Objective: By producing instability in mouse knee joints, we attempted to determine the involvement of runt‐related transcription factor 2 (RUNX‐2), which is required for chondrocyte hypertrophy, in the development of osteoarthritis (OA).Methods: An experimental mouse OA model was created by surgical transection of the medial collateral ligament and resection of the medial meniscus of the knee joints of heterozygous RUNX‐2–deficient (Runx2^+/−) mice and wild‐type littermates. Cartilage destruction and osteophyte formation in the medial tibial cartilage were compared by histologic and radiographic analyses. Localization of type X collagen and matrix metalloproteinase 13 (MMP‐13) was examined by immunohistochemistry. Localization of RUNX‐2 was determined by X‐Gal staining in heterozygous RUNX‐2–deficient mice with the lacZ gene insertion at the Runx2‐deletion site (Runx2^+/lacZ). Messenger RNA levels of type X collagen, MMP‐13, and RUNX‐2 were examined by real‐time reverse transcriptase–polymerase chain reaction analysis.Results: RUNX‐2 was induced in the articular cartilage of wild‐type mice at the early stage of OA, almost simultaneously with type X collagen but earlier than MMP‐13. Runx2^+/− and Runx2^+/lacZ mice showed normal skeletal development and articular cartilage; however, after induction of knee joint instability, they exhibited decreased cartilage destruction and osteophyte formation, along with reduced type X collagen and MMP‐13 expression, as compared with wild‐type mice.Conclusion: RUNX‐2 contributes to the pathogenesis of OA through chondrocyte hypertrophy and matrix breakdown after the induction of joint instability.

Growing evidence shows that microRNAs (miRNAs) regulate various developmental and homeostatic events in vertebrates and invertebrates. Osteoblast differentiation is a key step in proper skeletal development and acquisition of bone mass; however, the physiological role of non-coding small RNAs, especially miRNAs, in osteoblast differentiation remains elusive. Here, through comprehensive analysis of miRNAs expression during osteoblast differentiation, we show that miR-206, previously viewed as a muscle-specific miRNA, is a key regulator of this process. miR-206 was expressed in osteoblasts, and its expression decreased over the course of osteoblast differentiation. Overexpression of miR-206 in osteoblasts inhibited their differentiation, and conversely, knockdown of miR-206 expression promoted osteoblast differentiation. In silico analysis and molecular experiments revealed connexin 43 (Cx43), a major gap junction protein in osteoblasts, as a target of miR-206, and restoration of Cx43 expression in miR-206-expressing osteoblasts rescued them from the inhibitory effect of miR-206 on osteoblast differentiation. Finally, transgenic mice expressing miR-206 in osteoblasts developed a low bone mass phenotype due to impaired osteoblast differentiation. Our data show that miRNA is a regulator of osteoblast differentiation.

Bone metastatic lesions are classified as osteoblastic or osteolytic lesions. Prostate and breast cancer patients frequently exhibit osteoblastic-type and osteolytic-type bone metastasis, respectively. In metastatic lesions, tumor cells interact with many different cell types, including osteoblasts, osteoclasts, and mesenchymal stem cells, resulting in an osteoblastic or osteolytic phenotype. However, the mechanisms responsible for the modification of bone remodeling have not been fully elucidated. MicroRNAs (miRNAs) are transferred between cells via exosomes and serve as intercellular communication tools, and numerous studies have demonstrated that cancer-secreted miRNAs are capable of modifying the tumor microenvironment. Thus, cancer-secreted miRNAs can induce an osteoblastic or osteolytic phenotype in the bone metastatic microenvironment. In this study, we performed a comprehensive expression analysis of exosomal miRNAs secreted by several human cancer cell lines and identified eight types of human miRNAs that were highly expressed in exosomes from osteoblastic phenotype-inducing prostate cancer cell lines. One of these miRNAs, hsa-miR-940, significantly promoted the osteogenic differentiation of human mesenchymal stem cells in vitro by targeting ARHGAP1 and FAM134A. Interestingly, although MDA-MB-231 breast cancer cells are commonly known as an osteolytic phenotype-inducing cancer cell line, the implantation of miR-940–overexpressing MDA-MB-231 cells induced extensive osteoblastic lesions in the resulting tumors by facilitating the osteogenic differentiation of host mesenchymal cells. Our results suggest that the phenotypes of bone metastases can be induced by miRNAs secreted by cancer cells in the bone microenvironment. Significance Prostate cancer is one of most common cancers in men worldwide, and osteoblastic bone metastasis is frequently observed in prostate cancer patients. However, the mechanisms responsible for the predominantly osteoblastic phenotype have not been fully elucidated. Cancer-secreted microRNAs (miRNAs) were recently shown to be significant in the modification of the tumor microenvironment. Here, hsa-miR-940, which was highly secreted by prostate cancer cells, promoted osteogenic differentiation of human mesenchymal stem cells in vitro, and induced extensive osteoblastic lesions in the bone metastatic microenvironment in vivo. Our study provides a demonstration that osteoblastic bone metastasis can be induced by miRNAs secreted by cancer cells in the bone microenvironment.

Significance: Current genomic and biochemical analysis revealed mutations in isocitrate dehydrogenase ( IDH ) genes associated with several neoplasms and a novel enzymatic activity of IDH mutations to catalyze α-ketoglutarate to d -2-hydroxyglutarate, contributing to tumorigenesis. We identified a broad range of IDH1 mutations, including a previously unidentified IDH1 -R132Q mutation, in cartilage tumors. Cartilage-specific Col2a1-Cre/ERT2;Idh1-R132 mutant knock-in mice developed multiple enchondroma-like lesions. These data show that mutant Idh in growth-plate cells causes persistence of chondrocytes, giving rise to enchondromas adjacent to the growth cartilage in bone.