摘要
目的:研究甲嘎松汤对氧化应激损伤L-02的影响,并从Nrf2/Bach1-ARE抗氧化酶途径研究其抗氧化的机制。方法:MTT法评价建立葡萄糖氧化酶肝损伤细胞模型的方法。实验共分9组,分别为空白组,模型组,20%甲嘎松汤含药血清组,5μmol·L-1全反式视黄酸(ATRA)组,80μmol·L-1染料木黄铜(Genstein)组,80μmol·L-1叔丁基对苯二酚(t-BHQ)组,t-BHQ+20%甲嘎松汤含药血清组,ATRA+20%甲嘎松汤含药血清组,Genstein+20%甲嘎松汤含药血清组,以正常培养液或含t-BHQ,ATRA,Genistein的培养液干预L-02细胞12 h,再用20%甲嘎松汤含药血清培养液预保护12 h,最后采用100 U·L-1葡萄糖氧化酶制备氧化损伤模型。MTT法测定细胞活性,测定培养上清的丙氨酸氨基转移酶(ALT),天门冬氨酸氨基转移酶(AST)活性,丙二醛(MDA)含量,谷胱甘肽过氧化物酶(GSH-Px)活性,利用免疫荧光法和Western blot法观察细胞内核因子相关因子(Nrf2)和转录因子BTB-CNC异体同源体(Bach1)的分布及核内两者的表达量。结果:与空白组比较,模型组ALT,AST活性及MDA含量明显升高,GSH-Px活性明显降低(P<0.01),细胞核内Nrf2及Bach1的表达明显升高(P<0.01);与模型组比较,甲嘎松汤能升高氧化损伤以及3种干预剂下的细胞活性和GSH-Px活性,降低ALT,AST活性及MDA含量(P<0.05,P<0.01);可协同t-BHQ升高核内Nrf2表达,抑制ATRA降低核内Nrf2表达,抑制Genistein升高核内Nrf2表达,均具有统计学差异(P<0.01);在3种干预剂作用下,甲嘎松汤可促进Bach1出核(P<0.01)。结论:甲嘎松汤可保护葡萄糖氧化酶诱导氧化应激损伤肝细胞,其抗氧化保肝作用与调节Nrf2与Bach1表位位置以及核内蛋白水平有关。
Objective： To discuss the effects of Jiagasong decoction （JGSD） on oxidative damaged liver cells and to explore its possible Nrf2/Bachl-ARE-antioxidant enzymes pathway mechanism. Method： The liver injury cells model was induced by 100 U · L^-1 glucose oxidase （GO） and evaluated by MTT. The experiment included 9 groups： the control group, the model group, 20% JGSD contained serum group, 5 μ mol · L^-1 all-trans retinoic acid （ATRA） group, 80 μmol · L^-1 genstein group, 80 μ mol · L^-1 tert-butylhydroquinone （t-BHQ） group, t-BHQ ＋ 20% JGSD contained serum group, ATRA ＋ 20% JGSD contained serum group, genstein ＋ 20% JGSD contained serum group. The activities of alanine aminotransferase （ALT）, aspartate aminotransferase （AST）, malonaldehyde （MDA） levels of Nrf2 and Bach1 glutathione peroxidase （GSH-Px） were tested. The location in cells and nucleus protein were tested by immunofluorescence and Western blot. Result： Compared to the control group, ALT, AST and MDA contents incearsed, GSH-Px activity and cell proliferation decreased, and levels of Nrf2 and Baehl increased in the model group （P 〈 0.01 ）. The above indexes had good improvement in JGSD- contained groups （P 〈0.05, P 〈 0.01 ）. Moreover, there was coordinated effects between JGSY and the other three agents on Nrf2 expression （P 〈0. 01）. Meanwhile, JGSD coule promote the Bachl export from cell nuclear （P 〈 0.01 ）. Conclusion： JGSD could protect GO-induced cells damage from oxidative stress, and its antioxidation mechanism is related to regulating Nrf2 and Bachl protein levels in nuclear.
出处
《中国实验方剂学杂志》
CAS
CSCD
北大核心
2015年第3期118-124,共7页
Chinese Journal of Experimental Traditional Medical Formulae
基金
国家自然科学基金项目(81202997)
广东省科技计划项目(2011B031700075)
广东省自然科学基金项目(S2011010003925)
广州中医药大学科学研究基金项目(2004D25)
