| Objective To explore the expression pattern of miR-133a in patients with osteoporotic fractures (OPF) and its effects on osteoblast activity, differentiation, apoptosis, and the underlying molecular mechanism. Methods Bone tissue and serial blood samples were collected from 20 OPF patients and 20 patients with non-osteoporotic traumatic fractures (non-OPF group). The expression levels of miR-133a and RUNX2 mRNA in bone tissue, blood, and isolated osteoblasts were detected by quantitative real-time PCR (qRT-PCR). RUNX2 protein level was assessed by Western Blot. The mouse osteoblastic cell line MC3T3-E1 was cultured in vitro and transfected with miR-133a mimics, miR-133a inhibitor, or their corresponding negative controls. Rescue experiments were performed using the groups: miR-133a inhibitor + si-RUNX2 and miR-133a mimics + RUNX2 overexpression plasmid. The effects of miR-133a on osteoblast proliferation, apoptosis, differentiation, and the expression of osteogenic-related genes (RUNX2, BMP2, ALP, COL-I) and apoptosis-related genes (Bax, Bcl-2) were systematically evaluated using CCK-8 assay, flow cytometry, ALP/Alizarin Red S staining, qRT-PCR, and Western Blot. The targeting relationship between miR-133a and RUNX2 was verified by dual-luciferase reporter assay. Results The expression of miR-133a in bone tissue, blood, and osteoblasts of the OPF group was significantly higher than that in the non-OPF group (P<0.05), while the expression of RUNX2 mRNA and protein was significantly lower (P<0.05). The level of miR-133a in the blood of OPF patients decreased significantly over time after treatment. The dual-luciferase assay confirmed that miR-133a could directly bind to the 3'UTR of RUNX2. Cell experiments showed that overexpression of miR-133a inhibited osteoblast proliferation and differentiation, promoted apoptosis, downregulated the mRNA and protein expression of RUNX2, BMP2, ALP, and COL-I, and decreased the Bcl-2/Bax ratio. Inhibition of miR-133a produced the opposite effects. Rescue experiments confirmed that knocking down RUNX2 partially reversed the pro-osteogenic effects of the miR-133a inhibitor, while overexpressing RUNX2 partially reversed the anti-osteogenic effects of miR-133a mimics. Conclusion MiR-133a is highly expressed in OPF and negatively regulates the proliferation, differentiation, and anti-apoptotic capacity of osteoblasts by targeting and inhibiting RUNX2, thereby impeding the fracture healing process. |