胱抑素C对兔血管成形术后再狭窄中血管外膜的影响

Effect of cystatin C on adventitia in restenosis after rabbit aorta angioplasty

目的 探讨兔腹主动脉血管成形术后胱抑素C（CysC）水平变化对兔腹主动脉再狭窄中血管外膜的影响和机制.方法 将48只新西兰大白兔数字抽签随机分为损伤组（球囊扩张损伤腹主动脉）、治疗组（CysC单克隆抗体治疗）、对照组（穿刺股动脉和留置鞘管,不进行球囊扩张损伤腹主动脉和CysC单克隆抗体注射干预）,每组16只.分别于术前和术后8h、1d、1周、3周、6周取外周静脉血,检测血清中CysC水平.术后6周取腹主动脉行血管形态计量分析和血管外膜细胞计数;同时行平滑肌肌动蛋白（α-SM-actin）、增值细胞核抗原（PCNA）免疫组织化学染色,观察各项免疫指标在血管外膜上的表达情况;并且计数血管外膜PCNA阳性细胞核数量,计算PCNA增殖指数.以血管重构指数、血管外弹力板围绕面积（EELA）、内弹力板围绕面积（IEIA）变化评价血管重构情况,以剩余狭窄率、血管腔面积变化衡量血管狭窄情况.结果 损伤组兔球囊损伤8h后血清CysC水平即开始升高,术后1周达到高峰,持续增高5周,术后3周和6周均高于治疗组和对照组（均P＜0.05）.治疗组和对照组CysC水平无上述变化规律.损伤组PCNA阳性细胞数个高于治疗组和对照组（均P＜0.05）.损伤组血管腔面积、血管EELA、IELA较对照组缩小（P＜0.05）,血管重塑指数为0.871,剩余狭窄率为33.1％（P＜0.05）.给予CysC单克隆抗体干预后,治疗组CysC水平较损伤组下降,血管腔面积、血管的EELA、IELA缩小程度较损伤组减轻（P＜0.05）,血管重塑指数为0.748,剩余狭窄率为19.7％（P＜0.05）.相关分析显示,CysC水平与血管腔面积、新生内膜面积、IEIA、EELA及PCNA阳性细胞数呈正相关（r值分别为0.812、0.797、0.876、0.932、0.822,P＜0.01）.结论 CysC水平与动脉狭窄程度呈正相关,CysC诱导了血管外膜的成纤维细胞激活、增殖、表型转化及迁移,加速了动脉粥样硬化及狭窄的过程,是发生腹主动脉狭窄的独立危险因素.

Objective To investigate the effect of cystatin C （Cys C） on adventitia in rabbit abdominal aorta restenosis after angioplasty and its mechanism.Methods 48 New Zealand white rabbits were randomly divided into injury group （receiving balloon dilation of abdominal aorta）,the treatment group （taking Cys C monoclonal antibody therapy） and the control group （receiving femoral artery puncture and catheter sheath without balloon dilation and intervention of Cys C monoclonal antibody injection）,and each group had 16 rabbits.Peripheral vein blood was drawn to measure the serum level of cystatin C before and 8 h,1 day,1 week,3 weeks,6 weeks after the operation in all rabbits.After 6 weeks of operation,the abdominal aorta were taken and stained with HE.Vascular morphometry analysis and adventitial cell count were conducted.Smooth muscle actin （SM-actin） and proliferating cell nuclear antigen （PCNA） expressions in the adventitia were observed by immunohistochemical staining.The number of PCNA positive cell in the adventitia was counted and the PCNA proliferation index was calculated.The vascular remodeling index,vascular external elastic lamina area （EELA）,internal elastic lamina area （IEIA） were used to evaluate the vascular remodeling and the residual stenosis and vascular cavity area was used to measure the vascular stenosis.Results Plasma Cys C level began to rise at 8h after operation and reached the peak at 1 week after operation,and continuously increased for 5 weeks in injury group,and reached to respectively at 3 weeks and 6 weeks after operation.The Cys C levels were significantly higher in injury group than in the treatment and control groups at different time points （all P〈0.05）.There were no significant differences in Cys C levels at different time points between the treatment group and the control group.The injury group showed that the number of PCNA positive cells was higher in injury group than in treatment and control groups,both P〈0.05）.Compared with the control group,the vascular luminal area,EELA and IELA were significantly increased （all P〈0.05）.After treated with the monoclonal antibody Cys C intervention,the treatment group showed that lumen area,vascular EELA,IELA was significantly decreased （P〈0.05）,and the vascular remodeling index and residual stenosis rate were decreased as compared with the injury group （0.871 vs.0.784,33.1% vs.19.7 %,both P〈0.05）.Correlation analysis showed that Cystatin C level was positively correlated with the vascular lumen area,neointimal area,internal elastic lamina area,external elastic lamina area and the number of PCNA positive cells （r=0.812,0.797,0.876,0.932 and 0.822 respectively,all P〈0.01）.Conclusions Plasma Cys C level is increased in rabbit after abdominal aorta balloon injury and has a positive correlation with the severity of arterial stenosis.High Cys C level can induce adventitial fibroblast activation,proliferation,phenotype transformation and migration,and accelerate the processes of atherosclerosis and stenosis.Cys C level is the independent risk factor for abdominal aortic stenosis.