大鼠微粒体谷胱甘肽S-转移酶1对苯丁酸氮芥体外致肿瘤细胞毒性的影响

Rat microsomal glutathione S-transferase 1 alters cytotoxic effects of chlorambucil on PC-3,K562,HepG2 and P388D1 cell lines

目的:基于苯丁酸氮芥(chlorambucil,CHB)激活微粒体谷胱甘肽S-转移酶1(microso-mal glutathione S-transferase1,mGST1)可显著加快催化CHB-GSH结合效应,探讨大鼠mGST1激活对CHB致不同来源肿瘤细胞株毒性的影响。方法:取雄性Sprague-Dawley大鼠肝脏,制备微粒体,去除胞浆GST的污染,获得纯化mGST1并测其活性。分别设CHB组,CHB+GSH对照组以及CHB+GSH+mGST1组。分别与PC-3、K562、HepG2和P388D1细胞株培养24h、48h、72h后,以IC50作为细胞损伤指标;作用8h后以JC-1染色评价线粒体膜电位(ΔΨm)改变作为损伤早期指标;作用48h以后以AO/EB染色差异作为细胞核损伤后期指标。结果:在mGST1存在环境下,24h、48h、72h PC-3、K562、HepG2和P388D1细胞株IC50均有显著增加。72h CHB+GSH对照组PC-3、K562、HepG2和P388D1细胞株的IC50分别为(6.13±2.42)μmol/L、(3.77±0.95)μmol/L、(5.36±3.06)μmol/L和(9.42±1.77)μmol/L;CHB+GSH+mGST1组显著高于CHB+GSH对照组,相应IC50(15.01±3.47)μmol/L(P<0.001)、(8.33±1.72)μmol/L(P<0.001),(26.74±5.45)μmol/L(P<0.001)和(16.93±0.85)μmol/L(P<0.001)。并伴有线粒体ΔΨm降低和核损伤减轻,呈明显的时效关系。结论:mGST1在富含GSH溶液中,可显著降低CHB对上述肿瘤细胞株的细胞毒性作用。

Objective： To explore the possible association between activation of rat microsomal glutathione S-transferase 1 （mGST1） and chlorambucil toxicity on selected cancer cell lines. Methods： Hepatic microsomes were prepared from male Sprague-Dawley rats and washed to remove cytosolic contamination, mGST1 was purified and its activity was measured. PC-3,K562, HepG2 and P388D1 cell lines were exposured to chlorambucil （CHB） alone or to CHB with mGST1 at concentations of 0～100 μmol/L for 8,24,48,72 h. Cytotoxic effects of CHB were determined by cell growth inhibition （MTT assay）, mitochondrial transmembrane potential （△ψm） ,and fluorescence morphological examination （AO/EB staining）. Results： The decreased cytotoxic effects of CHB on the cell lines altered by mGST1 were demonstrated in concentrationand time-dependant manners. The CHB-induced apoptosis on PC-3 and K562 cell lines altered by mGST1 was confirmed using △ψm examination,JC-1 or AO/EB staining. Conclusion： mGST1 can reduce the cytotoxic effects of CHB in selected cancer cell lines.