Spatial analysis of Alcoholic Hepatitis Liver Tissue

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SUMMARY Excessive alcohol consumption is a major global public health issue, with alcoholic liver disease (ALD) being a leading cause of liver-related death in the United States. Alcoholic hepatitis (AH) is a particularly serious form of ALD with high short term mortality rates, ~35% after one month. Although anti-inflammatory steroid treatments and other general therapies exist, patient response to such therapies varies widely. There is a need to understand both the mechanisms of AH disease onset and progression as well as identify markers of treatment efficacy. To this end, we aim to apply more precise analytical methods to move beyond gross tissue analytics and towards more cell specific and spatial protein abundances. Specifically, by more precisely analyzing the spectrum of AH liver tissue, we will gain a better understanding of how hepatocytes progress to apoptotic ballooning and the effect of these cells on adjacent hepatocytes particularly in moderate or confounded AH cases. We have developed a breakthrough technology, termed nanoPOTS (Nanodroplet Preparation in One pot for Trace Samples) which currently enables effective analysis of <10 mammalian cells with extensive proteome coverage. Coupling nanoPOTS with laser capture microdissection (LCM) and downstream high throughput and high sensitivity mass spectrometry analysis platforms now enables the technical capability to capture global protein abundances at the micro scale level. Our objective is to apply nanoPOTS to spatially map with high resolution heterogenous AH biopsy tissues, guided by histologically defined markers of AH. We hypothesize that through the analysis of precise intra-liver protein abundance variations, we will gain greater resolution of protein level mechanisms in the control and dysregulation of the liver in AH. The overall specific aims include 1) Correlate histological events associated with AH pathology, i.e., cell ballooning, steatosis, infiltration, with their overlapping spatially resolved protein abundances utilizing the LCM-nanoPOTS MS platform. 2) Identify spatial protein abundance variations across AH, Alcoholic cirrhotic, and normal liver tissues.