosis in OA chondrocytes.
Osteogenic differentiation and osteolysis after hip replacement are both associated with bone metabolism. Interaction between the long non-coding RNA (lncRNA) prostate cancer non-coding RNA 1 (PRNCR1) and miR-211-5p was analyzed to illuminate their roles in osteogenic differentiation and osteolysis. The expression of PRNCR1, miR-211-5p and C-X-C chemokine receptor-4 (CXCR4) protein in tissues and mesenchymal stem cells (MSCs) were determined by qRT-PCR and western blot, separately. The osteogenic differentiation was assessed with Alkaline phosphatase (ALP) activity detection and ARS staining. The endogenous expressions of genes were modulated by recombinant plasmid and cell transfection. Combination condition and interaction between RNA and protein were determined with RIP and RNA pull-down assay, respectively. Interaction between miR-211-5p and CXCR4 was examined with Dual luciferase reporter assay. PRNCR1 and CXCR4 were up-regulated in wear particles around prosthesis and in MSCs incubated with Polymethylmethacrylate (PMMA), while miR-211-5p was down-regulated. Repression of PRNCR1 weakened the inhibitory effect of wear particles on osteogenic differentiation. PRNCR1 positively regulated CXCR4 through inhibiting miR-211-5p. Wear particles regulated CXCR4 level through miR-211-5p to affect osteogenic differentiation of MSCs. Wear particles regulated the miR-211-5p level through PRNCR1 to affect osteogenic differentiation of MSCs. LncRNA PRNCR1 up-regulates CXCR4 through inhibiting miR-211-5p, which inhibits osteogenic differentiation and thereby leading to osteolysis after hip replacement.
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