贵州医科大学学报

2015, v.40;No.177(06) 603-607

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高磷、高钙诱导人血管平滑肌细胞钙化的研究
Influence of High Phosphate and High Calcium on Calcification of Human Vascular Smooth Muscle Cells

傅碧玲;邹世海;彭鑫;翁俊雄;邓柳霞;
FU Biling;ZOU Shihai;PENG Xin;WENG Junxiong;DENG Liuxia.;Department of Nephrology,the People's Hospital of Baoan District;

摘要(Abstract):

目的:探讨高磷及高钙诱导血管平滑肌细胞发生钙化的分子机制。方法:以人血管平滑肌细胞(VSMCs)为模型,采用不同钙磷浓度的培养基与细胞共培养12 d,将细胞分为正常钙、磷组(对照组)、高磷组、高钙组和高磷+高钙组,分别采用免疫印迹(western-blot)技术、逆转录-聚合酶链反应(RT-PCR)技术检测人血管平滑肌细胞核心结合因子α1(Cbfα1)、骨钙素、α-平滑肌肌动蛋白(α-SMA)蛋白质和mRNA的表达;高磷+高钙条件孵育下的细胞加入ZVAD-FMK干预后,再次采用western-blot技术检测Cbfα1、骨钙素蛋白质表达的情况。结果:试验组Cbfα1、骨钙素表达增强(P=0.013;P=0.007),而肌成纤维细胞的特异性标志物α-SMA表达下降(P=0.009),对照组上述3个指标无明显变化(P=0.055、P=0.074、P=0.63);加入ZVAD-FMK与高磷+高钙组细胞共培养后,Cbfα1、骨钙素蛋白质的表达受到明显抑制。结论:高磷或(和)高钙通过使血管平滑肌细胞转化为成骨样细胞而导致钙化的发生;ZVAD-FMK可以在一定程度上抑制这一病理生理进程,提示凋亡和成骨分化均参与了钙化过程。
Objective: To investigate the effect and molecular mechanism of high phosphate and high calcium on calcification of human vascular smooth muscle cells( VSMCs). Methods: The human VSMCs were co-cultured with culture medium of different dose of phosphate and calcium in vitro for 12 days and according to dosage of calcium and phosphate the cells were divided into 5 groups as follows:normal calcium and phosphate group( control group,P 1. 4 mmol / L,Ca 2. 0 mmol / L),high phosphate group( P 2. 0 mmol / L,Ca2. 0 mmol / L),high calcium group( P 1. 4 mmol / L,Ca 2. 8 mmol /L),high calcium and high phosphate group( P 2. 0mmol / L,Ca 2. 8mmol / L). Western-blot and RTPCR were adopted to detect the protein and mRNA expression of Cbfα1,osteocalcin,α-SMA respectively. In high calcium and high phosphate group,the nonspecific inhibitor ZVAD-FMK of caspase was added in. After this intervention,western-blot was adopted again to detect protein expression of Cbfα1and osteocalcin. Results: Compared with control group,the expression of Cbfα1 and osteocalcin increased in all experimental groups while the expression of α-SMA,which was the specific marker of myofibroblast,decreased( P < 0. 05). There were no obvious changes in the above-mentioned 3 indicators in the control group( P > 0. 05). After ZVAD-FMK was added in and co-cultured with cells of high calcium and high phosphate group,the protein expression of Cbfα1and osteocalcin were inhibited obviously. Conclusion: High concentration of phosphate or( and) calcium can induce calcification of human VSMCs through turning VSMCs into osteoblast like cell( OLCs). ZVAD-FMK can inhibit this pathophysiological process to a certain extent,which reveals that both apoptosis and osteogenic differentiation of VSMCs involve in the calcification process.

关键词(KeyWords): 肌细胞,平滑肌;血管;高磷;高钙;核心结合因子α1(Cbfα1);骨钙素;α-平滑肌肌动蛋白(α-SMA)
myocytes,smooth muscle;blood vessels;high phosphate;high calcium;vascular calcification;core binding factor α1;osteocalcin;α-smooth muscle actin

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作者(Author): 傅碧玲;邹世海;彭鑫;翁俊雄;邓柳霞;
FU Biling;ZOU Shihai;PENG Xin;WENG Junxiong;DENG Liuxia.;Department of Nephrology,the People's Hospital of Baoan District;

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DOI: 10.19367/j.cnki.1000-2707.2015.06.014

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