Esfandiari Farah, Villanueva Jesus A, Wong Donna H, French Samuel W, Halsted Charles H
Department of Internal Medicine, University of California-Davis, Davis, California 95616, USA.
Am J Physiol Gastrointest Liver Physiol. 2005 Jul;289(1):G54-63. doi: 10.1152/ajpgi.00542.2004. Epub 2005 Feb 10.
Previously, we showed that feeding micropigs ethanol with a folate-deficient diet promoted the development of hepatic injury while increasing hepatic levels of homocysteine and S-adenosylhomocysteine (SAH) and reducing the level of S-adenosylmethionine (SAM) and the SAM-to-SAH ratio. Our present goals were to evaluate mechanisms for hepatic injury using liver specimens from the same micropigs. The effects of ethanol feeding or folate-deficient diets, singly or in combination, on cytochrome P-450 2E1 (CYP2E1) and signal pathways for apoptosis and steatosis were analyzed using microarray, real-time PCR, and immunoblotting techniques. Apoptosis was increased maximally by the combination of ethanol feeding and folate deficiency and was correlated positively to liver homocysteine and SAH. Liver CYP2E1 and the endoplasmic reticulum stress signals glucose-regulated protein 78 (GRP78), caspase 12, and sterol regulatory element binding protein-1c (SREBP-1c) were each activated in pigs fed folate-deficient or ethanol diets singly or in combination. Liver mRNA levels of CYP2E1, GRP78, and SREBP-1c, and protein levels of CYP2E1, GRP78, nuclear SREBP, and activated caspase 12 each correlated positively to liver levels of SAH and/or homocysteine and negatively to the SAM-to-SAH ratio. The transcripts of the lipogenic enzymes fatty acid synthase, acetyl-CoA carboxylase, and stearoyl-CoA desaturase were elevated in the ethanol-fed groups, and each was positively correlated to liver homocysteine levels. The induction of abnormal hepatic methionine metabolism through the combination of ethanol feeding with folate deficiency is associated with the activation of CYP2E1 and enhances endoplasmic reticulum stress signals that promote steatosis and apoptosis.
此前，我们发现给小型猪喂食叶酸缺乏饮食的同时给予乙醇会促进肝损伤的发展，同时增加肝脏中同型半胱氨酸和S-腺苷同型半胱氨酸（SAH）的水平，降低S-腺苷甲硫氨酸（SAM）水平以及SAM与SAH的比值。我们目前的目标是使用来自相同小型猪的肝脏标本评估肝损伤的机制。采用微阵列、实时PCR和免疫印迹技术分析单独或联合给予乙醇或叶酸缺乏饮食对细胞色素P-450 2E1（CYP2E1）以及凋亡和脂肪变性信号通路的影响。乙醇喂养与叶酸缺乏联合作用时凋亡增加最为显著，且与肝脏同型半胱氨酸和SAH呈正相关。单独或联合给予叶酸缺乏或乙醇饮食的猪肝脏中的CYP2E1以及内质网应激信号葡萄糖调节蛋白78（GRP78）、半胱天冬酶12和固醇调节元件结合蛋白-1c（SREBP-1c）均被激活。肝脏中CYP2E1、GRP78和SREBP-1c的mRNA水平，以及CYP2E1、GRP78、细胞核SREBP和活化的半胱天冬酶12的蛋白水平均与肝脏中SAH和/或同型半胱氨酸水平呈正相关，与SAM与SAH的比值呈负相关。在乙醇喂养组中，生脂酶脂肪酸合酶、乙酰辅酶A羧化酶和硬脂酰辅酶A去饱和酶的转录本升高，且各自均与肝脏同型半胱氨酸水平呈正相关。乙醇喂养与叶酸缺乏联合导致的肝脏蛋氨酸代谢异常与CYP2E1的激活相关，并增强了促进脂肪变性和凋亡的内质网应激信号。
