ABSTRACT
Chronic ethanol exposure is associated with gut dysbiosis, impaired immunity, and the development of organ
dysfunction leading to a rising appreciation for inter-organ crosstalk for ethanol-induced pathologies. First hit by
ethanol ingestion, the intestine and gut microbiome play a central role in immune system homeostasis. Chronic
ethanol decreases gut bacterial abundance and diversity and destabilizes the intestinal epithelial barrier, which
is associated with reduced luminal butyrate and intestinal all-trans retinoic acid (atRA) levels, systemic endotoxin,
and inflammation. Intestinal microvascular endothelial cells (EC), located within the intestinal lamina propria, are
critical for mucosal immune function by recruiting and activating leukocytes and regulating gut vascular perfusion.
Following ethanol-induced epithelial barrier disruption and endotoxin translocation, proinflammatory molecules
interact with intestinal microvascular EC and immune cells within the lamina propria. Endothelial and immune
cell activation by bacterial endotoxins leads to their dysfunction. The intestinal microvascular endothelial barrier
serves as second defensive barrier for a disrupted epithelium, providing additional protection against
macromolecule and microbe translocation. Butyrate and atRA are intestinal-generated immune nutrients known
to promote epithelial barrier function, and immune homeostasis via T-lymphocyte regulation. We have shown
butyrate (tributyrin) supplementation mitigates ethanol-induced gut-liver injury by preventing disassembly of
epithelial tight junction proteins, reducing oxidative stress, and promoting expression of intestinal microvascular
endothelial associated junctional proteins and immune cells in mouse proximal colon. Here we propose the
hypothesis that through ethanol’s disruption of the gut microbiome, depletion of butyrate and all-trans retinoic
acid, and subsequent destabilization of the intestinal epithelial barrier and translocation of endotoxin, intestinal
microvascular endothelium become functionally disrupted and activated which induces intestinal T-cell
dysregulation and inflammation. We will test two specific aims. Aim 1: Test the hypothesis that ethanol-induced
gut dysbiosis, resultant butyrate depletion and epithelial barrier destabilization disrupts intestinal microvascular
endothelial junctional protein integrity. Using both in vivo and ex vivo approaches, we will study how butyrate
mitigates the effects of ethanol on endothelium disassembly of barrier junctional proteins. Aim 2: Test the
hypothesis that ethanol activates and induces alterations in T-cell homing and Treg expansion within the
intestinal lamina propria which causes intestinal microvascular endothelial dysregulation in a butyrate and atRA
dependent manner. We will utilize ex vivo co-culture system, and in vivo wild-type and Foxp3DTR mice to test for
ethanol-induced activation of intestinal microvascular EC, T-cell homing receptors, and Treg expansion and
cytokine responses in lamina propria and splenocytes. Importantly, intestinal microvascular EC has not been
studied in the context of ethanol, butyrate and atRA, and therefore new mechanistic insights may bring novel
therapeutics to protect against ethanol’s disruption in intestinal homeostasis.
