Transduction pathways regulating the trophic effects of Saccharomyces boulardii in rat intestinal mucosa.

Abstract Saccharomyces boulardii is a probiotic yeast that is widely prescribed in lyophilized form; it determines several effects in human and rat small intestine including endoluminal secretion of enzymes and of polyamines, stimulation of microvillous enzymes, of sIgA, increased production of the receptor for polymeric immunoglobulins by crypt cells, and enhanced d-glucose uptake. Aim. The objective of this study was to determine the pathway(s) by which these effects generated by the yeast are transduced into mucosal cells. Methods. Litters of six growing Wistar rats each were treated with S. boulardii (50 mug/gram body weight) or with saline between days 30 and 34 postpartum. For each animal, the cytosol was prepared from the whole mucosa after the fat cake was discarded. Several known intestinal substrates were immunoprecipitated and immunoblotted using specific antibodies recognizing the non-, mono-, or diphosphorylated forms of these substrates. The signals were detected using Echochemiluminoscence (ECL) and were measured by optodensitometry. Results. Treatment with S. boulardii markedly enhanced the RAS-GAP-RAF-ERK(1,2) pathway with participation of growth receptor bound 2 protein, SHC, SOS, and CRKII. Unit p85alpha of phosphatidylinositol 3 kinase, tested in its phosphorylated form, was also enhanced by the probiotic compared to control samples. In rats treated with an inhibitor of RAF-1 and of ERK(1,2) (PD098059) the expression of mucosal disaccharidases was inhibited by about 50%. Conclusion. The probiotic S. boulardii generates in vivo mitogen and metabolic signals that are transduced into intestinal mucosal cells, downstream from the apical membrane to the nuclei, using recruitment substrates and serine, threonine, or tyrosine kinases

Interaction of Saccharomyces boulardii With Intestinal Brush Border Membranes: Key to Probiotic Effects?

The probiotic Saccharomyces boulardii exerts beneficial effects in humans, which include trophic effects, anti-inflammatory effects, antisecretory effects, inhibition of toxins, immunostimulatory effects, and resistance to bacterial overgrowth. This short communication discusses the interactions of the probiotic with brush border membrane (BBM) constituents because most of these effects are BBM mediated. The use of bacterial and yeast probiotics has increased dramatically in more and more clinical states, but their exact mechanisms of action remain largely unknown. The present communication focuses on the interactions of a confirmed yeast probiotic (S boulardii) on the constituents of BBMs

Effects of Saccharomyces boulardii on intestinal mucosa.

Saccharomyces boulardii (S. boulardii) is a non-pathogenic biotherapeutic agent, widely prescribed in a lyophilized form in many countries over the world. S. boulardii acts as a shuttle liberating effective enzymes, proteins and trophic factors during its intestinal transit that improve host immune defenses, digestion, and absorption of nutrients. In addition, S. boulardii secretes during its intestinal transit polyamines, mainly spermine and spermidine that regulate gene expression and protein synthesis. In this review, we will focus on the interactions of the yeast with the host intestinal mucosa

Cellular adaptation of the rat small intestine after proximal enterectomy: changes in microvillous enzymes and in the secretory component of immunoglobulins.

To investigate the adaptation of functions expressed by the villous and crypt cell of the intestinal mucosa after intestinal resection, a 50% proximal enterectomy or a single transection was performed in 16 growing rats weighing 175-200 g. Ten days following the enterectomy, we determined the mucosal mass parameters (weight, protein, and DNA content), the activity of microvillous enzymes (lactase, sucrase, and aminopeptidase) in villus cells, and the concentration of the secretory component of immunoglobulins in crypt cells isolated from the proximal intestinal remnant. Mucosal hyperplasia was attested by the finding that mucosal weight, protein, and DNA content per cm of intestinal length were significantly (p less than 0.01) higher (+29 to +48%) in the resected group than in transected controls. The specific activity of lactase, sucrase, and aminopeptidase were significantly (p less than 0.05) lower (-23 to -56%) in villous cells isolated from the intestinal remnant of resected rats compared to controls. Sucrase activity was depressed in each cell fraction of the entire villous-crypt unit resulting in a lower villous to crypt gradient of enzyme activity. Km for the enzyme determined in villous cells was similar in both groups but the Vmax was reduced proportionally to the enzyme activity in the resected group indicating less enzyme per cell. By contrast, the concentration of secretory component measured by an immunoradiometric assay in both villous and crypt cells was significantly (p less than 0.05) increased (+37 to 45%) following proximal enterectomy. Our data indicate that the response of the epithelial cell to intestinal resection varies according to the metabolic function and that the mechanism of adaptation at the cellular level is complex

Intestinal development in the suckling rat: effect of insulin on the maturation of villus and crypt cell functions.

The influence of insulin on the postnatal development of intestinal functions linked to villus cells (sucrase, lactase, maltase and aminopeptidase) and crypt cells (secretory component of immunoglobulins, SC) has been studied in suckling and weanling rats. At 9 days of age, the animals received a daily injection of insulin 12.5 mU g-1 body weight day-1 for 4 days. Compared with saline-treated controls, insulin had no effect on the development of the intestinal mucosal mass parameters determined in the jejunum, ileum and colon. A premature appearance of sucrase was noted in isolated jejunal villus and crypt cells, the level of activity reached by the enzyme being dependent of the amount of insulin injected. By 6 and 12 h after a single injection of the hormone (12.5 mU g-1 body weight), sucrase activity was detected in all the cell fractions along the villus-crypt axis. In villus cells of insulin-treated rats, maltase, lactase and aminopeptidase activities were significantly (P less than 0.001) increased (+201%, +50%, +207%, respectively, vs. controls), whereas the concentration of SC measured by a sensitive immunoradiometric assay was enhanced over the controls by 75% in villus cells, 83% in crypt cells and 172% in the liver. Weanling rats treated from day 10 to day 20 postpartum with 12.5 mU insulin also exhibited a higher intestinal production of SC (+93%, P less than 0.01) than did saline controls.(ABSTRACT TRUNCATED AT 250 WORDS

Saccharomyces boulardii enhances rat intestinal enzyme expression by endoluminal release of polyamines.

Saccharomyces boulardii is a yeast widely used in humans for the prevention and treatment of infectious enteritis and Clostridium difficile-associated enterocolopathies. After oral administration to human volunteers or growing rats, S. boulardii enhances markedly the expression of intestinal enzymes as well as the production of the polymeric immunoglobulin receptor by mechanisms that remain unknown. We have analyzed the role of the yeast polyamines as potential mediators in the intestinal trophic response. In weanling rats (d 20 to d 30), a daily dose of 100 mg of lyophilized S. boulardii produced significant (p < 0.025) increases in sucrase (157%) and maltase (47%) activities. This dose corresponded to a total oral load of 678 nmol of polyamines per day (spermidine; 376 +/- 32, spermine: 293 +/- 26, putrescine: 9.5 +/- 1.4 nmol/100 mg). Spermine, given orally to growing rats at doses nearly equivalent (500 nmol) to the load of polyamines provided by the yeast (678 nmol), reproduced similar enzymatic changes, including a 2.5-fold induction of sucrase, and enhanced maltase activity (+24%). Spermidine and spermine concentrations measured in the jejunal mucosa of treated rats were increased over matched controls by 21.4% (p < 0.005) and 21.9%, respectively (p < 0.002). After being centrifuged and filtered to discard residual yeast cells, 2-mL samples of jejunal and ileal fluid collected from S. boulardii-treated rats by intestinal flushing contained higher levels of spermidine (48 and 60%) and spermine (150 and 316%) than did control rats. Our data indicate that lyophilized S. boulardii exerts trophic effects on the small intestine that are likely mediated by the endoluminal release of spermine and spermidine

Changes in serum and intestinal diamine oxidase (DAO) activity after proximal enterectomy in rats. Correlation of DAO activity with mucosal mass parameters.

To determine whether serum and mucosal DAO activity reflects quantitative changes in the small bowel mucosal mass, we have chosen an experimental model of mucosal hyperplasia which is known to occur in the rat after enterectomy. A 50% proximal enterectomy or a single transection was performed in 20 growing rats weighing 145-160 g. Ten days following surgery, we determined mucosal mass parameters (weight, protein, and DNA content), sucrase activity, and DAO activity in the duodenum (segment A), proximal ileum (segment B), and distal ileum (segment C) of the remaining small intestine. Mucosal hyperplasia was demonstrated by the finding that in each segment, mucosal weight, protein, and DNA content per centimeter of gut length were significantly (P less than 0.01) higher (+38 to + 78%) in the resected group than in transected controls. In segments B and C of resected rats, the changes in DAO activity expressed per gram of mucosa paralleled the changes in mucosal mass, the activity being increased by +69% and +49% (P less than 0.05) compared to the values recorded in transected controls. Expressed per centimeter of gut length, total DAO activity was also enhanced by +141% in segment B (P less than 0.05 vs controls) and by +87% in segment C (P less than 0.01 vs controls) of resected rats. In the duodenum, the changes in DAO activity were small (+36%) and not significant.(ABSTRACT TRUNCATED AT 250 WORDS

Hormonal regulation of the rat small intestine: responsiveness of villus and crypt cells to insulin during the suckling period and unresponsiveness after weaning.

To further document the effect of insulin on intestinal maturation, suckling rats were treated either with exogenous insulin (12.5 mU.g body wt, intraperitoneally, twice daily) or with saline from d 8 to 12 postpartum. Sucrase activity in brush border membrane extracts was precociously induced by insulin, whereas the activities of other brush border membrane enzymes (maltase, aminopeptidase, and neutral lactase) were enhanced (+ 30 to + 131%, p less than 0.01 versus controls). The lysosomal enzyme, N-acetyl-beta-glucosaminidase, which normally declines at weaning was significantly (p less than 0.025) decreased in both villus (-51%) and crypt cells (-57%) isolated from the jejunum of insulin-treated rats. The microsomal enzyme, sulfatase C, and the cytosolic enzyme, lactate dehydrogenase, were also sensitive to insulin with decreases in activity ranging from -37 to -63% (p less than 0.05) compared to saline-treated control rats. Insulin at doses of 0.5 or 12.5 mU did not influence plasma total corticosterone levels, which were about 9-fold lower in suckling than in 25-d-old weaned rats. In weaned rats (from d 25 to 32) insulin treatment (12.5 mU) failed to influence the activity of brush border membrane hydrolases or of lysosomal, microsomal, and cytosolic enzymes. The synthesis rate of mature sucrase-isomaltase, measured in weaned rats (32 d) by the incorporation of 14C-leucine into the enzyme precursor protein, was equivalent in both groups. These data demonstrate that the immature enterocyte of the suckling rat is responsive to insulin, whereas the mature enterocyte of the weaned rat is unresponsive.(ABSTRACT TRUNCATED AT 250 WORDS

Characterization of alpha,alpha-trehalase released in the intestinal lumen by the probiotic Saccharomyces boulardii.

Objective. Trehalose intolerance due to alpha,alpha-trehalase deficiency has scarcely been studied. The purpose of this study was to measure alpha,alpha-trehalase activity in intestinal biopsy samples from 200 consecutive patients over a period of 6 months, and to characterize alpha,alpha-trehalase released by the probiotic Saccharomyces boulardii (S. boulardii). Material and methods. Enzyme activities were measured in human and rat intestinal mucosal samples using the micromethod of Messer & Dalqvist. alpha,alpha-trehalase from S. boulardii was immunoprecipitated and Western blotted using an IgG purified antibody raised against a 23 amino acid peptide of alpha,alpha-trehalase of S. cerevisiae. Results. Among 200 patients, most of whom complained of abdominal symptoms and diarrhoea, 18 (9%) had total alpha,alpha-trehalase deficiency (0-12 U/g mucosa) and 39 had partial deficiency (3-12 U/g mucosa). Only 4 patients (2%) presented selective alpha,alpha-trehalase deficiency with otherwise normal disaccharidases. Expressed per gram of powder, alpha,alpha-trehalase from S. boulardii delivered in vitro an activity 175 times higher than that of human trehalase per gram of intestinal mucosa. V(max) (22+/-0.43 micromol) and K(m) (5 mM) were close to that of the human enzyme, whereas Western blot revealed a signal of two subunits of 82 kDa. Finally, treatment of rats with S. boulardii resulted in increases in alpha,alpha-trehalase activities of 25 to 45% (p<0.01) in endoluminal fluid and intestinal mucosa compared with in controls. Conclusions. Our data suggest that alpha,alpha-trehalase deficiency is more common than is believed and that oral administration of S. boulardii could be beneficial in patients with digestive symptoms caused by trehalose intolerance