Osteoarthritis (OA), the most prevalent aging-related joint disease, is characterized by insufficient extracellular matrix synthesis and articular cartilage degradation, mediated by several proteinases, including Adamts-5. miR-140 is one of a very limited number of noncoding microRNAs (miRNAs) specifically expressed in cartilage; however, its role in development and/or tissue maintenance is largely uncharacterized. To examine miR-140 function in tissue development and homeostasis, we generated a mouse line through a targeted deletion of miR-140. miR-140^−/− mice manifested a mild skeletal phenotype with a short stature, although the structure of the articular joint cartilage appeared grossly normal in 1-mo-old miR-140^−/− mice. Interestingly, miR-140^−/− mice showed age-related OA-like changes characterized by proteoglycan loss and fibrillation of articular cartilage. Conversely, transgenic (TG) mice overexpressing miR-140 in cartilage were resistant to antigen-induced arthritis. OA-like changes in miR-140-deficient mice can be attributed, in part, to elevated Adamts-5 expression, regulated directly by miR-140. We show that miR-140 regulates cartilage development and homeostasis, and its loss contributes to the development of age-related OA-like changes.

Objective: Several microRNA, which are ∼22‐nucleotide noncoding RNAs, exhibit tissue‐specific or developmental stage–specific expression patterns and are associated with human diseases. The objective of this study was to identify the expression pattern of microRNA‐146 (miR‐146) in synovial tissue from patients with rheumatoid arthritis (RA).Methods: The expression of miR‐146 in synovial tissue from 5 patients with RA, 5 patients with osteoarthritis (OA), and 1 normal subject was analyzed by quantitative reverse transcription–polymerase chain reaction (RT‐PCR) and by in situ hybridization and immunohistochemistry of tissue sections. Induction of miR‐146 following stimulation with tumor necrosis factor α (TNFα) and interleukin‐1β (IL‐1β) of cultures of human rheumatoid arthritis synovial fibroblasts (RASFs) was examined by quantitative PCR and RT‐PCR.Results: Mature miR‐146a and primary miR‐146a/b were highly expressed in RA synovial tissue, which also expressed TNFα, but the 2 microRNA were less highly expressed in OA and normal synovial tissue. In situ hybridization showed primary miR‐146a expression in cells of the superficial and sublining layers in synovial tissue from RA patients. Cells positive for miR‐146a were primarily CD68+ macrophages, but included several CD3+ T cell subsets and CD79a+ B cells. Expression of miR‐146a/b was markedly up‐regulated in RASFs after stimulation with TNFα and IL‐1β.Conclusion: This study shows that miR‐146 is expressed in RA synovial tissue and that its expression is induced by stimulation with TNFα and IL‐1β. Further studies are required to elucidate the function of miR‐146 in these tissues.

Objective: MicroRNA (miRNA) are a class of noncoding small RNAs that act as negative regulators of gene expression. MiRNA exhibit tissue‐specific expression patterns, and changes in their expression may contribute to pathogenesis. The objectives of this study were to identify miRNA expressed in articular chondrocytes, to determine changes in osteoarthritic (OA) cartilage, and to address the function of miRNA‐140 (miR‐140).Methods: To identify miRNA specifically expressed in chondrocytes, we performed gene expression profiling using miRNA microarrays and quantitative polymerase chain reaction with human articular chondrocytes compared with human mesenchymal stem cells (MSCs). The expression pattern of miR‐140 was monitored during chondrogenic differentiation of human MSCs in pellet cultures and in human articular cartilage from normal and OA knee joints. We tested the effects of interleukin‐1β (IL‐1β) on miR‐140 expression. Double‐stranded miR‐140 (ds–miR‐140) was transfected into chondrocytes to analyze changes in the expression of genes associated with OA.Results: Microarray analysis showed that miR‐140 had the largest difference in expression between chondrocytes and MSCs. During chondrogenesis, miR‐140 expression in MSC cultures increased in parallel with the expression of SOX9 and COL2A1. Normal human articular cartilage expressed miR‐140, and this expression was significantly reduced in OA tissue. In vitro treatment of chondrocytes with IL‐1β suppressed miR‐140 expression. Transfection of chondrocytes with ds–miR‐140 down‐regulated IL‐1β–induced ADAMTS5 expression and rescued the IL‐1β–dependent repression of AGGRECAN gene expression.Conclusion: This study shows that miR‐140 has a chondrocyte differentiation–related expression pattern. The reduction in miR‐140 expression in OA cartilage and in response to IL‐1β may contribute to the abnormal gene expression pattern characteristic of OA.

Mohawk ( Mkx ) is a member of the Three Amino acid Loop Extension superclass of atypical homeobox genes that is expressed in developing tendons. To investigate the in vivo functions of Mkx, we generated Mkx ^−/− mice. These mice had hypoplastic tendons throughout the body. Despite the reduction in tendon mass, the cell number in tail tendon fiber bundles was similar between wild-type and Mkx ^−/− mice. We also observed small collagen fibril diameters and a down-regulation of type I collagen in Mkx ^−/− tendons. These data indicate that Mkx plays a critical role in tendon differentiation by regulating type I collagen production in tendon cells.

The transcriptional activation by SRY-type high mobility group box 9 (SOX9) and the transforming growth factor β (TGF-β) signals are necessary for chondrogenic differentiation. We have previously shown that CREB-binding protein (CBP/p300) act as an important SOX9 co-activator during chondrogenesis. In the present study, we investigated the relationship between TGF-β-dependent Smad2/3 signaling pathways and the SOX9-CBP/p300 transcriptional complex at the early stage of chondrogenesis. Overexpressed Smad3 strongly induced the primary chondrogenesis of human mesenchymal stem cells. In addition, Smad3 enhanced the transcriptional activity of SOX9 and the expression of α1(II) collagen gene (COL2A1), and small interference RNA against Smad3 (si-Smad3) inhibited them. We observed that Smad2/3 associated with Sox9 in a TGF-β-dependent manner and formed the transcriptional complexes with SOX9 on the enhancer region of COL2A1. Interestingly, the association between Sox9 and CBP/p300 was increased by Smad3 overexpression and was suppressed by si-Smad3. Our findings indicate that Smad3 has a stronger potential to stimulate the SOX9-dependent transcriptional activity by modulating the interaction between SOX9 and CBP/p300, rather than Smad2. This study suggests that the Smad3 pathway presents a key role for the SOX9-dependent transcriptional activation in primary chondrogenesis.