细胞间黏附分子1在红霉素抗炎机制中的作用.docVIP

细胞间黏附分子1在红霉素抗炎机制中的作用 作者：李梅华 钟小宁 柳广南 何志义 朱敏 周刚 【摘要】 目的 从分子水平上探讨红霉素(EM)的抗炎作用机制。方法 体外培养人支气管上皮细胞(16HBE)，将细胞随机分为8组，先加入EM干预，后加入肿瘤坏死因子α(TNFα)刺激。分组如下：空白对照组;TNFα(20 ng/ml,16 h)组;EM(0.3 μg/ml，24 h)+TNFα(20 ng/ml，16 h)组;EM(3 μg/ml，24 h)+TNFα(20 ng/ml，16 h)组;EM(30 μg/ml，24 h)+TNFα(20 ng/ml，16 h)组;EM(0.3 μg/ml，48 h)+TNFα(20 ng/ml，16 h)组;EM(3 μg/ml，48 h)+TNFα(20 ng/ml，16 h)组;EM(30 μg/ml，48 h)+TNFα(20 ng/ml，16 h)。然后收集各组细胞分别提取RNA，应用逆转录聚合酶链反应(RTPCR)方法检测细胞间黏附分子1(ICAM1) mRNA的表达。因ICAM1 RTPCR产物分子大小约为700 bp，与原设计产物分子大小490 bp不相符，进一步作基因克隆测序了解基因之间是否具有同源性。结果 ICAM1的RTPCR产物即为目的基因：ICAM1基因。TNFα刺激人支气管上皮细胞(16HBE)后，细胞ICAM1mRNA表达增高。先加入不同浓度及作用时间的EM，后加入TNFα刺激人支气管上皮细胞(16HBE)，各组细胞ICAM1mRNA表达均降低，且提示有浓度与时间依赖性。结论 TNFα能激活人支气管上皮细胞使其ICAM1基因表达增高，促进炎症的发生 发展 。EM能抑制人支气管上皮细胞ICAM1基因表达，可能是EM的抗炎作用机制之一。 【关键词】 人支气管上皮细胞;细胞间黏附分子1;肿瘤坏死因子α;红霉素 　　【Abstract】 Objective To investigate the antiinflammatory molecular mechanism of erythromycin (EM) in order to find a new way to prevent and treat chronic obstructive pulmonary disease (COPD). Methods Human bronchial epithelial cell line 16HBE was cultured with 10% serum DMEM (high glucose). The cells were randomly divided into Eight groups: blank control group, TNFα (20 ng/ml,16 h), EM(0.3 μg/ml，24 h)+TNFα (20 ng/ml，16 h), EM(3 μg/ml，24 h)+TNFα (20 ng/ml，16 h), EM(30 μg/ml，24 h)+TNFα(20 ng/ml，16 h), EM(0.3 μg/ml，48 h)+TNFα (20 ng/ml，16 h), EM(3 μg/ml，48 h)+TNFα(20 ng/ml，16 h), and EM(30 μg/ml，48 h)+TNFα (20 ng/ml，16 h). Then total cellular RNA was extracted to detect the expression of ICAM1 mRNA by RTPCR. The molecular size of ICAM1 RTPCR production was not correspondent for what it was designed，therefore，clone sequencing was applied to confirm it. Results ICAM1 RTPCR production was ICAM1 mRNA. ICAM1mRNA was increased by the addition of TNFα,which was inhibited by the prEIncubation with EM in dosedependent and timedependent fasion. Conclusions ICAM1mRNA can be increased by the stimulation of TNFα, which indicate that EM modify inflammation presumably by suppre