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Ceramidase

Supplementary Materials Appendix EMBJ-37-e97537-s001

Supplementary Materials Appendix EMBJ-37-e97537-s001. and reveal an NKT cellCDC crosstalk mainly because a key mechanism for the regulation AZD-0284 of gut homeostasis. (SFB) (Ivanov whether CD1d expression on CD11c+ cells is required to induce Nur77 upregulation in iNKT cells in response to commensal\derived antigens. Single\cell suspensions from the mLN of Cre? and AZD-0284 Cre+ CD1dfl/flCD11cCre mice were prepared and incubated with commensal bacteria, and iNKT cell activation was detected as upregulation of Nur77 expression by intracellular staining. While commensal bacteria induced Nur77 upregulation in iNKT cells from Cre? CD1dfl/flCD11cCre cultures, iNKT Rabbit Polyclonal to CDC25C (phospho-Ser198) cell activation was absent in Cre+ CD1dfl/flCD11cCre cultures (Fig?1I). Thus, altogether our data suggest that CD1d expression in CD11c+ cells is necessary to mediate iNKT cell responses to intestinal lipids. CD1d\dependent presentation of intestinal lipids by CD11c+ cells controls the homeostasis and activation of intestinal iNKT cells We next investigated whether lipid presentation by CD11c+ cells controls intestinal iNKT cell homeostasis by analysing the iNKT cell population in the intestinal compartment of CD1dfl/flCD11cCre mice (Fig?2A and B, Appendix?Figs S3 and S4). AZD-0284 Analyses of tissues from WT (C57BL/6) and Cre? CD1dfl/flCD11cCre mice revealed that, as previously reported (Lee ?0.0001, two\tailed unpaired and phyla. To identify bacterial taxa that are significantly affected by NKT cells, we used the Wilcoxon test to compare the relative abundance of specific taxa colonizing CD1d\KO and littermate control mice. To avoid false positives as the result of multiple comparisons, the BenjaminiCHochberg false discovery rate (FDR) was put on those taxa that differed considerably ((OTU04, OTU33, OTU58 and OTU123) in the intestinal lumen of Compact disc1d?/? mice vs. littermate settings (Fig?3C). Especially considerable was the decrease noticed for OTU4 (unclassified (UC) as well as the family members (Fig?3A and D). No variations were within SFB (that are recognized to colonize the ileum wall structure) between WT and KO mice, by deep sequencing or qPCR (Appendix?Fig S7C). Put into this, no significant variations had been within the Shannon variety index between KO and WT mice, suggesting that Compact disc1d/NKT cells usually do not impact the total diversity of the intestinal microbiota (Appendix?Fig S7B). In the caecum, we measured a decrease in the total number of bacteria in CD1d?/? mice, but we did not detect any significant differences in any bacterial taxa between CD1d?/? and CD1d+/? mice (Appendix?Fig S7D and E). Open in a separate window Physique 3 CD1d and NKT cells regulate the intestinal microbiota A Principal coordinates analysis (PCoA) using the YueCClayton distances obtained for bacterial samples from the ileum content and ileum wall of CD1d+/? and CD1d?/? mice. The axes show the percentage of variation explained by PC1 and PC2. Each dot corresponds to one mouse.B Average relative abundance of the most frequent ( ?1%) operational taxonomic units (OTUs) of the ileum content and ileum wall from CD1d+/? and CD1d?/? mice. Bacterial taxa (at the genus level, or the closest level of classification) are shown, grouped by phylum and labelled with different colours as indicated. UC, unclassified.C, D Relative abundance of specified OTUs in the ileum content (C) and of specified taxa in the ileum wall (D) from CD1d+/? and CD1d?/? mice.ECG C57BL/6 mice were orally administered GalCer, and faecal bacteria were analysed before (d0) and 10?days (d10) after the treatment. (E) PCoA using the YueCClayton distances obtained among faecal samples at d0 and d10. The axes show the percentage of variation explained by PC1 and PC2. Each dot corresponds to one mouse. (F) Average relative abundance of the most frequent ( ?1%) OTUs at d0 and d10. Taxa are shown and labelled with different colours as indicated. (G) Relative abundance of the specified phyla, before and 10?days after GalCer treatment.Data information: In the boxplots, lines indicate the median, boxes show the 75th and the 25th percentiles and whiskers indicate the maximum and minimum values. *and and a decrease in and (Fig?3G). Accordingly, at the OTU level, we detected a significant decrease in bacteria belonging to the phylum (i.e. UC Lachnospiraceae) and an increase in OTUs belonging to the phylum (i.e. UC Bacteroidales; Appendix?Fig S7F). It is worth noting that while CD1d\dependent iNKT cell activation resulted in an increase in (Fig?3F and G), mice deficient in CD1d/NKT cells showed the opposite phenotype with a decrease in OTUs belonging to the phylum (Fig?3B and C). Hence, our data demonstrate that entirely.