昆明山海棠提取物对血管平滑肌细胞增殖及凋亡的影响

Effect of Extract from Tripterygium Hypoglaucum on Proliferation and Apoptosis of Vascular Smooth Muscle Cells

目的探讨昆明山海棠提取物 (THW 4 )对离体血管平滑肌细胞 (VSMC)增殖及凋亡的影响。方法培养家兔主动脉平滑肌细胞 ,加入不同浓度的THW 4进行处理 ,用MTT法测定细胞生长曲线 ,以AnnexinV FITC/PI双标记、透射电镜、原位末端标记法检测细胞凋亡。结果THW 4可以明显抑制VSMC增殖 ,呈剂量和时间依赖性 ,作用 4 8h的半有效抑制浓度 (IC50 )为 15 .6 μg/L ;VSMC在含 10～ 10 0 μg/LTHW 4的培养液中孵育 5 6h ,可检测到细胞早期凋亡 ,且凋亡率随THW 4浓度增加而升高 ;延长孵育时间至 72h ,透射电镜下可观察到典型的凋亡细胞形态学改变 ,原位末端标记法显示DNA片段化增多 ,有核碎裂、固缩等改变 ;细胞周期时相分布显示THW 4主要阻滞VSMC于G2 /M期。结论THW 4能有效抑制体外培养的VSMC增殖及诱导其凋亡 ,提示THW

Objective To study the effect of THW 4, an extract from Trypterygium hypoglaucum on proliferation and apoptosis of vascular smooth muscle cells (VSMC). Methods VSMC derived from rabbit aorta were cultured in vitro and different concentrations of THW 4 were added in experimental groups. Cell proliferation was evaluated by MTT and apoptosis by transmission electron microscopy (TEM), TUNEL assay and Annexin V FITC/PI double labelled assay. Results The inhibitory effects of THW 4 on proliferation of VSMC displayed dose time dependently, with the IC 50 value of 15.6 μg/L at 48 hrs. Incubated with THW 4 (10-100 μg/L) for 56 hrs, VSMC mainly appeared early stage apoptosis and the percentage of apoptosis was found to raise along with the increase of the THW 4 concentration. Typical images of apoptosis could be observed under TEM and TUNEL assay showed increase of DNA segments with karyorrhexis and pyknosis after THW 4 treatment for 72 hrs. Analysis of cell cycle indicated the THW 4 mainly lead to the blockage of VSMC in G 2/M stage. ConclusionTHW 4 could inhibit the proliferation and induce the apoptosis of VSMC in vitro, suggesting that THW 4 is a potential agent for prevention of restenosis following angioplasty.