尿毒症血清诱导人脐动脉平滑肌细胞成骨样转化和钙化的研究  

Uremic serum induces osteogenic transition and calcification of human umbilical artery smooth muscle cells

作　　者：孙明妹[1] 郭永平[1] 张敏芳[1] 顾乐怡[1] 戴慧莉[1] 钱家麒[1] 

机构地区：[1]上海交通大学医学院附属仁济医院肾内科,200001

出　　处：《中华肾脏病杂志》2008年第4期265-270,共6页Chinese Journal of Nephrology

基　　金：国家自然科学基金（30370653）;上海市教委基金（04BC35）

摘　　要：目的研究尿毒症血清对原代培养人脐动脉平滑肌细胞（HUASMC）成骨样转化和细胞外基质钙沉积的影响。方法收集40例健康人、40例非透析尿毒症患者和45例透析尿毒症患者血清，检测有关生化指标。组织植块法原代培养HUASMC，分别给予健康人血清（对照组）、透析（透析组）和非透析（非透析组）尿毒症血清孵育。茜素红S钙沉积染色及甲氧．酚酞络合酮法检测细胞层钙盐含量。实时定量PCR、荧光定量法及WeStem印迹分别检测骨特异性碱性磷酸酶（BAP）、骨桥蛋白（OPN）和骨形成蛋白2（BMP2）基因以及蛋白表达。结果血清生化检测结果显示尿毒症血清较健康对照存在高磷、高脂、高全段甲状旁腺激素（iPTH）、高C反应蛋白（CRP）、自细胞介素6（IL-6）和低胎球蛋白A（P〈0．05），而透析组血清的三酰甘油、CRP、IL-6较非透析组血清高（P〈0．05）。与健康对照组相比，透析组和非透析组尿毒症血清均使HUASMC钙盐沉积更显著，BAP、OPN以及BMP2的表达上调（P〈0．05），且透析组血清作用更明显。结论尿毒症血清体外能够诱导HUASMC成骨样转化和钙化，提示可能和尿毒症相关的血管钙化有关。透析并发的微炎性反应状态可能加速了这一过程。Objective To study the effect of uremic serum on the calcification and osteogenic transition of cultured human umbilical artery smooth muscle ceils （HUASMC）. Methods Sera from 40 healthy controls （control group）, 40 nondialysis uremic patients （nondialysis group） and 45 uremic patients on dialysis （dialysis group） were detected for biochemical indexes concerned and used to treat the cultured HUASMC. Alizarin red S stain was applied to examined calcium deposition in the cell layer. Calcium concentration was determined calorimetrically by the O- cresolphthalein complexone method, and corrected by total cell proteins. The mRNA expression of bone specific alkaline phosphatase （BAP）, osteopontin （OPN） and bone morphogenetic protein 2 （BMP2） was estimated by realtime PCR. OPN and BMP2 protein expression was assessed by Western blotting and＇fluorometry method was used to check the BAP concentration. Results Serum biochemical detection revealed that both uremic groups had higher levels of phosphate, triglyseride, iPTH, C-reactive protein （CRP） and IL-6, and lower level of fetuin-A than healthy control （P〈0.05）. Furthermore, dialysis serum had higher levels of triglyseride, CRP and IL-6 than nondialysis serum（P〈0.05）. Compared with control group, both uremic sera induced more cell layer calcium deposition and higher mRNA and protein expression levels of BMP2, BAP and OPN （P〈 0.05 ）. Higher mRNA and protein expression levels of above factors were found in dialysis group as compared to nondialysis group （P〈0.05）. Conclusions Uremic serum can induce HUASMC calcification and osteogenic transition in vitro, which may be one of the mechanisms involved in vascular calcification of ESRD patients. Microinflammatory state may promote the osteogenic transition and vascular calcification in dialysis patients.

关 键 词：尿毒症 血清 肌细胞 平滑肌 脐动脉 成骨细胞样转化 血管钙化 

分 类 号：R282.71[医药卫生—中药学] R692.5[医药卫生—中医学]