beta(2 -> 1) chicory and beta(2 -> 1)-beta(2 -> 6) agave fructans protect the human intestinal barrier function in vitro in a stressor-dependent fashion

Dietary fibers such as fructans can protect the intestinal epithelial barrier integrity, but the mechanisms underlying this protection are not completely understood. We aimed to study the protective effect of β(2→1)-β(2→6) branched graminan-type fructans (GTFs) on gut epithelial barrier function that was disrupted by three different agents which impact the barrier function via different cellular mechanisms. The effects of GTFs were compared with those of linear β(2→1) inulin-type fructans (ITFs). T84 intestinal epithelial monolayers were incubated with GTFs and ITFs. Afterwards, the monolayers were challenged with the barrier disruptors calcium ionophore A23187, 12-myristate 13-acetate (PMA) and deoxynivalenol (DON). Transepithelial resistance was measured with an electric cell-substrate impedance sensing system. All fructans studied prevented the barrier disruption induced by A23187. ITF II protected from the disruptive effects of PMA. However, none of the studied fructans influenced the disruption induced by DON. As a measure of disruption-induced inflammation, interleukin-8 (IL-8) production by the intestinal epithelium was determined by ELISA. The production of IL-8 induced by A23187 was decreased by all fructans, whereas IL-8 production induced by DON decreased only upon pre-treatment with ITF II. None of the studied fructans prevented PMA induced IL-8 production. GTFs just like ITFs can influence the barrier function and inflammatory processes in gut epithelial cells in a structure-dependent fashion. These distinct protective effects are dependent on the different signaling pathways that lead to gut barrier disruption

Chicory inulin enhances fermentation of 2'-fucosyllactose by infant fecal microbiota and differentially influences immature dendritic cell and T-cell cytokine responses under normal and Th2-polarizing conditions

Scope: Non-digestible carbohydrates (NDCs) such as native chicory inulin and 2′-fucosyllactose (2′-FL) are added to infant formula to mimic some of the human milk oligosaccharide (HMO) functions. It is unknown whether combining inulin and 2′-FL influences their fermentation kinetics and whether the immune-modulatory effects of these NDCs are different under normal and inflammatory-prone Th2-polarizing conditions. Methods and results: We investigated the in vitro fermentation of 2′-FL and native chicory inulin, fermented individually and combined, using fecal inocula of 8-week-old infants. Native inulin was fermented in a size-dependent fashion and expedited the fermentation of 2′-FL. Fermentation of both native inulin and 2′FL increased the relative abundance of Bifidobacterium, which coincided with the production of acetate and lactate. The fermentation digesta of all fermentations differentially influenced both dendritic cell and T-cell cytokine responses under normal culture conditions or in presence of the Th2-polarizing cytokines IL-33 and TSLP, with the most pronounced effect for IL-1β in the presence of TSLP. Conclusions: Our findings show that native inulin can expedite the fermentation of 2′-FL by infant fecal microbiota and that these NDC fermentation digesta have different effects under normal and Th2-polarizing conditions, indicating that infants with different immune backgrounds might benefit from tailored NDC formulations

Effects of Different Human Milk Oligosaccharides on Growth of Bifidobacteria in Monoculture and Co-culture With Faecalibacterium prausnitzii

Human milk oligosaccharides (hMOs) are important bioactive components in mother’s milk contributing to infant health by supporting colonization and growth of gut microbes. In particular, Bifidobacterium genus is considered to be supported by hMOs. Approximately 200 different hMOs have been discovered and characterized, but only a few abundant hMOs can be produced in sufficient amounts to be applied in infant formula. These hMOs are usually supplied in infant formula as single molecule, and it is unknown which and how individual hMOs support growth of individual gut bacteria. To investigate how individual hMOs influence growth of several relevant intestinal bacteria species, we studied the effects of three hMOs (2′-fucosyllactose, 3-fucosyllactose, and 6′-sialyllactose) and an hMO acid hydrolysate (lacto-N-triose) on three Bifidobacteria and one Faecalibacterium and introduced a co-culture system of two bacterial strains to study possible cross-feeding in presence and absence of hMOs. We observed that in monoculture, Bifidobacterium longum subsp. infantis could grow well on all hMOs but in a structure-dependent way. Faecalibacterium prausnitzii reached a lower cell density on the hMOs in stationary phase compared to glucose, while B. longum subsp. longum and Bifidobacterium adolescentis were not able to grow on the tested hMOs. In a co-culture of B. longum subsp. infantis with F. prausnitzii, different effects were observed with the different hMOs; 6′-sialyllactose, rather than 2′-fucosyllactose, 3-fucosyllactose, and lacto-N-triose, was able to promote the growth of B. longum subsp. infantis. Our observations demonstrate that effects of hMOs on the tested gut microbiota are hMO-specific and provide new means to support growth of these specific beneficial microorganisms in the intestine.</p

Endo-1,3(4)-β-Glucanase-Treatment of Oat β-Glucan Enhances Fermentability by Infant Fecal Microbiota, Stimulates Dectin-1 Activation and Attenuates Inflammatory Responses in Immature Dendritic Cells

Background: Non-digestible carbohydrates are added to infant formula to mimic the effects of human milk oligosaccharide by acting as prebiotics and stimulating the immune system. Although not yet used in infant formulas, β-glucans are known to have beneficial health effects, and are therefore of potential interest for supplementation. Methods and results: We investigated the in vitro fermentation of native and endo-1,3(4)-β-glucanase-treated oat β-glucan using pooled fecal inocula of 2-and 8-week-old infants. While native oat β-glucan was not utilized, both inocula specifically utilized oat β-glucan oligomers containing β(1→4)-linkages formed upon enzyme treatment. The fermentation rate was highest in the fecal microbiota of 2-week-old infants, and correlated with a high lactate production. Fermentation of media supplemented with native and enzyme-treated oat β-glucans increased the relative abundance of Enterococcus and attenuated proinflammatory cytokine production (IL-1β, IL-6, TNFα) in immature dendritic cells. This attenuating effect was more pronounced after enzyme treatment. This attenuation might result from the enhanced ability of fermented oat β-glucan to stimulate Dectin-1 receptors. Conclusion: Our findings demonstrate that endo-1,3(4)-β-glucanase treatment enhances the fermentability of oat β-glucan and attenuates pro-inflammatory responses. Hence, this study shows that especially enzyme-treated oat β-glucans have a high potential for supplementation of infant formula.</p

Structure-Specific Fermentation of Galacto-Oligosaccharides, Isomalto-Oligosaccharides and Isomalto/Malto-Polysaccharides by Infant Fecal Microbiota and Impact on Dendritic Cell Cytokine Responses

SCOPE: Next to galacto-oligosaccharides (GOS), starch-derived isomalto-oligosaccharide preparation (IMO) and isomalto/malto-polysaccharides (IMMP) could potentially be used as prebiotics in infant formulas. However, it remains largely unknown how the specific molecular structures of these non-digestible carbohydrates (NDCs) impact fermentability and immune responses in infants. METHODS AND RESULTS: In vitro fermentation of GOS, IMO and IMMP using infant fecal inoculum of 2- and 8-week-old infants showed that only GOS and IMO were fermented by infant fecal microbiota. The degradation of GOS and IMO coincided with an increase in Bifidobacterium and production of acetate and lactate, which was more pronounced with GOS. Individual isomers with an (1↔1)-linkage or di-substituted reducing terminal glucose residue were more resistant to fermentation. GOS, IMO and IMMP fermentation digesta attenuated cytokine profiles in immature dendritic cells (DCs), but the extent was dependent on the infants age and NDC structure. CONCLUSION: The IMO preparation, containing reducing and non-reducing isomers, showed similar fermentation patterns as GOS in fecal microbiota of 2-week-old infants. Knowledge obtained on the substrate specificities of infant fecal microbiota and the subsequent regulatory effects of GOS, IMO and IMMP on DC responses might contribute to the design of tailored NDC mixtures for infants of different age groups. This article is protected by copyright. All rights reserved

Bridging the immune gap between human milk and infant formula : non-digestible oligosaccharides

Cow-milk based infant formulas are commonly supplemented with non-digestible carbohydrates (NDCs) to resemble the functional properties of oligosaccharides present in human milk. NDCs support the development of a balanced immune system through their direct interaction with immune cells, but also through their fermentation by the gut microbiota. In this thesis, it was investigated how the structure of the NDCs affects the fermentability and the microbiota-dependent immune effects of the NDCs. First, the use of pooled infant faecal inoculum in in vitro fermentation experiments was validated. This fermentation set-up enabled us to efficiently judge the fermentability of 7 structurally different NDCs by faecal microbiota of 2- and 8-week-old infants; galacto-oligosaccharides (GOS), fructo-oligosaccharides, native inulin, enzyme-treated and native oat &beta;-glucan, isomaltose-oligosaccharides (IMO) and isomalto/malto-polysaccharides. The fermentability was dependent on the age of the infants and was negatively correlated with the size of the NDCs. In addition, a characterization method was developed based on ultra-high performance liquid chromatography-mass spectrometry (UHPLC-MS) with a stationary phase of porous graphitic carbon (PGC) to study the fate of individual isomers in the infant gut. The application of this method to digesta of GOS and IMO fermentation indicated that e.g. the type of glycosidic linkage, anomeric configuration, monomer composition and substitution of the terminal reducing residue all affect the fermentability of these complex NDCs by infant faecal microbiota. Finally, the incubation of the fermentation digesta with dendritic cells derived from the umbilical vein demonstrated the immune-attenuating effects of the NDCs, which could be linked to the bacteria-derived components and metabolites. The extent of attenuation by the NDCs depended on the age and consequent microbiota composition of the infants as well as the structure of the NDCs. These findings support the use of tailored NDC mixtures for substitution of infant formula to meet the needs of infants of different age

In Vitro Fermentation of Porcine Milk Oligosaccharides and Galacto-oligosaccharides Using Piglet Fecal Inoculum

In this study, the in vitro fermentation by piglet fecal inoculum of galacto-oligosaccharides (GOS) and porcine milk oligosaccharides (PMOs) was investigated to identify possible preferences for individual oligosaccharide structures by piglet microbiota. First, acidic PMOs and GOS with degrees of polymerization 4-7 were depleted within 12 h of fermentation, whereas fucosylated and phosphorylated PMOs were partially resistant to fermentation. GOS structures containing β1-3 and β1-2 linkages were preferably fermented over GOS containing β1-4 and β1-6 linkages. Upon in vitro fermentation, acetate and butyrate were produced as the main organic acids. GOS fermentation by piglet inoculum showed a unique fermentation pattern with respect to preference of GOS size and organic acids production.</p

Pooled faecal inoculum can predict infant fiber fermentability despite high inter-individual variability of microbiota composition

Infants are known for their high inter-individual variability in gut microbiota composition in the first months of life. This poses difficulties when predicting the fate of non-digestible carbohydrates (NDCs) in the infant gut using in vitro models, since numerous experiments with individual faecal inocula of different infants are required. In this study an in vitro fermentation experiment was performed with galacto-oligosaccharides (GOS) using both individual and pooled faecal inocula of five two-week-old infants. It was shown that pooled faecal inoculum can be used to judge the fermentability of GOS, as a similar trend in total organic acid production and relative increase in Bifidobacterium was observed for the pooled faecal inoculum and four out of five individual faecal inocula. An additional fermentation using pooled faecal inoculum of five other infants of the same age confirmed these findings. Additionally, we provided evidence for both size- and isomer-specific fermentation of GOS by infant microbiota, which reveals the potential for the production of tailored NDC mixtures to meet the needs of specific subgroups of infants. Hence, the use of pooled faecal inocula contributes to increasing our knowledge on structure-specific effects of NDCs in infants efficiently.</p

Chicory inulin enhances fermentation of 2’-fucosyllactose by infant fecal microbiota and differentially influences immature dendritic cell and T-cell cytokine responses under normal and Th2-polarizing conditions

Scope: Non-digestible carbohydrates (NDCs) such as native chicory inulin and 2’-fucosyllactose (2’-FL) are added to infant formula to mimic some of the human milk oligosaccharide (HMO) functions. It is unknown whether combining inulin and 2’-FL influences their fermentation kinetics and whether the immune-modulatory effects of these NDCs are different under normal and inflammatory-prone Th2-polarizing conditions. Methods and results: We investigated the in vitro fermentation of 2’-FL and native chicory inulin, fermented individually and combined, using fecal inocula of 8-week-old infants. Native inulin was fermented in a size-dependent fashion and expedited the fermentation of 2’-FL. Fermentation of both native inulin and 2’FL increased the relative abundance of Bifidobacterium, which coincided with the production of acetate and lactate. The fermentation digesta of all fermentations differentially influenced both dendritic cell and T-cell cytokine responses under normal culture conditions or in presence of the Th2-polarizing cytokines IL-33 and TSLP, with the most pronounced effect for IL-1β in presence of TSLP. Conclusions: Our findings show that native inulin can expedite the fermentation of 2’-FL by infant fecal microbiota and that these NDC fermentation digesta have different effects under normal and Th2-polarizing conditions, indicating that infants with different immune backgrounds might benefit from tailored NDC formulations

Chicory inulin enhances fermentation of 2’-fucosyllactose by infant fecal microbiota and differentially influences immature dendritic cell and T-cell cytokine responses under normal and Th2-polarizing conditions

Scope: Non-digestible carbohydrates (NDCs) such as native chicory inulin and 2’-fucosyllactose (2’-FL) are added to infant formula to mimic some of the human milk oligosaccharide (HMO) functions. It is unknown whether combining inulin and 2’-FL influences their fermentation kinetics and whether the immune-modulatory effects of these NDCs are different under normal and inflammatory-prone Th2-polarizing conditions. Methods and results: We investigated the in vitro fermentation of 2’-FL and native chicory inulin, fermented individually and combined, using fecal inocula of 8-week-old infants. Native inulin was fermented in a size-dependent fashion and expedited the fermentation of 2’-FL. Fermentation of both native inulin and 2’FL increased the relative abundance of Bifidobacterium, which coincided with the production of acetate and lactate. The fermentation digesta of all fermentations differentially influenced both dendritic cell and T-cell cytokine responses under normal culture conditions or in presence of the Th2-polarizing cytokines IL-33 and TSLP, with the most pronounced effect for IL-1β in presence of TSLP. Conclusions: Our findings show that native inulin can expedite the fermentation of 2’-FL by infant fecal microbiota and that these NDC fermentation digesta have different effects under normal and Th2-polarizing conditions, indicating that infants with different immune backgrounds might benefit from tailored NDC formulations