Figure S1.) Tslp-/- mice have normal Treg cells, numbers and activation of DCs. (A) Cells from the spleen and MLN of naïve Tslp+/+ and Tslp-/- mice were ...
Immunity, Volume 35
Supplemental Information Thymic Stromal Lymphopoetin-Induced Expression of the Endogenous Inhibitory Enzyme SLPI Mediates Recovery from Colonic Inflammation Colin Reardon, Matthias Lechmann, Anne Brüstle, Mélanie G. Gareau, Naomi Shuman, Dana Philpott, Steven F. Ziegler, and Tak W. Mak
Inventory of Supplementary Items: Figure S1. TSLP-/- mice have Treg cells, normal numbers and activation of DCs. Flow cytometry, and qRT-PCR comparing Treg cell, and DC numbers and activation status in Tslp+/+ and Tslp-/- mice. Figure S2. Onset of colitis, severity, and sensitivity to DSS is not enhanced in Tslp-/- or Crlf2/-
mice. (related to Figure 1)
Demonstrates lack of enhanced inflammation: at a time point earlier post-DSS, with a lower dose of DSS, and a head to head comparison of WT, Tslp-/- and Crlf2-/- mice. Data presented are weight loss, summary of histological damage scores, and survival curve for head to head comparison of three genotypes.
Figure S3 in TSLP does not alter the response to OVA immunization. (related to Figure 3) Indicates no enhanced Th1 immune skewing, and an intact Th2 response in a T-cell dependent antibody response by assaying for OVA specific serum antibodies by ELISA.
Figure S4 Intestinal barrier function and the microbiota are not altered in Tslp-/- mice. Data presented assess the intestinal barrier function by determining bacterial translocation, detection of gavaged dextran probe in serum, IgA+ B-cells in Peyer’s patches, goblet cell numbers, and composition of the microbiota.
Figure S5 Figure S5.) Impact of TSLP deficiency on colonic gene expression. (related to Figure 4) qRT-PCR data for genes involved in various aspects of wound repair.
Figure S6 IEC apoptosis and proliferation after neutrophil elastase inhibition. (related to Figure 5) Data presented include: nascent collagen synthesis quantified by Sircol assay, summarized IHC findings for cleaved caspase3 (apoptosis), Ki67 (proliferation), PRGN protein in the colon of WT or Tslp-/- mice treated with vehicle or the NE inhibitor SSR69071. Figure S7 Progranulin and collagen expression in Tslp-/- mice or with NEi treatment. (related to Figure 5) qRT-PCR data for PRGN and various intestinal predominant collagen genes in naïve, DSS (8 days post), or DSS + SSR69071 treated WT or Tslp-/- mice.
Table S1 (related to Figure 1) Provides results of Erythrocyte and Thrombocyte analysis.
Table S2 Provides all primer sequences used in this study for qRT-PCR.
Tslp-/-
B.)
2.6±0.26
2.9±0.29
4.1±0.23
4.1±0.13
Tslp
CD4 APC
*
Tslp+/+
CON DSS CON DSS +/+ -/-
Tslp+/+
% MAX
Gate: R3
R3 R4
Tslp-/-
% MAX
CD40 FITC
CD11b APC
% MAX
CD80 FITC
CD86 FITC
Figure S1.
Tslp-/-
2.67±0.26
PDCA-1 PE
Spleen
D.)
MHCII PE
*
C.)
CD11b APC
Tslp+/+
Foxp3 Fold expressio on
Fo oxp3 Alexa488
MLN
Spleen n
A.)
MLN Gate: R4
Gate: R3
Gate: R4
2.16±0.21
Figure S1.) Tslp-/- mice have normal Treg cells, numbers and activation of DCs. (A) Cells from the spleen and MLN of naïve Tslp+/+ and Tslp-/- mice were assessed for CD4+ Foxp3+ T-cells T cells by flow cytometry. cytometry (B) Colonic Foxp3 expression was evaluated by qRT-PCR on colonic tissue from naïve mice or 8 days post-DSS. (C) Splenic plasmacytoid DC (7aad- FSChi CD11c+/low, PDCA-1+ CD11b-) were assessed by flow cytometry. (D) DC in the spleen or MLN (7aad- FSChi CD11c+MHCII+ CD11b- or 7aadFSChi CD11c+ MHCII+ CD11b+) and their activation status were assayed. Left panel; representative dot plot of splenic DCs, cells contained in “Gate R3” are: 7aad- FSChi CD11c+ MHCII+ CD11b-, “Gate Gate R4 R4” are: 7aad- FSChi CD11c+ MHCII+ CD11b+. Middle panels activation status (CD40, CD80, CD86) of cells contained in the respective gate from the spleen, right panels are cells isolated from MLN. (n=3 mice/genotype, mean ± SD, *P
