controller 22  (Genovis Inc)

Journal: Cancer Discovery

Article Title: Aryl Hydrocarbon Receptor Ligands Drive Pancreatic Cancer Initiation and Progression through Protumorigenic T-cell Polarization

doi: 10.1158/2159-8290.CD-25-0377

Figure Lengend Snippet: Cigarette smoke and TCDD increase IL22 production in an AHR-dependent manner. A − C, In vitro naïve CD4 + T-cell polarization schema ( A ). CYP1A1 mRNA expression, IL22 mRNA expression, and IL22 protein concentration in culture supernatant after in vitro naïve CD4 + T-cell polarization with CSE ( B and C, n = 3/group), 6-formylindolo(3,2-b)carbazole (FICZ) and TCDD ( D, n = 2–3/group), and TCDD with AHR inhibitor CH-223191 ( E, n = 3/group). F, Experimental schema of naïve CD4 + T-cell polarization with human splenocytes. G, CYP1B1 and IL22 mRNA expression and IL22 protein concentration in the culture supernatant after naïve CD4 + T-cell polarization with TCDD and CH-223191 ( n = 3 technical replicates/group). H, Experimental schema, wherein the culture supernatant of polarized naïve CD4 + T cells was placed upon 7940b PDAC cells for 15 minutes, and protein was then isolated for Western blot. I, Western blot showing pSTAT3, total STAT3, and GAPDH expression among 7940b PDAC cells treated with unpolarized Th0 T-cell media, TCDD-polarized T-cell media, TCDD-polarized media with IL22-binding protein (IL22bp), TCDD alone, negative control, and IL22 alone. J, Schema of orthotopic tumor model with CSE or TCDD. K, IL22 mRNA expression in orthotopic tumors treated with CSE ( n = 3–5/group) or TCDD ( n = 4/group). Data represented as the mean ± SD unless otherwise noted (portion of the figure created with BioRender.com ).

Article Snippet: Controls included 1 nmol/L TCDD in complete DMEM (TCDD alone), negative control with DMEM media alone, and positive control with IL22 (20 ng/mL, R&D Systems, #582-ML-010) in complete DMEM.

Techniques: In Vitro, Expressing, Protein Concentration, Isolation, Western Blot, Binding Assay, Negative Control

Journal: Cancer Discovery

Article Title: Aryl Hydrocarbon Receptor Ligands Drive Pancreatic Cancer Initiation and Progression through Protumorigenic T-cell Polarization

doi: 10.1158/2159-8290.CD-25-0377

Figure Lengend Snippet: TCDD induced PDAC growth is mediated by IL22 signaling and Treg infiltration. A, Genetic makeup of the IL22-tdTomato reporter mouse (Catch-22) and experimental schema showing 3-week treatment course with TCDD. B and C, Representative mfIHC ( B ) and quantification of IL22 + immune cells ( C ) in the duodenum of Catch-22 mice treated with vehicle or TCDD for 3 weeks, n = 3 mice/group, n = 25–26 regions of interest/group. D, Genetic makeup of the IL22 eGFP reporter mouse, with quantification of the proportion of T cells, IL22 + T cells, ILCs, and IL22 + ILCs among pancreata of mice treated with 3 weeks of vehicle or TCDD, n = 3/group. E and F, Experimental schema, n = 6–9/group, and tumor weight and volume for treatment of WT mice with TCDD, IL22 −/− mice with TCDD, and IL22 −/− mice with vehicle for 19 days. G and H, Representative mfIHC ( G, scale bars, 50 μm, inlaid white arrows identifying CD3 + FOXP3 + Tregs), and quantification of T cell, Th cell, CD8 + T cell, and Treg infiltration via mfIHC ( H ), n = 4–5 mice/group, and quantification of 17 regions of interest/group. Phenotypes were identified as follows: T cells (CD3 + ), Th cells (CD3 + CD8 − FOXP3 − ), CD8 + T cells (CD3 + CD8 + ), and Tregs (CD3 + FOXP3 + CD8 − ). I, Quantification of T cells, CD4 + T cells, CD8 + T cells, and Tregs via flow cytometry of orthotopic tumors of WT mice treated with vehicle or TCDD, n = 3–4/group. J, Experimental design, n = 4–8/group, and tumor weight and volume of the orthotopic model of WT mice or FOXP3- DTR mice, allowing for inducible depletion of Tregs, treated with vehicle or TCDD for 16 days. Groups: WT treated with vehicle, WT treated with TCDD, and FOXP3- DTR treated with TCDD. K, Experimental schema and representative gating strategy on CD45 + CD3 + intratumoral T cells from orthotopic tumors of WT mice treated with vehicle or TCDD for 19 days. L, Quantification of IL22 + cells among CD4 + T cells, FOXP3 + cells among CD4 + T cells, and IL22 + cells among FOXP3 + Tregs. Data are represented as the mean ± SD unless otherwise noted. DAPI, 4′6-diamidino-2-phenylindole.

Article Snippet: Controls included 1 nmol/L TCDD in complete DMEM (TCDD alone), negative control with DMEM media alone, and positive control with IL22 (20 ng/mL, R&D Systems, #582-ML-010) in complete DMEM.

Techniques: Flow Cytometry

Journal: Cancer Discovery

Article Title: Aryl Hydrocarbon Receptor Ligands Drive Pancreatic Cancer Initiation and Progression through Protumorigenic T-cell Polarization

doi: 10.1158/2159-8290.CD-25-0377

Figure Lengend Snippet: TCDD-mediated IL22 production and Treg accumulation is dependent upon CD4 + T cell AHR signaling. A, CYP1A1 and CYP1B1 mRNA expression from CD4 + TILs isolated from orthotopic tumors of WT mice treated with vehicle or TCDD for 19 days. B, Genetic makeup of the CD4 Cre AHR mice with inducible CD4-specific deletion of AHR signaling. C, IL22 mRNA expression and protein concentration in the culture supernatant of naïve CD4 + T cells isolated from WT or CD4 Cre AHR mice and polarized under nonstimulating (Th0) conditions, with TCDD, or with TCDD and CH-223191, an AHR inhibitor. D − F, Experimental design ( D ), tumor weight and volume ( E ), and CYP1A1 and IL22 mRNA expression ( F ) from tumors of an orthotopic model utilizing WT mice and CD4-Cre AHR −/− mice treated with vehicle or TCDD, n = 4–7 mice/group. G and H, Representative mfIHC ( G, scale bars, 50 μm, inlaid white arrows identifying CD3 + CD8 + T cell), and quantification ( H ) of CD8 + T cell (CD3 + CD8 + FOXP3 − ) and Treg (CD3 + FOXP3 + CD8 − ) infiltration among orthotopic tumors of WT or CD4 Cre AHR mice treated with vehicle or TCDD, n = 17–39 regions of interest/group. I, Experimental design and ( J ) tumor weight and volume for WT mice treated with vehicle, AHR inhibitor CH-223191, TCDD, or TCDD and the AHR inhibitor for 19 days, n = 7–10/group. Data represented as the mean ± SD unless otherwise noted. 4′6-diamidino-2-phenylindole.

Article Snippet: Controls included 1 nmol/L TCDD in complete DMEM (TCDD alone), negative control with DMEM media alone, and positive control with IL22 (20 ng/mL, R&D Systems, #582-ML-010) in complete DMEM.

Techniques: Expressing, Isolation, Protein Concentration