Abstract
BackgroundDuring acute pancreatitis (AP), oxidative stress contributes to intestinal barrier failure. We studied actions of multispecies probiotics on barrier dysfunction and oxidative stress in experimental AP.Methodology/Principal FindingsFifty-three male Spraque-Dawley rats were randomly allocated into five groups: 1) controls, non-operated, 2) sham-operated, 3) AP, 4) AP and probiotics and 5) AP and placebo. AP was induced by intraductal glycodeoxycholate infusion and intravenous cerulein (6 h). Daily probiotics or placebo were administered intragastrically, starting five days prior to AP. After cerulein infusion, ileal mucosa was collected for measurements of E. coli K12 and 51Cr-EDTA passage in Ussing chambers. Tight junction proteins were investigated by confocal immunofluorescence imaging. Ileal mucosal apoptosis, lipid peroxidation, and glutathione levels were determined and glutamate-cysteine-ligase activity and expression were quantified. AP-induced barrier dysfunction was characterized by epithelial cell apoptosis and alterations of tight junction proteins (i.e. disruption of occludin and claudin-1 and up-regulation of claudin-2) and correlated with lipid peroxidation (r>0.8). Probiotic pre-treatment diminished the AP-induced increase in E. coli passage (probiotics 57.4±33.5 vs. placebo 223.7±93.7 a.u.; P<0.001), 51Cr-EDTA flux (16.7±10.1 vs. 32.1±10.0 cm/s10−6; P<0.005), apoptosis, lipid peroxidation (0.42±0.13 vs. 1.62±0.53 pmol MDA/mg protein; P<0.001), and prevented tight junction protein disruption. AP-induced decline in glutathione was not only prevented (14.33±1.47 vs. 8.82±1.30 nmol/mg protein, P<0.001), but probiotics even increased mucosal glutathione compared with sham rats (14.33±1.47 vs. 10.70±1.74 nmol/mg protein, P<0.001). Glutamate-cysteine-ligase activity, which is rate-limiting in glutathione biosynthesis, was enhanced in probiotic pre-treated animals (probiotics 2.88±1.21 vs. placebo 1.94±0.55 nmol/min/mg protein; P<0.05) coinciding with an increase in mRNA expression of glutamate-cysteine-ligase catalytic (GCLc) and modifier (GCLm) subunits.ConclusionsProbiotic pre-treatment diminished AP-induced intestinal barrier dysfunction and prevented oxidative stress via mechanisms mainly involving mucosal glutathione biosynthesis.
Highlights
Multi-organ-failure and systemic inflammatory response syndrome (SIRS) remain major causes of mortality at intensive care units [1]
Deitch et al [4] demonstrated for example, that shock-induced intestinal hypoperfusion leads to release of reactive oxygen species (ROS) and oxidative stress resulting in barrier failure and release of pro-inflammatory mediators, enhancing a subsequent SIRS
Five-day pre-treatment with these multispecies probiotics attenuated bacterial translocation and reduced the mortality in experimental acute pancreatitis (AP) in rats [23], but recently we demonstrated in a double-blind clinical study that these probiotics, contrary to any expectations, doubled the mortality compared with placebo in 298 patients with predicted severe AP [24]
Summary
Multi-organ-failure and systemic inflammatory response syndrome (SIRS) remain major causes of mortality at intensive care units [1]. Deitch et al [4] demonstrated for example, that shock-induced intestinal hypoperfusion leads to release of reactive oxygen species (ROS) and oxidative stress resulting in barrier failure and release of pro-inflammatory mediators, enhancing a subsequent SIRS. Evidence suggests that ROS disrupt epithelial tight junctions (TJs) [5,6] leading to barrier dysfunction [7]. ROS cause epithelial cell apoptosis [8] contributing to mucosal barrier failure [9,10,11] and associated mortality [12,13] in experimental studies. Taken together in critically ill patients, SIRS may be driven by an oxidative stress-induced disruption of the equilibrium of the otherwise symbiotic three-way partnership between intestinal microbiota, epithelium, and immune system. We studied actions of multispecies probiotics on barrier dysfunction and oxidative stress in experimental AP
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