Journal: Immunity

Article Title: Thymic stromal lymphopoetin-induced expression of the endogenous inhibitory enzyme SLPI mediates recovery from colonic inflammation

doi: 10.1016/j.immuni.2011.05.015

Figure Lengend Snippet: Inflammation was assessed in DSS-treated Tslp-/- mice by (A) macroscopic disease, (B) colon length, (C) serum SAA concentration, (D&E) histological damage, (F) weight loss, and (G) survival. Graphs indicate: control (open bars), DSS (solid bars), mean ± SD, n=10-15 mice/group, scale bar=50μm, *P<0.05, ANOVA.

Article Snippet: Cells were stimulated with recombinant mouse TSLP (R&D systems).

Techniques: Concentration Assay, Control

Journal: Immunity

Article Title: Thymic stromal lymphopoetin-induced expression of the endogenous inhibitory enzyme SLPI mediates recovery from colonic inflammation

doi: 10.1016/j.immuni.2011.05.015

Figure Lengend Snippet: FACS sorted CD4+CD45RBhi (Teff) ± CD4+CD45RBlo from Crlf2+/+ or Crlf2-/- were transferred to Rag1-/- recipients and monitored for (A) weight loss, (B) macroscopic disease, (C) colon length, and (D&E) histological damage (n=4-5 mice/group). (F) STAT5 phosphorylation was assessed following stimulation with rmIL-7 in CD4+ or CD8+ T-cells from Crlf2-/-, Tslp-/-, or WT (n=3 mice). (mean ± SD, scale bar =50μm)

Article Snippet: Cells were stimulated with recombinant mouse TSLP (R&D systems).

Techniques: Phospho-proteomics

Journal: Immunity

Article Title: Thymic stromal lymphopoetin-induced expression of the endogenous inhibitory enzyme SLPI mediates recovery from colonic inflammation

doi: 10.1016/j.immuni.2011.05.015

Figure Lengend Snippet: (A) Gene expression in colonic tissue was assessed by qRT-PCR (mean ± SD, n=10-15 mice/group, , control (open bars), DSS (solid bars), *P<0.05, ANOVA). (B) Mason's trichrome staining for collagen (blue) on colonic tissues from control and DSS-treated Tslp+/+ and Tslp-/- mice. (20× magnification is from red rectangle, 6 mice/group, scale bars=50μm). (C) SLPI expression was determined by immunoblotting colonic homogenates (n=6 mice/group). (D) MMP and (E) NE activity in colonic homogenates was assessed by a substrate cleavage assay (mean ± SD, n=4-9 mice/group, control (open symbols), DSS (solid symbols) *P<0.05 by ANOVA). (F) PGRN was determined in colonic homogenates by immunobloting (n=6 mice/group).

Article Snippet: Cells were stimulated with recombinant mouse TSLP (R&D systems).

Techniques: Gene Expression, Quantitative RT-PCR, Control, Staining, Expressing, Western Blot, Activity Assay, Cleavage Assay

Journal: Immunity

Article Title: Thymic stromal lymphopoetin-induced expression of the endogenous inhibitory enzyme SLPI mediates recovery from colonic inflammation

doi: 10.1016/j.immuni.2011.05.015

Figure Lengend Snippet: DSS-treated Tslp-/- or WT mice were gavaged with the NE inhibitor SSR69071 (3mg/kg) or vehicle (A) from day 6-14 and monitored for; (upper panel) weight loss, and (lower panel) mortality. In separate experiments, mice were gavaged with rSLPI (B) and assessed for (upper panel) weight loss, and (lower panel) mortality. (C&E) IEC proliferation as determined by colonic Ki67 staining after 4 days of treatment (day 10 post-DSS) with vehicle or SSR69071, and (D) apoptosis in Tslp+/+ and Tslp-/- DSS-treated mice. (F) Colonic PGRN was determined on full thickness tissue homogenates 10 days post-DSS. (mean ± SD, n=4 mice/group, *P<0.05, ANOVA).

Article Snippet: Cells were stimulated with recombinant mouse TSLP (R&D systems).

Techniques: Staining

Journal: Immunity

Article Title: Thymic stromal lymphopoetin-induced expression of the endogenous inhibitory enzyme SLPI mediates recovery from colonic inflammation

doi: 10.1016/j.immuni.2011.05.015

Figure Lengend Snippet: (A) Confocal microscopy was performed for TSLP (blue), SLPI (red) and E-cadherin+ IEC (green) in Tslp+/+ mice (*, upper panels) and Tslp-/- mice (†, lower panels), (arrows, “L”=lumen, scale bars=20μm). WT mice reconstituted with Crlf2+/+ or Tslp-/- BM were subjected to colitis and assessed by (B) weight loss (C) colon length (D) Mason's trichrome staining (E) histological damage (F) and SLPI expression (mean ± SD, scale bar=100μm, n=3-4 mice/group, control (open bars), DSS (solid bars), *P<0.05, **P<0.001, ANOVA).

Article Snippet: Cells were stimulated with recombinant mouse TSLP (R&D systems).

Techniques: Confocal Microscopy, Mouse Assay, Staining, Expressing, Control

Journal: Immunity

Article Title: Thymic stromal lymphopoetin-induced expression of the endogenous inhibitory enzyme SLPI mediates recovery from colonic inflammation

doi: 10.1016/j.immuni.2011.05.015

Figure Lengend Snippet: (A) Primary IEC were stained for flow cytometry for epithelial markers (E-cadherin+ CD3-, B220-, F4/80-) and TSLPR. (B) IHC was performed on colonic tissue sections with anti-TSLPR antibody. (C) TSLPR expression was assessed on fibroblasts (Vimentin+, alpha-smooth muscle actin- (α-SMA), E-cadherin- CD3- CD11b- B220- F4/80-) or myofibroblasts (Vimentin+ α-SMA+, E-cadherin- CD3- CD11b- B220- F4/80-) from Crlf2+/+ (grey fill black outline), and Crlf2-/- (negative control, red line) mice. (D) ERK and AKT phosphorylation in primary IEC from Crlf2+/+ and Crlf2-/- mice were assessed following stimulation with rmTSLP (50ng/mL, 30 min, mean ± SD, *P<0.05, ANOVA). (E) AKT Foxo3A and ERK phosphorylation were assessed in mICc12 cells by immunoblotting following stimulation with rmTSLP for the indicated times (50ng/mL). (F) Expression of anti-apoptotic genes including the Foxo3a regulated BCL2, and SLPI were determined following exposure to rmTSLP. (G) SLPI mRNA in mICcl2 cells following TSLP exposure was determined by qRT-PCR (n=3, mean ± SD, *P<0.05, ANOVA).

Article Snippet: Cells were stimulated with recombinant mouse TSLP (R&D systems).

Techniques: Staining, Flow Cytometry, Expressing, Negative Control, Phospho-proteomics, Western Blot, Quantitative RT-PCR

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Thymic stromal lymphopoietin contributes to endometriotic lesion proliferation and disease-associated inflammation.

doi: 10.1093/jimmun/vkae021

Figure Lengend Snippet: Figure 1. CRLF2 mRNA and TSLP protein are elevated in patient endometriotic lesions compared with control endometrial samples. RT-qPCR was run with RNA extracted from control endometria (n ¼ 7), patient eutopic endometria (n ¼ 5), and endometriotic lesions (n ¼ 8). The expression of (A) TSLP, (B) CRLF2, and (C) IL-7R was quantified relative to the GAPDH reference gene. A TMA with (D) control endometrial tissue (n ¼ 15) and matched (E) patient (n ¼ 15) eutopic endometria and (F) endometriotic lesions was stained with anti-TSLP Ab. TSLP-positive (G) stromal and (H) epithelial area was calculated as a fraction of total stromal and epithelial area, respectively, using HALO AI. Magnification provided at 8×. Scale bars ¼ 100 μm. A 1-way analysis of variance with Tukey post hoc test was used to assess statistical significance. P < 0.05, P < 0.01.

Article Snippet: Cells were treated in triplicates with vehicle control (PBS) or various concentrations of recombinant mouse TSLP (rmTSLP) (555-TS-010/CF; R&D Systems; 1, 10, 50 ng/mL) for 7 d at 37 �C with 5% CO2.

Techniques: Control, Quantitative RT-PCR, Expressing, Staining

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Thymic stromal lymphopoietin contributes to endometriotic lesion proliferation and disease-associated inflammation.

doi: 10.1093/jimmun/vkae021

Figure Lengend Snippet: Figure 2. TSLP treatment alters the proliferation and apoptosis of human endometriosis-representative cell lines. (A, B) 12Zs, (C, D) hESCs, and (E, F) THP-1 cells were treated with vehicle control (PBS) or various concentrations of rhTSLP (1, 10, 50 ng/mL) for 24 h. (A, C, E) WST-I proliferation and (B, D, F) caspase-3/7 apoptosis assays were performed. A 1-way analysis of variance with Tukey post hoc test was used to assess statistical significance. P < 0.05, P < 0.01, P < 0.001, P < 0.0001.

Article Snippet: Cells were treated in triplicates with vehicle control (PBS) or various concentrations of recombinant mouse TSLP (rmTSLP) (555-TS-010/CF; R&D Systems; 1, 10, 50 ng/mL) for 7 d at 37 �C with 5% CO2.

Techniques: Control

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Thymic stromal lymphopoietin contributes to endometriotic lesion proliferation and disease-associated inflammation.

doi: 10.1093/jimmun/vkae021

Figure Lengend Snippet: Figure 3. TSLP treatment influences the polarization of THP-1 cells and mouse BMDMs. THP-1 cells were treated with vehicle control (PBS) or various concentrations of rhTSLP (1, 10, 50 ng/mL) for 1 d, and mouse BMDMs were treated with vehicle control (PBS) or various concentrations of rmTSLP (1, 10, 50 ng/mL) for 7 d. (A) Flow cytometry was used to detect the expression of (B) CD11b, (C) CD163, and CD86 on THP-1 cells, as well as (D) CD11b, F4/80, (E) CD80, and CD206 on BMDMs. Quantification of results is provided in Figs. S2 and S3. Cell supernatant was collected from BMDMs and analyzed to detect the expression of several inflammation- and angiogenesis-related cytokines and chemokines, including (F) MCP-1, (G) IP-10, and (H) MIP-1α. A 1-way analysis of variance with Tukey post hoc test was conducted to assess statistical significance. P < 0.05, P < 0.01, P < 0.001.

Article Snippet: Cells were treated in triplicates with vehicle control (PBS) or various concentrations of recombinant mouse TSLP (rmTSLP) (555-TS-010/CF; R&D Systems; 1, 10, 50 ng/mL) for 7 d at 37 �C with 5% CO2.

Techniques: Control, Flow Cytometry, Expressing

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Thymic stromal lymphopoietin contributes to endometriotic lesion proliferation and disease-associated inflammation.

doi: 10.1093/jimmun/vkae021

Figure Lengend Snippet: Figure 4. TSLP treatment influences the presence and activation of immune cells in the peritoneal fluid of mice induced with endometriosis. Mice were treated with intraperitoneal PBS (n ¼ 5) or rmTSLP (2 μg or 0.01 mg/kg; n ¼ 4) every other day for 14 d. (A–E) Myeloid and (F–J) lymphoid cells, as well as their (B, D, G, J) TSLPR expression, (E) IRF4 expression, and (F, J) IRF-4/T-bet ratio was measured. Quantification of results is provided in Fig. S4. An unpaired Student’s t test was used to assess statistical significance. P < 0.05, P < 0.01, P < 0.001.

Article Snippet: Cells were treated in triplicates with vehicle control (PBS) or various concentrations of recombinant mouse TSLP (rmTSLP) (555-TS-010/CF; R&D Systems; 1, 10, 50 ng/mL) for 7 d at 37 �C with 5% CO2.

Techniques: Activation Assay, Expressing

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Thymic stromal lymphopoietin contributes to endometriotic lesion proliferation and disease-associated inflammation.

doi: 10.1093/jimmun/vkae021

Figure Lengend Snippet: Figure 6. TSLP treatment reduces the concentrations of prevalent proinflammatory cytokines in mouse peritoneal fluid. Mice were treated with intraperitoneal PBS (n ¼ 5) or rmTSLP (2 μg or 0.01 mg/kg; n ¼ 4) every other day for 14 d. Peritoneal fluid was collected to detect predominant inflammatory cytokines and chemokines. Key (A, B) proinflammatory and (C, D) anti-inflammatory cytokines are shown. An unpaired Student’s t test was conducted to assess statistical significance. P < 0.05.

Article Snippet: Cells were treated in triplicates with vehicle control (PBS) or various concentrations of recombinant mouse TSLP (rmTSLP) (555-TS-010/CF; R&D Systems; 1, 10, 50 ng/mL) for 7 d at 37 �C with 5% CO2.

Techniques:

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Thymic stromal lymphopoietin contributes to endometriotic lesion proliferation and disease-associated inflammation.

doi: 10.1093/jimmun/vkae021

Figure Lengend Snippet: Figure 5. TSLP treatment influences the presence and activation of immune cells in circulation. Mice were treated with intraperitoneal PBS (n ¼ 5) or rmTSLP (2 μg or 0.01 mg/kg; n ¼ 4) every other day for 14 d. (A–C) Myeloid and (D–H) lymphoid cells and their (B, E, H) TSLPR expression, (C) IRF-4 expression, and (F, J) IRF-4/T-BET ratio were measured. Quantification of results is provided in Fig. S4. An unpaired Student’s t test was used to assess statistical significance. P < 0.05, P < 0.01, P < 0.001.

Article Snippet: Cells were treated in triplicates with vehicle control (PBS) or various concentrations of recombinant mouse TSLP (rmTSLP) (555-TS-010/CF; R&D Systems; 1, 10, 50 ng/mL) for 7 d at 37 �C with 5% CO2.

Techniques: Activation Assay, Expressing

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Thymic stromal lymphopoietin contributes to endometriotic lesion proliferation and disease-associated inflammation.

doi: 10.1093/jimmun/vkae021

Figure Lengend Snippet: Figure 7. TSLP treatment significantly increases the presence of TSLP within endometriotic lesions and lesion proliferation. Endometriotic lesions from mice treated with intraperitoneal PBS (n¼ 5) or rmTSLP (2μg or 0.01mg/kg; n¼ 4) were embedded in paraffin and stained with (A, B) anti-TSLP, (C, D) anti-TSLPR, (E, F) anti-Ki67, and (G, H) anti-CD31 Abs. (I) TSLP-positive, (J) TSLPR-positive, (K) Ki67-positive, and (L) CD31-positive tissue was calculated with HALO AI. Magnification provided at 20×. Scale bars ¼ 50μm. An unpaired Student’s t test was used to assess statistical significance. P< 0.05, P< 0.001.

Article Snippet: Cells were treated in triplicates with vehicle control (PBS) or various concentrations of recombinant mouse TSLP (rmTSLP) (555-TS-010/CF; R&D Systems; 1, 10, 50 ng/mL) for 7 d at 37 �C with 5% CO2.

Techniques: Staining