甲醛对肝细胞极低密度脂蛋白分泌外排相关基因的影响
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摘要
目的观察甲醛对肝癌细胞株HepG2细胞内外甘油三酯(TG)含量及细胞内极低密度脂蛋白(VLDL)分泌外排相关基因表达水平的影响。方法以0、0.004、0.02、0.1 mmol/L甲醛分别处理人肝癌HepG2细胞24、48 h,采用甘油磷酸氧化酶法(GPO-POD)测定细胞内外TG含量,采用QRT-PCR法检测细胞内载脂蛋白B100(apo B100)、微粒体TG转运蛋白(MTTP)、载脂蛋白E(apo E)、TG水解酶(CES3)、二酰基甘油酰转移酶1(DGAT1)、包被蛋白Ⅱ(COPⅡ)、Sar1b、低密度脂蛋白受体(LDLR)基因的表达水平。结果与阴性对照组相比,0.004、0.1 mmol/L甲醛染毒48 h后HepG2细胞内TG含量减少,染毒24 h后上清中TG含量增加;染毒24 h后0.004、0.02、0.1 mmol/L甲醛组或染毒48 h后0.004 mmol/L甲醛组细胞内apo B100基因表达水平升高;染毒24 h后0.004 mmol/L甲醛组或染毒48 h后0.004、0.02、0.1 mmol/L甲醛组细胞内MTTP基因表达水平升高;染毒24 h后0.1 mmol/L甲醛组细胞内LDLR基因表达水平降低,染毒48 h后0.02、0.1 mmol/L甲醛组细胞内LDLR基因表达水平升高;上述差异均有统计学意义(P<0.05)。结论甲醛可能通过增加肝细胞内apo B100和MTTP表达来促进VLDL的分泌外排。
Objective To know the effects of formaldehyde(FA) on the intra and extra-hepatocellular triglyceride(TG) contents and the expression of very low density lipoprotein(VLDL) secretion-related genes in HepG2 cells. Methods HepG2 cells were treated with FA at the doses of 0.004, 0.02 and 0.1 mmol/L for 24 hours and 48 hours, respectively. The intra and extra-hepatocellular TG contents were determined by using triglycerideGPO-POD assay kit after FA exposure. The gene expression of apolipoprotein B100(apo B100), microsomal triglyceride transfer protein(MTTP),apolipoprotein E(apo E), carboxylesterase 3(CES3), diacylgycerol acyltransferase 1(DGAT1), coat protein comlexⅡ(COPⅡ), Sar1 b and low density lipoprotein receptor(LDLR) in the intracellular was detected by QRT-PCR method. Results Compared with negative control group, the intra TG levels in the hepatocytes significantly decreased after exposure to 0.004 and 0.1 mmol/L FA for 48 hours and the extra-hepatocellular TG contents significantly increased after exposure to 0.004 and 0.1 mmol/L FA for 24 hours(P<0.05). Apo B100 gene expression significantly increased in each FA group after 24 hours and in 0.004 mmol/L group after 48 hours exposure(P<0.05). MTTP gene expression significantly increased in 0.004 mmol/L group after 24 hours and in each FA group after 48 hours exposure(P <0.05). LDLR gene expression significantly decreased in 0.1 mmol/L group after 24 hours exposure,increased in0.02 and 0.1 mmol/L groups after 48 hours exposure(P <0.05). Conclusion Formaldehyde exposure may promote the hepatocellular VLDL secretion through increasing apo B100 and MTTP gene expressions.
Objective To know the effects of formaldehyde(FA) on the intra and extra-hepatocellular triglyceride(TG) contents and the expression of very low density lipoprotein(VLDL) secretion-related genes in HepG2 cells. Methods HepG2 cells were treated with FA at the doses of 0.004, 0.02 and 0.1 mmol/L for 24 hours and 48 hours, respectively. The intra and extra-hepatocellular TG contents were determined by using triglycerideGPO-POD assay kit after FA exposure. The gene expression of apolipoprotein B100(apo B100), microsomal triglyceride transfer protein(MTTP),apolipoprotein E(apo E), carboxylesterase 3(CES3), diacylgycerol acyltransferase 1(DGAT1), coat protein comlexⅡ(COPⅡ), Sar1 b and low density lipoprotein receptor(LDLR) in the intracellular was detected by QRT-PCR method. Results Compared with negative control group, the intra TG levels in the hepatocytes significantly decreased after exposure to 0.004 and 0.1 mmol/L FA for 48 hours and the extra-hepatocellular TG contents significantly increased after exposure to 0.004 and 0.1 mmol/L FA for 24 hours(P<0.05). Apo B100 gene expression significantly increased in each FA group after 24 hours and in 0.004 mmol/L group after 48 hours exposure(P<0.05). MTTP gene expression significantly increased in 0.004 mmol/L group after 24 hours and in each FA group after 48 hours exposure(P <0.05). LDLR gene expression significantly decreased in 0.1 mmol/L group after 24 hours exposure,increased in0.02 and 0.1 mmol/L groups after 48 hours exposure(P <0.05). Conclusion Formaldehyde exposure may promote the hepatocellular VLDL secretion through increasing apo B100 and MTTP gene expressions.
引文
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[17]Wilfling F,Thiam AR,Olarte MJ,et al.Arf1/COPI machinery acts directly on lipid droplets and enables their connection to the ER for protein targeting[J].Elife,2014,3:796-805.
[2]Wang HX,Li HC,Lv MQ,et al.Associations between occupational exposure to formaldehyde and semen quality,a primary study[J].Sci Rep,2015,5:708-710.
[3]Inci M,Zarars z I,Davarci M,et al.Toxic effects of formaldehyde on the urinary system[J].Turk J Urol,2013,39:48-52.
[4]Takeshita D,Nakajima-Takenaka C,Shimizu J,et al.Effects of formaldehyde on cardiovascular system in in situ rat hearts[J].Basic Clin Pharmacol,2009,105:271-280.
[5]曹凤华,蔡洁,刘志敏,等.甲醛对小鼠不同脑区的神经毒性作用[J].生理学报,2015,67(5):497-504.
[6]李亚琳.甲醛对小鼠肝脏毒性作用的研究[J].动物医学进展,2013,34(4):125-129.
[7]Swenberg JA,Moeller BC,Lu K,et al.Formaldehyde carcinogenicity research 30 years and counting for mode of action,epidemiology,and cancer risk assessment[J].Toxicol Pathol,2013,41:181-189.
[8]杨兰生,孙少华,高安敏,等.甲醛对小鼠机体毒性及其B淋巴细胞瘤-2基因表达的影响[J].兰州大学学报:医学版,2014,40(1):12-15.
[9]Stayner L,Smith AB,Reeve G,et al.Proportion mortality study of workers in the garment industry exposed to formaldehyde[J].Am J Ind Med,1985,7:229-240.
[10]Mashek DG.Hepatic fatty acid trafficking:multiple forks in the road[J].Adv Nutr,2013,4:697-710.
[11]Hussain MM,Shi J,Dreizen P.Microsomal triglyceride transfer protein and its role in apo B-lipoprotein in assembly[J].J Lipid Res,2003,44:22-32.
[12]Hooper AJ,Burnett JR,Watts GF.Contemporary aspects of the biology and therapeutic regulation of the microsomal triglyceride transfer protein[J].Circ Res,2015,116:193-205.
[13]Chen Z,Newberry EP,Norris JY,et al.Apo B100 is required for increased VLDL-triglyceride secretion by microsomal triglyceride transfer protein in ob/ob mice[J].J Lipid Res,2008,49:2013-2022.
[14]Dettlaff-Pokora A,Sledzinski T,Swierczynski J.Up-regulation mttp and apob gene expression in rat liver is related to post-lipectomy hypertriglyceridemia[J].Cell Physiol Biochem,2015,36:1767-1777.
[15]Nerurkar PV,Pearson L,Efird JT,et al.Microsomal triglyceride transfer protein gene expression and apo B secretion are inhibited by bitter melon in Hep G2 cells[J].J Nutr,2005,135:702-706.
[16]刘黎,欧阳冬生.微粒体甘油三酯转移蛋白的功能及临床意义[J].国际病理科学与临床,2009,29(2):1673-2588.
[17]Wilfling F,Thiam AR,Olarte MJ,et al.Arf1/COPI machinery acts directly on lipid droplets and enables their connection to the ER for protein targeting[J].Elife,2014,3:796-805.