A. Zhang1, P. Lu1, M. Wang1, E. M. O’Hare1, S. D. Brosten1, H. Ding3, C. L. Sears3, J. R. White4, D. J. Hackam1, S. M. Alaish1 1The Johns Hopkins University School Of Medicine,Surgery,Baltimore, MD, USA 3The Johns Hopkins University School Of Medicine,Medicine,Baltimore, MD, USA 4Resphera Biosciences,Baltimore, MD, USA
Introduction: Short bowel syndrome (SBS) is defined as massive small bowel loss resulting in malabsorption and an intestinal dysbiosis, a pathologic alteration in the microbiota. Lipocalin 2 (LCN2) is an antimicrobial protein, which reduces the availability of iron to proinflammatory bacteria and thus acts as a bacteriostatic agent. We hypothesized that the absence of LCN2 in SBS would result in greater relative amounts of pathologic bacteria and worse intestinal adaptation.
Methods: Under an ACUC-approved protocol, we performed 75% small bowel resection (SBR) or sham (SHA) operation on C57BL/6J wild type (WT) and LCN2 knockout (LCN2 -/-) mice. Body weight of the WT and LCN2 -/- mice were measured daily after SBR or sham operation. Intestinal tissue and cecal contents were collected on post-operative day 7 after euthanasia. Fecal DNA were isolated from cecal contents, 16s rRNA gene sequencing were performed and analyzed for all taxonomic groups on WT and LCN2 -/- mice. A 4% fecal slurry created with cecal contents from either WT or LCN2 -/- mice that previously underwent 75% SBR was gavage fed (fecal transplant) into germ-free mice. Body weight of the germ-free mice were measured before fecal transplantation and on post-transplant day 7 before euthanasia. Intestinal barrier integrity was assessed by permeability to FITC-Dextran in the serum following gavage feeding. The length of villi was assessed through histology. Statistical analysis was performed using ANOVA with p<0. 05 considered significant.
Results: A pronounced intestinal dysbiosis was observed in WT mice after 75% SBR, which was evidenced by significant relative increases of pro-inflammatory Proteobacteria and deceases of healthy Firmicutes and Bacteroidetes. Contrary to our hypothesis, LCN2-/- SBR mice had less pro-inflammatory Proteobacteria, and normal amounts of healthy Firmicutes and Bacteroidetes. Furthermore, LCN2-/- SBR mice had lower intestinal permeability and greater intestinal adaptation as compared to WT SBR mice. Germ-free mice that received fecal matter from LCN2 -/- SBR mice (GLR) gained 6% of their body weight and had decreased intestinal permeability and longer jejunal villi; whereas, germ-free mice that received fecal matter from WT SBR mice (GWR) lost 3% of their body weight and had increased intestinal permeability and shorter villi by comparison.
Conclusion:The presence of LCN2 results in a more pathologic intestinal dysbiosis in our mouse model of SBS. This altered microbiome leads to increased intestinal permeability and reduced adaptation. Inhibition of LCN2 may be a novel therapeutic target to improve intestinal adaptation and enteral tolerance in SBS patients.
