Hepatocellular carcinoma HCC) arises in men more than three times as often as women. Liver cancer also disproportionately affects male rodents, providing a valuable model to study mechanisms of sex-dependent hepatocarcinogenesis. We and others have shown that proinflammatory cytokines, not sex hormones, are the primary drivers of male tumor promotion. However, the process is not cytokine-specific. Interruption of either TNF-a, IFN-g or IL-6 reduces experimental liver cancer in male mice. Therefore, cancer predilection appears to be an innate characteristic of the masculinized liver. Sex-dependent hepatocyte programming, not upstream immunity, may explain gender-dimorphic tumor incidence. We have reported that HCC in male mice is associated with loss of a sex-specific genome-wide transcriptional signature, a process termed liver-gender disruption. Such pan-chromosomal alterations implicate chromatin disruption in hepatocellular transformation. Moreover, we and others have shown that male HCC susceptibility is maturationally imprinted, as there is a decreasing benefit of castration with advancing age. Liver masculinization is enacted by pulsatile growth hormone from the pituitary. Hepatocytes transduce this signal through Stat5b, which translocates to the nucleus and initiates the male transcription program. Liver masculinization involves the synchronous alteration of thousands of genes, suggesting involvement of root-level chromatin modifiers. Nevertheless, while nucleosomal relaxation may facilitate required Stat5b access to g-associated sequences (GAS) and other DNA motifs, an unintended consequence might be disproportionate binding of closely related proinflammatory transcription factors. Because of this, we hypothesize that masculine Stat5b-dependent epigenetic remodeling invokes chromatin hypersensitivity to tumor-promoting proinflammatory cytokines. Unfortunately, molecular studies of gender-specific hepatocyte function have been hampered by the lack of a suitable continuous cell line. We have developed a sexually dimorphic continuous hepatocyte culture system, and propose here to use it to probe epigenetic mechanisms that define a chromatin state as masculine or feminine. Next, we will test whether the masculine epigenetic profile increases sensitivity to proinflammatory cytokines by facilitating access of transcription factors (Stat1, Stat3, NF-kB) to cognate DNA sequences. Cell studies will be complemented with comprehensive in vivo epigenetic profiling of male and female liver in wild- type C57BL/6 and Stat5b-/- mice. A proinflammatory chemical injury model (DEN) will be used to assess transcription factor binding and acute-phase responses, with an emphasis on sexually dimorphic genes. Finally, we will determine whether Stat5b deficiency is sufficient to protect male mice from DEN-initiated HCC, suggesting a common downstream mechanism for susceptibility to inflammation-associated liver cancer. These studies are the first to define epigenetic modifiers of liver sexual differentiation, and to interrogate a novel mechanism of gender-specific tumor promotion predicated on chromatin sensitivity to inflammation.

Public Health Relevance

PUBLIC HEALTH SIGNIFICANCE: Hepatocellular carcinoma, the fastest rising cancer in the United States, exhibits marked gender disparity. These studies explore a cellular basis for liver sexual differentiation based on DNA and associated protein modifications, and show how inflammation may selectively increase cancer risk in men.

National Institute of Health (NIH)
National Cancer Institute (NCI)
Exploratory/Developmental Grants (R21)
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Cancer Genetics Study Section (CG)
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Johnson, Ronald L
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University of North Carolina Chapel Hill
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Chapel Hill
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Hartwell, Hadley J; Petrosky, Keiko Y; Fox, James G et al. (2014) Prolactin prevents hepatocellular carcinoma by restricting innate immune activation of c-Myc in mice. Proc Natl Acad Sci U S A 111:11455-60
Hines, Ian N; Hartwell, Hadley J; Feng, Yan et al. (2011) Insulin resistance and metabolic hepatocarcinogenesis with parent-of-origin effects in A×B mice. Am J Pathol 179:2855-65