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Principal Investigator: Lucy Mary Golden
Organization: UNIVERSITY OF SOUTHERN CALIFORNIA
Fiscal Year: 2022
Award: $371,250
Funding agency: National Institute of Diabetes and Digestive and Kidney Diseases
PROJECT SUMMARY /ABSTRACT
Non-alcoholic fatty liver disease (NAFLD) is the most common cause of chronic liver disease, affecting at least
a quarter of the world’s population, and therefore highly relevant to the NIH mission. From steatosis to fibrosis,
hepatic inflammation drives disease progression, and this study will elucidate the central roles of cholesterol.
The lack of evidence-based, FDA-approved treatment options for NASH underscores the need for further
research into understanding disease pathogenesis and identifying potential targets. Our compelling preliminary
and recently-published findings demonstrate innate immune cells (neutrophils and macrophages) are central to
the pathogenesis and progression of NAFLD. The overarching hypothesis of this proposal is that dysregulation
of innate immunity and inflammation is a dominant, indispensable feature of NAFLD, largely mediated by
cholesterol. We will test this hypothesis through completion of three Specific Aims performed in parallel using
murine NAFLD models and samples from patients with varying severity of NASH. In SA1, the PI will
comprehensively characterize fresh peripheral neutrophils from patients with different stages of NASH. This will
include NETosis, how cholesterol-induced NETs license responses from macrophages and differential effect of
neutrophils on liver endothelial cells. Murine models will focus on targeting of NETs to attenuate hepatic injury
in NASH. We will use the FPC diet, rich in fructose, palmitate, trans-fat, and cholesterol (1.2% by wt), which
has been shown to induce hepatic fibrosis-associated parameters within 16 weeks. SA2 will use live mice fed
different FPC-based diets with isolation of hepatic tissue by ultrasound-guided fine-needle aspiration, followed
by single-cell RNA sequencing (scRNA-seq) to measure genome-wide expression of macrophages from
progressing or regressing diet-induced NASH. Our data demonstrate that withdrawal of high cholesterol in the
diet downregulates macrophage-specific transcription as early as 4 weeks. The proposed experimental plan will
provide unprecedented insights into how dietary cholesterol affects different hepatic macrophage populations.
We will also characterize the functional consequences of different hepatic macrophages on co-cultured hepatic
stellate cells (HSCs). Moreover, we will use targeted sequencing of promoter regions in macrophage genes in
order to define key adaptive changes in epigenetic regulation. Finally, in SA3, the PI will focus on integrating
these robust analyses and utilizing Cre-Lox system to generate mice with myeloid-specific targeted mutants of
genes. As we have found that osteopontin (OPN)-encoding SPP1 expression was strongly induced in
macrophages in NASH, we will test an OPN nanoparticle in prevention and reversal studies of diet-induced
NASH. Mice carrying the Col1a1-GFP transgene will be used to directly assess the effect of different therapeutic
strategies on activation, differentiation, and fibrogenic responses of HSCs. Taken together, our multidisciplinary
approach and outstanding synergism of experts has the potential to fundamentally change our understanding of
how cholesterol shapes and dysregulates innate immunity in NASH and identify novel treatments.
Terms: <2aR phosphoprotein I><2ar peptide><Adipose tissue><Affect><Animal Model><Animal Models and Related Studies><Assay><Attenuated><Bioassay><Biologic Assays><Biological Assay><Blood Neutrophil><Blood Polymorphonuclear Neutrophil><Blood monocyte><Cell Body><Cells><Characteristics><Cholesterol><Chromatin><Cirrhosis><Cre Lox technology><Cre LoxP system><Cre lox recombination><Cre lox recombination system><Cre lox system><Cre recombinase/LoxP technology><Cre system><Crystallization><Cues><Data><Development><Diet><Dietary Cholesterol><Dietary Fats><Disease><Disease Progression><Disease model><Disorder><Endothelial Cells><Enzyme Gene><Enzymes><Epigenetic><Epigenetic Change><Epigenetic Mechanism><Epigenetic Process><Eta-1 protein><Eta-1-Op protein><FDA approved><FNA><Fatty Tissue><Fibrosis><Fine Needle Aspirate><Fine needle aspiration biopsy><Fine-Needle Aspiration><Fructose><Gene Expression><Gene Expression Monitoring><Gene Expression Pattern Analysis><Gene Expression Profiling><Gene Transcription><Genes><Genetic><Genetic Transcription><Goals><Hepatic><Hepatic Stellate Cell><Hepatic Tissue><Hepatic Transplantation><Hepatocarcinoma><Hepatocellular Carcinoma><Hepatocellular cancer><Hepatoma><Hexadecanoates><Histologic><Histologically><Human><Immune><Immunes><Inflammation><Inflammatory><Inflammatory Response><Injury><Injury to Liver><Innate Immune Response><Innate Immunity><Ito Cell><Kinetics><Levulose><Licensing><Lipids><Liver><Liver Cells Carcinoma><Liver Fibrosis><Liver Grafting><Liver Transplant><Marrow Neutrophil><Marrow monocyte><Measures><Mediating><Mice><Mice Mammals><Mission><Modern Man><Modernization><Murine><Mus><Myelogenous><Myeloid><Mφ><NAFLD><NASH><NIH><Nanotechnology><National Institutes of Health><Native Immunity><Natural History><Natural Immunity><Neutralization Tests><Neutrophilic Granulocyte><Neutrophilic Leukocyte><Non obese><Non-Specific Immunity><Nonobese><Nonspecific Immunity><OxLDL><Palmitates><Paper><Pathogenesis><Pathogenicity><Pathology><Patients><Pattern><Peripheral><Pharmacology><Phenotype><Polymorphonuclear Cell><Polymorphonuclear Leukocytes><Polymorphonuclear Neutrophils><Population><Prevention><Primary carcinoma of the liver cells><Process><Promoter Regions><Promotor Regions><Property><Protein-arginine deiminase><Publishing><RNA Expression><Research><Risk Factors><Rodent><Rodentia><Rodents Mammals><Role><Sampling><Severities><Shapes><Societies><Sterility><Stimulus><Testing><Therapeutic><Trans Fats><Transcript Expression Analyses><Transcript Expression Analysis><Transcription><Transgenes><United States National Institutes of Health><Withdrawal><adipose><antibody neutralization test><base><bisulfite><bone sialoprotein 1><bone sialoprotein I><chronic hepatic disease><chronic hepatic disorder><chronic liver disease><chronic liver disorder><cirrhotic><cohort><cytokine><developmental><dietary lipid><dietary manipulation><diets><disorder model><early T-lympocyte activation-1 protein><epigenetic regulation><evidence base><fibrotic liver><gene expression analysis><gene expression assay><genetic approach><genetic promoter element><genetic promoter sequence><genetic strategy><genome scale><genome-wide><genomewide><hepatic body system><hepatic damage><hepatic fibrosis><hepatic inflammation><hepatic injury><hepatic organ system><hydrogen sulfite><hydrosulfite><inflamed liver><injuries><insight><interdisciplinary approach><liver carcinoma><liver damage><liver inflammation><liver injury><liver transplantation><macrophage><model of animal><model organism><monocyte><mouse model><multidisciplinary approach><murine model><mutant><nano particle><nano tech><nano technology><nano-sized particle><nano-technological><nanoparticle><nanosized particle><nanotech><nanotechnological><neutrophil><new therapeutic approach><new therapeutic intervention><new therapeutic strategies><new therapy approaches><non-alcohol fatty liver disease><non-alcohol induced steatohepatitis><non-alcoholic fatty liver disease><non-alcoholic liver disease><non-alcoholic steato-hepatitis><non-alcoholic steatohepatitis><nonalcoholic fatty liver disease><nonalcoholic steato-hepatitis><nonalcoholic steatohepatitis><novel><novel therapeutic approach><novel therapeutic intervention><novel therapeutic strategies><novel therapy approach><osteopontin><ox-LDL><oxidized LDL><oxidized low density lipoprotein><peptidylarginine deiminase><prevent><preventing><promoter sequence><protein-L-arginine iminohydrolase><response><scRNA-seq><secreted phosphoprotein 1><sialoprotein 1><single cell RNA-seq><single cell RNAseq><single cell expression profiling><single cell transcriptomic profiling><single-cell RNA sequencing><social role><sterile><synergism><targeted drug therapy><targeted drug treatments><targeted sequencing><targeted therapeutic><targeted therapeutic agents><targeted therapy><targeted treatment><therapeutic target><transcriptional profiling><ultrasound><white adipose tissue><yellow adipose tissue>