Intestinal Alkaline Phosphatase Inhibits the Translocation of Bacteria of Gut-Origin in Mice with Peritonitis: Mechanism of Action

Abstract

Exogenous intestinal alkaline phosphatase (IAP), an enzyme produced endogenously at the brush edge of the intestinal mucosa, may mitigate the increase in aberrant intestinal permeability increased during sepsis. The aim of this study was to test the efficacy of the inhibitory effect of IAP on acute intestinal inflammation and to study the molecular mechanisms underlying IAP in ameliorating intestinal permeability. We used an in vivo imaging method to evaluate disease status and the curative effect of IAP. Two Escherichia coli (E.coli) B21 strains, carrying EGFP labeled enhanced green fluorescent protein (EGFP) and RFP labeled red fluorescent protein (RFP), were constructed as tracer bacteria and were administered orally to C57/B6N mice to generate an injection peritonitis (IP) model. The IP model was established by injecting inflammatory lavage fluid. C57/B6N mice bearing the tracer bacteria were subsequently treated with (IP+IAP group), or without IAP (IP group). IAP was administered to the mice via tail vein injections. The amount of tracer bacteria in the blood, liver, and lungs at 24 h post-injection was analyzed via flow cytometry (FCM), in vivo imaging, and Western blotting. Intestinal barrier function was measured using a flux assay with the macro-molecule fluorescein isothiocyanate dextran, molecular weight 40kD, (FD40). To elucidate the molecular mechanism underlying the effects of IAP, we examined the levels of ERK phosphorylation, and the expression levels of proteins in the ERK-SP1-VEGF and ERK-Cdx-2-Claudin-2 pathways. We observed that IAP inhibited the expression of Claudin-2, a type of cation channel-forming protein, and VEGF, a cytokine that may increase intestinal permeability by reducing the levels of dephosphorylated ERK. In conclusion, exogenous IAP shows a therapeutic effect in an injection peritonitis model. This including inhibition of bacterial translocation. Moreover, we have established an imaging methodology for live-animals can effectively evaluate intestinal permeability and aberrant bacterial translocation in IP models.

Tissue alkaline phosphatase activity and expression in an experimental infant swine model of cardiopulmonary bypass with deep hypothermic circulatory arrest

“…[71] iAP loss is thought to increase susceptibility to injury and worsen intestinal barrier function, both of which are improved with interventions to increase intestinal AP activity. [70][71][72] Contrary to our ndings, in both vascular clamping and colitis, loss of iAP activity was not balanced by an increase in iAP mRNA expression. [54,70] Several possibilities could explain this difference, including severity of injury (prolonged vascular clamping compared to the more translationally relevant DHCA), location of sampling (jejunum vs ileum), and complexity of the injury model and the subsequent tissue response.…”

Tissue alkaline phosphatase activity and expression in an experimental infant swine model of cardiopulmonary bypass with deep hypothermic circulatory arrest

“…Decreased intestinal AP activity has also been shown in models of colitis[64] and necrotizing enterocolitis [65]. iAP loss is thought to increase both susceptibility to injury and worsen intestinal barrier function, both of which are improved with interventions to increase intestinal AP activity [64][65][66]. Contrary to our findings, in both the vascular clamping model and the colitis model, loss of iAP activity was not balanced by an increase in iAP mRNA expression [49,64].…”

Effects of intestinal alkaline phosphatase on intestinal barrier function in a cecal ligation and puncture (CLP)‐induced mouse model for sepsis

“…54 In that study, bacterial translocation could be reduced with respectively 65% and 79% following oral IAP administration in a similar dose, but without the preventive administration. 54 The reason for the discrepancies between the results of these studies and our results is presumably multifactorial. Firstly, these models are characterized by different underlying pathophysiological mechanisms (eg, ischemia-reperfusion and peritonitis), in which the trigger for the sepsis is not continuously present.…”