Table S1. Analysis of Concentrations of 23 Cytokines and Chemokinesfrom Colonic Explant

Table S1. Analysis of concentrations of 23 cytokines and chemokinesfrom colonic explant and stimulated MLN cultures. The fold change in the median value and the P-value for each of the designated comparisons are shown .Statistics: Explant cultures: Mann-Whitney U-test; MLN cultures: Kruskal-Wallis test (KW) with Dunns post-hoc test.

Figure S1. Explants from the proximal and distal colons of BL/6 and Win mice (N=6, 20-24 wk) were cultured overnight, and cytokines measured in supernatants using a Bio-Plex assay. Concentrations are shown for all the cytokines/chemokines falling within the detectable range Statistics: median and IQR; Mann range. Mann-Whitney U test P values shown.

Figure S2. CD11c+ DC from LPMC (pooled from four 11-12 wk old mice) were sorted by flow cytometry and 2x105 cells/ml cultured with 100 ng/ml LPS. Culture supernatants were collected after 48 h and cytokines measured using a Bio-Plex assay. Concentrations are shown for all the cytokines/chemokines falling within the detectable range.

Figure S3 Immunophenotyping reveals no significant differences in the major leukocyte populations of primary and secondary lymphoid organs. (A) Enumeration of MLN leukocytes from wild type (BL/6) and Winnie (Win) mice (N=8-10, 10-14 wk). (B) Immunophenotyping by flow cytometry to enumerate lymphoid cells (T cells (CD4, CD8) and B cells (CD19)), myeloid cells (CD11b) and NKT cells (CD3+CD4+NK1.1+). The proportion of cells from the total live gate is shown. (C) Immunophenotyping of primary lymphoid tissues using the same markers as in (B). Bone marrow cells were divided into lymphoid and myeloid based on forward and side scatter and proportions shown are % of cells from either lymphoid ( CD4, CD8, B cells) or myeloid gating (CD11b). Thymic single cells were stained for CD4 and CD8. (D) Enumeration of Treg (CD4+CD25+Foxp3+) from MLN and spleen from wild type (BL/6) and Winnie (Win) mice. Representative staining for surface markers CD4 and CD25 followed by intra-cellular staining for Foxp3 in MLN. Statistics: median and IQR, Mann-Whitney U test P values shown.Statistics: (A) Median and IQR, Mann-Whitney U test; (B-C) mean ± SEM.

Figure S4. (A) Immunophenotyping for activated DC and memory T cells in MLN show no difference between BL/6 and Winnie. Leukocytes from MLN were stained for CD11c and MHC class II. The percentage of total double positive CD11c+MHC Class II hi cells are shown in the adjoining bar graph. (B) Phenotyping for the regulatory DC population in MLN was done by staining for CD11c and CD103. Total percentage of CD11c+ leukocytes showing CD103 positivity is represented in the adjoining bar graph. (C) Memory T cells (central) were classified as CD3+CD62Lhi and CD44+. The figure shows CD3+CD62Lhi cells gated for CD44 expression. Adjoining graph shows the percentage of CD3+CD62Lhi showing % of CD44+ cells.

Figure S5. Serum concentrations of cytokines and chemokines from BL/6 (N=4,10-12wk) and Winnie (N=6, 10-12 wk) assessed using a Bio-plex assay. Concentrations are shown for all the cytokines/chemokines falling within the detectable range. Statistics: median and IQR,Mann-Whitney U test.