Polystyrene beads were used as a control with common genes induced by the beads and the bacteria, suggesting that these genes may represent a gene signature Tanespimycin related to M-cell translocation. There were, however, genes that were specifically induced by the bacteria but not by the beads. These genes included the transcription factors that mediate the immediate-early response EGR1, FOS and JUN, the negative regulator ZFP36, and the phosphatase DUSP1.

These genes have previously been linked by cluster analysis in studies examining the response of human epithelial lung cells to avian influenza, endothelial cells stimulated with IL-1 and in tumour microarray analyses.30–32 The selective activation of these genes by the bacteria but not the beads indicates that the bacteria are activating and being sensed by the M cells, whereas the beads are merely being translocated non-specifically. Escherichia

coli and B. fragilis also activated more pro-inflammatory genes than L. salivarius, which again may be related to the lower translocation efficiency of L. salivarius. These inflammatory genes included the chemokines CXCL1, CXCL2 and IL8 which are potent chemoattractants for neutrophils and other immune cell types. Neutrophil recruitment is critical for clearance of bacteria once they have translocated the epithelium.33–35CXCL2 has Dorsomorphin manufacturer also been shown to be up-regulated in vivo in Peyer’s patches and mesenteric lymph nodes coincident with the accumulation of monocytes and neutrophils in these tissues.36 It is interesting to note that the increased

translocation efficiency of E. coli and B. fragilis compared with L. salivarius, was associated with a more potent induction of pro-inflammatory genes. NFKBIZ, NFKBIA and TNFAIP3 inhibit nuclear factor-κB signalling via a negative feedback loop.37–39NFKBIZ is induced by lipopolysaccharide, which could explain why it is induced by E. coli and B. fragilis but not L. salivarius.40 The microarray data were confirmed by qRT-PCR for selected genes, including IL8 and EGR1, and the same changes Resveratrol for each of the bacteria and the beads were observed, which validates our microarray findings. The gene ANKRD37, an HIF1α-inducible gene, was also confirmed, illustrating activation by all three bacterial species,41 whereas the gene, NR4A1, which is involved with fibronectin adhesion,42 was reduced in expression in C2-M cells treated with bacteria and the beads. It is also interesting to note that we observed differential expression of PRRs in C2-M cells compared with control differentiated C2 cells. MRC1 was highly expressed in C2-M cells compared with C2 cells and has previously been observed by Hase et al.43 to be increased in the follicle-associated epithelium overlying the murine Peyer’s patch.