Rong Shu, Zhao Xuezhi, Jin Xiucai, Zhang Zheng, Chen Lei, Zhu Yuxian, Yuan Weijie
Department of Nephrology, Shanghai Jiaotong University Affiliated First People's Hospital, Shanghai, China.
Cell Physiol Biochem. 2014;34(6):2049-60. doi: 10.1159/000366400. Epub 2014 Nov 28.
Vascular calcification (VC), in which vascular smooth muscle cells (VSMCs) undergo a phenotypic transformation into osteoblast-like cells, is one of the emergent risk factors for the accelerated atherosclerosis process characteristic of chronic kidney disease (CKD). Phosphate is an important regulator of VC.
The expression of different smooth muscle cell or osteogenesis markers in response to high concentrations of phosphate or exogenous bone morphogenetic protein 2 (BMP-2) was examined by qRT-PCR and western blotting in rat VSMCs. Osteocalcin secretion was measured by radioimmunoassay. Differentiation and calcification of VSMCs were examined by alkaline phosphatase (ALP) activity assay and Alizarin staining. Short hairpin RNA-mediated silencing of β-catenin was performed to examine the involvement of Wnt/β-catenin signaling in VSMC calcification and osteoblastic differentiation induced by high phosphate or BMP-2. Apoptosis was determined by TUNEL assay and immunofluorescence imaging.
BMP-2 serum levels were significantly higher in CKD patients than in controls. High phosphate concentrations and BMP-2 induced VSMC apoptosis and upregulated the expression of β-catenin, Msx2, Runx2 and the phosphate cotransporter Pit1, whereas a BMP-2 neutralization antibody reversed these effects. Knockdown of β-catenin abolished the effect of high phosphate and BMP-2 on VSMC apoptosis and calcification.
BMP-2 plays a crucial role in calcium deposition in VSMCs and VC in CKD patients via a mechanism involving the Wnt/β-catenin pathway.
血管钙化（VC）是慢性肾脏病（CKD）加速动脉粥样硬化进程的新兴危险因素之一，在此过程中血管平滑肌细胞（VSMC）会发生表型转变，成为成骨样细胞。磷酸盐是VC的重要调节因子。
通过qRT-PCR和蛋白质印迹法检测大鼠VSMC中不同平滑肌细胞或成骨标志物在高浓度磷酸盐或外源性骨形态发生蛋白2（BMP-2）作用下的表达。采用放射免疫分析法测定骨钙素分泌。通过碱性磷酸酶（ALP）活性测定和茜素染色检测VSMC的分化和钙化情况。利用短发夹RNA介导的β-连环蛋白沉默来研究Wnt/β-连环蛋白信号通路在高磷酸盐或BMP-2诱导的VSMC钙化和成骨细胞分化中的作用。通过TUNEL检测和免疫荧光成像确定细胞凋亡情况。
CKD患者的BMP-2血清水平显著高于对照组。高磷酸盐浓度和BMP-2诱导VSMC凋亡，并上调β-连环蛋白、Msx2、Runx2和磷酸盐共转运体Pit1的表达，而BMP-2中和抗体可逆转这些作用。敲低β-连环蛋白可消除高磷酸盐和BMP-2对VSMC凋亡和钙化的影响。
BMP-2通过涉及Wnt/β-连环蛋白途径的机制在CKD患者的VSMC钙沉积和VC中起关键作用。
