The expression and function of microRNAs in chondrogenesis and osteoarthritis

Abstract

Objective. To use an in vitro model of chondrogenesis to identify microRNAs (miRNAs) with a functional role in cartilage homeostasis.Methods. The expression of miRNAs was measured in the ATDC5 cell model of chondrogenesis using microarray and was verified using quantitative reverse transcription-polymerase chain reaction. MicroRNA expression was localized by in situ hybridization. Predicted miRNA target genes were validated using 3-untranslated region-Luc reporter plasmids containing either wild-type sequences or mutants of the miRNA target sequence. Signaling through the Smad pathway was measured using a (CAGA) 12 -Luc reporter.Results. The expression of several miRNAs was regulated during chondrogenesis. These included 39 miRNAs that are coexpressed with miRNA-140 (miR-140), which is known to be involved in cartilage homeostasis and osteoarthritis (OA). Of these miRNAs, miR-455 resides within an intron of COL27A1 that encodes a cartilage collagen. When human OA cartilage was compared with cartilage obtained from patients with femoral neck fractures, the expression of both miR-140-5p and miR-455-3p was increased in OA cartilage. In situ hybridization showed miR-455-3p expression in the developing limbs of chicks and mice and in human OA cartilage. The expression of miR-455-3p was regulated by transforming growth factor ␤ (TGF␤) ligands, and miRNA regulated TGF␤ signaling. ACVR2B, SMAD2, and CHRDL1 were direct targets of miR-455-3p and may mediate its functional impact on TGF␤ signaling.Conclusion. MicroRNA-455 is expressed during chondrogenesis and in adult articular cartilage, where it can regulate TGF␤ signaling, suppressing the Smad2/3 pathway. Diminished signaling through this pathway during the aging process and in OA chondrocytes is known to contribute to cartilage destruction. We propose that the increased expression of miR-455 in OA exacerbates this process and contributes to disease pathology.Osteoarthritis (OA) is a degenerative joint disease characterized by degradation of articular cartilage, thickening of subchondral bone, and formation of osteophytes (1). The etiology of OA is complex, with the contribution of genetic, developmental, biochemical, and biomechanical factors. Chondrocytes are the only cells in cartilage and are responsible for the synthesis and turnover of extracellular matrix (ECM), which is crucial to tissue function.During development, mesenchymal cells aggregate and differentiate into chondrocytes, which undergo a series of differentiation events: proliferation, hypertrophy, terminal differentiation, mineralization, and programmed cell death. Blood vessels penetrate the calcified matrix, bringing in osteoblasts that build new bone. The cartilage model grows by rounds of chondrocyte cell

MiR‐455‐3p regulates early chondrogenic differentiation via inhibiting Runx2

“…MiRNAs play an important role in differentiation and development across a wide range of organisms and tissue types, including bone and cartilage development [1,5,6,8,[24][25][26]38]. To explore the specific involvement of miRNAs in the chondrogenesis of MSCs, we previously used the miRNA microarray technique to profile miRNA expression and screen miRNAs with significant differences in expression before and after chondrogenic induction.…”

“…Furthermore, Runx2 regulates the differentiation of MSCs as the target of miRNAs [22,23]. Recently, miR-455-3p was found to be expressed in the ATDC5 cell model of chondrogenesis and in adult articular cartilage, as shown in human MSCs, and it regulated TGFb signaling by suppressing the Smad2/3 pathway, which contributes to cartilage destruction [5]. In this study, we investigated the function of miR-455-3p in early chondrogenesis to explore its relationship with Runx2.…”

“…MiRNAs function as regulators in gene silencing by binding to the 3 0 -untranslated region (3 0 UTR) of target mRNAs, leading to translational repression. miRNAs have been reported to have important roles in multiple biological processes, including the process of cartilage differentiation and degradation [1,[3][4][5][6][7][38][39][40]. Dicer is an essential component for the biogenesis of miRNAs, and Dicer-null growth plates showed a progressive reduction in the proliferation of chondrocytes and the acceleration of hypertrophy, leading to severe skeletal growth defects and premature death in mice [8].…”

“…MiR-455-3p was first sequenced in 250 small RNA libraries from 26 different organ systems and types of cells in humans and rodents that were enriched in neuronal as well as normal and malignant hematopoietic cells and tissues [11]; it was then found in human cancer cells [12,13]. Recently, miR-455-3pwas found to be involved in chondrogenesis, and its actions against the DPP homologs in the (Smad)-2/3 pathway were suppressed by regulating TGFb signaling [1,5]. Based on our continuing investigation on the exact biological function and mechanisms of these miRNAs, we found that miR-455-3p was an interesting regulator during chondrogenesis and might have a role in osteoarthritis by potentially targeting Runt-related transcription factor 2 (Runx2) as we previously reported after computational analysis [1].…”

“…Deletion of Runx2 in chondrocytes causes failure of endochondral ossification and lethality at birth, and it directly regulates a set of cell cycle genes Gpr132, Sfn, c-Myb, and Cyclin A1 to control the proliferative capacity of chondrocytes [20]. Runx2 also mediates chondrogenesis and ossification as a co-regulator with Smad, TGF-b, and CCAAT/enhancer binding protein beta (C/EBPb) [5,21]. Furthermore, Runx2 regulates the differentiation of MSCs as the target of miRNAs [22,23].…”