Immunomodulatory effect of non-viable components of probiotic culture stimulated with heat-inactivated Escherichia coli and Bacillus cereus on holoxenic mice

Abstract

Background: Competition of probiotic bacteria with other species from the intestinal microbiota involves different mechanisms that occur regardless of probiotics' viability. The objective of this paper was to assess the cytokine serum levels in holoxenic mice after oral administration of non-viable components (NVC) of Enterococcus faecium probiotic culture stimulated with heat-inactivated Escherichia coli and Bacillus cereus in comparison to NVC of unstimulated E. faecium probiotic culture.

Methods: Probiotic E. faecium CMGb 16 culture, grown in the presence of heat-inactivated cultures of E. coli and B. cereus CMGB 102, was subsequently separated into supernatant (SN) and heat-inactivated cellular sediment (CS) fractions by centrifugation. Each NVC was orally administered to holoxenic mice (balb C mouse strain), in three doses, given at 24 hours. Blood samples were collected from the retinal artery, at 7, 14, and 21 days after the first administration of the NVC. The serum concentrations of IL-12 and tumor necrosis factor-alpha (TNF-α) interleukins were assessed by ELISA method.

Results: After the oral administration of SN component obtained from the probiotic culture stimulated with heat-inactivated cultures of B. cereus CMGB 102 and E. coli O28, the serum concentrations of IL-12 were maintained higher in the samples collected at 7 and 14 days post-administration. No specific TNF-α profile could be established, depending on stimulated or non-stimulated probiotic culture, NVC fraction, or harvesting time.

Conclusion: The obtained results demonstrate that non-viable fractions of probiotic bacteria, stimulated by other bacterial species, could induce immunostimulatory effects mediated by cytokines and act, therefore, as immunological adjuvants.

Keywords: Enterococcus faecium; immunomodulation; interleukins; intestinal microbiota; non-viable components; probiotic.

Fig. 1

Graphic representation of IL-12 concentration in the serum of holoxenic mice, collected after: I – 7 days from the first oral administration of NVCs; II – 14 days from the first oral administration of NVCs; III – 21 days from the first oral administration of NVCs. IC, integral culture; CS, cellular suspension; SN, supernatant; A, serum collected from holoxenic mice after oral administration of NVC; B, serum collected from holoxenic mice after oral administration of NVC stimulated with E. coli O28C; C, serum collected from holoxenic mice after oral administration of NVC stimulated with B. cereus; MC, media control – serum collected from holoxenic mice after oral administration of MRS; NC, negative control – serum collected from holoxenic mice control.

Fig. 2

Graphic representation of TNF α concentration in the serum of holoxenic mice, collected after: I – 7 days from the first oral administration of NVCs; II – 14 days from the first oral administration of NVCs; III – 21 days from the first oral administration of NVCs. IC, integral culture, CS, cellular suspension; SN, supernatant; A, serum collected from holoxenic mice after oral administration of NVC; B, serum collected from holoxenic mice after oral administration of NVC stimulated with E. coli O28C; C, serum collected from holoxenic mice after oral administration of NVC stimulated with B. cereus; MC, media control – serum collected from holoxenic mice after oral administration of MRS; NC, negative control – serum collected from holoxenic mice control.