The endoplasmic reticulum (ER) chaperone protein glucose-regulated protein 78 (GRP78/BiP) is a master regulator of ER homeostasis and stress response which are implicated in pathogenesis of metabolic disorders. By applying the LoxP-Cre strategy, we generated mice with liver specific GRP78 loss. Our studies using this novel mouse model revealed that liver GRP78 was required for neonatal survival and more than 50% loss of GRP78 in the adult liver caused ER stress response and dilation of ER compartment, accompanied by onset of apoptosis, suggesting critical involvement of GRP78 in maintaining hepatocyte ER homeostasis and viability. Further, these mice exhibited elevation of serum alanine aminotransferase and fat accumulation in the liver, and were sensitized to a variety of acute and chronic hepatic disorders by alcohol, high fat diet, drugs and toxins, which were alleviated by simultaneous administration of the molecular chaperone, 4-phenylbutyrate. Analysis by microarray followed by 2D protein profile revealed major perturbation of unfolded protein response (UPR) targets and common enzymes/factors in lipogenesis as well as new factors including liver major urinary proteins, fatty acid binding proteins, adipose differentiation-related protein, cysteine-rich with EGF-like domains 2 (Creld2), nuclear protein 1, and growth differentiation factor 15 (gdf15) possibly contributing to liver steatosis or fibrosis under ER stress. Our findings underscore the importance of GRP78 in managing the physiological client protein load and suppressing apoptosis in hepatocytes, supporting the pathologic role of ER stress in the evolution of fatty liver and disease under adverse conditions of drugs, diet, toxins and alcohol.

A portion of HIV-infected patients under therapy with protease inhibitors (HIV PIs) concomitantly consume or abuse alcohol leading to hepatic injury. The underling mechanisms are not known. We hypothesize that HIV PI aggravates alcohol-induced liver injury through an endoplasmic reticulum (ER) stress mechanism. To address this, we treated mice and primary mouse and human hepatocytes (PMH and PHH respectively) with alcohol and the two HIV PIs: ritonavir and lopinavir. In mice, ritonavir and lopinavir (15 mg/kg body weight each) induced mild ER stress and inhibition of Sarco/ER Calcium-ATPase (SERCA) without significant increase in serum ALT levels. However, a single dose of alcohol by gavage (5g/kg body weight) plus the two HIV PIs caused a greater than 5-fold increase in serum ALT, a synergistic increase in alcohol-induced liver lipid accumulation and ER stress response, and a decrease of SERCA. Mice treated with chronic HIV PIs and alcohol developed moderate liver fibrosis. In PMH, the HIV drugs plus alcohol also inhibited SERCA expression and increased expression of GRP78, CHOP, SREBP1c and phosphorylated JNK2, which were accompanied by a synergistic increase in cell death compared to alcohol or the HIV drugs alone. In PHH, ritonavir and lopinavir or alcohol alone treatment increased mRNA of spliced Xbp1 and decreased SERCA, which were accompanied by reduced levels of intracellular calcium. Alcohol combined with the HIV drugs significantly reduced intracellular calcium levels and potentiated cell death, which was comparable to the cell death caused by the SERCA inhibitor-thapsigargin. Our findings suggest the possibility that HIV PIs potentiate alcohol-induced ER stress and injury through modulation of SERCA and maintaining calcium homeostasis should be a therapeutic aim for a better care of HIV patients.

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