摘要
为了解脂质过氧化损伤能否诱导内皮细胞表达巨噬细胞炎性蛋白 1α ,用不同浓度 (1、5和 10 μmol L)的联胺刺激培养人脐静脉内皮细胞脂质过氧化损伤 ,用硫代巴比妥酸荧光微量测定法测定样品中的丙二醛含量 ;用一步法提取RNA ,反转录多聚酶链反应检测巨噬细胞炎性蛋白 1αmRNA ;用免疫细胞化学法检测巨噬细胞炎性蛋白 1α蛋白的表达。结果发现 ,随联胺浓度增高 ,各组丙二醛含量依次增高 (P <0 .0 5 ) ,分别为对照组的 2 .0、5 .3和8.6倍 (F =138.6 4,P <0 .0 1)。正常内皮细胞表达较低水平的巨噬细胞炎性蛋白 1αmRNA ,加联胺后各组巨噬细胞炎性蛋白 1αmRNA表达明显增加 ,分别是对照组的 4、6和 3.5倍。免疫细胞化学显示 ,巨噬细胞炎性蛋白 1α蛋白为棕黄色颗粒 ,主要分布于胞浆的核周区。图像分析结果显示各组细胞平均吸光度值增加 (P <0 .0 5 ) ,分别是对照组的 1.4、1.9和 1.3倍 (F =6 8.6 0 ,P <0 .0 1)。以上结果提示 ,联胺能引起内皮细胞脂质过氧化损伤 ,并诱导其产生巨噬细胞炎性蛋白 1α,吸引单核细胞粘附于内皮 ,并向内皮下间隙迁移 ,在动脉粥样硬化发生过程中可能起重要作用。
Aim To investigate whether lipid peroxidation injury to cultured endothelial cells (ECs) induces the expression of macrophage inflammatory protein-1α(MIP-1α). Methods The cultured human umbilical vein ECs were randomly divided into the A,B and C groups,cultured in the media containing 1 μmol/L,5 μmol/L and 10 μmol/L diamide respectively and the control group cultured in the media without diamide.After exposure of ECs to diamide for 4 h,the lipopeeroxide in the ECs was determined by thiobarbiuric acid flurophotometry;toral cellular RNA was extracted from the culrured ECS by the single step method, amd the expression of MIP-1α mRNA was determined by RT-PCR;the MIP-1α protein in ECs was determined by immunocytochemistry. Results The malomdialdehyde content in ECs im group A,B and C were 2.0-fold and 8.6-fold as much as that there was significant difference between experiment group and control group (F=138.64,P<0.01), and Q test also revealed statistical difference among the experiment groups (P<0.05). RT-PCR showed that the MIP-1α mRNA expression in ECs im group A, B and C were 4-fold, 6-fold and 3.5-fold as much as that in the control group. Immunocytochemistry assay showed that MIP-1α mRNA Protein was granular brown substance which located mainly in the cytoplasm around the nuclei.Diamide induced stronger expression of MIP-1α protein in the experimental groups. Image analysis showed that the expression of MIP-1α protein in ECs was 1.9-fold B,1.4-fold in group A and 1.3-fold in group C as mush as that in the control group.The analysis of variance proved that there were significant differences between the experimental groups and the control group (F=68.60,P<0.01),and there were also significant differences among group A,B and C which were proved by Q test (P<0.05). Conclusion Lipid peroxidation injury may play an important role in recuitment of monocytes into intima in atherogenesis through inducing ECs to produce increased MIP-1α.
出处
《中国动脉硬化杂志》
CAS
CSCD
2001年第2期96-99,共4页
Chinese Journal of Arteriosclerosis
基金
国家自然科学基金资助项目 (项目编号 39730 2 2 0 )
关键词
脂质过氧化
血管内皮
巨噬细胞炎性蛋白1Α
逆转录
多聚酶链反应
动脉粥样硬化
Lipid Peroxidation
Endothelium, Vascular
Macrophage Inflammatory Protein-1α
Transcription, Genetic
Polymerase Chain Reaction
Atherosclerosis
