Zhang Chun-Ling, Wu Li-Jun, Tashiro Shin-ichi, Onodera Satoshi, Ikejima Takashi
China-Japan Research Institute of Medical and Pharmaceutical Sciences, Shenyang Pharmaceutical University, China.
Biol Pharm Bull. 2004 Oct;27(10):1527-31. doi: 10.1248/bpb.27.1527.
Oridonin, an active component isolated from Rabdosia rubescences, has been reported to exhibit antitumor effects, but little is known about its molecular mechanisms of action. In this study, the growth-inhibitory activity of oridonin for L929 cells is in time- and dose-dependent manner. After treatment with various concentrations of oridonin for 12 h, the majority of L929 cells underwent apoptosis as measured by an LDH activity-based assay. Although apoptotic bodies were observed in oridonin-treated L929 cells, DNA fragmentation as a hallmark of apoptosis was not found. The pan-caspase inhibitor, z-VAD, and caspase-3 inhibitor, z-DEVD, sensitized L929 cells to oridonin, however, a PARP inhibitor (DPQ) effectively blocked oridonin-induced cell death. After 12 h treatment, PARP proenzyme was significantly cleaved. This result indicated that oridonin-induced L929 cell death required PARP degradation in a caspase-independent manner. In addition, an MEK/ERK inhibitor (PD98059) markedly blocked oridonin-induced cell death, whereas a p38 inhibitor (SB203580) and JNK inhibitor (SP600125) weakly protected the cells against death. Treatment with 41.2 microM oridonin for 12 h induced significant and persistent ERK activation and p38 inactivation in L929 cells without evident changes in the protein levels. The responsiveness of ERK and p38 to oridonin suggests the involvement of these kinases in this apoptotic process. Moreover, oridonin increased the ratio of Bax/Bcl-2 protein expression, whereas it had no effect on the expression of Bcl-xL. These results indicate that regulation of the Bcl-2 and MAPK families maybe the effector mechanisms of oridonin-induced L929 cell death, independent of the caspase pathway.
冬凌草甲素是从冬凌草中分离出的一种活性成分，据报道具有抗肿瘤作用，但其分子作用机制尚不清楚。在本研究中，冬凌草甲素对L929细胞的生长抑制活性呈时间和剂量依赖性。用不同浓度的冬凌草甲素处理12小时后，通过基于乳酸脱氢酶（LDH）活性的检测方法测定，大多数L929细胞发生凋亡。虽然在经冬凌草甲素处理的L929细胞中观察到凋亡小体，但未发现作为凋亡标志的DNA片段化。泛半胱天冬酶抑制剂z-VAD和半胱天冬酶-3抑制剂z-DEVD使L929细胞对冬凌草甲素敏感，然而，聚（ADP-核糖）聚合酶（PARP）抑制剂（DPQ）有效地阻断了冬凌草甲素诱导的细胞死亡。处理12小时后，PARP前体酶被显著切割。该结果表明，冬凌草甲素诱导的L929细胞死亡需要以不依赖半胱天冬酶的方式降解PARP。此外，MEK/ERK抑制剂（PD98059）显著阻断了冬凌草甲素诱导的细胞死亡，而p38抑制剂（SB203580）和JNK抑制剂（SP600125）对细胞死亡的保护作用较弱。用41.2微摩尔/升的冬凌草甲素处理12小时可诱导L929细胞中ERK的显著且持续激活以及p38的失活，而蛋白质水平无明显变化。ERK和p38对冬凌草甲素的反应性表明这些激酶参与了这一凋亡过程。此外，冬凌草甲素增加了Bax/Bcl-2蛋白表达的比率，但对Bcl-xL的表达没有影响。这些结果表明，Bcl-2和丝裂原活化蛋白激酶（MAPK）家族的调节可能是冬凌草甲素诱导L929细胞死亡的效应机制，且不依赖于半胱天冬酶途径。
