中文摘要：

目的 探讨CD137分子信号是否通过活化T细胞核因子c1（NFATc1）调控血管平滑肌细胞（VSMC）表型转变.方法 将小鼠VSMC采用组织块贴块法原代培养,细胞分为对照组、CD137刺激组和CD137抑制组,以CD137L重组蛋白及抗CD137阻断型抗体分别激动和抑制CD137信号.在小干扰RNA试验中,将小鼠VSMC分为对照序列组与si-NFATc1组并分别转染对照序列与si-NFATc1序列.采用荧光定量PCR（qRT-PCR）及Westem blot检测VSMC中NFATc1和表型蛋白、肌动蛋白α（α-SMA）、平滑肌肌球蛋白重链（SM-MHC）、波形蛋白的表达;免疫荧光和Western blot检测核蛋白中NFATc1活化水平.采用Transwell迁移实验检测穿膜细胞数目,比较细胞迁移能力.结果 与对照组比较,CD137刺激组中VSMC中NFATc1（5.07 ±0.36比1.00±0.00）及波形蛋白（3.23±0.27比1.00±0.00）表达较高（P均＜0.05）,而α-SMA（0.73±0.15比1.00±0.00）、SM-MHC（0.45±0.05比1.00±0.00）表达较低（P均＜0.05）.与CD137刺激组比较,CD137抑制组NFATc1（1.56±0.27比5.07±0.36）和波形蛋白（1.21±0.17比3.23±0.27）表达较低（P均＜0.05）,而α-SMA（2.01±0.43比0.73 ±0.15）和SM-MHC（2.85±0.32比0.45±0.05）表达较高（P均＜0.05）.与对照序列组比较,siNFATc1组VSMC的SM-MHC、α-SMA的mRNA及蛋白水平较高,而波形蛋白表达较低（P均＜0.05）.细胞迁移实验显示,CD137L刺激组细胞迁移数量高于对照组（3.85±0.31比1.00±0.00,P＜0.05）,而抑制NFATc1后迁移细胞数与对照组比较,差异无统计学意义（P＞0.05）.结论 CD137信号通过NFATc1参与VSMC表型调控.

英文摘要：

Objective To investigate whether CD137 induces primary vascular muscle cells （VSMCs） phenotype transformation through activating nuclear factor of activated T-cells 1 （NFATc1）signaling.Methods VSMCs were obtained from aorta of C57BL/6J mice （8 weeks,male） through tissuepiece inoculating.Cells were divided into control group,CD137 agonist group （treated with CD137L recombinant protein） and anti-CD137 group （treated with anti-CD137 antibody）.In si-RNA transfection assay,cells were divided into si-control group and si-NFATc1 group which were transfected with control or si-NFATc1 sequence respectively.The levels of NFATc1 and other phenotype related protein such as α-smooth muscle actin （α-SMA）,smooth muscle myosin heavy chain （SM-MHC）,vimentin were detected by Q-PCR and Western blot.Nuclear protein expression and activity of NFATc1 were detected by immunofluorescence and Western blot.Transwell assay was performed to measure the migration of VSMCs.Results According to Western blot,the expression of NFATc1 and vimentin was significantly upregulated （5.07 ± 0.36 vs.1.00 ± 0.00,P ＜ 0.05;3.23 ± 0.27 vs.1.00 ± 0.00,P ＜ 0.05） while α-SMA and SM-MHC expressions was significantly downregulated （0.73 ± 0.15 vs.1.00 ± 0.00,P ＜ 0.05;0.45 ± 0.05 vs.1.00 ± 0.00,P ＜ 0.05） in CD137 agonist group compare to control group.Compared with CD137 agonist group,the expression of NFATc1 and vimentin was significantly downregulated （1.56 ± 0.27 vs.5.07 ± 0.36,P ＜ 0.05;1.21 ± 0.17 vs.3.23 ± 0.27,P ＜ 0.05）,but the levels of α-SMA and SM-MHC were significantly upregulated （2.01 ± 0.43 vs.0.73 ± 0.15,P ＜ 0.05;2.85 ± 0.32 vs.0.45 ± 0.05,P ＜ 0.05） in anti-CD137 group.Compared with si-con group,the expression of SM-MHC and α-SMA was significantly upregulated while the expression of vimentin was significantly downregulated in si-NFATc1 group.Transwell assay results demonstrated that migration cell numbers was significantly higher in CD137L group compared with control gr