Promising immunomodulatory effects of selected strains of dairy propionibacteria as evidenced in vitro and in vivo

Abstract

Immunomodulatory properties of 10 dairy propionibacteria, analyzed on human peripheral blood mononuclear cells (PBMCs), revealed a highly strain-dependent induction of anti-inflammatory cytokine interleukin 10 (IL-10). Two selected strains of Propionibacterium freudenreichii showed a protective effect against two models of colitis in mice, suggesting a probiotic potential predicted by immune-based selection criteria for these cheese starter bacteria.

FIG. 1.

In vitro immunomodulatory-based screening. Comparative anti-inflammatory IL-10 (A) and proinflammatory TNF-α (B), IL-12 (C), and IFN-γ (D) cytokine responses of human PBMCs for four reference and five propionibacterium strains at a bacterial density corresponding to a multiplicity of infection (MOI) of 10. (E) IL-10 response at distinct dosing (MOI of 5, 10 and 50). (F) Anti-inflammatory role of surface layer protein(s). Transmission electron microscopy appearance of untreated (left panel) and guanidium chloride-treated P. freudenreichii BIA118 and SI48, showing removal of specific surface layer. (G) IL-10 and IL-12 response of P. freudenreichii BIA118 and SI48 and the corresponding extracted bacteria (MOI of 10 for all strains). Immunocompetent cells were stimulated by bacteria for 24 h and collected supernatants analyzed by ELISA. Data are expressed in pg/ml as means and standard deviations of results for four distinct healthy blood donors (the dashed lines indicate the cutoff in vitro threshold of the corresponding strain previously shown to be protective in vivo [9]; dashed window for panel D corresponds to a ×25 magnification).

FIG. 2.

Protective effect of 5 days of oral treatment with P. freudenreichii SI48 (5 × 10⁸ CFU per day) on TNBS colitis. (A) 2-day body weight loss (percentage of initial weight) for healthy control mice (empty circles), vehicle-TNBS-treated animals (black circles), and SI48-fed mice (gray circles). (B) Individual macroscopic disease scores (Wallace score). (C) Macroscopic and histological damage scores (Ameho score). (D) Changes in colon length (cm). (E) Colonic MPO activities. Results for panels B to E were all recorded 48 h after induction of colitis. Results are expressed as means ± standard errors of the mean (n = 10 per group); *, P < 0.05; **, P < 0.01; ***, P < 0.001 versus vehicle by Mann-Whitney U test or the Student t test where appropriate. (F) Representative histological sections from an untreated mouse (upper panel) and from mice after induction of TNBS colitis following treatment either with vehicle (middle panel) or with P. freudenreichii SI48 (lower panel); May-Grünwald- and Giemsa-stained 5-μm paraffin sections, ×40.

FIG. 3.

Protective effect of 5 days of oral treatment with P. freudenreichii BIA118 and SI48 (5 × 10⁸ CFU per day) on 10-day C. rodentium (strain DBS120)-induced colitis. (A) Time course of pathogen enumeration in pooled feces following the distinct treatments. (B) Weight changes (percentage of initial weight) for healthy control mice (empty circles) and infected control (full circles), BIA118-fed (squares), and SI48-fed mice (triangles). (C to F) Spleen weight (C), blood serum amyloid A protein (mg/ml) (D), colonic MPO activities (E), and crypt length (μm) (F) measured on at least 10 well-orientated sections per animal per group. The results are expressed as mean ± SEM (n = 8 mice per group), except for weight changes in panel B, where error bars were eliminated for clarity. nd, not detected. Bars with no asterisks, 0.05 < P < 0.1; *, P < 0.05; **, P < 0.01; ***, P < 0.001 versus infected animals by Mann-Whitney U test or the Student t test where appropriate.