K. Gee1, M. Isani1, K. Maselli1, T. Grikscheit1 1Children’s Hospital Los Angeles,Surgery,Los Angeles, CA, USA
Introduction: Intestinal failure associated liver disease (IFALD) is associated with short bowel syndrome (SBS). The pathogenesis may include bile acid dysmetabolism linked to changes in the microbiome. We previously showed that prolonged antibiotic administration with intestinal resection durably alters the intestinal microbiome in neonates. We hypothesized that administration of broad spectrum antibiotics might reduce intestinal adaptation and affect liver steatosis and components of IFALD in a SBS zebrafish model.
Method: Adult male zebrafish comprised five groups: starvation(n=25), sham laparotomy(n=29), SBS(n=29), sham+ antibiotics(n=37), and SBS+ antibiotics(n=29). Sham fish had a laparotomy only. SBS surgery consisted of laparotomy with distal intestinal ligation and proximal stoma creation. Antibiotic groups received 100μg/ml ampicillin, 5μg/ml kanamycin, and 250 ng/ml amphotericin B every 3 days. Weekly weights were recorded until harvest at 2 weeks. Villus height, epithelial perimeter, intestinal circumference, and liver steatosis droplets per high power field were measured on H&E slides. BrdU injection prior to harvest followed by immunofluorescent detection assessed proliferating cells. cyp7a1, fxr, and tnfa expression were determined by RT-PCR.
Results: SBS+ antibiotics fish lost a significant percentage of initial weight compared to sham+ antibiotics at 14 days (95% ± 3% v 73.3% ± 2.7%, p<0.0001). There was a decrease in villus height (279.2μm ± 50.8μm v 175μm ± 31.8μm, p=0.0016) and epithelial perimeter (589.8μm ± 155.9μm v 374.1μm ± 79μm, p=0.0005) between SBS and SBS+ antibiotics fish. There was no significant change in intestinal circumference between SBS groups. Additionally, SBS fish had significantly more steatosis in their liver compared to all other cohorts (p<0.0001). BrdU+ cells/villi were increased in SBS fish compared to sham (8.9 ± 3.7 v 2 ± 0.7, p=0.02) as well as SBS + antibiotics fish vs sham+ antibiotics (9.59 ± 2.86 v 1.95 ± 1.8, p=0.02). On RT-PCR analysis, there was increased gene expression of hepatic cyp7a1 (p=0.01) and decreased gene expression of intestinal fxr in SBS fish when compared to SBS+ antibiotic fish (p=0.03). SBS fish had increased expression of intestinal tnfa when compared to sham fish (p=0.04).
Conclusion: Administration of broad spectrum antibiotics to SBS zebrafish significantly reduces villus height and epithelial perimeter, classic indicators of successful intestinal adaptation, when compared to SBS fish that were not exposed to antibiotics. There was increased expression of hepatic cyp7a1 and decreased expression of intestinal fxr accompanied by liver steatosis when compared to SBS + antibiotics fish. While avoidance of IFALD is critical in patients with SBS, additional research to manipulate the FXR pathway without dysregulation of the microbiome is needed.