miRNA-30b-5p Targets P4HA2 to Regulate Osteosarcoma Cell Proliferation, Invasion, Migration, and Apoptosis

Objective: This study aims to elucidate the mechanism by which the microRNA miR-30b-5p inhibits the proliferation, invasion, and migration of osteosarcoma cells while inducing apoptosis through the regulation of P4HA2. Methods: The MG63 osteosarcoma cell line was transfected with miR-NC and miR-30b-5p mimics to establish negative control and miR-30b-5p up-regulated expression models. Subsequently, various assays were conducted, including the CCK8 assay, Transwell assay, flow cytometry, dual luciferase reporter assay, real-time PCR (RT-PCR), and Western blot analysis. Results: Following the transfection of MG63 cells with miR-30b-5p mimics, the expression of miR-30b-5p was significantly up-regulated compared to the control group (t = 77.53, p = 0.0002). After transfecting the cells with miR-NC and miR-30b-5p mimics for 24, 48, and 72 hours, the CCK8 assay results indicated a marked reduction in proliferative ability relative to the control group (24 h: t = 16, P < 0.001; 48 h: t = 5.318, P < 0.0001; 72 h: t = 10.58, P < 0.0001). Additionally, the Transwell assay results demonstrated that the invasive ability of cells in the miR-30b-5p mimics group was significantly lower than that in the miR-NC group (t = 8.92, P < 0.0001), and the migration level was also significantly reduced (t = 23, P < 0.0001). Flow cytometry analysis revealed that the apoptosis level of MG63 cells in the miR-30b-5p mimics group was significantly increased compared to the miR-NC group (t = 14.06, P = 0.0001). Dual-luciferase reporter assay demonstrated that P4HA2 is a direct target molecule of miR-30b-5p. Furthermore, RT-PCR and Western blot experiments confirmed that the expression of P4HA2 in osteosarcoma cells was significantly down-regulated at both the mRNA and protein levels following miR-30b-5p transfection. In contrast, the expression of the pro-apoptotic protein BAX was elevated, while the expression of the anti-apoptotic protein BCL-2 was decreased. Conclusion: miR-30b-5p plays a role in inhibiting the proliferation, invasion, and migration of osteosarcoma cells while also inducing apoptosis by targeting and down-regulating P4HA2 expression. However, the experiments were constrained, as they were validated in a single cell line and lacked the most direct evidence obtained through “functional recovery assays. experiments”.