【摘要】 目的 观察铁缺乏（ID）对主动脉中层退行性变（AMD）小鼠主动脉壁、人主动脉血管平滑肌细胞（HASMCs）骨架结构的影响，探讨其可能机制。方法 （1）ApoE^-/-小鼠48只随机分为AMD组、ID组、ID+AMD组和对照组各12只。AMD组皮下泵注血管紧张素Ⅱ（AngⅡ）1μg/（kg·min）连续24 d制备AMD模型，同时饲喂正常铁含量饲料；ID组饲喂低铁含量饲料构建ID模型；ID+AMD组AMD模型制备同AMD组，ID模型制备同ID组；对照组饲喂正常铁含量饲料。造模第4天测量4组小鼠收缩压（SBP）;造模第24天处死小鼠，应用血清铁检测试剂盒检测铁含量，采用免疫组织化学SP法检测主动脉转铁蛋白（TF）、转铁蛋白受体1（TFR1）、磷酸化肌球蛋白轻链（p-MLC）、细胞分裂周期因子42（Cdc42）、Ras相关的C3肉毒素底物1（Rac1）表达。（2）取对数生长期HASMCs分为AngⅡ组（0.1μmol/L AngⅡ）、ID细胞组（20μmol/L去铁胺）、ID+AngⅡ组（0.1μmol/L AngⅡ+20μmol/L去铁胺）和对照细胞组（正常培养基）。处理48 h取4组细胞，采用Western blot法检测TF、TFR1、p-MLC、Cdc42、Rac1蛋白相对表达量。结果 （1）造模第4天AMD组、ID+AMD组小鼠SBP[（171.17±7.73）、（168.00±3.41）mmHg]高于ID组[（115.00±2.00）mmHg]和对照组[（105.17±10.57）mmHg]（P<0.05）,对照组与ID组，AMD组与ID+AMD组SBP比较差异均无统计学意义（P>0.05）。（2）4组小鼠血清铁含量及主动脉TF、TFR1、p-MLC、Cdc42、Rac1表达量比较差异均有统计学意义（P<0.05）。ID组小鼠血清铁含量[（3.67±1.37）μmol/L]及主动脉TF（3.17±0.75）、p-MLC（3.83±1.17）、Cdc42（4.00±1.27）表达量均低于AMD组[（7.67±1.37）μmol/L、8.50±0.55、8.17±2.23、8.83±1.94]和对照组[（9.83±1.72）μmol/L、10.33±1.86、9.83±1.72、9.67±1.75]（P<0.05）,TFR1（9.67±1.86）及Rac1（6.17±2.04）表达量均高于AMD组（6.00±1.79、3.83±1.84）和对照组（4.50±1.23、2.50±1.05）（P<0.05）;AMD组小鼠血清铁含量及主动脉TF、p-MLC、Cdc42表达量均高于ID+AMD组[（3.17±1.60）μmol/L、2.83±0.75、3.67±1.21、3.83±1.17)（P<0.05）、TFR1、Rac1表达量均低于ID+AMD组（10.33±1.86、8.67±1.97）（P<0.05）;AMD组与对照组，ID组与ID+AMD组血清铁含量及主动脉TF、TFR1、p-MLC、Cdc42、Rac1表达量比较差异均无统计学意义（P>0.05）。（3）4组细胞TF、TFR1、p-MLC、Cdc42、Rac1蛋白相对表达量比较差异有统计学意义（P<0.05）。ID细胞组TF（0.427±0.014）、p-MLC（0.223±0.002）、Cdc42（0.525±0.006）蛋白相对表达量均低于AngⅡ组（1.264±0.023、0.529±0.004、0.957±0.014）和对照细胞组（1.000±0.007、1.000±0.006、1.000±0.169）,TFR1（1.446±0.054）、Rac1（1.417±0.073）蛋白相对表达量均高于AngⅡ组（0.606±0.146、1.163±0.016）和对照细胞组（1.000±0.033、1.000±0.059）（P<0.05）;AngⅡ组TF、p-MLC及Cdc42蛋白相对表达量均高于ID+AngⅡ组（0.578±0.027、0.354±0.017、0.644±0.010）（P<0.05）,TFR1及Rac1蛋白相对表达量均低于ID+AngⅡ组（1.168±0.037、1.593±0.070）（P<0.05）;ID细胞组TF、TFR1、p-MLC、Cdc42、Rac1蛋白相对表达量与ID+AngⅡ组比较差异均无统计学意义（P>0.05）。结论 泵注AngⅡ可促进小鼠AMD形成，增高血压；ID可进一步破坏AMD小鼠的血管平滑肌细胞骨架结构，加重AMD损伤，其机制可能是通过Cdc42/Rac1通路诱导血管平滑肌细胞骨架结构破坏，促进AMD形成。更多还原

【Abstract】 Objective To observe the influence of iron deficiency（ID） on the aortic wall of mice with aortic medial degeneration（AMD） and on the cytoskeletal structure of human aortic smooth muscle cells（HASMCs）, and to explore the potential mechanism. Methods（1） Forty-eight ApoE^-/- mice were randomly divided into AMD group, ID group, ID+AMD group and control group, with 12 mice in each group. In AMD group, the mice were pumped subcutaneously 1 μg/（kg·min） of angiotensin Ⅱ（AngⅡ） continuously for 24 days to establish AMD models, and meanwhile they were fed with a normal-iron diet. In ID group, the mice were fed with a low-iron diet to establish ID models. In ID+AMD group, the AMD models and ID models were established with the methods in AMD group and ID group, respectively. The mice in control group were fed with a normal-iron diet. The systolic blood pressure（SBP） was detected in four groups by day 4 after modeling. The mice were sacrificed by day 24 after modeling. The iron levels were detected by the serum iron detection kit, the expressions of transferrin（TF）, transferrin receptor 1（TFR1）, phosphorylated myosin light chain（p-MLC）, recombinant cell division cycle protein 42（Cdc42） and Rac-related C3 botulinum toxin substrate 1（Rac1） in the aortic tissues were detected by immunohistochemical SP method.（2） HASMCs in logarithmic growth phase were divided into AngⅡ group（0.1 μmol/L AngⅡ）, ID HASMCs group（20 μmol/L deferoxamine）, ID+AngⅡ group（0.1 μmol/L AngⅡ +20 μmol/L deferoxamine）, and HASMCs control group（normal culture medium）. The relative expressions of TF, TFR, p-MLC, Cdc42 and Rac1 proteins were detected by Western blot after 48-h treatment. Results（1） The SBP was higher in AMD group [（171.17±7.73） mmHg] and ID+AMD group [（168.00±3.41） mmHg] than that in ID group [（115.00±2.00） mmHg] and control group [（105.17±10.57） mmHg] by day 4 after modeling（P<0.05）, and showed no significant difference between control group and ID group, and between AMD group and ID+AMD group（P>0.05）.（2） There were significant differences in the serum iron content and the expressions of TF, TFR1, p-MLC, Cdc42 and Rac1 among four groups（P<0.05）. The serum iron content and expressions of TF, p-MLC and Cdc42 were lower in ID group [（3.67±1.37） μmol/L, 3.17±0.75, 3.83±1.17, 4.00±1.27] than those in AMD group [（7.67±1.37） μmol/L, 8.50±0.55, 8.17±2.23, 8.83±1.94] and control group [（9.83±1.72） μmol/L, 10.33±1.86, 9.83±1.72, 9.67±1.75]（P<0.05）, and the expressions of TFR1 and Rac1 were higher in ID group（9.67±1.86, 6.17±2.04） than those in AMD group（6.00±1.79, 3.83±1.84） and control group（4.50±1.23, 2.50±1.05）（P<0.05）. The serum iron content and expressions of TF, p-MLC and Cdc42 were higher in AMD group than those in ID+AMD group [（3.17±1.60） μmol/L, 2.83±0.75, 3.67±1.21, 3.83±1.17]（P<0.05）, and the expressions of TFR1 and Rac1 were lower in AMD group than those in ID+AMD group（10.33±1.86, 8.67±1.97）（P<0.05）. There were no significant differences in the serum iron content and expressions of TF, TFR1, p-MLC, Cdc42 and Rac1 between AMD group and control group, and between ID group and ID+AMD group（P>0.05）.（3） The relative expressions of TF, TFR, p-MLC, Cdc42 and Rac1 showed significant differences among four groups（P<0.05）. The realtive expressions of TF, p-MLC and Cdc42 proteins were lower in ID HASMCs group（0.427±0.014, 0.223±0.002, 0.525±0.006） than those in AngⅡ group（1.264±0.023, 0.529±0.004, 0.957±0.014） and HASMCs control group（1.000±0.007, 1.000±0.006, 1.000±0.169）, and the relative expressions of TFR1 and Rac1 proteins were higher in ID HASMCs group（1.446±0.054, 1.417±0.073） than those in AngⅡ group（0.606±0.146, 1.163±0.016） and HASMCs control group（1.000±0.033, 1.000±0.059）（P<0.05）. The relative expressions of TF, p-MLC and Cdc42 were higher in AngⅡ group than those in ID+AngⅡ group（0.578±0.027, 0.354±0.017, 0.644±0.010）（P<0.05）, and the relative expressions of TFR1 and Rac1 were lower in AngⅡ group than those in ID+AngⅡ group（1.168±0.037, 1.593±0.070）（P<0.05）. There were no significant differences in the relative expressions of TF, TFR1, p-MLC, Cdc42 and Rac1 between ID HASMCs group and ID+AngⅡ group（P>0.05）. Conclusion Pumping AngⅡ can promote the formation of AMD and increase the blood pressure of mice, while ID can further destroy the cytoskeletal structure of vascular smooth muscle cells and aggravate the AMD injury, probably by inducing the destruction of cytoskeletoal structure of vascular smooth muscle cells via Cdc42/Rac1 pathway and promoting the formation of AMD.更多还原