Mir-381-3p aggravates ovariectomy-induced osteoporosis by inhibiting osteogenic differentiation through targeting KLF5/Wnt/β-catenin signaling pathway.

Increasing evidence shows the pivotal significance of miRNAs in the pathogenesis of osteoporosis. miR-381-3p has been identified as an inhibitor of osteogenesis. This study explored the role and mechanism of miR-381-3p in postmenopausal osteoporosis (PMOP), the most common type of osteoporosis. Bilateral ovariectomy (OVX) rat model was established and miR-381-3p antagomir was administrated through the tail vein in vivo. The pathological changes in rats were assessed through the evaluation of serum bone turnover markers (BALP, PINP, and CTX-1), hematoxylin and eosin (H&E) staining, as well as the expression of osteoblast differentiation biomarkers. Moreover, isolated bone marrow mesenchymal stem cells from OVX-induced rats (OVX-BMMSCs) were utilized to explore the impact of miR-381-3p on osteoblast differentiation. In addition, the target gene and downstream pathway of miR-381-3p were further investigated both in vivo and in vitro. miR-381-3p expression was elevated, whereas KLF5 was suppressed in OVX rats. miR-381-3p antagomir decreased serum levels of bone turnover markers, improved trabecular separation, promoted osteoblast differentiation biomarker expression in OVX rats. ALP activity and mineralization were suppressed, and levels of osteoblast differentiation biomarkers were impeded after miR-381-3p overexpression during osteoblast differentiation of OVX-BMMSCs. While contrasting results were found after inhibition of miR-381-3p. miR-381-3p targets KLF5, negatively affecting its expression as well as its downstream Wnt/β-catenin pathway, both in vivo and in vitro. Silencing of KLF5 restored Wnt/β-catenin activation induced by miR-381-3p antagomir. miR-381-3p aggravates PMOP by inhibiting osteogenic differentiation through targeting KLF5/Wnt/β-catenin pathway. miR-381-3p appears to be a promising candidate for therapeutic intervention in PMOP.

Zhao Y ，Liu J ，Zhang Y ，Liang M ，Li R ，Song Y ，Wang Y ... -  《Journal of Orthopaedic Surgery and Research》

MiR-23b-3p promotes postmenopausal osteoporosis by targeting MRC2 and regulating the Wnt/β-catenin signaling pathway.

Postmenopausal osteoporosis (PMOP) is one of the most common metabolic bone diseases in postmenopausal women. Increasing evidence has indicated that microRNAs (miRNAs) play vital regulatory roles during osteoporosis progression. This study aimed to investigate the potential function of miR-23b-3p in the osteogenic differentiation of human bone marrow mesenchymal stem cells (hMSCs). PMOP was induced in mice by bilateral ovariectomy. X-ray absorptiometry was applied to detect BMD and BMC in PMOP mice. Luciferase reporter assay and RIP assay were utilized to investigate the relationship between miR-23b-3p and MRC2. We found the upregulation of miR-23b-3p in bone tissues of PMOP mice. Silencing of miR-23b-3p relieved PMOP in mice. Moreover, miR-23b-3p knockdown facilitated the osteogenic differentiation of hMSCs by increasing the expression of Runx2, OCN, Osterix and promoting ALP activity. Mechanistically, MRC2 is a downstream target gene of miR-23b-3p. MRC2 knockdown significantly rescued the promoting effect of lenti-miR-23b-3p inhibitor on osteogenic differentiation of hMSCs. Furthermore, miR-23b-3p targeted MRC2 to inhibit the Wnt/β-catenin pathway during the osteogenic differentiation of hMSCs. In summary, inhibition of miR-23b-3p alleviates PMOP by targeting MRC2 to inhibit the Wnt/β-catenin signaling, which may provide a novel molecular insight for osteoporosis therapy.

Li R ，Ruan Q ，Yin F ，Zhao K ... -  《-》

Foxf1 knockdown promotes BMSC osteogenesis in part by activating the Wnt/β-catenin signalling pathway and prevents ovariectomy-induced bone loss.

Forkhead box protein f1 (Foxf1) is associated with cell differentiation, and may be a key player in bone homoeostasis. However, the effect of Foxf1 on osteogenesis of bone marrow-derived mesenchymal stem cells (BMSCs) and ovariectomy-induced bone loss, as well as its clinical implications, is unknown. By quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR) and western blotting, we assayed Foxf1 expression in bone tissue, BMSCs, and bone marrow-derived macrophages (BMMs), derived from ovariectomised (OVX) mice, and during osteogenic differentiation and osteoclast differentiation. Using a loss-of-function approach (small interfering RNA [siRNA]-mediated knockdown) in vitro, we examined whether Foxf1 regulates osteoblast differentiation of BMSCs via the Wnt/β-catenin signalling pathway. Furthermore, we assessed the anabolic effect of Foxf1 knockdown (siFoxf1) in OVX mice in vivo. We also assayed the expression of Foxf1 in bone tissue derived from postmenopausal osteoporosis (PMOP) patients and its link with bone mineral density (BMD). Finally, we examined the effect of Foxf1 knockdown on the osteoblastic differentiation of human BMSCs. Foxf1 expression was significantly increased in bone extract and BMSCs from OVX mice and gradually decreased during osteoblastic differentiation of BMSCs but did not differ significantly in OVX mouse-derived BMMs or during osteoclast differentiation. In vitro, Foxf1 knockdown markedly increased the expression of osteoblast specific genes, alkaline phosphatase (ALP) activity, and mineralisation. Moreover, siFoxf1 activated the Wnt/β-catenin signalling pathway. The siFoxf1-induced increase in osteogenic differentiation was partly rescued by inhibitor of Wnt signalling (DKK1). In OVX mice, Foxf1 siRNA significantly reduced bone loss by enhancing bone formation. Foxf1 expression levels negatively correlated with reduced bone mass and bone formation in bone tissue from PMOP patients. Finally, Foxf1 knockdown significantly promoted osteogenesis by human BMSCs. Our findings indicate that Foxf1 knockdown promotes BMSC osteogenesis and prevents OVX-induced bone loss. Therefore, Foxf1 has potential as a biomarker of osteogenesis and may be a therapeutic target for PMOP.

Shen G ，Ren H ，Shang Q ，Zhao W ，Zhang Z ，Yu X ，Tang K ，Tang J ，Yang Z ，Liang D ，Jiang X ... -  《EBioMedicine》

Down-regulated microRNA-199a-3p enhances osteogenic differentiation of bone marrow mesenchymal stem cells by targeting Kdm3a in ovariectomized rats.

Osteoporosis is a prevalent systemic skeletal disorder entailing bone fragility and increased fracture risk, often emerging in post-menopausal life. Emerging evidence implicates the dysregulation of microRNAs (miRNAs or miRs) in the progression of osteoporosis. This study investigated the effect of miR-199a-3p on osteoporosis and its underlying mechanism. We first examplished an ovariectomized (OVX)-induced rat osteoporosis model, and then isolated mesenchymal stem cells (MSCs) from bone marrow of the model rats. The overexpression and knock down of miR-199a-3p were conducted in OVX rats and MSCs to verify the role of miR-199a-3p on MSC differentiation. Calcium nodules were measured using alizarin red S (ARS) staining. RT-qPCR and Western blot assay were performed to measure the expression of miR-199a-3p, Kdm3a and osteogenic differentiation-related markers in rat tissues and cells. The correlation between miR-199a-3p and Kdm3a was confirmed using dual-luciferase reporter assay. The enrichment of Kdm3a at the Erk2 and Klf2 promoter was assessed using chromatin immunoprecipitation (ChIP) assay. Isolated MSCs were positive for CD29, CD44, CD90, and CD45, suggesting successful isolation of MSCs. There was increased expression of miR-199a-3p and inhibited osteogenic differentiation in OVX rats. Kdm3a was negatively targeted by miR-199a-3p. Our results also demonstrated that Kdm3a elevated the expression of Erk2 and Erk2 by promoting Erk2 and Klf2 demethylation, which further contributed to osteogenic differentiation. Overall, our results revealed a regulatory network of miR-199a-3p in osteogenic differentiation, highlighting miR-199a-3p as a potential target for therapeutic interventions in osteoporosis.

Wu JC ，Sun J ，Xu JC ，Zhou ZY ，Zhang YF ... -  《-》

CircRNA hsa_circ_0006859 inhibits the osteogenic differentiation of BMSCs and aggravates osteoporosis by targeting miR-642b-5p/miR-483-3p and upregulating EFNA2/DOCK3.

Hsa_circ_0006859 has been found as a possible biomarker for postmenopausal osteoporosis (PMOP) with an effect on the osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs), but the underlying mechanism is unclear. Bioinformatics analysis was used to identify dysregulated RNAs involved in osteoporosis based on public datasets. Function assays were used to determine the functions of hsa_circ_0006859 on cell proliferation and osteogenic differentiation in vitro. It was found that hsa_circ_0006859 was upregulated in OVX mice-derived BMSCs, but lowly expressed during osteogenic differentiation. Overexpressing hsa_circ_0006859 inhibited the cell proliferation and osteogenesis of BMSCs and hFOB 1.19 cells, vice versa. Bilateral ovariectomy (OVX) was used to induce PMOP in mice. The interactions among circ_0006859, miR-642b-5p/miR-483-3p, and EFNA2/DOCK3 were determined using the RIP assay. Silencing circ_0006859 relieved PMOP in mice. Mechanistically, circ_0006859 bound to miR-642b-5p/miR-483-3p directly, while miR-642b-5p and miR-483-3p respectively targeted EFNA2 and DOCK3. Hsa_circ_0006859 downregulated the expression of miR-642b-5p/miR-483-3p to upregulate EFNA2/DOCK3. Additionally, miR-642b-5p/miR-483-3p targeted EFNA2/DOCK3 to inhibit BMSCs osteogenic differentiation and facilitate osteoporosis progression by inactivating the Wnt signaling. In conclusion, hsa_circ_0006859 is involved in PMOP by targeting miR-642b-5p/EFNA2 and miR-483-3p/DOCK3 axes to maintain the Wnt-signaling pathway, which may be a novel possible therapeutic targets and biomarkers for PMOP.

Yin P ，Xue Y 《-》