MO679EFFECTS OF ALANYL-GLUTAMINE SUPPLEMENTED PD FLUID ON THE PLASMA METABOLOME AND GUT MICROBIOME IN EXPERIMENTAL PD*

Abstract

Abstract Background and Aims Peritoneal dialysis (PD) is associated with morphological and also functional changes to the peritoneum limiting the long-term use. PD and CKD lead to vasculopathy, disrupt the endothelial and peritoneal barrier, but also influence the gut microbiome. Microbial dysbiosis in return is speculated to drive inflammation as an additional risk factor for cardiovascular disease. New PD-fluids that could slow or prevent these processes are needed. Alanyl-glutamine (AlaGln) has immunomodulatory and cytoprotective properties and has been shown to also improve endothelial barrier functions. Our aim is to investigate the effects of AlaGln-supplementation to PD-fluid on the gut microbiome and plasma and effluent metabolome and their interplay. Method Mice (C57/BI6N) underwent a subtotal nephrectomy (5/6 nephrectomy) to induce uraemia. Chronic exposure to PD-fluid was performed for 9 weeks via subcutaneously implanted peritoneal catheters. Mice were exposed daily to 2 ml of commercially available glucose-based PD-fluid (3.86% glucose) without or with the addition of 8 mM AlaGln. Uremic and healthy mice, kept in parallel were used as controls. All mice were fed standard chow and tap water ad libitum. On the last day, blood and an effluent (after a 30 minute dwell) were collected and faecal matter was collected from 3 different sites of the gut (ileum, caecum and colon). The plasma and effluent metabolome were analysed using a targeted approach. 180 metabolites were analysed with a mass spectrometry based kit (Biocrates) in both samples types. Following microbial DNA isolation, the microbiome has been analysed by 16S rRNA sequencing. The experiment was approved by the local animal ethics committee. Results Significantly elevated creatinine values in mice following 5/6 nephrectomy confirmed their uremic status. Significantly different plasma and effluent levels of alanine and glutamine were found in mice exposed to AlaGln supplemented PD-fluid. A correlation analysis of the plasma revealed the uremic status of the mice as main driver of differences whereas the effluent metabolome was mainly changed by PD fluid exposure. Plasma of uremic mice also showed significantly increased levels of a toxic non-proteinogenic amino acid symmetric dimethyl arginine (SDMA) and citrulline. Microbiome analysis yielded over 2800 amplicon sequence variants. Microbiome composition was location specific and influenced by the different treatments. The microbiome data were also correlated with over 130 metabolites in both plasma and PD effluent. Our data showed increased abundance in bacterial family Pseudomonadacea in caecum of uremic mice and positive correlation with the plasma level of trans-tetra-hydroxyproline. Mice exposed to conventional PD fluid had higher abundance of the bacterial class Clostridia in the colon compared to mice with no PD exposition. Conclusion CKD itself and PD-fluid exposure both affect the gut microbiome and the AlaGln-supplementation effects are likely reflected at the level of microbial diversity and functional interaction with metabolites. As next step specific cellular and molecular mechanisms of these effects are analysed. Preservation of mesothelial and also endothelial cellular barrier function could be a clinically important benefit for patients treated with PD, by preventing increased PD-associated pathomechanisms.