Enhanced iNOS Gene Expression in the Steatotic Rat Liver after Normothermic Ischemia

Abstract

Background: Impaired hepatic microcirculation in the steatotic liver has been identified as a considerable factor for increased vulnerability after ischemia/reperfusion (I/R). Changes in regulation and synthesis of vasoactive mediators, such as nitric oxide (NO) and endothelin (ET-1), may result in functional impairment of postischemic sinusoidal perfusion. The aim of the current study was to assess the impact of I/R injury on postischemic gene expression of NO and ET-1 in steatotic livers. Materials and Methods: Male Sprague-Dawley rats with or without hepatic steatosis (induced by carbon tetrachloride treatment) were subjected to normothermic I/R injury. Steady-state mRNA levels were assessed using RT-PCR to study the expression of genes encoding ET-1, NO synthase (endothelial cell NO synthase and inducible NO synthase, iNOS). Immunohistochemistry was performed for detection of iNOS. Results: I/R injury was followed by increased iNOS gene expression (RT-PCR/immunohistochemistry) in animals with hepatic steatosis, predominately in hepatocytes with fatty degeneration. A mild increase in mRNA levels for ET-1 was found in steatotic rat livers. I/R induced a further increase in ET-1 gene expression in some but not all reperfused steatotic livers. Conclusions: We show an enhanced gene expression of iNOS in postischemic steatotic rat livers. Hepatocytes with fatty degeneration appear to be the major source for NO generation. Furthermore, I/R may also induce ET-1 gene expression. Dysregulation of sinusoidal perfusion by NO and ET-1 is therefore likely to contribute to I/R injury of the steatotic liver.

Mitochondrial dysfunction and oxidative stress in the pathogenesis of alcohol- and obesity-induced fatty liver diseases

“…In normal healthy liver iNOS levels are very low or undetectable [91], whereas during inflammation and other disease states iNOS levels are increased in liver due to infiltrating inflammatory cells and via induction in Kupffer cells (resident liver macrophage), hepatocytes, and biliary epithelial cells [92][93][94]. Studies also report increased iNOS expression in liver of ob/ob mice [53], CCl 4 -induced steatotic liver following normothermic ischemia [95], and in the obese fa/fa rat exposed to binge alcohol [5]. This is important for hepatotoxicity because NO and peroxynitrite (ONOO − ) are known to mediate mitochondrial dysfunction [96,97] and increases in iNOS expression in fatty liver are correlated with nitration of mitochondrial proteins [53,98].…”

Molecular Mediators of Liver Ischemia and Reperfusion Injury: A Brief Review

“…Results from a porcine liver transplant model suggest that IRI may be triggered by iNOS in Kupffer cell and PMNs (56). Similarly, steatotic rat livers subjected to IR show upregulation of iNOS and endothelin (ET-1), suggesting an important role in regulation of sinusoidal perfusion (57). Inhibition of iNOS may exert beneficial biological effects.…”

Methods to Reduce Liver Ischemia/Reperfusion Injury

“…This enzyme is not expressed under physiological conditions but is up-regulated in many cell types during IRI, including hepatocytes, Kupffer cells and neutrophils (169)(170)(171). In the setting of liver IRI, the main source of NO after the upregulation of iNOS appears to be hepatocytes (169,172,173).…”