An organoid model to study the effect of bacterial metabolites on the intestinal epithelium
Authors: Pearce Sarah C. , Ferguson Nicholas J. , Karl James P. , Arcidiacono Steven M. , Soares Jason W. , Breault David T. , Racicot Kenneth 
Published: June 2021

Short‐chain fatty acids (SCFA) are bacterialmetabolites produced in the large intestine (LI), and are thought to bebeneficial to the host. Of high interest is butyrate (BUT) which is the mainfuel source for colonocytes. Effects of BUT on intestinal physiology have been studied in several in vitro models but data in primary cells, including intestinal stem cells (ISCs) and theirdistinct progenies is still not fully characterized. Recent advances in technology which allow for culturing of primary intestinal cells (organoids) that recapitulate epithelial lineage development, have allowed these mechanisms to be studied in a more physiologically relevant model. To date only a few studies have utilized organoids to study host response to BUT. Therefore, our objective was to examine effects of BUT on biomarkers of ISC activity, differentiation, barrier function and epithelial defense in small intestinal (SI) and LI organoids. We utilized murine colonic (colonoids) and duodenal (enteroids) organoids to test the effects of 5 mM BUT compared to untreated controls. In colonoids and enteroids, gene expression of intestinal stem cell marker, Lgr5decreased 7‐fold and 16‐fold, respectively due to BUT treatment (P < 0.05). Proliferating cells, labeled by Edu were also significantly decreased indicating decreased ISC proliferation. BUT treatment significantly decreased enter ocyte biomarker Alpi in both colonoids (1.7‐fold) and enteroids (>20‐fold). Secretory mucins Muc2and Muc3 were significantly decreased in both types of organoids (P < 0.05) indicating that butyrate did not increase mucus secretion. However, membrane‐bound Muc1 was increased in both organoid types. Interestingly, tuft cells which are involved in chemosensing and immune response were upregulated 5‐fold due to BUT in colonoids, and ~2‐fold in enteroids (Dckl1; P < 0.05). Hormone secreting enteroendocrine cell marker Chga was decreased 1.6‐fold due to BUT treatment in colonoids but was significantly elevated (1.7‐fold) due to BUT treatment in enteroids indicating a potential regional difference between SI and LI. Gene expression of tight junction (TJ) markers occludin (Ocln),and claudin‐3 (Cldn3) were significantly decreased by BUT treatment in both colonoids and enteroids possibly indicating disruption of the TJ (P< 0.05). Microbial defense was also affected as Defb1, anepithelial anti‐microbial peptide was decreased due to SCFA treatment incolonoids, however was increased two‐fold in enteroids (P < 0.05). Another anti‐microbialpeptide, Reg3b was significantly elevated 2‐fold and 30‐fold in colonoids and enteroids treated with BUT, respectively. In conclusion, BUT causes a decrease in stem cell proliferation and consequently alters proportions of progeny cell types. It also may alterepithelial barrier integrity. Additionally, anti‐microbial defense may be enhanced by BUT. These results support previous organoid data and add new insight to the effects of BUT on intestinal cell physiology in SI and LI.Support or Funding InformationOrganoids were kindly provided by Dr. David B reault. Funding was provided by the Department of Defense Combat Feeding Research and Engineering Program through project “Gut Microbiome and Performance Nutrition (TB 15–19)”.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.