葛根素调节血管平滑肌细胞凋亡相关基因在斑块组织中表达的研究

 目的差异筛选葛根素促进VSMC凋亡相关基因，观察其在动脉粥样硬化组织中的表达，探讨葛根素防治动脉粥样硬化的可能机制。方法葛根素诱导培养的兔血管平滑肌细胞凋亡，提取总RNA,荧光标记差异显示反转录-PCR法筛选差异基因片段，克隆测序及同源性分析，Northern印记杂交鉴定，组织原位杂交观察该基因在兔斑块组织中的表达水平。结果 差异筛选出全长733 bp的基因片断与兔葡萄糖调节蛋白94基因高度同源，Northern印记杂交显示该基因表达量随着葛根素浓度而增加；兔斑块组织原位杂交显示该基因片段在动脉粥样硬化斑块组织中表达水平增高。结论①证实葡萄糖调节蛋白94基因在葛根素促进增殖血管平滑肌细胞凋亡过程中表达增高,可能是葛根素作用靶点之一；②证实该基因片段在动脉斑块细胞质中表达增高，提示如能上调该基因促进需管平滑肌细胞凋亡，将有可能对动脉粥样硬化治疗产生一定作用。

Study on expression of vascular smooth muscle cell apoptosis gene in atherosclerosis tissue induced by puerarin

OBJECTIVE To investigate the puerarin induced VSMC apoptosis gene and the molecular mechanism of this effect on anti-atherosclerosis.METHODS The RNA of both the control VSMCs and those stimulated by puerarin were isolated and analyzed by differential display RT-PCR (DDRT-PCR).These differentially expressed fragments were recovered,cloned,sequenced and compared with GeneBank databases.One EST (expressional Sequence Tag) was selected to further analyzed by northern hybridization and tissue situ hybridization to observe the gene expression of the GRP94 in VSMC of apoptosis and AS.RESULTS One EST,733 bp,was isolated from VSMC stimulated by puerarin,similar to the gene of glucose regulated protein GRP94.The expression of the EST in VSMC was correlated to the concentration of puerarin and the level of expression was significantly higher in high cholesterol group than in control group.CONCLUSION The GRP94 gene expressed in both stressed model of puerarin-induced apoptosis and high cholesterol induced AS.Puerarin possesses the function of promoting apoptosis of VSMC.Its molecular mechanism may be related to promoting the gene expression of the GRP94 of VSMC.