Oxidative stress induced by reactive oxygen species (ROS) plays a causative role in the pathogenesis of alcohol induced cellular injury. The highly conserved heat shock response, resulting in production of heat shock proteins has been implicated in protective mechanisms against increased oxidative stress as well as other types of cellular injuries including TNFa cytotoxicity. Heat shock proteins (hsp) such as the hsp-70, 27 and hsp-90 not only inhibit apoptosis but also block increased inflammatory responses generated by oxidative stress. Therefore, these properties of heat shock proteins make it an excellent system for manipulation resulting in opening of new avenues of anti-oxidant therapies. We hypothesize that acute and chronic alcohol exposure of liver macrophages (Kupffer cells) as well as hepatic cells (hepatocytes) may activate the heat shock response and regulate the increased inflammatory responses and apoptosis resulting in alcohol induced oxidative stress. Our preliminary data shows that chronic alcohol increases inflammatory responses in monocytes and macrophages. Further, alcohol exposure also increases NFkB, an anti-apoptotic molecule in hepatic cells. Therefore, we propose that acute alcohol may induce hsps but chronic alcohol may inhibit the hsps to regulate the inflammatory response. We also propose that in hepatic cells chronic alcohol exposure may induce insufficient hsps to inhibit apoptosis, leading to alcohol-induced liver injury. NFkB in these cells may be up regulated to counteract the alcohol-induced alterations and promote survival though with less efficiency.
The specific aims of this proposal are - 1) To investigate the effect of acute and chronic alcohol exposure on expression of heat shock proteins such as hsp-70, hsp-90 hsp-60 and small hsps like hsp-27 in inflammatory cells and hepatocytes by evaluating -a) Protein levels of the hsps in nucleus and cytoplasm of the cells by Western blotting, b) Alterations in gene expression by determining mRNA levels by Northern Blotting. c) Activation of the heat shock transcription factor. 2) To evaluate the effect of acute and chronic alcohol on - Induction of inflammation and related genes in macrophages by the microarray method and induction of apoptosis by DNA fragmentation, caspase-3, -8 activation and pro-/anti-apoptotic genes in HEPA 1-6 cells by the microarray method and correlate changes with hsp alterations.
|Mandrekar, Pranoti; Catalano, Donna; Jeliazkova, Valentina et al. (2008) Alcohol exposure regulates heat shock transcription factor binding and heat shock proteins 70 and 90 in monocytes and macrophages: implication for TNF-alpha regulation. J Leukoc Biol 84:1335-45|
|Mandrekar, Pranoti (2007) Signaling mechanisms in alcoholic liver injury: role of transcription factors, kinases and heat shock proteins. World J Gastroenterol 13:4979-85|