丹参酮ⅡA对兔动脉粥样硬化病灶凋亡相关基因表达的影响

Effect of Tanshinone ⅡA on expression of apoptosis related gene in rabbit atherosclerosis

目的差异筛选丹参酮ⅡA诱导血管平滑肌细胞(VSMC)凋亡相关基因葡萄糖调节蛋白78(GRP78),观察其在动脉粥样硬化(AS)病灶中的表达情况,探讨丹参酮ⅡA抗AS的可能机制。方法建立同型半胱氨酸(HCY)诱导的兔VSMC增殖模型,与不同浓度丹参酮ⅡA共同培养48 h后,MTT法检测细胞增殖能力;RT-PCR筛选差异基因片段,克隆、鉴定、测序,组织原位杂交观察该基因在正常及AS病变组织中的表达水平。结果不同浓度的丹参酮ⅡA都可抑制HCY所诱导的VSMC增殖;筛选出全长648 bp的基因片段与兔GRP78基因高度同源,组织原位杂交显示该基因在AS病灶VSMC胞质中高表达。结论 GRP78基因参与AS病变形成,上调其表达可能是丹参酮ⅡA诱导VSMC凋亡作用机制之一。

Objective To select the differentially expressed vascular smooth.muscle cells （VSMCs） apoptosis gene induced by Tan-shinone Ⅱ A, and observe the expression of GRP78 in atheroselerotic tissue, investigate the mechanism of anti-atherosclerosis of Tanshinone Ⅱ A. Methods Proliferation of VSMCs were established by using DL-homocysteine in vitro. The VSMCs was incubated with various con-centrations of Tanshinone Ⅱ A for 48 hours. The apoptosis of cells was detected by MTF. GRP78 gene expression was measured by PCR, re-combinant plasmid were constructed and transfered into E. coli. , positive clonings were selected and confirmed by restriction endonucleas a- nalysis, and sequenced. In situ hybridization was used to observe GRP78 expression in atherosclerotic tissue in rabbit model of atheroselero-sis. Results Tanshinone Ⅱ A promoted VSMCs apoptosis. Furthermore, the 648 bp fragment was similar to the gene of GRIt78 from VSMCs and the level of expression was significantly higher in high cholesterol group than that of control group. Conclusions The GRP78 may play a key role in atheroselerotic lesions. The mechanism of anti-atherosclerosis of Tanshinone Ⅱ A might be partly due to promoting apoptosis of VSMCs.