Immunohistochemistry pattern of hepatic inflammatory and insulin resistance markers in experimental model of nonalcoholic steatohepatitis

Abstract

Introduction: The pathophysiology of nonalcoholic steatohepatitis (NAS) includes, basically, insulin resistance, inflammation and oxidative stress. Thus, a study of immunostaining for liver insulin, adiponectin, tumor necrosis factor alpha (TNF-α), and inducible nitric oxide synthase (iNOS) receptors was conducted. Objective: To expand the knowledge about the pathophysiological and molecular mechanisms underlying the experimental model of steatohepatitis in rats fed a high-fat diet. Method: Twenty Wistar rats were divided into two groups: G1 (control, fed a standard diet), and G2 (fed a high-fat diet containing 58% of energy derived from fat, 18% from protein and 24% from carbohydrate). After eight weeks the animals were sacrificed. Blood glucose, insulin, total cholesterol, high-density lipoprotein (HDL), the very low-density lipoproteins (VLDL), triglycerides, aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP) and gamma-glutamyl transferase (GGT) were determined. The liver tissue was submitted to histopathological analysis, using a NAS score. In immunohistochemistry, we studied the expression of the insulin receptor, adiponectin, TNF-α and iNOS by tissue microarray method. Results and conclusion: There was marked cytoplasmic immunostaining for TNF-α and iNOS mediators in the group on a fat diet. Regarding insulin and adiponectin molecular markers, a reduction of cytoplasmic immunoreactivity of these antigens was observed in the group on a fat diet, reflecting, respectively, the state of hepatocellular inflammation (steatohepatitis) and insulin resistance in this experimental model of fat liver disease.

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“…The interpretation of immunoreactivity was performed in a semiquantitative manner by analyzing the extent of the staining positivity of the liver and Kupffer cells. Immunostaining of <10% was scored +1; 10% to 50% positivity scored +2; 51% to 90% scored +3; while >90% positivity scored +4 …”

“…Immunostaining of <10% was scored +1; 10% to 50% positivity scored +2; 51% to 90% scored +3; while >90% positivity scored +4. 19 2.6 | Assessment of liver enzymes and lipid profile Serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were measured using a commercial kit supplied by Diamond Diagnostics, Egypt. Serum cholesterol (TC), triglycerides (TG), and high-density lipoprotein cholesterol (HDL-c) were estimated using kit supplied by Spinreact, Barcelona, Spain.…”