目的 研究冬凌草甲素诱导人黑色素瘤 A375-S2细胞凋亡的作用机制。方法 MTT法检测冬凌草甲素对 A375-S2细胞生长的影响及各种 Caspase抑制剂、佛波酯 (PMA)、PKC抑制剂 H- 7对冬凌草甲素作用的影响 ;DNA凝胶电泳法检测细胞凋亡 ;L DH法测定乳酸脱氢酶 (L DH)活力。结果 冬凌草甲素对 A375-S2细胞生长有显著抑制作用,并能显著诱导 A375-S2细胞发生凋亡,其作用呈明显的量效关系和时间依赖性。形态学观察可见凋亡小体的形成,琼脂糖凝胶电泳可见凋亡典型的 DNA梯带 ;Caspase家族抑制剂,Caspase- 9抑制剂能显著抑制冬凌草甲素诱导 A375-S2细胞的凋亡。大剂量 PMA(4 0 0 ng/m L)显著抑制 34.3μmol/L 冬凌草甲素诱导 A375-S2细胞凋亡,而小剂量 PMA(10 ng/m L)与冬凌草甲素有协同杀死细胞的作用,且 PKC抑制剂 H- 7也可下调这种凋亡作用。 Caspase- 3的抑制剂可抑制 Caspase- 3的活力升高。结论 适宜浓度的冬凌草甲素 (34.3μm ol/L )诱导A375-S2细胞凋亡,这种作用是通过线粒体调节 Caspase的激活来实现的。

Object To study the mechanisms of oridon in-induced A 3752S2 cell apoptosis. Methods MTT assay, morphological observation, agarose gelelect rophoresis, LDH release and Caspase inhibitors,PMA, PKC were used. Results Oridon in had significant apoptotic effect on A 3752S2 cells in a do se-andatime-dependen t manners. The marked morphological changes including condensed chromatin, nuclear fragmentation and apoptotic bodies; and DNA ladder in agarosegel were observed. The activity of Cas-pase-3 was increased after treatment with oridonin about 6 times as much as the control value, since high dose PMA inhibited PKC activity, low dose PMA activated PKC. In this study, 400 ng/m L PMA blocked 34.3 Lmol/L oridon in-induced A 375-S2 cell apoptosis and PKC inhibitor H-7 down-regulated the apoptotic effect, however, low dose PMA (10 ng/m L ) more sensitized A 375-S2 cell to oridonin. Conclus ion Ori-donin (34. 3 Lmol/L ) can induce A 375-S2 cell apoptosisvia mitochondria-regulated Caspase pathway acti-vation, PKC activation is involved in this mechanism.