摘要
目的:研究雌激素受体α(ER-α)在流体剪切力促进细胞外信号调节激酶5(ERK5)活化以及ER-α-ERK5信号通路在成骨细胞增殖中的具体分子机制。方法:给予成骨MC3T3-E1细胞孵育特异性ER-α抑制剂MPP(methyl-piperidinopyrazole,1μmol/L),孵育高选择性ERK5活性抑制剂XMD8-92(5μmol/L),和/或加载12 dyn/cm^2流体剪切力处理。采用MTT实验检测成骨细胞的增殖活性,采用蛋白免疫印记实验检测ER-α、P-ERK5、ERK5、Cyclin D1(细胞周期蛋白)和CDK4(细胞周期蛋白依赖性激酶)蛋白的表达水平。结果:生理强度(12 dyn/cm^2)的流体剪切力作用于成骨MC3T3-E1细胞60 min后能显著促进成骨细胞增殖,促进Cyclin D1和CDK4的表达;同时ER-α和P-ERK5的表达显著增加。成骨细胞孵育MPP抑制ER-α后,P-ERK5的表达显著下降,Cyclin D1和CDK4表达也显著降低。孵育XMD8-92抑制ERK5活性后,流体剪切力促进成骨细胞中Cyclin D1和CDK4表达水平显著下降。结论:流体剪切力通过ER-α-ERK5信号通路上调Cyclin D1和CDK4的表达水平,最终导致成骨MC3T3-E1细胞增殖。
Objective To study on the effect of estrogen receptor-alpha（ER-α） on the activation of extracellular signal-regulated kinase 5（ERK5） and the specific molecular mechanism of ER-α-ERK5 signaling pathway in the proliferation of osteoblasts promoted by fluid shear stress（FSS）. Methods MC3T3-E1 cells were treated with specific ER-α antagonist MPP（methylpiperidino-pyrazole, 1 μmol/L）, highly selective ERK5 inhibitor XMD8-92（5 μmol/L）, and or loading FSS of 12 dyn/cm^2. The proliferation activity of osteoblasts was detected by MTT assay, and the protein expression levels of ER-α, ERK5, phosphoERK5（P-ERK5）, Cyclin D1 and cyclin-dependent kinase 4（CDK4） were detected by Western blotting. Results Physiological FSS（12 dyn/cm^2） exposure for 60 min promoted the proliferation of MC3T3-E1 cells significantly and increased the expression of Cyclin D1 and CDK4. Meanwhile, the expression of ER-α and P-ERK5 increased significantly. After being treated with ER-α antagonist MPP, the expression of P-ERK5 decreased significantly, so were the expression of Cyclin D1 and CDK4. After inhibiting the ERK5 activity by incubating with XMD8-92, the expression levels of Cyclin D1 and CDK4 in osteoblasts promoted by FSS were significantly decreased. Conclusion FSS up-regulates the expression levels of Cyclin D1 and CDK4 via ER-α-ERK5 signaling pathway, resulting in the proliferation of MC3T3-E1 cells.
出处
《中国医学物理学杂志》
CSCD
2017年第3期291-296,共6页
Chinese Journal of Medical Physics
基金
国家自然科学基金(81450042
81071478
81672207)
甘肃省自然科学基金(1506RJZA240)
兰州市科技计划项目(2014-2-27)
