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Oliva J.,LABioMed | French S.W.,LABioMed
Experimental and Molecular Pathology | Year: 2012

Mallory-Denk body (MDB) formation is a component of alcoholic and non alcoholic hepatitis. Proteins of the TLR pathway were shown to be involved in the formation of MDBs, in mice fed DDC. TLR genes are upregulated and SAMe supplementation prevents this up regulation and prevented the formation of MDBs. DNA of livers from control mice, from mice fed DDC 10weeks, refed 1week with DDC and with DDC+SAMe were extracted and used to study the methylation pattern of genes involves in the TLR pathway. A PCR array was used to analyze it. Using PCR arrays for the mouse TLR pathway,24 genes were found whose expression of IL12A was regulated by the methylation of its gene. DDC fed for 10weeks reduced the methylation of the IL12A gene expression. This expression was also reduced when DDC was refed. However, when SAMe was fed, the intermediate level methylation of IL12A was up regulated to the intermediate level and the methylation of the promoter decreased compared to DDC refeeding or DDC 10weeks. IL12A is known to induce the production of IFNg by NK and L T. We showed in a previous publication that IFNg is one of the major cytokines involved in the induction of MDB formation. The low expression of IL12A associated with the intermediate methylation of its promoter could explain one step in the mechanism which leads to the formation of MDBs. © 2012 Elsevier Inc. Source

Oliva J.,LABioMed | Bardag-Gorce F.,LABioMed | Lin A.,LABioMed | French B.A.,LABioMed | French S.W.,LABioMed
Experimental and Molecular Pathology | Year: 2010

Mallory-Denk bodies (MDBs) are found in chronic liver diseases. Previous studies showed that diethyl-1,4-dihydro-2,4,6-trimethyl-3,5-pyridinedicarboxylate (DDC) induced formation of MDBs and the up regulation of UbD expression in mouse liver. UbD is a protein over expressed in hepatocellular carcinomas. It is a potential preneoplastic marker in the mouse. It is hypothesized that inflammatory cytokines play a critical role in UbD up regulation and MDB formation. TNFa and IFNg treatment of HCC cell line Hepa 1-6, induced the expression of UbD and the expression of genes coding for the immunoproteasome (LMP2, LMP7, and MECL-1 subunits). TNFa and IFNg induced the activity of the UbD promoter, using a luciferase assay. The cotreatment with TNFa and IFNg induced the activity of the UbD promoter through an Interferon Sequence Responsive Element (ISRE). In addition, long term treatment with TNFa and IFNg induced the formation of MDB-like aggresomes in Hepa 1-6 cells, which emphasizes the role of inflammation in the formation of MDBs leading to the formation of liver tumors, in the mouse. Identifying the mechanism that regulates gene expression of UbD supports the hypothesis that down regulation of UbD and the proinflammatory gene expression would prevent MDB and HCC formations. Previous studies indicate that S-adenosylmethionine or betaine prevented IFNg induced UbD and MDB formations. © 2010 Elsevier Inc. Source

Oliva J.,LABioMed | Bardag-Gorce F.,LABioMed | French B.A.,LABioMed | Li J.,LABioMed | French S.W.,LABioMed
Experimental and Molecular Pathology | Year: 2010

Mice fed DDC (0.1%) for 10. weeks, and then withdrawn from the drug for 1. month, retain the ability to form Mallory-Denk bodies (MDBs) when the drug is refed for 7. days. The number of liver cells that form MDBs increased and partially replaced normal liver cells, at the end of 7. days of refeeding DDC. The MDBs that formed were associated with increased expression of UbD (also called FAT10) in the Mallory-Denk body forming cells. UbD is over expressed in 70% of human HCCs, but its cellular localization is not well established. UbD belongs to the UbL family (ubiquitin-like), and can be linked to others proteins with their 2 C-terminal glycine to lysine. By Western Blot, UbD was found to be covalently linked with proteins. We performed immunohistochemistry on tissue from mouse liver and found that UbD was located in the cytoplasm and in one or two nuclei of the same hepatocyte. However, in primary cell culture, UbD formed speckles within the cytoplasm of the liver cell. A similar pattern of cytoplasmic localization was observed in the Hepa 1-6 cell lines, which over expressed UbD fused with GFP at the C-Terminal. The localization and the control of UbD localization remain unclear. The identification of proteins that interact with UbD and the post translational modification of UbD would help to determine the regulation of this localization and function. © 2010. Source

French B.A.,University of California at Los Angeles | Oliva J.,LABioMed | Bardag-Gorce F.,LABioMed | Li J.,University of California at Los Angeles | And 3 more authors.
Experimental and Molecular Pathology | Year: 2012

EZH2/H3K27me3 and polycomb group complex (PcG) play a major role in regulating global gene expression including tumor suppressor genes. EZH2 is linked to cell cycle regulated EZH2 phosphorylation by CDK1, a mitotic kinase which increases in arrested mitosis compared to S phase. CDK1 phosphorylation of EZH2 accelerates the degradation of pEZH2. Phospho-EZH2 is subjected to ubiquitination. The half-like of pEZH2 is shorter when compared to total EZH2. In the present study, pEZH2 was found concentrated together with ubiquitin in the Mallory-Denk bodies (MDB) that were formed in hepatocytes in the livers of drug primed mice refed DDC and humans with alcoholic hepatitis or hepatocellular carcinoma. The cells that formed MDBs in the mice livers studied were associated with a growth advantage and a high proliferative index. However, the livers from patients with alcoholic hepatitis showed evidence of cell cycle arrest where PCNA, cyclin D1 and p27 positive nuclei were numerous but Ki-67 positive nuclei were scarce. It is concluded that MDB formation is linked to the cell cycle and global gene expression (i.e. loss of gene silencing) through its association with the regulation of the polycomb group PRC2/EZH2/H3K27me3 complex. © 2012 Elsevier Inc. Source

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