miR-21对骨质疏松小鼠骨髓基质细胞增殖的影响

目的探讨miR-21对骨质疏松小鼠骨髓基质细胞(BMSCs)增殖的影响。方法采用双侧卵巢切除法构建骨质疏松小鼠模型(VOX),分离、培养、纯化小鼠BMSCs并采用si PORT Neo FX转染pre-miR-21、pre-miR-negative control(pre-miR-NC)、antmiR-21、ant-miR-negative control(ant-miR-NC)并进行RT-PCR验证,MTT法检测小鼠BMSCs增殖情况、茜素红与碱性磷酸酶染色法检测小鼠BMSCs成骨能力、Western-blotting检测细胞增殖、成骨分化相关蛋白水平。结果骨质疏松症小鼠BMSCs中miR-21相对表达水平低于Ctrl组(P0.05),OVX-pre-miR-21组BMSCs中miR-21相对表达水平、细胞增殖、PCNA水平、Ki-67水平、ALP染色程度、ALP活性、茜素红染色程度、Runx2水平、Osterix水平均高于OVX-pre-miR-NC组(P0.05),OVX-premiR-NC组BMSCs中miR-21相对表达水平、细胞增殖、PCNA水平、Ki-67水平、ALP染色程度、ALP活性、茜素红染色程度、Runx2水平、Osterix水平均显著低于Ctrl-pre-miR-NC(P0.05); OVX-ant-miR-21组BMSCs中miR-21相对表达水平、细胞增殖、PCNA水平、Ki-67水平、ALP染色程度、ALP活性、茜素红染色程度、Runx2水平、Osterix水平均显著低于OVX-ant-miR-NC组(P0.05),OVX-ant-miR-NC组BMSCs中miR-21相对表达水平、细胞增殖、PCNA水平、Ki-67水平、ALP染色程度、ALP活性、茜素红染色程度、Runx2水平、Osterix水平均显著低于Ctrl-ant-miR-NC(P0.05)。结论提高miR-21表达水平可促进骨质疏松小鼠BMSCs增殖能力与成骨分化能力。

Effect of miR-21 on the proliferation of bone marrow stromal cells in osteoporosis mice

Objective To explore the effect of miR-21 on the proliferation of bone marrow stromal cells (BMSCs) in osteoporosis mice. Methods The osteoporosis mice model was constructed with bilateral ovariectomy (OVX) method. BMSCs were separated, cultured, and purified. They were transfected with pre-miR-21, pre-miR-negative control (pre-miR-NC), ant-miR-21, and ant-miR-negative control (ant-miR-NC), respectively, with siPORT NeoFX. RT-PCR verification was conducted. The proliferation of BMSCs was detected with MTT. The osteogenic ability of BMSCs was detected with Alizarin red staining and alkaline phosphatase staining. The levels of cells proliferation and osteogenic differentiation-related proteins were detected with Western blotting. Results The relative expression level of miR-21 in BMSCs of osteoporosis mice was lower than that in Ctrl group (P<0.05). The relative expression levels of miR-21, cell proliferation, PCNA, Ki67, ALP staining, ALP activity, Alizarin red staining, Runx2, and Osterix in OVX- pre-miR-21 group were higher than those in OVX-pre-miR-NC group (P<0.05), and were significantly lower in OVX-pre-miR-NC group than in Ctrl-pre-miR-NC (P<0.05). The relative expression levels of miR-21, cell proliferation, PCNA, Ki67, ALP staining, ALP activity, Alizarin red staining, Runx2, and Osterix in OVX-ant-miR-21 group were significantly lower than those in OVX-ant-miR-NC group (P<0.05), and were significantly lower in OVX-ant-miR-NC group than in Ctrl-ant-miR-NC (P<0.05). Conclusion Increasing the expression level of miR-21 promotes BMSCs proliferation and osteogenic differentiation in osteoporosis mice.