中文摘要：

目的探讨CDl37信号是否通过核因子-KB（NF—KB）调控小鼠血管平滑肌细胞（VSMC）活化T细胞核因子cl（mclear factor of activated T cells cl，NFATcl）表达。方法采用组织块贴壁法对正常C57BL／6J小鼠的VSMC进行原代培养。以每孔2×105^个VSMC接种于6孔板中培养，待其贴壁后，饥饿24h，用含10％FBS＋肿瘤坏死因子（TNF-a，终浓度10ng／m1）的DMEM培养基分别培养细胞0、4、8、12．24、36、48h后，收获细胞用于CDl37mRNA和蛋白的检测，选取CDl37表达最多的时间点进行下一步实验。细胞实验分为3组，即对照组[含10％FBS、TNF-α（终浓度10ng／m1）、DMEM培养基共同干预]、刺激组[对照组干预基础上加CDl37mAb激动剂（终浓度10μg/m1）]和抑制组[对照组干预基础上加PDTC（终浓度30μmol／L），然后加CDl37mAb激动剂（终浓度10μ/m1）]。加入不同干预因素后继续培养，分别于90rain时收集核内外磷酸化（P）-NF—KBp65、核因子KB抑制蛋白（IKB-α），24h收集细胞总RNA和NFATcl蛋白备检。逆转录定量聚合酶链反应检测CDl37和NFATclmRNA表达水平。流式细胞术检测CDl37膜蛋白表达水平。蛋白印迹法检测IKB—α、NF．KBp65、p-NF—KBp65和NFATcl蛋白表达水平。结果（1）TNF-α诱导VSMC表达CDl37的结果：TNF．d刺激VSMC4、8、12、24、36、48h后，细胞CDl37mRNA表达均上调。RT—PCR结果示，以24h组CDl37mRNA升高最为显著，以未刺激（即0h）组的细胞为对照进行比较，差异有统计学意义（40．00±2．83比1，P〈0．05）。流式细胞术检测亦显示24h组CDl37膜蛋白升高最为显著。（2）各组VSMC中p-NF-KBp65蛋白表达水平的检测结果：刺激组VSMC胞浆中P-NF—KBp65蛋白表达水平高于对照组（8．34±0．28比1，P〈0．05），而IkB一仅蛋白表达水平低于对照组（1比2．70±0．28，P〈0．05）。抑制组VSMC胞浆中p-NF—KBp65蛋白表达水平低于刺激组（1．15±0．14比8．3

英文摘要：

Objective To observe whether CD137 signaling could affect the nuclear factor of activated T cells cl （ NFATcl ） expression through nuclear factor-KB （ NF-KB ） pathway in mice aortic vascular smooth muscle cells （VSMCs）. Methods Adherence methods for tissues explants were used for primary culture of mouse aortic VSMCs. The mRNA expression of CD137 and NFATcl was detected by real- time quantitative PCR （RT-qPCR）. The VSMCs protein expression of IKB-α, NF-KB 1365, phospo-p65 and NFATcl was determined by Western blot. The level of CD137 was measured by Flow Cytometry （FCM）. Results （ 1 ）The mRNA and protein expression of CD137 in VSMCs was significantly upregulated at 24 h after co-culture with TNF-α （10 ng/ml, all P 〈 0.05 ）. （2）Compared with the control group, the level of p-NF-KB p65 in cytoplasm and nucleus was significantly increased （ 8.34 ± 0. 28 vs. 1, P 〈 0. 05, and 2. 64 ± 0. 42 vs. 1, P 〈 0. 05 ） while the level of IKB-α was reduced （ 1 vs. 2.70 ± 0. 28, P 〈 0. 05 ） after co-treatment with agonist-CD137 mAb, above effects were partly blocked by adding specific NF-KB inhibitor PDTC （30 μmol/L： 1. 15 ±0. 14 vs. 8.34±0. 28 ,P 〈0.05 ,and 2. 09 ±0. 12 vs. 2. 64 ±0.42, P 〈0.05, and 1.78±0.74 vs. 1, P〈0.05）. （3）The mRNA （2.07 ±0.09 vs. 1, P〈0.05） and protein （1.75± 0. 07 vs. 1, P 〈 0.05 ） expression of NFATcl was significantly upregulated by agonist CD137mAb compared with the control group, and these effects could be reversed by PDTC （ 1.15 ± 0. 07 vs. 2. 07 ± 0. 09, P 〈 0. 05, and 0. 90 ± 0. 11 vs. 1.75 ± 0. 07, P 〈 0.05 ）. Conclnsion CD137 signaling could affect the NFATclexpression in VSMCs through NF-kappaB pathway.