DMP1、E11和Sclerostin在高磷诱导钙化的血管平滑肌细胞中的表达

Expression of DMP1,E11 and Sclerostin on High Phosphorus Induced Calcification of Vascular Smooth Muscle Cells

目的：观察DMP1、E11和Sclerostin在高磷诱导钙化的血管平滑肌细胞（VSMC）中的表达,探讨VSMC钙化的可能机制。方法：组织块贴壁法原代培养VSMC,倒置相差显微镜观察细胞形态及α-SMA免疫组化鉴定;3~8代VSMC用于实验,分为空白对照组、正常磷组和高磷组（Pi分别为0.9、1.3和2.6 mmol/L）。培养第6天,茜素红染色检测钙沉积,RT-PCR或Real-time PCR检测DMP1、E11、Sclerostin mRNA表达,Western Blot检测E11蛋白表达。结果：与空白对照组和正常磷组相比,高磷组VSMC钙盐沉积明显增多;DMP1、E11和Sclerostin mRNA表达明显上调（P〈0.05）;E11蛋白表达明显增加（P〈0.05）。结论：骨细胞标志性蛋白DMP1、E11和Sclerostin在高磷诱导钙化的VSMC中表达增加,提示血管钙化过程中VSMC最终向骨细胞样细胞转分化。

Objective： To observe the expression of DMP1,E11 and Sclerostin involved inhigh phosphorus induced calcification of vascular smooth muscle cells（ VSMC）. Methods： The explants derived from thoracic aorta were used for primary culture of VSMC and identification of cells was carried on by morphological identification and direct immunohistochemical staining of α- SMA,Passage 3 to 8 of VSMC were used for experiments. VSMC were divided in three groups： normal control group（ Pi 0. 9 mmol / L）,normal phosphorus group（ Pi 1. 3 mmol / L） and high phosphorus group（ Pi 2. 6 mmol / L）. Calcium deposition was visualized by Alizarin stain method at day 6. DMP1,E11 and Sclerostin mRNA levels were determined by RT- PCR or Real- Time PCR. E11 expression was semi- quantified by Western Blot. Results： Compared with normal control group and normal phosphorus group,the calcium deposition was increased in high phosphorus group. DMP1,E11 and Sclerostin mRNA expression was significantly increased in high phosphorus group（ P 〈0. 05）. The expression of E11 in high phosphorus group was significantly increased compared with normal control groupand normal phosphorus group（ P〈 0. 05）. Conclusion： The increase of expression of DMP1,E11 and Sclerostin in calcified VSMCshowsvascularcalcificationinvolves a VSMCtransition to an osteocyte- like phenotype.