摘要
目的 观察细胞因子白细胞介素 1(IL 1) β、肿瘤坏死因子 (TNF)α和脂多糖 (LPS)是否诱导人脐静脉内皮细胞表达单核细胞趋化蛋白 1(MCP 1)mRNA及蛋白。方法 选取生长汇合的人脐静脉内皮细胞 ,在其培养基中分别加入终浓度为 2ng ml的IL 1β、2 0ng ml的TNFα和 10 0ng ml的LPS ,37℃共育 4h后 ,按照一步法提取其总RNA ,用γ 32 P标记的寡核苷酸探针进行dotblot分析 ,观察其MCP 1mRNA表达 ,另一部分培养 2 4h ,收集长满细胞的盖玻片 ,用免疫细胞化学法及图像分析系统检测细胞内的MCP 1蛋白质表达。结果 培养的人脐静脉内皮细胞能低水平地表达MCP 1mRNA和蛋白质。IL 1β、TNFα及LPS均能增加MCP 1mRNA的表达 (分别是对照组的 7.8倍、2 .6倍及 1.2倍 )。免疫细胞化学显示 ,各组细胞对MCP 1多克隆抗体均显示阳性反应。图像扫描显示 ,LPS组、TNFα组及IL 1β组细胞的平均吸光度 (A)值分别为 0 .0 78± 0 .113、0 .10 2± 0 .0 0 5和 0 .117± 0 .0 10 ,均明显高于对照组 (0 .0 5 1± 0 .0 0 4) ,差异有显著性 (F =193.2 5 ,P <0 .0 1)。结论 IL 1β和TNFα均能显著增强内皮细胞MCP 1mRNA和蛋白的表达 ,提示它们通过诱导内皮细胞产生MCP 1,吸引单核细胞迁入动脉内膜而参与动脉粥样硬化的发生和发展。
Objective To investigate whether the proinflammatory cytokines IL 1β, TNF α and lipopolysaccharide (LPS) enable to induce the expression of MCP 1 mRNA and protein in human umbilical vein endothelial cells (HUVECs). Methods After a four hour exposure to 2 ng/ml IL 1β, 20 ng/ml TNF α or 100 ng/ml LPS, total RNA of HUVECs was extracted by single step method. The expression of MCP 1 mRNA in HUVECs was examined by dot blot analysis using a probe of γ 32 P end labelled 35 mer oligonucleotide. Meanwhile, MCP 1 protein in the cytoplasm was detected by SABC immunostaining. Results Dot blot analysis showed that cultured HUVECs were able to express MCP 1 mRNA at a low level. Exposure to IL 1β, TNF α and LPS resulted in a 7.8 fold, 2.6 fold and 1.2 fold induction of MCP 1 mRNA expression in HUVECs, respectively. The cells on the coverglass in all groups revealed MCP 1 immunoreactivity. Densitometry scans showed that the mean absorbance ( A ) values of the cells in LPS, TNF α and IL 1β groups were 0.078±0.113, 0.102±0.005 and 0 117±0.010, respectively; whereas the absorbance values of the control group was 0.051±0.004. There were significant differences between all the experimental groups and the control group ( F =193.25, P <0.01). Conclusions IL 1β and TNF α induce a strong expression of MCP 1 mRNA and protein in HUVECs. Both cytokines may be involved in the atherogenesis by inducing the liberation of MCP 1 by endothelial cells and increasing the recruitment of monocytes into the subendothelial space.
出处
《中华病理学杂志》
CSCD
北大核心
2000年第1期46-48,共3页
Chinese Journal of Pathology
基金
国家自然科学基金资助项目!(39470289)
