Based on the well accepted two-hit model for ALD, Chronic Ethanol (EtOH)/LPS generated endotoxins activate NF?B that up regulates TNF?, and IL1 the potent proinflammatory cytokines causing hepatocyte injury. Further, EtOH induced Cyp2E1 and ADH lead to metabolic generation of acetaldehyde and increased reactive oxygen species (ROS), which activate quiescent HSC to myofibroblasts resulting in up regulation of fibrogenic genes, platelet derived growth factor -receptor (PDGFr), ?-smooth muscle actin (?SMA) and extracellular matrix proteins (ECM), collagen I, III, and fibronectin and epigenetic repressor gene, methyl-CpG binding protein 2 (MeCP2). In contrast, adipogenic genes, peroxisome proliferator-activated receptor ? (PPAR?), and sterol regulatory element-binding protein 1c (SREBP1c) are suppressed resulting in the loss of their vitamin A stores and their transdifferentiation from quiescent lipid storing phenotype to active myofibroblastic phenotype. Significantly, thymosin 4 (T4), a bioactive peptide, is reported to prevent inflammation and fibrosis in many extra-hepatic tissues. Based on these, PI hypothesizes that T4's anti-inflammatory and anti-fibrogenic actions against EtOH/LPS liver injury are mediated by (i) inhibiting the activation of NF?B by blocking the phosphorylation and dissociation of I?B and thereby prevent the up regulation of TNF?, and IL1 the potent proinflammatory cytokines and consequent liver injury, and (ii) suppressing the up regulated MeCP2, that coordinately reverses (a) the down regulated adipogenic genes and (b) up regulated fibrogenic genes and thereby prevent the trans-differentiation of HSC from lipid-storing pericytes to myofibroblasts. We also hypothesize that T4 elicits its above actions by overexpressing miR132 that suppresses MeCP2 overexpression caused by EtOH/LPS. PI has the following encouraging preliminary results in the EtOH/LPS mouse model to reassure the feasibilities of his novel exploratory approaches: T4 protects against EtOH/LPS induced 1. Up-regulation of NF?B signaling cascade, pI?B, TNF?, IL1 and consequent serum markers for liver injury; 2. up regulation of hepatic MeCP2, PDGFr, ?SMA, Col1?1 & proteins; and 3. down-regulation of adipogenic gene, PPAR?. As a result, we propose that T4 blocks the (i) activation of NF?B, TNF? and inflammatory cascade and (ii) transdifferentiation of quiescent HSC to fibrogenic HSC; yet, T4 maintains hepatocyte regeneration. Thus, the major goals of our innovative proposal are to accomplish the following specific aims:
Specific Aim 1. What are the possible mechanism/s of action/s of T4 Pre- and Post-treatment to protect/alleviate EtOH/LPS-mediated up regulation of NF?B signaling cascade, TNF? & IL1, and consequent serum and liver markers for liver injury? Specific Aim 2: What are the possible mechanism/s of action/s of T4 Pre- and Post-treatment to protect/alleviate against EtOH/LPS-mediated (a) up regulation of hepatic fibrogenic genes and their products? and (b) down regulation of adipogenic genes and their products? Are there corresponding morphological changes in liver cell architecture as well as in HSC phenotype? PI plans to approach using established EtOH/LPS two-hit mouse in vivo & in vitro model utilizing biochemistry, molecular biology, immuno- & histo-chemistry techniques. PI has a strong molecular biology and biochemical group that has the expertise in all aspects of this proposal. This may lead to novel therapy for ALD.
Alcoholic Liver Disease is characterized by alcohol- and endotoxin-induced oxidative stress leading to liver inflammation due to NF?? activation associated with activation of Stellate cells to transdifferentiate into fibrogenesis. Since Thymosin ?4 (T?4) has both anti-inflammatory and antifibrogenic properties, it is important to test whether this bioactive peptide can effectively prevent chronic alcohol/LPS-induced liver injury, T?4 would be established as a novel therapeutic agent in alcoholic liver injury and fibrogenesis. Thus, T?4 could be a potential therapeutic agent targeted for future preclinical and clinical trials for the treatment of alcoholic liver disease (ALD).
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