Project Summary: The mechanisms that drive the behavior of hepatic macrophages in homeostasis and disease are poorly understood. The liver X receptor (LXR) transcription factors are highly expressed in hepatic macrophages and hepatocytes. While in hepatocytes LXR signaling controls cholesterol metabolism, macrophage LXR signaling regulates both cholesterol metabolism and inflammation, processes that are central to the pathogenesis of non-alcoholic steatohepatitis (NASH). Activation of LXRs represses inflammation by interfering with nuclear factor κ (NFκ) activation. NFκ is a downstream target of multiple NASH signaling pathways, including toll-like-receptors, interleukin-1, and tumor-necrosis factor The central hypothesis of this proposal is that loss of LXR signaling in liver macrophages will block LXR mediated inhibition of inflammatory signaling and worsen NASH progression. Prior research by the Glass lab has demonstrated that LXR is highly expressed in Kupffer cells (the resident macrophages of the liver) and rapidly upregulated in bone marrow derived macrophages upon entry into both healthy and NASH livers. This led to the hypothesis that loss of LXR in Kupffer cells would worsen NASH phenotypes in mice with metabolic disease via increased NFκ signaling. A pilot study was performed to examine the effect of myeloid-lineage-specific LXR deletion in NASH. In this study mice lacking myeloid LXR fed a NASH- model diet for 20 weeks demonstrated increased hepatic fibrosis, and hepatic inflammatory gene expression. However, whether this effect is due to loss of LXR in Kupffer cells or other hepatic macrophage populations present during NASH is unknown. Aim 1 will test whether loss of LXR signaling in Kupffer cells increases hepatic fibrosis and worsens NASH progression. Serum markers of inflammation, liver damage, and cholesterol metabolism and hepatic histopathology will be used to assess the effect of myeloid-specific or Kupffer cell-specific LXR knockout during NASH. ChIP-seq, ATAC-seq, and RNA-seq will be used on two novel mouse strains to mechanistically assess whether loss of LXR signaling alters inflammation in different hepatic macrophage populations during NASH. Aim 2 will test whether hepatic desmosterol is the native LXR ligand in Kupffer cells. Studies in this aim will utilize a newly developed transgenic mouse to assess the effects of hepatic desmosterol depletion on Kupffer cell transcription both at homeostasis and during NASH. If successful, these studies could identify a new myeloid specific molecular target for the treatment or prevention of NASH.