Krüppel样因子4基因修饰的间充质干细胞抑制颈动脉再狭窄的研究  

Inhibition of carotid restenosis by Krüppel like factor 4 gene-modified bone marrow mesenchymal stem cells

作　　者：艾合顺 孙忠东 任冠桦 夏斌 刘天宇 裴宝成 程显峰 Ai Heshun;Sun Zhongdong;Ren Guanhua;Xia Bin;Liu Tianyu;Pei Baocheng;Cheng Xianfeng(School of Clinical Medicine,Shandong Second Medical University,Weifang 261053,China;Department of Cardiac Surgery,the First Affiliated Hospital of Shandong Second Medical University,Weifang 261000,China)

机构地区：[1]山东第二医科大学临床医学院,潍坊261053 [2]山东第二医科大学第一附属医院心外科,潍坊261000

出　　处：《中华实验外科杂志》2024年第5期929-931,共3页Chinese Journal of Experimental Surgery

基　　金：潍坊市科学发展计划项目(医学类)(2021YX003)。

摘　　要：目的:构建携带含Krüppel样因子4(KLF4)基因的重组慢病毒载体,建立KLF4基因过表达的骨髓间充质干细胞株(BMSCs),并观察颈动脉损伤后血管增生的变化。方法:以慢病毒为载体介导KLF4基因转染BMSCs。选用10周龄SD大鼠24只,随机分为3组,分别为假手术组、模型组、KLF4组(注入KLF4修饰的BMSCs细胞)。采用球囊导管构建大鼠颈动脉内皮损伤的动物模型,于14 d后股动脉放血处死大鼠,并采用苏木精-伊红(HE)染色法观察各组颈总动脉血管壁厚度变化及形态学变化;采用Griess法检测各组实验大鼠血清中一氧化氮(NO)的含量;采用蛋白质印迹法(Western blot)检测大鼠颈动脉内皮型一氧化氮合酶(eNOS)、KLF4蛋白的表达水平。组间比较采用单因素方差分析。结果:模型组内皮细胞增生高于假手术组(1.68±0.29比0.01±0.01,F=244.345,P<0.05),差异有统计学意义,而KLF4组内皮细胞增生低于模型组(0.14±0.01比1.68±0.29,F=244.345,P<0.05)。模型组血清中NO浓度明显低于假手术组(7.22±1.40比21.29±1.83,F=171.000,P<0.05),差异有统计学意义,而KLF4组NO浓度明显高于模型组(16.34±1.33比7.22±1.40,F=171.000,P<0.05),差异有统计学意义。Western blot显示,KLF4/β-肌动蛋白(β-actin)在假手术组、模型组和KLF4组中的比值分别为0.564±0.015、0.626±0.023、0.916±0.042,差异有统计学意义(F=125.121,P<0.05);eNOS/β-actin在假手术组、模型组和KLF4组中的比值分别为0.852±0.043、0.383±0.017、0.707±0.014,差异有统计学意义(F=223.879,P<0.05)。结论:KLF4可正性调节eNOS的表达,从而增加血管中NO的浓度,通过抑制血管内皮细胞的迁移和增殖来抑制颈动脉损伤造成的血管狭窄。Objective To construct a recombinant lentiviral vector carrying the Krüppel like factor 4(KLF4)gene,establish bone marrow mesenchymal stem cells(BMSCs)overexpressing the KLF4 gene,and observe the changes in vascular proliferation after carotid artery injury.Methods The lentivirus was used as a vector to mediate KLF4 gene transfection into BMSCs.Totally,2410-week-old SD rats were randomly divided into three groups:sham surgery group,model group,and KLF4 group(injected with KLF4-modified BMSCs).An animal model of carotid artery endothelial injury in rats was constructed using a balloon catheter.After 14 days,the animals were euthanized by femoral artery bleeding,and hematoxylin and eosin(HE)staining was used to observe the changes in the thickness and morphology of the common carotid artery vessel walls in each group.The Griess method was used to detect the content of nitric oxide(NO)in the serum of experimental animals in each group.Western blotting was used to detect the expression levels of endothelial nitric oxide synthase(eNOS)and KLF4 proteins in rat carotid arteries.One-way analysis of variance was used for comparison between groups.Results The endothelial cell hyperplasia in the model group was severer than that in the sham operation group(1.68±0.29 vs.0.01±0.01,F=244.345,P<0.05),and the difference was statistically significant,while the endothelial cell hyperplasia in the KLF4 group was milder than that in the model group(0.14±0.01 vs.1.68±0.29,F=244.345,P<0.05).The serum NO concentration in the model group was significantly lower than that in the sham group(7.22±1.40 vs.21.29±1.83,F=171.000,P<0.05),and that in the KLF4 group was significantly higher than that in the model group(16.34±1.33 vs.7.22±1.40,F=171.000,P<0.05).Western blotting showed that the ratios of KLF4/β-actin in the sham operation group,model group and KLF4 group were 0.564±0.015,0.626±0.023 and 0.916±0.042,respectively,and the differences were statistically significant(F=125.121,P<0.05),and the ratios of eNOS/β-actin in the s

关 键 词：Krüppel样因子4 基因治疗 慢病毒 内皮型一氧化氮合酶 颈动脉 

分 类 号：R654.3[医药卫生—外科学]