Visium spatial transcriptomics and proteomics identifies novel hepatic cell populations and transcriptomic signatures of alcohol-associated hepatitis.

Background: Alcohol-associated hepatitis (AH) is the clinical manifestation of alcohol-associated liver disease (ALD). AH is a complex disease encompassing the dysregulation of many cells and cell subpopulations. This study used a hepatic spatial transcriptomic and proteomic approach (10X Genomics Visium) to identify hepatic cell populations and their associated transcriptomic and proteomic alterations in human AH.

Methods: Formalin-fixed paraffin-embedded liver tissue from AH patients (n = 2) and non-ALD controls (donors) (n = 2) were used for Visium spatial transcriptomic and proteomic analysis.

Results: AH cell clusters and cell markers were drastically different in regard to tissue pattern and number of cell types compared to non-ALD controls. Cholangiocytes, endothelial cells, macrophages, and stellate cells were more profuse in AH relative to non-ALD controls. Transcriptionally, proliferating cell nuclear antigen-positive (PCNA⁺) hepatocytes in AH more closely resembled cholangiocytes suggesting they were non-functional hepatocytes derived from cholangiocytes. Furthermore, mitochondria protein-coding genes were reduced in AH versus non-ALD control hepatocytes, suggesting reduced functionality and loss of regenerative mechanisms. Macrophages in AH exhibited elevated gene expression involved in exosomes as compared to non-ALD controls. The most upregulated macrophage genes observed in AH were those involved in exosome trafficking. Gene and protein signatures of disease-associated hepatocytes (ANXA2⁺/CXCL1⁺/CEACAM8⁺) were elevated in AH and could visually identify a pre-malignant lesion.

Conclusions: This study identified global cell type alterations in AH and distinct transcriptomic changes between AH and non-ALD controls. These findings characterize cellular plasticity and profuse transcriptomic and proteomic changes that are apparent in AH and contribute to the identification of novel therapeutics.