Er, not each significant distinction on graphs is marked with an asterisk.RESULTSIn earlier perform, we observed that EPEC infection resulted inside the release of adenosine in to the intestinal lumen at concentrations as much as 30 to 40 M in rabbits (11). We wondered if adenosine was the end in the line metabolically or if nucleoside catabolism continued additional with production of inosine or other metabolites (Fig. 1). Although wanting to create assays for inosine, we were surprised to discover that higher levels of uric acid had been readily detectable inside the supernatants of cultured cells infected with EPEC (Fig. 2). Figure 2A shows that in response to infection with EPEC E2348/69, uric acid levels in the supernatant medium rose to 200 M concentrations within six h of infection. The plasmidcured derivative of that strain, JPN15, that is defective in adherence and ability to inflict host cell harm, showed considerably less uric acid release, as did the normal commensal E. coli strain HS. Figure 2B shows that the uric acid release enhanced with escalating multiplicity of infection (MOI) for EPEC (strain JCP88) as well as for Salmonella enterica serotype Enteritidis. E. coli HS failed to release significantly uric acid even at high MOIs. We also observed that infection with Aeromonas hydrophila or treatment of cells with a. hydrophila culture supernatants containing aerolysin also released uric acid from host cells at levels related to these for S. enterica (data not shown). A. hydrophila was tested mainly because Fujii et al. had previously shown that this pathogen also releases ATP from host cells (12). Figure 2C shows that the EPEC-induced uric acid release was inhibited by allopurinol, displaying that the catalytic activity of xanthine oxidase was involved within the uric acid generation. OxypurinolFIG two Release of uric acid into supernatant medium of cultured T84 cells in response to EPEC infection. (A) Comparison of uric acid release amongst commensal E. coli strain HS, wild-type EPEC strain E2348/69, and also the plasmid-cured derivative of E2348/69, JPN15. *, significantly greater than JPN15. (B) Impact of increasing multiplicity of infection (MOI) on uric acid released from cultured T84 monolayers by infection with Salmonella enterica serotype Enteritidis, EPEC JCP88, and E.250674-51-2 web coli HS.Buy5-Fluoro-2-iodobenzoic acid methyl ester (C) Impact of the xanthine oxidase inhibitors allopurinol and oxypurinol on EPEC-induced uric acid release from T84 cells. *, considerably enhanced when compared with the uninfected control; **, drastically decreased compared to JCP88 with out allopurinol. uninf, uninfected.also inhibited uric acid with potency related to that of allopurinol (data not shown). To further pursue the findings shown in Fig.PMID:24635174 two, we assayed uric acid levels in the intestinal loop fluid recovered from rabbits infected with EPEC. In this model, 10-cm loops of ileum are ligated and then infected by injection of bacteria directly into the loop. AApril 2013 Volume 81 Numberiai.asm.orgCrane et al.FIG three Release of uric acid and xanthine oxidase activity into intestinal loopfluids and serum immediately after infection of rabbits with EPEC E22 or rabbit STEC E22-stx2. (A) Comparison with the uric acid contents of uninfected and EPEC E22-infected ligated rabbit intestinal loops after a 20-h infection. Each and every line segment represents the uric acid from an uninfected and an infected intestinal loop fluid in the identical animal. (B) Enhance in serum uric acid in nonsurgically altered rabbits infected orally with strain E22 for 7 days compared with levels.
