溶酶体cathepsin B参与大黄素诱导HK-2细胞凋亡

目的:研究大黄素对人肾小管上皮HK-2细胞的细胞毒性及其机制。方法:用MTT法检测经0～100μmol·L-1浓度大黄素作用后HK-2细胞的活力,透射电镜观察细胞核形态的改变,流式细胞仪检测亚二倍体细胞比例,用特异性的底物分别测定caspase 3和cathepsin B的活性。结果:大黄素可引起HK-2细胞死亡,并伴有亚二倍体细胞比例的增加和核浓缩、染色体边集等形态的改变,在引起HK-2细胞凋亡的浓度下caspase 3活性增加,而用caspase 3特异性抑制剂Ac-DEVD-CHO可抑制上述改变。在诱导HK-2细胞凋亡的剂量下大黄素使cathepsin B表达及其活性升高,cathepsin B特异性抑制剂CA-074能拮抗大黄素诱导的caspase 3活性升高,恢复HK-2的细胞活力。结论:大黄素在体外主要以caspase 3依赖方式引起HK-2细胞凋亡,cathepsin B参与了此过程。

Participation of cathepsin B in emodin-induced apoptosis in HK-2 cells

Objective To Investigated cytotoxic effects of emodin on HK-2 cell (a human proximal tubular epithelial cell line)and relative mechanism in vitro.Methods Cells treated with a series of concentration of emodin were detected their viabilities by MTT assay.Morphological changes of nuclear were observed under a transmission electron microscope.The ratio of hypodiploid cells was analyzed by flow cytometry.The activities of caspase 3 and cathepsin B were detected by incubation with specific substrates respectively.Results Emodin induceed the death of HK-2 cell death,accompanied by the increase in the ratio of hypodiploid cells,and nuclear condensation and chromatin margination.Emodin at apoptosis-inducing concentrations caused an increase of caspase 3 activity.The caspase 3 inhibitor,Ac-DEVD-CHO,recovered HK-2 cell viability,inhibited the ratio of hypodiploid cells.Emodin increased the expression of cathepsin B protein and the activity of cathepsin B also.CA-074 N-(1-3-trans-propyl-carbam oyloxirane-2-carbonyl)-1-isoleucyl-1-proline],a cathepsin B inhibitor,inhibited the increase of caspase 3 activity induced by emodin,and recovered HK-2 cell viability.Conclusion Emodin impairs HK-2 cell by inducting apoptosis in a caspase 3-dependent manner,and cathepsin B may be involved in this process.