膦甲酸钠干预高磷诱导的血管平滑肌细胞钙化的研究

目的　观察不同浓度的膦甲酸钠 (PFA)对高磷诱导的牛主动脉血管平滑肌细胞钙沉积和骨钙素 (OC)表达的影响。方法　用不同磷浓度 (正常磷Pi1.5mmol·L-1、高磷Pi2 0mmol·L-1)及含不同浓度膦甲酸钠的培养液 ,体外培养牛主动脉平滑肌细胞 ,观察血管平滑肌细胞钙沉积及骨钙素表达。用甲ο 酚酞络合酮方法测定钙含量 ,BCA法测定蛋白含量。培养上清液中骨钙素浓度用放射免疫法测定 ,用蛋白含量标化钙含量、骨钙素的浓度 ,RT PCR观察骨钙素mR NA的表达。结果　①高磷组较正常磷组平滑肌细胞钙沉积增加 :细胞培养 6d后 ,高磷组 (77 187± 11 6 92 )mg·g-1Pro ,正常磷组 (2 5 76 8± 1 75 0 )mg·g-1Pro ,P <0 0 1;②膦甲酸钠能有效地抑制钙沉积 :培养 6d ,高磷 +PFA 1 0mmol·L-1组 (37 72 9± 5 899)mg·g-1Pro ,与高磷未干预组相比 ,P <0 0 1;③高磷组骨钙素表达明显增高。高磷组与正常磷组相比 ,培养上清液中骨钙素水平 :(1 5 0 3× 10 -2 ±2 6 0 1× 10 -3 )mg·g-1Pro对 (2 981× 10 -3 ± 8 382× 10 -4)mg·g-1Pro ,P <0 0 1;平滑肌细胞骨钙素mRNA表达 (OC/GAPDH) :1 886± 0 16 5对 0 75 2± 0 0 5 2 1,P <0 0 1;④膦甲酸钠能有效地抑制骨钙素的表达。高磷 +PFA 1 0mmol·L-1组与高磷组?

Effect of phosphonoformic acid on vascular smooth muscle cell calcification induced by elevated phosphate

AIM To observe the effect of phosphonoformic acid (PFA) in different concentration on vascular calcification induced by elevated phosphate. METHODS The cell culture of bovine aortic smooth muscle were performed. Calcium deposition in different phosphate and PFA concentrations were determined by O cresolphthalein complexone method, and osteocalcin expression by radioimmunity and RT PCR. RESULTS Compared to Pi 1 5 mmol·L -1 group,bovine smooth muscle cells (BSMC) cultured in medium containing Pi 2 0 mmol·L -1 phosphate level increased calcium deposition 〔On day 6, (77 187±11 692) mg·g -1 Pro versus (25 768±1 750) mg·g -1 Pro, P <0 01〕. Elevated phosphate treatment of BSMCs also enhanced osteocalcin expression 〔after 72 hour, Pi 2 0 mmol·L -1 group versus Pi 1 5 mmol·L -1 group: (1 503×10 -2 ±2 601×10 -3 ) mg·g -1 Pro versus (2 981×10 -3 ±8 382×10 -4 ) mg·g -1 Pro, P <0 01〕. PFA decreased ciacium deposition and osteocalcin expression statistically: ciacium deposition, (Pi 2 0+PFA1 0) mmol·L -1 group 〔(37 729±5 899) mg·g -1 Pro〕 versus Pi 2 0 mmol·L -1 group, P <0 01; osteocalcin: (Pi 2 0+PFA1 0) mmol·L -1 group 〔(4 529×10 -3 ±1 250×10 -3 ) mg·g -1 Pro〕 versus Pi 2 0 mmol·L -1 group, P <0 01, the same as osteocacin mRNA expression. CONCLUSION Elevated phosphate may directly stimulate calcium deposition and osteocalcin expression induced by elevated phosphate in vascular smooth muscle cell culture, and PFA had a negative regulation about this. PFA may be a new agent to treat vascular calcification induced by elevated phosphate.