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Background Probiotic Lactobacillus reuteri DSM 17938 (LR 17938) is beneficial to infants with colic. To understand its mechanism of action, we assessed ultrasonic vocalizations (USV) and brain pain/stress genes in newborn mice exposed to maternal separation stress. Methods Pups were exposed to unpredictable maternal separation (MSU or SEP) or MSU combined with unpredictable maternal stress (MSU + MSUS or S + S), from postnatal days 5 to 14. USV calls and pain/stress/neuroinflammation-related genes in the brain were analyzed. Results We defined 10 different neonatal call patterns, none of which increased after MSU. Stress reduced overall USV calls. Orally feeding LR 17938 also did not change USV calls after MSU. However, LR 17938 markedly increased vocalizations in mice allowed to stay with their dams. Even though LR 17938 did not change MSU-related calls, LR 17938 modulated brain genes related to stress and pain. Up-regulated genes following LR 17938 treatment were opioid peptides, kappa-opioid receptor 1 genes, and CD200, important in anti-inflammatory signaling. LR 17938 down-regulated CCR2 transcripts, a chemokine receptor, in the stressed neonatal brain. Conclusions USV calls in newborn mice are interpreted as "physiological calls" instead of "cries." Feeding LR 17938 after MSU did not change USV calls but modulated cerebral genes favoring pain and stress reduction and anti-inflammatory signaling. Impact We defined mouse ultrasonic vocalization (USV) call patterns in this study, which will be important in guiding future studies in other mouse strains. Newborn mice with maternal separation stress have reduced USVs, compared to newborn mice without stress, indicating USV calls may represent "physiological calling" instead of "crying." Oral feeding of probiotic Lactobacillus reuteri DSM 17938 raised the number of calls when newborn mice continued to suckle on their dams, but not when mice were under stress. The probiotic bacteria had a dampening effect on monocyte activation and on epinephrine and glutamate-related stress gene expression in the mouse brain.

Recovery of the ability to digest and absorb lipids is essential to the maintenance of normal nutrition in infants with bowel damage. Two intrinsic microsomal enzymes, monoacylglycerol acyltransferase (MGAT) and diacylglycerol acyltransferase (DGAT), catalyze the major pathway for intestinal triacylglycerol biosynthesis. This study describes the effects of intestinal ischemia on epithelial DGAT and MGAT activities and their recovery in response to two luminal treatments: L-glutamine (Gln), the primary intestinal fuel, and transforming growth factor-alpha (TGF-alpha), a mitogenic hormone similar to epidermal growth factor present in breast milk. Ischemic damage and recovery were analyzed in mucosa from Thiry-Vella loops in the mid-ileum of 7-wk-old pigs. Loops were subjected to 2-h occlusion of local mesenteric arteries, followed by 6 or 72 h of recovery in the presence of luminal glucose (control), Gln, or TGF-alpha. Ischemic tissue followed by 6-h recovery exhibited an approximate 50% decrease in both MGAT and DGAT activities compared with nonischemic loop tissue. At 72 h, MGAT and DGAT recovery in Gln plus TGF-alpha-treated loops was significantly greater than their corresponding 6-h peak damage levels (p < 0.05). From 6 to 72 h, MGAT increased 4-fold and DGAT increased 3.6-fold after Gln plus TGF-alpha treatment. With other treatments, MGAT and DGAT activities increased

Arginine (Arg) is deficient in the serum of the preterm neonate and is lower in those developing intestinal ischemia. We investigated whether Arg or its precursor, citrulline (Cit), protects intestinal tight junctions (TJs) from hypoxia (HX) and determined whether inducible nitric oxide (NO) plays a role.Neonatal piglet jejunal IPEC-J2 cell monolayers were treated with Arg or Cit, reversible and irreversible NO synthetase (NOS) inhibitors, and were exposed to HX. TJs were assessed by serial measurements of transepithelial electrical resistance (TEER), flux of inulin-fluorescein isothiocyanate, and immunofluorescent staining of TJ proteins.We found that Arg and Cit were protective against HX-related damage. At the final time point (14 h), the mean TEER ratio (TEER as compared with baseline) for Arg + HX and Cit + HX was significantly higher than that for HX alone. Both Arg and Cit were associated with decreased inulin flux across hypoxic monolayers and qualitatively preserved TJ proteins. Irreversible inhibition of NOS blocked this protective effect. Lipid peroxidation assay showed that our model did not produce oxidant injury.Arg and Cit, via a mechanism dependent on NO donation, protected intestinal epithelial integrity.

Cryptosporidium parvum produces a prolonged watery diarrhea unresponsive to conventional antimicrobials. Because of reported efficacy of antibody-based immunotherapy, we studied the effect of inexpensive, commercially available oral bovine serum concentrate (BSC) in experimental cryptosporidiosis. Twenty-four calves were treated with 57 g/d BSC (n = 12) or soy protein (n = 12) added to their standard whey protein-based milk replacer (227 g/2 L twice daily). Of the 24, 9 were also treated with L-glutamine (GLN), 8 g/L (50 mM) in the milk (5 calves in the BSC group and 4 in the soy group). Animals were inoculated with 10(8) cryptosporidium oocysts per os on d 8 of life and received oral rehydration on d 12-14. Eight uninfected controls were treated with BSC or soy protein. Fecal and urine volume and urinary Cr-EDTA excretion were measured. Animals were killed on d 18 of life. Cryptosporidiosis induced severe watery diarrhea lasting9 d and produced a 25% increase in intestinal permeability, a 33% decrease in villous surface area, and a 40% reduction in mucosal lactase specific activity. Glutamine treatment had no effect on the diarrhea or any of the intestinal tests; and therefore pooled data were used to compare the 12 calves treated with BSC with the 12 treated with soy. In animals receiving BSC, peak diarrheal volume and intestinal permeability were reduced 33%, fewer oocysts were shed, intestinal crypts were significantly deeper, and villous surface area returned to normal by 9 d after infection (all por= 0.05). BSC should be studied as a treatment for human cryptosporidiosis.

A randomized, investigator-masked trial determined the effects of oral recombinant human transforming growth factor-alpha (TGF alpha) on jejunal mucosal recovery in 75 piglets with rotavirus diarrhea. Rotavirus inoculation of artificially reared piglets induced subtotal (approximately 50%) villus atrophy and watery diarrhea. Dietary TGF alpha was associated with significant restoration of villus surface area by 4 d postinoculation (p.i.) and complete restoration by 8 d p.i., whereas saline-treated animals required 12 d for recovery. Jejunal segments from clinically recovered TGF alpha-treated piglets showed an increase in electrical resistance across the epithelial barrier in vitro which was proportional to villus height. TGF alpha treatment for 12 d also produced a 30-50% increase in jejunal mucosal mass (protein content and wet weight), compared with the corresponding values in saline-treated piglets and in uninfected controls. However, oral TGF alpha did not hasten the resolution of diarrhea, enhance the specific activities of jejunal mucosal digestive enzymes, or increase jejunal glucose-stimulated Na+ absorption in vitro. We conclude that dietary TGF alpha stimulates jejunal mucosal hypertrophy, improves barrier function, and enhances regrowth of villi in rotavirus enteritis; however, it does not facilitate the restoration of functional activity or mucosal digestive enzymes. Oral TGF alpha can facilitate intestinal epithelial recovery in diseases associated with mucosal damage.

sential nutrients, they do not contain glutamine, the most abundant amino acid in the body, considered by some to be an essential amino acid for preterm infants.1 Not only does GLN function as a major precursor for the endogenous synthesis of arginine by way of citrulline, it also plays a role in nucleotide synthesis and in providing “fuel” to intestinal and muscle cells.2 GLN increases protein synthesis and inhibits protein degradation in skeletal muscle, thereby promoting positive nitrogen balance under catabolic conditions. GLN is also essential for intestinal crypt cell proliferation and enhances the cellular response to growth factors.3

Glutamine Protects Intestinal Cells Via Heat Shock Protein (hsp)-Dependent and Metabolic Effects

We measured the effect of pharmacological doses of glucocorticoid on piglet jejunal structure and function during acute viral diarrhea. Weaned piglets, infected experimentally with transmissible gastroenteritis virus, a coronavirus that induces a diarrheal illness similar to human rotavirus infection, received methylprednisolone (30 mg/kg) or saline intramuscularly at 48 and 72 h after infection; noninfected littermate controls were similarly injected with methylprednisolone. Animals were killed at 96 h, at the height of diarrhea, and jejunal epithelium was studied in vitro. Transmissible gastroenteritis, as expected, induced structural, enzyme, and Na transport abnormalities. Methylprednisolone did not affect small intestinal structure or function of noninfected control piglets. In transmissible gastroenteritis-infected piglets, jejunal villi were longer and glucose-facilitated Na absorption was greater after methylprednisolone than after saline treatment. Increased glucose stimulation of Na flux in vitro in the methylprednisolone-treated infected group was not attributable to enhanced Na+-K+-ATPase activity and occurred despite persistence of the virus within mucosal cells, shown by immunofluorescense microscopy. In this piglet model of viral diarrhea, early regeneration of absorptive surface that precedes recovery of disaccharidase function is accelerated by glucocorticoid therapy.