S-Adenosylhomocysteine Inhibits NF-κ B-Mediated Gene Expression in Hepatocytes and Confers Sensitivity to TNF Cytotoxicity.

Autor:	Watson, Walter H., Burke, Tom J., Doll, Mark A., McClain, Craig J.
Předmět:	ADENOSINES ALCOHOLISM APOPTOSIS BIOLOGICAL assay CELL culture CELL physiology CELLULAR signal transduction GENE expression LIVER POLYMERASE chain reaction PROTEINS RESEARCH funding TOXICITY testing TUMOR necrosis factors WESTERN immunoblotting HOMOCYSTEINE REVERSE transcriptase polymerase chain reaction DATA analysis software DESCRIPTIVE statistics
Zdroj:	Alcoholism: Clinical & Experimental Research; Apr2014, Vol. 38 Issue 4, p889-896, 8p
Abstrakt:	Background Chronic alcohol exposure results in liver injury that is driven in part by inflammatory cytokines such as tumor necrosis factor-α ( TNF). Hepatocytes are normally resistant to the cytotoxic effects of TNF, but they become sensitized to TNF by chronic alcohol exposure. Recently, we reported that the decrease in the ratio of S-adenosylmethionine ( SAM) to S-adenosylhomocysteine ( SAH) that occurs with alcoholic liver injury renders hepatocytes sensitive to TNF cytotoxicity. The purpose of this study was to determine whether inhibition of the transcription factor nuclear factor-kappaB ( NF-κ B) contributed to TNF-induced cell death in hepatocytes with high levels of SAH. Methods Primary human hepatocytes or Hep G2 cells were pre-incubated with a combination of adenosine plus homocysteine to increase SAH levels. Following exposure to TNF, viability was determined by the MTT assay, and activation of the NF-κ B pathway was assessed by measuring degradation of cytosolic Iκ B-α, phosphorylation and translocation of NF-κ B to the nucleus, and expression of NF-κ B-dependent genes. TNF-induced apoptotic signaling pathways were assessed by monitoring levels of the anti-apoptotic protein, A20, and cleavage products of the caspase-8 substrate, RIP1. Results NF-κ B-mediated gene expression was inhibited in cells with high SAH, despite the fact that TNF-induced degradation of the cytoplasmic inhibitor Iκ B-α and accumulation of NF-κ B in the nucleus persisted for much longer. In contrast to control cells, the NF-κ B that accumulated in the nucleus of cells with high SAH levels was not phosphorylated at serine 536, a modification associated with activation of the transactivation potential of this transcription factor. The inhibition of transactivation by NF-κ B resulted in lower mRNA and protein levels of the anti-apoptotic protein A20 and increased cleavage of RIP1. Conclusions High SAH levels inhibited NF-κ B-mediated gene expression and sensitized primary hepatocytes and Hep G2 cells to the cytotoxic effects of TNF. It is likely that crosstalk with other transcription factors is perturbed under these conditions, resulting in still other changes in gene expression. [ABSTRACT FROM AUTHOR]
Databáze:	Complementary Index
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