Title: Human pregnane X receptor and sexual dimorphism in alcoholic liver disease. Abstract: Alcoholic liver disease (ALD) develops from excessive alcohol use and is more severe in both human and rodent females than their male counterparts. Female alcoholics also have higher death rate than men. Several mechanisms have been proposed to account for more severe ALD in females, including sex differences in ethanol (EtOH) pharmacokinetics, estrogen levels, and alcohol elimination rates. However, the precise mechanism(s) is/are not well understood, although there are recent indications that suggest complexities in both ALD sexual dimorphism and ALD pathogenesis. For example, the upregulation of murine hepatic fat-specific protein 27 (Fsp27) and its human ortholog, cell death-inducing DNA fragmentation factor alpha-like effector c (CIDEC), which have been previously implicated in metabolic disorders and cell apoptosis, promote a progressive form of ALD, alcoholic steatohepatitis (ASH) in both mice and human patients with ASH. However, whether EtOH hepatotoxicity in females involves Fsp27 upregulation is unknown. FSP27 is a target gene of the lipogenic transcription factor peroxisome proliferator-activated receptor γ (PPARγ) and inhibition of PPARγ ameliorates ASH in mice. Upstream, PPARγ is transcriptionally regulated by pregnane X receptor (PXR), a xenobiotic nuclear receptor that induces genes involved in the detoxification of drugs and other foreign chemicals. We have previously shown that mouse PXR exacerbates EtOH-induced hepatotoxicity. Therefore, we asked whether human PXR signaling similarly modulates the PPARγ-FSP27 signaling axis in EtOH hepatotoxicity, particularly, focusing on sex differences in ALD. Unexpectedly, we found that chronic-plus-binge EtOH ingestion upregulated both PPARγ and FSP27 in a sex-dependent manner in a PXR-humanized (hPXR) mice, lacking the mouse Pxr gene, but carrying the full human PXR gene. Furthermore, the nuclear constitutive androstane receptor (CAR) and PXR target gene, Cyp2b10, known to generate reactive oxygen species, was also increased in the livers of EtOH-fed female hPXR mice. Thus, our central hypothesis is that the female hPXR gene mediates sexual dimorphism in EtOH hepatotoxicity via increased lipogenesis and synergistic generation of reactive oxygen species (ROS) by enhanced Fsp27 and Cyp2b10 gene expression. To address these hypotheses, we will determine the contribution of the PXR-PPARγ-FSP27 signaling axis to female-specific ALD pathology (Aim 1). determine whether CYP2B6 induction is involved in female-specific EtOH-induced hepatotoxicity (Aim 2). determine the contribution of PXR, PPARγ, CYP2B6, and CIDEC genetic polymorphisms to sex and ethnic differences in ALD (Aim 3). A strength of this proposal is that, it makes use of humanized mice and human tissues from both sexes, expediting application to clinical studies. Based on our strong preliminary data, this proposal will provide valuable insights into how sex and species-specific regulation of PXR target genes, as well as specific biomarkers from metabolomics contribute to sex difference in EtOH-induced hepatotoxicity, which may identify therapeutic targets for the treatment of ALD.

PROJECT NARRATIVE Title: Human pregnane X receptor and sexual dimorphism in alcoholic liver disease. Alcoholic liver disease (ALD) is a disease that disproportionately affect women and African Americans (AAs). We have shown that the nuclear receptor pregnane X receptor (PXR) in mice promotes the development of ALD. Unexpectedly, we found that alcohol produced a more severe liver injury in female mice which carry the human PXR gene than female wild type mice. This study will use humanized animal models and human liver and blood samples of both sexes to provide important new insights into the mechanisms by which the human PXR gene promotes in ALD, especially in women and treatment options for ALD.