Bifidobacterium longum and Lactobacillus helveticus Synergistically Suppress Stress-related Visceral Hypersensitivity Through Hypothalamic-Pituitary-Adrenal Axis Modulation.

BACKGROUND/AIMS: Visceral pain and hypothalamic-pituitary-adrenal axis (HPA) dysregulation is a common characteristic in irritable bowel syndrome (IBS) patients. Previously, we reported that a probiotic formulation (Lactobacillus helveticus R0052 and Bifidobacterium longum R0175) prevents chronic stress-mediated brain function abnormalities by attenuating the HPA axis response. Here, we compared the effect between different probiotic treatments on the perception of visceral pain during colorectal distension (CRD) following a chronic stress and the consequences to the activity of the HPA axis. METHODS: After a 2-week treatment with a combined probiotic formulation, or L. helveticus or B. longum alone in stressed mice, the visceral pain in response to CRD was recorded. The expression of glucocorticoid receptors was determined in the different brain areas involved in the stress response (hypothalamus, hippocampus, and prefrontal cortex). The plasma levels of stress hormones were also measured. RESULTS: A pretreatment using the combination of probiotic formulation significantly reduces the chronic stress-induced visceral hypersensitivity respectively at 0.06, 0.08, and 0.10 mL CRD volume. However, a single probiotic (B. longum or L. helveticus) administration is less effective in reducing visceral pain in stressed mice. Moreover, the expression of the glucocorticoid receptor mRNA was consistently up-regulated in several brain areas after pretreatment with a combined probiotic, which correlated with the normalization of stress response compared to the inconsistent effects of a single probiotic. CONCLUSION: The combination of L. helveticus and B. longum is more effective in regulating glucocorticoid negative feedback on the HPA axis than probiotic alone and subsequently in treating stress-induced visceral pain.

The role of mucoprotectants in the management of gastrointestinal disorders.

INTRODUCTION: The intestinal barrier controls the absorption of nutrients and water whilst helping to prevent the entry of toxins and pathogenic micro-organisms from the lumen into the tissues. Deficiencies in the barrier are associated with various gastrointestinal and extra digestive disorders. Areas covered: This review provides an overview of the relationship between increased intestinal permeability and disease, and considers the role of mucosal protectants (mucoprotectants) in restoring normal intestinal barrier function, with a particular focus on diarrheal disorders. Expert commentary: Impairment of the intestinal barrier characterizes a variety of diseases, and there is ongoing interest in the development of pharmacological approaches targeting the reduction of intestinal permeability. These include corticosteroids, aminosalicylates and anti-tumor necrosis factor-α (TNF-α), which act by reducing inflammation; probiotics, which modulate the production of mucin and epithelial tight junction proteins; and mucoprotectants, which form a protective film over the epithelium. Recently, preclinical and clinical data highlight, the ability of new mucoprotectants, such as gelatin tannate and xyloglucan, to protect the intestinal mucosa and to exert anti-diarrheal effects. In the future the ability of these substances to enhance the intestinal barrier may extend their use in the management of a variety of gastro-intestinal diseases associated with 'leaky gut'.

Gut dysbiosis and impairment of immune system homeostasis in perinatally-exposed mice to Bisphenol A precede obese phenotype development.

Epidemiology evidenced the Bisphenol A (BPA), a chemical found in daily consumer products, as an environmental contributor to obesity and type II diabetes (T2D) in Humans. However, the BPA-mediated effects supporting these metabolic disorders are still unknown. Knowing that obesity and T2D are associated with low-grade inflammation and gut dysbiosis, we performed a longitudinal study in mice to determine the sequential adverse effects of BPA on immune system and intestinal microbiota that could contribute to the development of metabolic disorders. We observed that perinatal exposure to BPA (50 µg/kg body weight/day) induced intestinal and systemic immune imbalances at PND45, through a decrease of Th1/Th17 cell frequencies in the lamina propria concomitant to an increase of splenic Th1/Th17 immune responses. These early effects are associated with an altered glucose sensitivity, a defect of IgA secretion into faeces and a fall of faecal bifidobacteria relative to control mice. Such BPA-mediated events precede infiltration of pro-inflammatory M1 macrophages in gonadal white adipose tissue appearing with ageing, together with a decreased insulin sensitivity and an increased weight gain. Our findings provide a better understanding of the sequential events provoked by perinatal exposure to BPA that could support metabolic disorder development in later life.

Paneth Cell Defects Induce Microbiota Dysbiosis in Mice and Promote Visceral Hypersensitivity.

BACKGROUND & AIMS: Separation of newborn rats from their mothers induces visceral hypersensitivity and impaired epithelial secretory cell lineages when they are adults. Little is known about the mechanisms by which maternal separation causes visceral hypersensitivity or its relationship with defects in epithelial secretory cell lineages. METHODS: We performed studies with C3H/HeN mice separated from their mothers as newborns and mice genetically engineered (Sox9flox/flox-vil-cre on C57BL/6 background) to have deficiencies in Paneth cells. Paneth cell deficiency was assessed by lysozyme staining of ileum tissues and lysozyme activity in fecal samples. When mice were 50 days old, their abdominal response to colorectal distension was assessed by electromyography. Fecal samples were collected and microbiota were analyzed using Gut Low-Density Array quantitative polymerase chain reaction. RESULTS: Mice with maternal separation developed visceral hypersensitivity and defects in Paneth cells, as reported from rats, compared with mice without maternal separation. Sox9flox/flox-vil-Cre mice also had increased visceral hypersensitivity compared with control littermate Sox9flox/flox mice. Fecal samples from mice with maternal separation and from Sox9flox/flox-vil-cre mice had evidence for intestinal dysbiosis of the microbiota, characterized by expansion of Escherichia coli. Daily gavage of conventional C3H/HeN adult mice with 109 commensal E coli induced visceral hypersensitivity. Conversely, daily oral administration of lysozyme prevented expansion of E coli during maternal separation and visceral hypersensitivity. CONCLUSIONS: Mice with defects in Paneth cells (induced by maternal separation or genetically engineered) have intestinal expansion of E coli leading to visceral hypersensitivity. These findings provide evidence that Paneth cell function and intestinal dysbiosis are involved in visceral sensitivity.

Evaluation of reticulated gelatin-hibiscus-propolis against intestinal commensal species commonly associated with urinary tract infections.

AIM: Reticulated gelatin (RG), hibiscus and propolis (RGHP) is a medical device that can reduce the bacterial adherence to epithelial cultured cells and invasion by enteropathogens, thus gathering relevant properties to decrease the risk of urinary tract infections (UTIs). We aimed at evaluating in Wistar rats the efficacy of RGHP, RG and vehicle against intestinal commensals commonly involved in UTIs. METHODS: Animals received orally (with supplemental Na2CO3): RGHP 1540 mg/day/rat; RG 500 mg/day/rat or vehicle. RESULTS: RGHP significantly reduced fecal Escherichia coli and Enterococcus spp. levels without affecting other targeted Enterobacteriaceae. The antagonistic property of RGHP was confirmed in streptomycin-pretreated rats highly colonized with a human commensal E. coli strain with uropathogenic potential. CONCLUSION: RGHP may decrease the risk of UTIs by reducing colonization by opportunistic uropathogens.

Perinatal exposure to a low dose of bisphenol A impaired systemic cellular immune response and predisposes young rats to intestinal parasitic infection.

Perinatal exposure to the food contaminant bisphenol A (BPA) in rats induces long lasting adverse effects on intestinal immune homeostasis. This study was aimed at examining the immune response to dietary antigens and the clearance of parasites in young rats at the end of perinatal exposure to a low dose of BPA. Female rats were fed with BPA [5 µg/kg of body weight/day] or vehicle from gestational day 15 to pup weaning. Juvenile female offspring (day (D)25) were used to analyze immune cell populations, humoral and cellular responses after oral tolerance or immunization protocol to ovalbumin (OVA), and susceptibility to infection by the intestinal nematode Nippostrongylus brasiliensis (N. brasiliensis). Anti-OVA IgG titers following either oral tolerance or immunization were not affected after BPA perinatal exposure, while a sharp decrease in OVA-induced IFNγ secretion occurred in spleen and mesenteric lymph nodes (MLN) of OVA-immunized rats. These results are consistent with a decreased number of helper T cells, regulatory T cells and dendritic cells in spleen and MLN of BPA-exposed rats. The lack of cellular response to antigens questioned the ability of BPA-exposed rats to clear intestinal infections. A 1.5-fold increase in N. brasiliensis living larvae was observed in the intestine of BPA-exposed rats compared to controls due to an inappropriate Th1/Th2 cytokine production in infected jejunal tissues. These results show that perinatal BPA exposure impairs cellular response to food antigens, and increases susceptibility to intestinal parasitic infection in the juveniles. This emphasized the maturing immune system during perinatal period highly sensitive to low dose exposure to BPA, altering innate and adaptative immune response capacities in early life.

Food intolerance at adulthood after perinatal exposure to the endocrine disruptor bisphenol A.

The food contaminant bisphenol A (BPA) is pointed out as a risk factor in development of food allergy and food intolerance, two adverse food reactions increasing worldwide. We evaluated the consequences of perinatal exposure to low doses of BPA on immune-specific response to the food antigen ovalbumin (OVA) at adulthood. Perinatal exposure to BPA (0.5, 5, or 50 µg/kg/d) from 15th day of gravidity to pups weaning resulted in an increase of anti-OVA IgG titers at all BPA dosages in OVA-tolerized rats, and at 5 µg/kg/d in OVA-immunized rats compared to control rats treated with vehicle. In BPA-treated and OVA-tolerized rats, increased anti-OVA IgG titers were associated with higher IFNγ secretion by the spleen. This result is in accordance with the increase of activated CD4(+)CD44(high)CD62L(low) T lymphocytes observed in spleen of BPA-exposed rats compared to controls. Finally, when BPA-treated OVA-tolerized rats were orally challenged with OVA, colonic inflammation occurred, with neutrophil infiltration, increased IFNγ, and decreased TGFß. We show that perinatal exposure to BPA altered oral tolerance and immunization to dietary antigens (OVA). In summary, the naive immune system of neonate is vulnerable to low doses of BPA that trigger food intolerance later in life.

Multispecies probiotic protects gut barrier function in experimental models.

AIM: To investigate the effect of the probiotic combination Lactibiane Tolerance(®) (LT) on epithelial barrier function in vitro and in vivo. METHODS: The effect of the multispecies probiotic LT was assessed on several models of epithelial barrier function both in vitro (in basal and inflammatory conditions) and in vivo [visceral hypersensitivity induced by chronic stress or by colonic perfusion of a fecal supernatant (FSN) from patients with irritable bowel syndrome (IBS)]. In vitro, we measured the permeability of confluent T84 cell monolayers incubated with or without LT by evaluating the paracellular flux of macromolecules, in basal conditions and after stimulation with lipopolysaccharide (LPS) or with conditioned medium of colonic biopsies from IBS patients (IBS-CM). In vivo, male C57/Bl6 mice received orally NaCl or LT for 15 d and were submitted to water avoidance stress (WAS) before evaluating visceral sensitivity by measuring the myoelectrical activity of the abdominal muscle and the paracellular permeability with (51)Cr-EDTA. Permeability and sensitivity were also measured after colonic instillation of FSN. Tight-junctions were assessed by immunoblotting and TLR-4 expression was evaluated by immunohistochemistry RESULTS: Incubation of T84 cell monolayers with LT in basal conditions had no significant effect on permeability (P > 0.05 vs culture medium). By contrast, addition of LT bacterial bodies (LT) completely prevented the LPS-induced increase in paracellular permeability (P < 0.01 vs LPS 10 ng/mL (LPS 10); P < 0.01 vs LPS 100 ng/mL (LPS 100), P > 0.05 vs culture medium). The effect was dose dependent as addition of 10(9) LT bacterial bodies induced a stronger decrease in absorbance than 10(6) LT (10(9) LT + LPS 10: -20.1% ± 13.4, P < 0.01 vs LPS 10; 10(6) LT + LPS 10: -11.6% ± 6.2, P < 0.01 vs LPS 10; 10(9) LT + LPS 100: -14.4% ± 5.5, P < 0.01 vs LPS 100; 10(6) LT + LPS 100: -11.6% ± 7.3, P < 0.05 vs LPS 100). Moreover, the increase in paracellular permeability induced by culturing T84 cells with conditioned medium of colonic biopsies from IBS patients (IBS-CM) was completely inhibited in the presence of 10(9) LT (P < 0.01 vs IBS-CM). LT also significantly prevented the epithelial disruption induced by intracolonic infusion of fecal supernatant from IBS patients (P < 0.01 vs IBS FSN) or water avoidance stress P < 0.01 vs WAS) in C57/Bl6 mice and increased the expression of occludin in vitro and in vivo, as assessed by immnunoblotting. The WAS-induced effect on visceral sensitivity was prevented by LT treatment since values obtained for all steps of colorectal distension were significantly (P < 0.01) different from the WAS group. Finally, LT down-regulated the response mediated through TLR-4 in vitro (decrease in tumor necrosis factor α secretion in response to LPS: -65.8% for 10(9) LT and -52.5% for 10(6) LT, P < 0.01 vs LPS) and in vivo (inhibition of WAS induced an increase in TLR-4 expression in the LT treated mice colon, P < 0.01 vs WAS). CONCLUSION: The probiotic LT mix prevented the disruption to the epithelial barrier induced by LPS, stress or colonic soluble factors from IBS patients and prevented visceral hypersensitivity.