[1]唐梦灵,胡治强,袁思琪,等.类鼻疽杆菌通过抑制自噬调控HepG2细胞胞内脂质代谢[J].第三军医大学学报,2019,41(18):1722-1729.
 TANG Mengling,HU Zhiqiang,YUAN Siqi,et al.Burkholderia pseudomallei regulates lipid metabolism in HepG2 cells by inducing autophagy suppression[J].J Third Mil Med Univ,2019,41(18):1722-1729.
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《第三军医大学学报》[ISSN:1000-5404/CN:51-1095/R]

卷:
41卷
期数:
2019年第18期
页码:
1722-1729
栏目:
基础医学
出版日期:
2019-09-30

文章信息/Info

Title:
Burkholderia pseudomallei regulates lipid metabolism in HepG2 cells by inducing autophagy suppression
作者:
唐梦灵胡治强袁思琪饶承龙张雨谢建平李倩毛旭虎
西南大学生命科学学院,三峡地区生态环境与生物资源国家重点实验室培育基地,现代生物制药研究所;陆军军医大学(第三军医大学)药学与检验医学系临床微生物及免疫学教研室
Author(s):
TANG Mengling HU Zhiqiang YUAN Siqi RAO Chenglong ZHANG Yu XIE Jianping LI Qian MAO Xuhu

Institute of Modern Biopharmaceuticals, State Key Laboratory Breeding Base of Eco-Environment and Bio-Resource of the Three Gorges Area, Key Laboratory of Eco-environments in Three Gorges Reservoir Region of Ministry of Education, School of Life Sciences, Southwest University, Chongqing, 400715; Department of Clinical Microbiology and Immunology, Faculty of Pharmacy and Medical Laboratory, Army Medical University (Third Military Medical University), Chongqing, 400038, China

关键词:
类鼻疽杆菌HepG2细胞自噬脂代谢
Keywords:
Burkholderia pseudomallei HepG2 cells autophagy lipid metabolism
分类号:
R322.47; R329.28; R378.99
文献标志码:
A
摘要:

目的 探讨类鼻疽杆菌感染HepG2细胞后诱导的自噬抑制对胞内脂质代谢的调控作用。方法 类鼻疽杆菌感染HepG2细胞6、12、24 h为感染组,以未感染组作为对照。激光共聚焦显微镜观测各组细胞内脂滴和GFP-自噬点的数量;定量试剂盒检测各组细胞内甘油三酯及胆固醇水平;Western blot检测各组细胞中自噬相关蛋白LC3Ⅱ、p62表达水平。利用自噬抑制剂3-MA、Baf-A1以及自噬激活剂Rapa处理细胞,检测改变宿主细胞自噬水平后,对类鼻疽杆菌感染HepG2细胞后胞内脂质代谢的影响;外源加入游离脂肪酸三醋酸甘油酯,通过细菌胞内计数实验检测类鼻疽杆菌胞内存活情况。 结果 类鼻疽杆菌感染HepG2细胞6、12、24 h后,与未感染组比较,胞内脂滴数量增多,脂质成分甘油三酯和胆固醇水平也随感染时间延长逐渐升高,且在感染24 h后达到最高水平(P<0.01);自噬检测发现,与未感染组比较,LC3Ⅱ蛋白表达随感染时间逐渐下降,p62蛋白表达水平随感染时间逐渐蓄积,胞内自噬小体的数量也随感染时间逐渐减少,且感染24 h后数量最低(P<0.01),提示类鼻疽杆菌感染HepG2细胞后自噬受抑;使用自噬抑制剂3-MA、Baf-A1以及自噬激活剂Rapa处理细胞后,与对照组比较,抑制自噬导致胞内脂质蓄积加重(P<0.01),而激活自噬可以减轻胞内脂质的蓄积(P<0.01);此外,外源补充三醋酸甘油酯促进宿主细胞脂质代谢速率能明显降低类鼻疽杆菌胞内存活量(P<0.01)。 结论 类鼻疽杆菌感染HepG2细胞后,通过诱导自噬抑制调控HepG2细胞胞内脂质代谢。

Abstract:

Objective To investigate the regulatory effect of Burkholderia pseudomallei (B. pseudomallei)-induced autophagy suppression on lipid metabolism in HepG2 cells. MethodsHepG2 cells infected by B. pseudomallei were observed for changes in intracellular lipid droplets and GFP-autophagy spots using confocal laser microscopy at 6, 12, 24 h following the infection. The levels of triglyceride and cholesterol in the cells were determined using a quantitative detection kit, and the expression levels of autophagy-related proteins LC3Ⅱ and p62 were detected with Western blotting. The effects of pretreatment with the autophagy inhibitors 3-MA and Baf-A1 or the autophagy activator Rapa prior to B. pseudomallei infection were examined on lipid metabolism in HepG2 cells. The changes in the intracellular survival of B. pseudomallei in response to exogenous free fatty acid triacsin C were examined by counting the viable bacteria in the cells. ResultsB. pseudomallei infection resulted in significantly increased number of intracellular lipid droplets at 6, 12 and 24 h after the infection and progressively increased levels of triglyceride and cholesterol in HepG2 cells over time, which reached the highest level at 24 h after the infection (P<0.01). The cells infected by B. pseudomallei showed progressively lowered expression of LC3Ⅱ protein and increased expression of p62 protein over time with gradually decreased number of autophagosomes, which reached the lowest level at 24 h after the infection (P<0.01), suggesting suppressed autophagy in HepG2 cells after B. pseudomallei infection. Inhibition of autophagy by pretreatment of the cells with 3-MA and Baf-A1 caused significantly increased intracellular lipid accumulation (P<0.01), whereas activation of autophagy by Rapa obviously reduced the accumulation of intracellular lipids (P<0.01). The addition of exogenous triacsin C significantly reduced the intracellular survival of B. pseudomallei (P<0.01). ConclusionB. pseudomallei infection regulates intracellular lipid metabolism in HepG2 cells by inducting autophagy inhibition.

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更新日期/Last Update: 2019-09-21