Unidad de Investigación del Hospital Universitario Reina Sofía, Instituto Maimónides de Investigación Biomédica, Córdoba 14004, Spain.
Lab Invest. 2010 Jan;90(1):116-27. doi: 10.1038/labinvest.2009.104. Epub 2009 Oct 5.
Cholestatic liver injury following extra- or intrahepatic bile duct obstruction causes nonparenchymal cell proliferation and matrix deposition leading to end-stage liver disease and cirrhosis. In cholestatic conditions, nitric oxide (NO) is mainly produced by a hepatocyte-inducible NO synthase (iNOS) as a result of enhanced inflow of endotoxins to the liver and also by accumulation of bile salts in hepatocytes and subsequent hepatocellular injury. This study was aimed to investigate the role of NO and S-nitrosothiol (SNO) homeostasis in the development of hepatocellular injury during cholestasis induced by bile duct ligation (BDL) in rats. Male Wistar rats (200-250 g) were divided into four groups (n=10 each), including sham-operated (SO), bile duct-ligated (BDL), tauroursodeoxycholic acid (TUDCA, 50 mg/kg) and S-methylisothiourea (SMT, 25 mg/kg) treated. After 7 days, BDL rats showed elevated serum levels of gamma-glutamiltranspeptidase, aspartate aminotransferase, alanine aminotransferase, LDH, and bilirubin, bile duct proliferation and fibrosis, compared with the SO group. TUDCA treatment did not significantly alter these parameters, but the iNOS inhibitor SMT ameliorated hepatocellular injury, as shown by lower levels of circulating hepatic enzymes and bilirubin, and a decreased grade of bile duct proliferation and fibrosis. Both TUDCA and SMT treatments reversed Mrp2 canalicular pump expression to control levels. However, only SMT treatment significantly lowered the increased levels of plasma NO and S-nitrosation (S-nitrosylation) of liver proteins in BDL rats. Moreover, BDL resulted in a reduction of the S-nitrosoglutathione reductase (GSNOR/Adh5) enzymatic activity and a downregulation of the GSNOR/Adh5 mRNA expression that was reverted by SMT, but not TUDCA, treatment. A total of 25 liver proteins, including S-adenosyl methionine synthetase, betaine-homocysteine S-methyltransferase, Hsp90 and protein disulfide isomerase, were found to be S-nitrosated in BDL rats. In conclusion, the inhibition of NO production during induced cholestasis ameliorates hepatocellular injury. This effect is in part mediated by the improvement of cell proficiency in maintaining SNO homeostasis.
肝外或肝内胆管阻塞导致胆汁淤积性肝损伤会引起非实质细胞增殖和基质沉积，从而导致终末期肝病和肝硬化。在胆汁淤积的情况下，一氧化氮 (NO) 主要由肝细胞诱导型一氧化氮合酶 (iNOS) 产生，这是由于内毒素流入肝脏增加，以及胆汁盐在肝细胞内积聚和随后的肝细胞损伤所致。本研究旨在探讨一氧化氮和 S-亚硝基硫醇 (SNO) 平衡在胆管结扎 (BDL) 诱导的大鼠胆汁淤积性肝损伤发展中的作用。雄性 Wistar 大鼠 (200-250 g) 分为四组 (每组 10 只)，包括假手术 (SO)、胆管结扎 (BDL)、牛磺熊脱氧胆酸 (TUDCA，50 mg/kg) 和 S-甲基异硫脲 (SMT，25 mg/kg) 处理。7 天后，与 SO 组相比，BDL 大鼠血清γ-谷氨酰转肽酶、天冬氨酸转氨酶、丙氨酸转氨酶、乳酸脱氢酶和胆红素水平升高，胆管增生和纤维化。TUDCA 治疗并未显著改变这些参数，但 iNOS 抑制剂 SMT 改善了肝细胞损伤，表现为循环肝酶和胆红素水平降低，胆管增生和纤维化程度降低。TUDCA 和 SMT 治疗均使 Mrp2 管腔泵表达恢复至对照水平。然而，只有 SMT 治疗可显著降低 BDL 大鼠血浆 NO 和肝蛋白 S-亚硝化 (S-亚硝基化) 水平升高。此外，BDL 导致还原型谷胱甘肽 S-亚硝基转移酶 (GSNOR/Adh5) 酶活性降低和 GSNOR/Adh5 mRNA 表达下调，SMT 治疗可逆转这种降低，但 TUDCA 治疗无效。在 BDL 大鼠中发现 25 种肝脏蛋白发生 S-亚硝化，包括 S-腺苷甲硫氨酸合成酶、甜菜碱同型半胱氨酸 S-甲基转移酶、热休克蛋白 90 和蛋白二硫键异构酶。总之，在诱导性胆汁淤积期间抑制 NO 生成可改善肝细胞损伤。这种作用部分是通过改善细胞维持 SNO 平衡的能力来介导的。
