The combination of increased secretion of triglyceride (TG)-rich apolipoprotein B (apoB) lipoproteins (Lps) and hepatic steatosis, are characteristic concomitants of insulin resistance, the metabolic syndrome, and type 2 diabetes mellitus. The basis for increased secretion of apoB-Lps is complex, and includes altered regulation of apoB synthesis and transport in hepatocytes, increased delivery of fatty acids (FA) to the liver, and increased de novo hepatic lipogenesis (DNHL). Increased hepatic uptake of FA and increased DNHL are also critical for the development of hepatic steatosis. However, recent evidence from our laboratory, and others, indicate that endoplasmic reticulum (ER) stress and aberrant hepatic expression of PPAR32 may both play important roles in apoB-Lp secretion and the development of hepatic steatosis. The studies proposed in this application focus mainly on our very recent and exciting findings linking disordered FA metabolism to both ER stress and hepatic expression of PPAR32. Specifically, we have demonstrated that oleic acid (OA)-loading of cultured hepatoma cells can induce ER stress that can have, at lower doses of FA for shorter durations, a """"""""selective"""""""" effect on apoB100 secretion without affecting total protein synthesis or secretion of albumin or apoA-I. We have comparable in vivo data in mice. Higher doses or longer exposure to OA cause """"""""global"""""""" ER stress with inhibition of secretion of several proteins and reduced total protein synthesis. We propose a series of experiments to extend, in detail, these exciting findings. We will compare ER stress effects on apoB by OA and tunicamycin, characterize the pathways by which apoB is degraded during ER stress, examine the effects of palmitic acid (PA) and docosahexanoic acid (DHA), extend our investigations regarding the particular sensitivity of apoB100 to ER stress, demonstrate in vivo that inhibition of ER stress results in increased secretion of VLDL and reversal of steatosis, and examine the important link between ER stress and increased lipogenesis. In a parallel series of studies we will extend observations related to the role of aberrant hepatic expression of PPAR32 in steatosis. In particular, we will determine the role of C/EBPs in the expression of PPAR32 in liver, investigate the signaling pathways for insulin stimulation of PPAR32 expression, and compare the effects of different FAs. We will extend initial studies indicating that hepatic expression of PPAR32 may both increase DNHL and, by suppressing a neutral TG lipase, TGH2, reduce the incorporation of hepatic TG into apoB-containing Lps. Finally, we will assess recent data suggesting that PPAR32 expression in liver can be a response to ER stress, possibly via increased expression of C/EBP. Overall, our proposed studies should provide the basis for a unified model to explain the common association of increased (but not maximal) apoB secretion, increased hepatic DNHL, and steatosis in mice and people with insulin resistance. The combination of increased secretion of triglycerides and apolipoprotein B (apoB), and hepatic steatosis (fatty liver), are characteristic concomitants of insulin resistance, the metabolic syndrome, and type 2 diabetes mellitus. Recent evidence from our laboratory indicate that endoplasmic reticulum (ER) stress and aberrant hepatic expression of the nuclear receptor, PPAR2, which is typically found in fat, may both play important roles in the increased secretion of triglycerides and apoB, and in the development of hepatic steatosis.
The studies proposed in this application will focus on these new discoveries and extend our knowledge about the roles of ER stress and PPAR2 in the regulation of hepatic lipid metabolism. We will use cell culture models and mice, conducting in vitro and in vivo studies. Overall, our proposed studies should provide evidence for a unified model to explain the common association of increased apoB secretion, increased hepatic DNHL, and steatosis in mice and people with insulin resistance.
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