CAJ | 학술논문

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。
목적：탐토고린급고개유도혈관평활기세포발생개화적분자궤제。방법：이인혈관평활기세포（ VSMCs）위모형，채용불동개린농도적배양기여세포공배양12 d，장세포분위정상개、린조（대조조）、고린조、고개조화고린﹢고개조，분별채용면역인적（ western-blot）기술、역전록-취합매련반응（ RT-PCR）기술검측인혈관평활기세포핵심결합인자α1（Cbfα1）、골개소、α-평활기기동단백（α-SMA）단백질화mRNA적표체；고린﹢고개조건부육하적세포가입ZVAD-FMK간예후，재차채용western-blot기술검측Cbfα1、골개소단백질표체적정황。결과：시험조Cbfα1、골개소표체증강（ P=0.013；P=0.007），이기성섬유세포적특이성표지물α-SMA표체하강（P=0.009），대조조상술3개지표무명현변화（P=0.055、P=0.074、P=0.63）；가입ZVAD-FMK여고린﹢고개조세포공배양후，Cbfα1、골개소단백질적표체수도명현억제。결론：고린혹（화）고개통과사혈관평활기세포전화위성골양세포이도치개화적발생；ZVAD-FMK가이재일정정도상억제저일병리생리진정，제시조망화성골분화균삼여료개화과정。
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 phos-phate 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 RT-PCR were adopted to detect the protein and mRNA expression of Cbfα1,osteocalcin,α-SMA respec-tively. 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 indica-tors 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 obvi-ously. Conclusion:High concentration of phosphate or( and)calcium can induce calcification of hu-man 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 differ-entiation of VSMCs involve in the calcification process.