# Project 3 - Human colonoids as a model for the pathobiology of EHEC

> **NIH NIH P01** · UNIVERSITY OF MARYLAND BALTIMORE · 2020 · $261,109

## Abstract

Abstract
The proposed studies will advance the understanding of enteric disease caused by food-borne, Shiga-toxin-
producing enterohemorrhagic E. coli (EHEC), whichis the major cause of life-threatening hemorrhagic colitis
and hemolytic uremic syndrome in the US. While Shiga toxin 2a is the main virulence factor for intestinal and
extra-intestinal disease manifestations, clinical data strongly suggest that Stx-toxemia occurs early in illness
and is short-lived. Thus, our studies will focus on early EHEC-host interactions, particularly the complex
interaction between EHEC and the human colonocyte, goblet cells and immune cells, including macrophages
and neutrophils. Our exciting preliminary data are gained using a novel model of EHEC infection, termed
human colonoid monolayers (HCM), which are primary colonic epithelial cultures derived from adult stem cells
isolated from colonic biopsies of healthy donors. Upon differentiation, HCM are composed of all major types of
epithelial cells, including colonocyte, goblet and enteroendocrine cells. They develop mature microvilli and
produce a thick layer of mucus similar to that which is normally present in the human colon. Using these HCM
we are demonstrating here that EHEC infection results in the destruction of the mucus layer allowing EHEC to
gain access to the colonocyte surface. The serine protease EspP plays a role in EHEC colonization and also
induces the ion secretion, indicating its role in EHEC-induced watery diarrhea. These data suggest that human
colonoids recapitulate EHEC infection and provide unique insights into pathogenesis that differ from other
studies performed in human colon cancer cell lines, allowing improved appreciation of the roles of virulence
factors and assessment of novel therapeutic targets. Relevant to this pathogen, we will test the hypothesis that
HCM, which uniquely represent ex vivo human colonic epithelium, recapitulate human EHEC infection and
provide unique insights in pathogenesis. To test this hypothesis and enhance our understanding of EHEC
intestinal pathogenesis, we propose the following Aims: 1. Determine the role of serine protease EspP in
EHEC colonization of human colonic epithelium and virulence development. 2. Determine the molecular
mechanisms of EHEC induced watery diarrhea. 3. Determine the role of macrophages and neutrophils in
EHEC-induced immune response. The results gained from the proposed experiments will further elucidate the
molecular mechanisms for EHEC interactions with human colonic epithelium, test the role of EspP serine
protease in EHEC colonization and development of watery diarrhea as well as establish the role of neutrophil
and macrophages in EHEC clearance.

## Key facts

- **NIH application ID:** 9982183
- **Project number:** 5P01AI125181-05
- **Recipient organization:** UNIVERSITY OF MARYLAND BALTIMORE
- **Principal Investigator:** JAMES B KAPER
- **Activity code:** P01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $261,109
- **Award type:** 5
- **Project period:** — → 2021-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9982183

## Citation

> US National Institutes of Health, RePORTER application 9982183, Project 3 - Human colonoids as a model for the pathobiology of EHEC (5P01AI125181-05). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/9982183. Licensed CC0.

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