Modulation of intestinal inflammation by yeasts and cell wall extracts: strain dependence and unexpected anti-inflammatory role of glucan fractions.

Yeasts and their glycan components can have a beneficial or adverse effect on intestinal inflammation. Previous research has shown that the presence of Saccharomyces cerevisiae var. boulardii (Sb) reduces intestinal inflammation and colonization by Candida albicans. The aim of this study was to identify dietary yeasts, which have comparable effects to the anti-C. albicans and anti-inflammatory properties of Sb and to assess the capabilities of yeast cell wall components to modulate intestinal inflammation. Mice received a single oral challenge of C. albicans and were then given 1.5% dextran-sulphate-sodium (DSS) for 2 weeks followed by a 3-day restitution period. S. cerevisiae strains (Sb, Sc1 to Sc4), as well as mannoprotein (MP) and β-glucan crude fractions prepared from Sc2 and highly purified β-glucans prepared from C. albicans were used in this curative model, starting 3 days after C. albicans challenge. Mice were assessed for the clinical, histological and inflammatory responses related to DSS administration. Strain Sc1-1 gave the same level of protection against C. albicans as Sb when assessed by mortality, clinical scores, colonization levels, reduction of TNFα and increase in IL-10 transcription. When Sc1-1 was compared with the other S. cerevisiae strains, the preparation process had a strong influence on biological activity. Interestingly, some S. cerevisiae strains dramatically increased mortality and clinical scores. Strain Sc4 and MP fraction favoured C. albicans colonization and inflammation, whereas β-glucan fraction was protective against both. Surprisingly, purified β-glucans from C. albicans had the same protective effect. Thus, some yeasts appear to be strong modulators of intestinal inflammation. These effects are dependent on the strain, species, preparation process and cell wall fraction. It was striking that β-glucan fractions or pure β-glucans from C. albicans displayed the most potent anti-inflammatory effect in the DSS model

Colonic inflammation in mice is improved by cigarette smoke through iNKT cells recruitment.

Cigarette smoke (CS) protects against intestinal inflammation during ulcerative colitis. Immunoregulatory mechanisms sustaining this effect remain unknown. The aim of this study was to assess the effects of CS on experimental colitis and to characterize the intestinal inflammatory response at the cellular and molecular levels. Using the InExpose® System, a smoking device accurately reproducing human smoking habit, we pre-exposed C57BL/6 mice for 2 weeks to CS, and then we induced colitis by administration of dextran sodium sulfate (DSS). This system allowed us to demonstrate that CS exposure improved colonic inflammation (significant decrease in clinical score, body weight loss and weight/length colonic ratio). This improvement was associated with a significant decrease in colonic proinflammatory Th1/Th17 cytokine expression, as compared to unexposed mice (TNF (p=0.0169), IFNγ (p<0.0001), and IL-17 (p=0.0008)). Smoke exposure also induced an increased expression of IL-10 mRNA (p=0.0035) and a marked recruitment of iNKT (invariant Natural Killer T; CD45+ TCRβ+ CD1d tetramer+) cells in the colon of DSS-untreated mice. Demonstration of the role of iNKT cells in CS-dependent colitis improvement was performed using two different strains of NKT cells deficient mice. Indeed, in Jα18KO and CD1dKO animals, CS exposure failed to induce significant regulation of DSS-induced colitis both at the clinical and molecular levels. Thus, our study demonstrates that iNKT cells are pivotal actors in the CS-dependent protection of the colon. These results highlight the role of intestinal iNKT lymphocytes and their responsiveness to environmental stimuli. Targeting iNKT cells would represent a new therapeutic way for inflammatory bowel diseases

Schematic diagram of glycan preparation from <i>S. cerevisiae</i>.

<p>Schematic diagram of glycan preparation from <i>S. cerevisiae</i>.</p

Comparison of the probiotic potential of <i>S. cerevisiae</i> var. <i>boulardii</i> and <i>S. cerevisiae</i> 1-1 strain.

<p>(<b>A</b>)<b> Experimental design</b>. Low doses of DSS were used for 2 weeks followed by a 3-day restitution period. Either <i>S. cerevisiae</i> strains or β-glucan fractions were inoculated daily by oral gavage starting 3 days after <i>C. albicans</i> challenge. Three days post-<i>C. albicans</i> challenge was chosen to start <i>S. cerevisiae</i> treatment as both <i>Candida</i> colonization and colonic inflammation are well established at this time point. (<b>B</b>)<b> Percentage survival of mice</b>. The results are shown as percent survival from the time of <i>C. albicans</i> challenge and DSS treatment. The survival data were significantly different by the log-rank test. A total of 140 mice were divided into five control groups: water (n = 16 mice), Ca (n = 16 mice), Sb (n = 16 mice), Sc1-1 (n = 16 mice) and DSS alone (n = 16 mice), and three experimental groups: CaD (n = 20 mice), CaDSb (n = 20 mice), and CaDSc1-1 (n = 20 mice). (*<i>P</i><0.05 for CaDSb mice vs. CaD mice; and ‡<i>P</i><0.05 for CaDSc1-1 mice vs. CaD mice.) (<b>C</b>)<b> Number of </b><b><i>C. albicans</i></b><b> colony forming units recovered from stools</b>. Each data set represents the mean values of 10 mice/group. (*<i>P</i><0.05 for CaDSb mice vs. CaD mice; and ‡<i>P</i><0.05 for CaDSc1-1 mice vs. CaD mice.) (<b>D</b>)<b> Clinical analysis of DSS-induced colitis in mice</b>. Both <i>S. cerevisiae</i> var. <i>boulardii</i> and <i>S. cerevisiae</i> Sc1-1 significantly reduced the clinical score. (*<i>P</i><0.05 for CaDSb mice vs. D mice; ‡<i>P</i><0.05 for CaDSb mice vs. CaD mice; †<i>P</i><0.05 for CaDSc1-1 mice vs. D mice; and **<i>P</i><0.05 for CaDSc1-1 mice vs. CaD mice.) (<b>E</b>)<b> Histological score</b>. The histological score was determined by two independent, blinded examiners (degree of inflammation: 0, no changes, to 6, extensive cell infiltration and tissue damage). The histological score increased in both DSS and CaDSS groups. The histological score was significantly lower in both CaDSb and CaDSc1-1 groups when compared to CaDSS and DSS-treated groups. (*<i>P</i><0.05 for CaDSb mice vs. D mice; ‡<i>P</i><0.05 for CaDSb mice vs. CaD mice; †<i>P</i><0.05 for CaDSc1-1 mice vs. D mice; and **<i>P</i><0.05 for CaDSc1-1 mice vs. CaD mice.) (<b>F and G</b>)<b> Relative expression levels, determined by real-time quantitative PCR, of TNF-</b><b><i>α</i></b><b> and IL-10 mRNA in the colon</b>. Data are expressed as the mean ± SE of five mice in each group. (*<i>P</i><0.05 for CaDSb mice vs. D mice; †<i>P</i><0.05 for CaDSb mice vs. CaD mice; ‡<i>P</i><0.05 for CaDSc1-1 mice vs. D mice; and **<i>P</i><0.05 for CaDSc1-1 mice vs. CaD mice.)</p

Schematic diagram of β-glucan preparation from <i>C. albicans</i>.

<p>Schematic diagram of β-glucan preparation from <i>C. albicans</i>.</p

Yeast strains used in the investigation.

<p>Yeast strains used in the investigation.</p

Summary of the effects of <i>S. cerevisiae</i> strains or glycan fractions on <i>Candida</i> DSS-treated mice.

<p>Radar blots showed the effect of each <i>S. cerevisiae</i> strain or glycan fraction on: (1) mortality; (2) body weight loss; (3) clinical score; (4) histological score; and (5) <i>C. albicans</i> colonization. Sb, Sc1-1 and Sc3 strains improved the intestinal damage due to DSS-induced colitis and <i>C. albicans</i> colonization. Sc1-2, Sc2 and Sc4 strains had a dramatic effect on all inflammation signs including <i>Candida</i> colonization. Regarding the glycan fractions, both β-glucans and F2 reduced all colitis parameters and eliminated <i>C. albicans</i> colonization, while MP was less effective at controlling these parameters.</p

Effect of glycan fractions derived from <i>S. cerevisiae</i> on <i>Candida</i> DSS-treated mice.

<p>(<b>A</b>)<b> Percentage survival of mice</b>. The results are displayed as percent survival from the time of <i>C. albicans</i> challenge and DSS treatment. Three days after <i>C. albicans</i> challenge, mice were given either <i>S. cerevisiae</i> (Sc2), mannoproteins (Sc2MP) or β-glucan fractions (Sc2GP) for 2 weeks. The survival data were significantly different by the log-rank test. A total of 40 mice were divided into four experimental groups. (*<i>P</i><0.05 for CaDSc2GP mice vs. CaD mice; and ‡<i>P</i><0.05 for CaDSc2MP mice vs. CaD mice.) (<b>B</b>)<b> Difference in body weight loss in mice</b>. Each data set represents the mean value for each body weight. (*<i>P</i><0.05 for CaDSc2GP mice vs. CaD mice, and †<i>P</i><0.05 for CaDSc2MP mice vs. CaD mice.) (<b>C</b>)<b> Number of </b><b><i>C. albicans</i></b><b> colony forming units recovered from stools</b>. Each data set represents the mean value of 10 mice/group. (*<i>P</i><0.05 for CaDSc2GP mice vs. CaD mice.) (<b>D</b>)<b> Histological analysis of DSS-induced colitis in mice</b>. Panel (a) shows a colon section from a <i>C. albicans</i>+mannoprotein DSS-treated mouse; panel (b) shows a colon section from a <i>C. albicans</i>+β-glucan DSS-treated mouse. Inflammatory cell infiltrates were insignificant in both CaDSc2MP and CaDSc2GP mice. (<b>E</b>)<b> Clinical analysis of DSS-induced colitis in mice</b>. Clinical score was determined by assessing weight loss, change in stool consistency and the presence of gross bleeding. The clinical score ranged from 0 to 8. (*<i>P</i><0.05 for CaDSc2 mice vs. CaD mice; †<i>P</i><0.05 for CaDSc2MP mice vs. CaD mice; and ‡<i>P</i><0.05 for CaDSc2GP mice vs. CaD mice.) (<b>F</b>)<b> Histological score</b>. The histological score decreased significantly in CaDSc2GP mice. (*<i>P</i><0.05 for CaDSc2 mice vs. CaD mice; †<i>P</i><0.05 for CaDSc2MP mice vs. CaD mice; and ‡<i>P</i><0.05 for CaDSc2GP mice vs. CaD mice.)</p

Histological analysis of DSS-induced colitis in mice.

<p>Colon sections (4 µm thick) were stained with haematoxylin/eosin. Panel (a and b) show colon sections from a mouse colonized with <i>C. albicans</i> and treated with DSS; panel (c) shows colon sections from a <i>C. albicans</i>+Sc1-1 DSS-treated mouse; and panel (d), colon sections from a <i>C. albicans</i>+Sb DSS-treated mouse. Colon sections from the <i>C albicans</i>+DSS-treated mouse showed an inflammatory cell infiltrate in colonic wall structures (arrow, asterisk). Colon sections from the mouse with DSS-induced colitis colonized with <i>C. albicans</i>+either Sb or Sc1-1 showed an attenuated inflammatory infiltrate in the submucosa with the presence of occasional leukocytes in the lamina propria of the mucosa (panels c and d). The scale bars represent 50 µm (panels a, c, and d) and 10 µm (panel b).</p