Hexosamine biosynthesis pathway flux promotes endoplasmic reticulum stress, lipid accumulation, and inflammatory gene expression in hepatic cells

Abstract

There is increasing evidence that endoplasmic reticulum (ER) stress contributes to the development of atherosclerosis in diabetes mellitus. The purpose of this study was to determine the effects of increased hexosamine biosynthesis pathway (HBP) flux on ER stress levels and the complications of ER stress associated with diabetes and atherosclerosis in hepatic cells. Glutamine:fructose-6-phosphate amidotransferase (GFAT), the rate-limiting enzyme of the HBP, was overexpressed in HepG2 cells by use of an adenoviral expression system. The ER stress response and downstream effects, including activation of lipid and inflammatory pathways, were determined using real-time PCR, immunoblot analysis, and cell staining techniques. GFAT overexpression resulted in increased expression of ER stress markers, including Grp78, Grp94, calreticulin, and GADD153, relative to cells infected with an empty adenoviral vector. In addition, GFAT overexpression promoted lipid, but not cholesterol, accumulation in hepatic cells as well as inflammatory pathway activation. Treatment with 6-diazo-5-oxo-norleucine, a GFAT antagonist, blocked the effects of GFAT overexpression. Consistent with our in vitro data, hyperglycemic mice presented with elevated markers of hepatic ER stress, glucosamine and lipid accumulation. Together, these data suggest that HBP flux-induced ER stress plays a role in the development of hepatic steatosis and atherosclerosis under conditions of hyperglycemia.

Protein kinase R-like endoplasmic reticulum kinase and glycogen synthase kinase-3α/β regulate foam cell formation

“…Elevated levels of ER stress have been observed in animal models of hyperglycemia, obesity, and dyslipidemia ( 6,7,30,37,38 ). In a hyperglycemic state, GLN, a metabolite of glucose, accumulates within cells and acts as a potent inducer of ER stress (39)(40)(41). In addition, lipids such as PA and unesterifi ed cholesterol are thought to disturb ER function by disrupting the composition of the ER membrane ( 11,42 ).…”

Endoplasmic Reticulum Stress and the Unfolded Protein Response in Nonalcoholic Fatty Liver Disease

“…The hexosamine biosynthetic pathway plays a key role in glycosylation and increased flux through this pathway, which can occur under conditions of hyperglycemia and/or hyperlipidemia, has been linked to PERK-dependent ER stress and attenuation of ApoB100 synthesis (128,136). Overexpression of glutamine:fructose-6-phosphate amidotransferase (GFAT), the rate-limiting enzyme of the hexosamine biosynthetic pathway induced ER stress, lipid accumulation, and upregulation of genes associated with inflammatory pathways in HepG2 cells (136). Treatment of cells with a GFAT antagonist blocked these responses to GFAT overexpression.…”

VAMP-associated Proteins (VAP) as Receptors That Couple Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) Proteostasis with Lipid Homeostasis

“…Overall, these observations suggest that protein misfolding in the ER leads to global effects on lipid homeostasis. Indeed aberrant cholesterol distribution is observed during general perturbation of protein folding in the ER; drugs that perturb ER N-glycosylation or oxidative folding similarly lead to cholesterol accumulation in LE (11)(12)(13). The mechanisms that enable the coupling of protein biogenesis in the ER with cholesterol flow remain undefined.…”