中文摘要：

为探讨E1A激活基因阻遏子（cellular repressor of E1A-stimulated genes，CREG）对人血管平滑肌细胞（vascular smooth muscl ecells，VSMCs）凋亡的影响及调控机制，应用正、反义重组逆转录病毒表达载体pLNCX，（＋）／CREG及pLXSN（-）／CREG制备稳定感染人胸廓内动脉平滑肌细胞克隆株（human internal thoracic artery-Shenyang，HITASY）细胞模型，观察CREG蛋白过表达及表达抑制对平滑肌细胞凋亡的影响。荧光显微镜下观察DAPI染色后凋亡细胞核形态，AnnexinV／PI流式细胞术检测细胞凋亡率，RT-PCR技术分析凋亡相关基因caspase-9mRNA的表达，蛋白质印迹法分析p38丝裂原活化蛋白激酶（p38 mitogen-activated protein kinase，p38MAPK）、磷酸化p38MAPK（phosphorylated p38 mitogen-activated proteinkinase，P-p38 MAPK）的表达变化。研究结果显示，CREG蛋白过表达明显抑制血清饥饿诱导的HITASY细胞凋亡的发生；同时细胞中p38MAPK、P-p38MAPK的表达增加。相反，抑制CREG蛋白表达则引起正常血清培养状态下VSMCs的自发凋亡明显增加，同时细胞内p38MAPK、P-p38MAPK表达显著下降。进一步研究发现，预先应用特异性抑制剂SB203580阻断p38MAPK信号转导通路后，CREG蛋白过表达引起的细胞凋亡抑制作用被明显减弱，血清饥饿后CREG蛋白过表达引起的HITASY细胞凋亡现象明显增加。上述结果提示，CREG蛋白过表达可以抑制体外培养的VSMCs凋亡，p38MAPK信号转导通路可能介导CREG蛋白对VSMCs凋亡的抑制作用。

英文摘要：

To investigate the effects and molecular mechanisms of the cellular repressor of E 1 A-stimulated genes （CREG） on the apoptosis of vascular smooth muscle cells （VSMCs）, the human internal thoracic artery-Shenyang （HITASY） cells were infected with sense-CREG [pLNCX2（＋）/CREG] and antisense-CREG [pLXSN（-）/CREG] retrovirus respectively. The stably infected cells were obtained by screening the G418-resistant clones. DAPI nuclei staining and Annexin V/PI FASC assay indicated that over-expression of CREG in HITASY cells infected with pLNCX2（＋）/CREG inhibited VSMC apoptosis induced by serum deprivation, accompanied with decreased expression of caspase-9 mRNA detected by RT-PCR. Furthermore, Western blot analysis showed that p38 mitogen activated protein kinase （p38 MAPK） expression and activation were significantly enhanced in HITASY cells infected with pLNCX2（＋）/CREG. The inhibition of CREG protein expression in cells infected with pLXSN（-）/CREG promoted the VSMC spontaneous apoptosis, as well as down-regulated p38 MAPK expression and activation, when cells were cultured with 10% fetal bovine serum （FBS） mediums. These results implicate that the CREG protein has the ability to regulate VSMC apoptosis in which the activation of p38 MAPK is possibly involved. To further identify the role of p38 MAPK in VSMC apoptosis, SB203580, a specific inhibitor of p38 MAPK, was used to inhibit p38 MAPK activity. When p38 MAPK signaling pathway was blocked, the effects that over-expression of CREG protein inhibited VSMC apoptosis disappeared. Taken together, the present work indicates that over-expression of CREG protein inhibits VSMC apoptosis, and this inhibitory effect is partly mediated by p38 MAPK signaling pathway.