br Experimental procedure br Acknowledgments
Acknowledgments This research was supported by funds provided by NIH/NIAAA grant P01 AA020683 and the Waggoner Center for Alcohol and Addiction Research.
Introduction Acute mesenteric ischemia (AMI) is still associated with a high mortality rate, and currently, an early diagnosis is the most effective way to improve the poor outcome.1, 2, 3 A common cause for AMI is an embolization of the mesenteric arterial circulation. In nearly all cases, the thrombus originates from the neomycin sulfate receptor due to either atrial tachyarrhythmia, endocarditis, valvular disorders, or myocardial ischemia.4, 5 Early clinical symptom is a prompt onset of severe abdominal pain. Histologically, already 30 min after arterial occlusion, an atrophy of the villi due to the anoxic state occurs. Subsequently, damage of Tunica mucosa and Tunica serosa can be observed. Reperfusion causes a systemic inflammatory response associated with activation of immune cells, endothelial cells, coagulation system, and thus could end up in multiple organ failure that leads to death.7, 8 In previous studies, our group identified glycine as a potent prophylactic agent against injury of the small intestine after ischemia-reperfusion (I/R) in rats, whereas the protective mechanism remained unclear. Since even a low dosage of 10 mg/kg body weight (BW) significantly diminished intestinal injury, a metabolic action was excluded as the responsible driving force. Indeed, glycine attenuated hypoxic injury in isolated hepatocytes by preventing an increase of intracellular sodium-level in vitro.10, 11 Unfortunately, much higher dosages were necessary, which might indicate a different, probably in vitro–limited, mechanism. In contrast, a beneficial effect via the pentameric glycine receptor (GlyR) may play a major role in ameliorating I/R-injury in vivo. Since this receptor is not exclusively present in the central nervous system but can be found additionally on immune cells, endothelial and tubular renal cells,12, 13, 14 glycine offers largely underestimated immunomodulatory properties by affecting the activity of macrophages on releasing proinflammatory cytokines.13, 15 Provided that this presumption is true, nearly identical capabilities should be evident for the simple amino acid β-alanine and taurine, which act as agonists of the GlyR. In the present study, the amino acid β-alanine was applied intravenously, and I/R-injury was induced by occluding and reopening the superior mesenteric artery in rats. Although the half maximal effective concentration (EC50) of glycine to its receptor was around 18 μM, the EC50 of β-alanine to the GlyR constituted 52 μM. Since there is additionally an eminent difference between the physiological plasma concentration of glycine (280 μM) and β-alanine (<5 μM),9, 18 there was some need to search for an optimal therapeutic dosage of the agonist. Therefore, the effect of three different dosages of β-alanine was compared: 10, 30, and 100 mg/kg BW.
Material and methods
Discussion In the present study, β-alanine, an agonist of the glycine receptor (GlyR), improved outcome after intestinal I/R in rats in a similar vein as glycine did. Thus, the evaluated data set very strongly evidenced that the GlyR was involved in the I/R-mediated injury. In this regard, the highest dosage of 100-mg β-alanine diminished tissue damage of the small intestine, as was demonstrated on both macroscopic and histologic level (Fig. 4A and B). Moreover, β-alanine caused an increased activity of saccharase, which deals as a marker for functional brush border (Fig. 4C). Interestingly, the amino acid depleted the accumulation of macrophages in the intestinal wall (Fig. 4D). Since there is evidence that Kupffer cells are involved in the mechanism of reperfusion injury of the liver, an undesired activation of macrophages in the intestine may also play a key role in the noxious mechanism against reperfusion injury. As the GlyR is also present on immune cells included macrophages,12, 13, 14 there should be possibilities available to downregulate the activity of macrophages in the intestine. In fact, via the pentameric receptor, glycine is able to modulate the activity of macrophages and to diminish their release of cytokines. Even this fact confirms the theory of a receptor-dependent effect of both glycine and its agonist β-alanine in intestinal I/R.13, 15