F. Adiliaghdam1, P. M. Cavallaro1, M. Najibi1, Y. Liu1, L. Rahme1, R. A. Hodin1 1Massachusetts General Hospital,Department Of Surgery/ Harvard Medical School,Boston, MASSACHUSETTS, USA
Introduction: Our previous data has shown that severe burn injury induces gut barrier dysfunction and leads to a gut-derived systemic inflammatory response.We hypothesized that there is a vicious cycle occurring in the gut after acute burn injury which progressively damages the gut barrier and transforms the gut into an inflammatory organ.Furthermore, we investigated how targeting this pro-inflammatory state with supplemental intestinal alkaline phosphatase (IAP) can prevent this vicious cycle.
Methods: Mice were subjected to a 30% total body surface area burn +/- burn site super-infection induced by injection of Pseudomonas aeruginosa into the dermis. IAP was given by gastric gavage.(2000unit)The pro-inflammatory characteristics of intestinal contents was measured in vitro by Caco2 trans-well epithelial electrical resistance (TEER).Primary mouse macrophages were incubated with ileocecal contents to evaluate the immune response.Peritoneal macrophages were evaluated for inflammatory gene expression.
Results: Our data suggest that burn injury leads to an increased expression of pro-inflammatory cytokines such as TNF, IL-6, IL-1B,and lipocalin in the intestine(P<0.01).Burn injury increased peritoneal macrophage pro-inflammatory cytokine expression compared to sham treatment(TNF-a and IL-6, 3-fold increase,P<0.01).Incubation of ileocecal contents of burned mice with Caco2 monolayer negatively affected the monolayer integrity and downregulated the tight junction protein(TJP)expression.There was a significant higher inflammatory response to the ileocecal content of burn-injured mice in both epithelial cells and macrophage cells compared to sham-treated ones(P<0.01).IAP supplementation after burn decreased the intestine inflammation(3-fold decrease,P<0.01).Peritoneal macrophages isolated from IAP-treated mice showed less inflammatory gene expression(P<0.01).Also, IAP treatment was associated with an improved monolayer integrity(3-fold improvement in TEER drop,P<0.05).IAP-treated ileocecal content caused a less inflammatory response in both epithelial cells and primary macrophages(P<0.01).Ex-vivo incubation of burn-injured ileocecal content with IAP decreased the pro-inflammatory characteristics of the contents(3-fold with primary macrophages,4-fold with epithelial cells,P<0.05).Furthermore, IAP treatment improved TJPs expression in in-vitro and in vivo burn models(ZO-1,P<0.05 and Occludin,P<0.01).Interestingly,the Pseudomonas strain(PA14) which was injected subcutaneously into the burn site was detectable in the ileum and stool of the burn infected mice, confirming the existence of a vicious inflammatory cycle in the gut after burn.
Conclusion:Acute burn injury causes an inflammatory response in the gut.IAP supplementation as a natural,anti-inflammatory enzyme significantly decreases gut inflammation and improves gut barrier function,and can represent a novel therapy to prevent a gut-induced systemic inflammation.