gw-4869 and Insulin-Resistance

We reported previously that increased acid sphingomyelinase (ASMase)-catalyzed hydrolysis of sphingomyelin, which leads to increases in ceramide and sphingosine 1 phosphate (S1P), played a key role in the synergistic upregulation of proinflammatory cytokines by palmitic acid (PA), a major saturated fatty acid, and lipopolysaccharide (LPS) in macrophages. Since macrophages are vital players in nonalcoholic steatohepatitis (NASH) and atherosclerosis, we assessed the effect of ASMase inhibition on NASH and atherosclerosis cooperatively induced by high-PA-containing high-fat diet (HP-HFD) and LPS in LDL receptor-deficient (LDLR-/-) mice. LDLR-/- mice were fed HP-HFD, injected with low dose of LPS and treated with or without the ASMase inhibitor amitriptyline. The neutral sphingomyelinase inhibitor GW4869 was used as control. Metabolic study showed that both amitriptyline and GW4869 reduced glucose, lipids, and insulin resistance. Histological analysis and Oil Red O staining showed that amitriptyline robustly reduced hepatic steatosis while GW4869 had modest effects. Interestingly, immunohistochemical study showed that amitriptyline, but not GW4869, strongly reduced hepatic inflammation. Furthermore, results showed that both amitriptyline and GW4869 attenuated atherosclerosis. To elucidate the underlying mechanisms whereby amitriptyline inhibited both NASH and atherosclerosis, but GW4869 only inhibited atherosclerosis, we found that amitriptyline, but not GW4869, downregulated proinflammatory cytokines in macrophages. Finally, we found that inhibition of sphingosine 1 phosphate production is a potential mechanism whereby amitriptyline inhibited proinflammatory cytokines. Collectively, this study showed that amitriptyline inhibited NASH and atherosclerosis through modulation of sphingolipid metabolism in LDLR-/- mice, indicating that sphingolipid metabolism in macrophages plays a crucial role in the linkage of NASH and atherosclerosis.

Topics: Amitriptyline; Aniline Compounds; Animals; Atherosclerosis; Benzylidene Compounds; Blood Glucose; Cytokines; Diet, High-Fat; Down-Regulation; Enzyme Inhibitors; Insulin Resistance; Lipopolysaccharides; Macrophages; Male; Mice; Mice, Knockout; Non-alcoholic Fatty Liver Disease; Receptors, LDL; Sphingolipids; Sphingomyelin Phosphodiesterase

Sphingolipids play an important role in the development of insulin resistance. Ceramides are the most potent inhibitors of insulin signal transduction. Ceramides are generated in response to stress stimuli and in old age. In this work, we studied the possible contribution of different pathways of sphingolipid metabolism in age-dependent insulin resistance development in liver cells. Inhibition of key enzymes of sphingolipid synthesis (serine palmitoyl transferase, ceramide synthase) and degradation (neutral and acidic SMases) by means of specific inhibitors (myriocin, fumonisin B1, imipramine, and GW4869) was followed with the reduction of ceramide level and partly improved insulin regulation of glucose metabolism in "old" hepatocytes. Imipramine and GW4869 decreased significantly the acidic and neutral SMase activities, respectively. Treatment of "old" cells with myriocin or fumonisin B1 reduced the elevated in old age ceramide and SM synthesis. Ceramide and SM levels and glucose metabolism regulation by insulin could be improved with concerted action of all tested inhibitors of sphingolipid turnover on hepatocytes. The data demonstrate that not only newly synthesized ceramide and SM but also neutral and acidic SMase-dependent ceramide accumulation plays an important role in development of age-dependent insulin resistance.

Topics: Aging; Aniline Compounds; Animals; Benzylidene Compounds; Ceramides; Fatty Acids, Monounsaturated; Fumonisins; Glucose; Glycogen; Hepatocytes; Imipramine; Insulin; Insulin Resistance; Male; Oxidoreductases; Rats; Serine C-Palmitoyltransferase; Sphingolipids; Sphingomyelin Phosphodiesterase