|dc.description.abstract||The human colon contains about 20 trillion commensal bacteria belonging to about 160 species per human. The mechanisms by which individual species of these bacteria evade host inflammatory immune responses and instead establish a mutually beneficial relationship with the host is not fully understood. Bacteria that colonise the gut during infancy may be important contributors to human health throughout life. Recent research revealed a unique anti-inflammatory mechanism activated by Bacteroides thetaiotaomicron, a commensal predominant in the adult gut. Recognition of an inflammatory stimulus by a pattern recognition receptor (PRR) leads to activation of the transcription factor ‘nuclear factor kappa B’ (NF-kB) which is translocated to the cell nucleus where it induces transcription of genes encoding pro-inflammatory cytokines. However, Bacteroides thetaiotaomicron initiated formation of a complex between the NF-κB subunit RelA and the nuclear hormone receptor, peroxisome proliferator activated receptor-γ (PPAR-γ). This complex was removed from the nucleus, resulting in the attenuation of NF-κB-mediated inflammatory gene expression.
In the present study, bacterial strains predominant in the infant gut were tested for their immuno-modulatory effects with a hypothesis, “Bacteria predominant in the infant gut contribute to innate immunological tolerance and homeostasis by regulating signalling cascades in epithelial cells.” Strains were selected based on pyrosequencing analysis done in house that compared faecal microbiota composition of infants fed breast milk, goat milk formula and cow milk formula. Based on these observations, Bifidobacterium species (B. longum, B. breve and B. bifidum), members of the Lachnospiraceae family (Ruminococcus gnavus and Blautia producta), and the Bacteroidaceae family (Bacteroides vulgatus, Bacteroides fragilis and Bacteroides thetaiotaomicron) were tested. Polyinosinic-polycytidylic acid [poly(I:C)], the synthetic analogue of viral double-stranded RNA (dsRNA) was used to induce an inflammatory transcriptional response. HT-29 intestinal epithelial cells were incubated with freeze-dried preparations of either non-heat-treated (NHT) or heat-treated (HT) bacteria with or without poly(I:C) to quantify the bacterial modulation of poly(I:C)- induced interleukin-8 (IL-8) gene trancription and protein synthesis. NHT, but not HT, B. thetaiotaomicron significantly down-regulated poly(I:C) induced IL-8 gene transcription and protein synthesis. This strain was used as a positive control throughout the study. Overall, significant modulation of poly(I:C) induced IL-8 gene expression and protein synthesis by the various bacteria tested was observed. The effect of heat-treatment on bacterial modulatory effects differed depending on the strain.
Bacterial modulation of the expression of 88 other genes involved in the poly(I:C) induced signalling pathway was investigated. Poly(I:C) increased transcription of the gene that codes for RIG-I, one of the PRRs that binds dsRNA. It also increased transcription of genes encoding NF-κB inhibitor, alpha (NFKBIA), IL-8, chemokine (C-X-C) motif ligand 10 (CXCL10), CXCL11 and tumour necrosis factor (TNF). Most of the bacterial strains tested decreased transcription of these poly(I:C)-induced genes.
The ability of Blautia producta to down-regulate pro-inflammatory gene transcription, has not been reported previously. Blautia producta belongs to the family Lachnospiraceae, which has greater abundance in the faeces of formula-fed infants compared to breast-fed babies. The finding that all three Bifidobacterium species attenuated expression of the caspase recruitment domain- containing protein 9 (CARD9) gene, a key transducer for several pathogen induced antimicrobial signalling pathways, is also new. It is therefore evident that bacterial species present in the infant bowel can regulate signalling cascades in epithelial cells, thus helping to maintain intestinal homeostasis.||