摘要
目的 Runx2基因在糖尿病合并骨质疏松患者成骨细胞分化,软骨细胞成熟中的促进作用.方法 构建pLko.1-shRunx2-puro以及pWPI-Runx2-绿色荧光蛋白（GFP）质粒,通过序列测定和Western blot验证这两个质粒的正确性.通过酶联免疫吸附试验（ELISA）法检测Runx2基因沉默和Runx2过表达后,肿瘤坏死因子（TNF）-α和血管内皮生长因子（VEGF）水平变化,探讨Runx2在成骨细胞分化和软骨细胞成熟中的作用.结果 pLko.1-shRunx2-puro以及pWPI-Runx2-GFP质粒构建成功.真核表达后Runx2基因正常组,沉默组和过表达组VEGF的表达水平分别为373.78、10.94、4 161.54 μg/L;而3组TNF-α表达水平分别为62.04、0.22、615.08 ng/L.两者表达水平随着随着Runx2基因的沉默和过表达而降低和升高.结论 在糖尿病合并骨质疏松患者破骨细胞修复过程中,Wnt/β-环连蛋白（β-catenin）信号通路通过激活Runx2表达调节骨质形成.
Objective To investigate the promoting role of Runx2 gene in the osteoblasts and chondrocytes maturation of diabetes mellitus patients with osteoporosis.Methods Constructed pLko.1-shRunx2-puro and pWPI-Runx2-green fluorescent protein（GFP） plasmids and verify the correctness of the two plasmids by sequencing and Western blotting.Enzyme linked immunosorbent assay （ELISA） was used to detect the levels of tumor necrosis factor （TNF）-α and vascular endothelial growth factor （VEGF） after overexpression and silencing of Runx2 gene, in order to explore the role of Runx2 in osteoblast differentiation and maturation of chondrocytes.Results pLko.1-shRunx2-puro and pWPI-Runx2-GFP plasmid was successfully constructed.After eukaryotic expression, the VEGF expression levels in the Runx2 normal expression group, the Runx2 silencing expression group and the Runx2 overexpression group were 373.78, 10.94, 4 161.54 μg/L, respectively;and the TNF-α expression levels were 62.04, 0.22,615.08 ng/L, respectively.The expression levels of VEGF and TNF-o expression levels lowered and raised as the decrease and increase of Runx2 gene.Conclusion In osteoclasts repair process of diabetes mellitus patients with osteoporosis, Wnt/β-catenin signaling pathway regulated bone formation by activating the expression of Runx2 gene.
出处
《中华实验外科杂志》
CAS
CSCD
北大核心
2015年第11期2789-2791,共3页
Chinese Journal of Experimental Surgery
