01.14 Enteral Antibiotics Stimulate Ordered Small Intestinal Mucosal Growth in Mice

M. P. Shaughnessy1, C. J. Park1, L. H. McCarthy1, N. A. Barry2, A. L. Goodman2, R. A. Cowles1  1Yale University School Of Medicine,Pediatric Surgery,New Haven, CT, USA 2Yale University School Of Medicine,Microbial Pathogenesis And Microbial Sciences Institute,New Haven, CT, USA

Introduction:  Previous work demonstrated enhanced enterocyte proliferation and mucosal growth in gnotobiotic mice colonized with a limited microbiome, supporting the notion that the intestinal flora participates in mucosal homeostasis. Prior studies have also shown that treatment with broad spectrum enteral antibiotics results in near germ-free (GF) conditions in mice with conventional flora (CF). We hypothesized that limiting the intestinal microbiome with enteral antibiotics would result in ordered small intestinal mucosal growth in CF mice but similar treatment would have no effect in GF mice lacking a microbiome.

Methods:  C57BL/6J CF mice and C57BL/6J GF mice were allowed ad libitum access to either an antibiotic solution (Ampicillin, Ciprofloxacin, Metronidazole, Vancomycin, Meropenem) mixed in artificial sweetener (n= 3CF, 6GF) or artificial sweetener alone (n=3CF, 6GF). After two weeks, segments from the proximal, middle, and distal small intestine were harvested, fixed, sectioned and stained with H&E. Villus height (VH) was measured and mucosal surface area (MSA) was calculated. Cellular composition of villi and crypts was investigated by standard immunofluorescent staining for chromogranin A and lysozyme. Enterochromaffin cells (ECC), Goblet Cells (GC), and Enterocytes (EC) were counted and calculated as a percentage of total cells per villus, and number of Paneth Cells (PC) per crypt was assessed. Data were analyzed with Student’s t-test and significance assumed for p<0.05.

Results: Antibiotic-treated CF (Abx-CF) mice had taller villi and greater MSA in the proximal, middle, and distal small intestine when compared to vehicle-treated CF mice (p<0.0001 for all regions). Conversely, antibiotic-treated GF (Abx-GF) mice demonstrated no significant increase in VH in any segment of the small intestine when compared to vehicle-treated GF mice. When combined, there was no difference in total mean VH between Abx-GF and vehicle-treated GF controls (p=0.77). Abx-CF mice had an increase in total cells per villus along the entire small bowel (p<0.003). The proportion of EC in villi was unchanged in the middle and distal small intestine (p>0.05), with a small decrease in the proximal small intestine in Abx-CF. The proportion of ECC and GC in villi and PC in crypts was unchanged in Abx-CF mice compared to vehicle-treated CF mice (p>0.05).

Conclusion: Enteral administration of broad-spectrum antibiotics to mice with a conventional microbiome stimulates ordered small intestinal mucosal growth, resulting in taller villi with retention of normal cellular composition. Mucosal growth was not seen in germ-free mice treated with antibiotics, confirming that the microbiome is likely the target for the stimulatory effects of antibiotics on intestinal mucosal growth. The relationship between the microbiome and mucosal growth warrants further study and may provide guidance for development of future therapies for patients with malabsorptive disorders.