A microRNA screen reveals the critical role of microRNA-23a-3p in maintaining cartilage homeostasis

Abstract

Purpose: Identification of biological therapeutic targets would be beneficial for patients with early osteoarthritis (OA) to prevent ongoing joint damage. In this study we identified and characterized a novel cartilage specific microRNA (miR) miR-23a-3p and investigated its role and therapeutic effect for OA. Methods: We explored the regulatory role of miR-23a-3p and their potential transcriptional and signaling targets using computational, in vitro and in vivo studies. miR-23a-3p levels in human normal and OA cartilage derived chondrocytes were assessed by polymerase chain reaction (PCR). Primary human chondrocytes were transfected with miR-23a-3p mimic or inhibitor, PCR and western blotting were performed to quantify catabolic and anabolic genes involved in cartilage. Ingenuity analysis was used to identify the potential target genes and respective functions. An OA model was induced surgically in rats, and subsequently treated with intra-articular injection of miR-23a-3p inhibitors and OA progression were evaluated macroscopically, histologically, and immunohistochemically in these rats. Results: Microarray profiling of 1,142 miRNAs identified miR-23a-3p as one of the most down-regulated miRNA in cartilage tissues sourced from patients with varying degrees of OA (mild to severe degeneration) and also in surgically induced meniscectomy OA in rats. Furthermore, IL-1β stimulated chondrocytes had lower expression of miR-23a-3p as well as chondrocyte-specific markers. Over-expression of miR-23a-3p suppressed the IL-1β induced catabolic effects, whereas the suppression of miR-23a-3p in OA chondrocytes enhanced the chondrocyte-specific marker expression. We identified Runt-related transcription factor 2 (RUNX2) as a direct target of miR-23a-3p and showed a down- and up-regulation of RUNX2 levels after transfection of, respectively with premiR- and antagomiR-23a-3p oligonucleotides. The levels of RUNX2 expression is highly expressed in the OA cartilage and inhibition of RUNX2 with lentiviral approach suppressed the chondrogenic gene expression but accelerated the hypertrophy and degradative marker expression. Pharmacological inhibition of miR-23a-3p in normal rats led to cartilage matrix loss and mimicked an OA phenotype. Conclusions: This is the first report to our knowledge that identifies miR-23a-3p as mediator of cartilage degradation in OA and could be a potential therapeutic target for stopping cartilage degradation.