Postprandial insulin resistance in hepato- and adipocytes

Over the last decades, there has been a dramatic change of life-style habits and human environment characterized by physical inactivity and overnutrition. These changes have led to escalting rates of insulin resistant states including type 2 diabetes. Despite the wealth of studies on insulin resistance and life-style, the pathophysiological mechanisms linking modern life-style habits to insulin resistance are incompletely understood. One consequence are prolonged and exaggerated phases of postprandial lipemia resulting from an ineffective removal process of dietary triglycerides since dietary fat exceeds the actual needs, and human tisues are faced to handle more nutrition than needed. Two studies from our laboratory, one performed in vivo and the other in vitro, have demonstrated for the first time that postprandial lipemia with its characteristic elevation of triglyceride-rich lipoproteins (TGRL), is indeed capable of inducing a state of insulin resistance. In our in vivo study on postprandial lipemia and insulin resistance, we could not define the principal organ(s) mediating lipemia-induced insulin resistance. From our in vitro study, we know that postprandial TGRL are capable of inducing insulin resistance in skeletal muscle. However, whether postprandial TGRL are capable of inducing insulin resistance also in liver and adipose tissue, the two other major sites of insulin resistance, is unknown. Therefore, the overall goal of the research in this proposal is to investigate the effect of postprandial TGRL on insulin sensitivity in hepatocytes and adipocytes and, second, to analyze the influence of postprandial TGRL on expression of adipocytokines and their receptors, which represent an important aspect of insulin resistance in these two tissues. To determine the effect of highly purified TGRL, we will isolate lipoproteins from plasma after ingestion of a standardized fat-rich meal by zonal ultracentrifugation. Then, we will determine the effects of postprandial TGRL on insulin-stimulated glucose metabolism, i.e. glucose uptake, glycogen content, glycogen synthase activity and glycogen phosphorylase-a activity, and on intracellular lipid accumulation. Moreover, effects of TGRL on crucial steps in insulin signalling, i.e. IRS-1 and IRS-2, phosphatidyl-inositol 3-kinase (PI-3K), Akt, and GSK-3, will be analyzed. TGRL-induced alterations of adipocytokine/receptor expression will be studied to elucidate the effects of postprandial lipoproteins on this endocrine pathway involved in the pathogenesis of insulin resistance. The proposed research project represents a logical continuation of our previous studies which yielded novel and intriguing insights regarding the pathogenesis of insulin resistance. Expanding our research to the other major tissues involved in the development of insulin resistance as well as insulin sensitivity-modifying factors, i.e. adipocytokines and their receptors, will greatly enhance our knowledge of the interactions among various tissues involved in the pathogenesis of insulin resistant states including type 2 diabetes mellitus. Our study may open new avenues for the prevention and treatment of insulin resistant states.

In the first part of project P 19821, we sought to answer the question whether postprandial lipemia through its characteristic triglyceride-rich lipoproteins (TGRL) may induce hepatic insulin resistance in cultured hepatocytes, and if so, to elucidate the underlying molecular mechanisms. Incubation of hepatocytes with purified TGRL particles induced hepatocellular triglyceride accumulation which was paralleled by an impairment of glucose metabolism and several important insulin signaling steps. The effects of TGRL were dependent on the presence of apolipoproteins in TGRL particles and were more pronounced for denser TGRL. These data suggest that postprandial lipemia is an important factor in the pathogenesis of non-alcoholic fatty liver disease and its associated hepatic insulin resistance. In the course of project P 19821, we also initiated a within-subject crossover study assessing the effect of postprandial lipemia, a state of acute insulin resistance, on plasma concentrations of several adipocytokines, i.e. adipocyte-fatty acid binding protein (A-FABP), retinol binding protein-4 (RBP-4) and visfatin. Several previous studies have indicated that changes in the plasma concentrations of these adipocytokines are associated with chronic states of insulin resistance. Our study revealed that postprandial lipemia has no significant effect on the plasma concentrations of visfatin, A-FABP or RBP-4 in relation to their postabsorptive plasma profiles. We concluded that prolonged states of insulin resistance are required to affect plasma concentrations of these adipocytokines. In another study of project P 19821, we tested the hypothesis that postprandial lipemia with its characteristic elevation of TGRL may affect pancreatic alpha cell function. The background of this study is the observation that insulin resistant states are associated with elevated plasma glucagon levels and inadequate insulin-induced suppression of glucagon secretion. In healthy humans, mouse pancreatic islets and cultured pancreatic alpha cells, we were able to show that TGRL may bring about the above mentioned changes in glucagon kinetics. The molecular basis of these changes in glucagon kinetics is that TGRL reduce GABA-A receptor translocation to the cell membrane in pancreatic alpha cells. The data of this study together with our previous studies on the effect of TGRL on skeletal muscle and liver insulin sensitivity as well as our data on insulin resistance during postprandial lipemia could be interpreted to suggest that postprandial TGRL represent a common factor in the etiology of the diverse defects in type 2 diabetes. At last, we performed a study aimed at analyzing the molecular mechanism by which Serotonin improves insulin resistance in skeletal muscle cells. Our data show that Serotonin increases glucose uptake and glycogen content and leads to a serotonylation of RAB and RHO-proteins in L6 skeletal muscle cells suggesting that serotonylation of RAB and RHO-proteins represents the underlying mechanism of the positive effect of Serotonin on glucose homeostasis.