蛇床子素对L02细胞的毒性作用及其机制研究

Study on Toxic Effects and Mechanism of Osthole on L02 Cells

关键词:  蛇床子素  L02细胞  毒性损伤  细胞凋亡

Key words:  osthole  L02 cells  toxicity  cell apoptosis

摘要:

目的 探究蛇床子素（osthole，Ost）对L02细胞的毒性损伤和作用机制，为中药蛇床子及其制剂的安全合理应用提供实验依据。方法 以不同浓度Ost作用于L02细胞，MTT法检测细胞活性；乳酸脱氢酶（LDH）试剂盒测定细胞LDH释放率；Hoechst 33342染色法检测细胞核形态；Annexin V/PI双染法检测细胞凋亡；Western blot检测Bcl-2、Bax、pro-caspase-3、cleaved-caspase-3（p17）、p-Histon H3（Ser10）的表达。结果 L02细胞在Ost作用下活性下降，LDH释放率提高，且呈浓度依赖；Hoechst 33342染色荧光下可见细胞核皱缩碎裂；AnnexinV/PI双染法结果表明凋亡率随浓度提高而上升。与对照组比较，50，100，200 μmol·L^-1 Ost作用24 h后，Bcl-2、pro-caspase-3、p-Histon H3（Ser10）表达水平降低，Bax、cleaved-caspase-3表达水平升高。结论 Ost对L02细胞有毒性损伤作用，呈一定的时间和浓度依赖性，可促进细胞凋亡，抑制细胞增殖。

Abstract:

OBJECTIVE To investigate the toxic effect of osthole (Ost) on L02 cells and explore its mechanism, which could provide an experimental evidence for the safe and rational application of fructus cnidii and its preparation. METHODS Different concentrations of Ost were applied to L02 cells, MTT assay was used to detect cell activity;the release rate of lactate dehydrogenase (LDH) was analyzed using LDH kits;Hoechst 33342 staining was used to detect cell nucleus morphology;Annexin V/PI staining was used to detect apoptosis;Western blot was utilized to detect the expression of Bcl-2, Bax, pro-caspase-3, cleaved-caspase-3(p17) and p-Histon H3(Ser10). RESULTS The activity of L02 cells decreased under the action of Ost, while the release rate of LDH increased and appeared to be concentration dependent. The cell nucleus treated by Ost were shrunken and fragmented under Hoechst 33342 fluorescence staining. The results of double staining of Annexin V/PI showed that the apoptosis rate increased with the increase of Ost concentration. compared with the control group, after Ost (50, 100, 200 μmol·L^-1) treatment for 24 h, the expression levels of Bcl-2, pro-caspase-3 and p-Histon H3(Ser10) decreased, and the expression levels of Bax and cleaved-caspase-3 increased. CONCLUSION Ost has toxic effects on L02 cells, which appeares a time and concentration dependent manner, as a result it will promote apoptosis and inhibit cell proliferation.