mil-17f  (R&D Systems)

Journal: Scientific Reports

Article Title: IL-17F induces inflammation, dysfunction and cell death in mouse islets

doi: 10.1038/s41598-020-69805-2

Figure Lengend Snippet: Pancreatic β-cells respond to IL-17F signals via IL-17RA and -RC. ( A ) Total RNA was isolated from Min6, NIT-1, pancreatic islets and colon tissue and the expression of Il17ra-e measured by Taqman quantitative RT-PCR. ( B ) NIT-1 cells and primary NOD islets were stained with antibodies against IL-17RA, RC and RD and cell surface expression quantified using flow cytometry (black line = antibody stained, grey line = unstained control). ( C ) NIT-1 cells were stimulated with IL-17F or IL-17A for 2 h and Nfkbiz expression measured by Taqman quantitative RT-PCR (n = 4 for all groups), p < 0.05 (*), p < 0.001 (***) One-Way ANOVA with Bonferonni’s Multiple Comparison test. ( D ) Min6 cells were stimulated with IL-17F or IL-17A for 4 h and Nfkbiz expression measured by Taqman quantitative RT-PCR (n = 4 for all groups), p < 0.001 (***) One-Way ANOVA with Bonferonni’s Multiple Comparison test. ( E ) NOD islets were stimulated with IL-17F or IL-17A for 4 h and Nfkbiz expression measured by Taqman quantitative RT-PCR (n = 3 for all groups), p < 0.001 (***) One-Way ANOVA with Bonferonni’s Multiple Comparison test. ( F ) IL-17RA knockout (left panel) and RC knockout (right panel) NIT-1 cells were generated with CRISPR/Cas9 and stained with antibodies against IL-17RA (left panel) and RC (right panel) (black = parental control cell line, grey = receptor knockout cell line, light grey = unstained control). ( E ) Wildtype, IL-17RA KO and IL-17RC KO NIT-1 cells were stimulated with IL-17F or IL-17A for 2 h and Nfkbiz expression measured by Taqman quantitative RT-PCR (n = 2 for all groups), p < 0.05 (*), p < 0.01 (**) Two-Way ANOVA with Bonferonni’s Multiple Comparison test.

Article Snippet: Mouse cytokines were used at the following concentrations: mTNFα (In Vitro Technologies) at 100 ng/ml, mIFNγ at 10 ng/ml (Australian Biosearch), hIL-1β at 50 ng/ml (Peprotech), mIL-17A at 25 ng/ml (R&D Systems) and mIL-17F at 100 ng/ml (R&D Systems).

Techniques: Isolation, Expressing, Quantitative RT-PCR, Staining, Flow Cytometry, Comparison, Knock-Out, Generated, CRISPR

Journal: Scientific Reports

Article Title: IL-17F induces inflammation, dysfunction and cell death in mouse islets

doi: 10.1038/s41598-020-69805-2

Figure Lengend Snippet: IL-17F induces chemokine expression in mouse β-cell lines and islets. Min6 cells were stimulated with IL-17F or IL-17A in the presence of TNFα + IFNγ (Cyto) for 24 h and ( A ) Nfkbiz , ( B ) Cxcl1 , ( C ) Cxcl2 , ( D ) Ccl20 expression measured by Taqman quantitative RT-PCR (n = 4 for all groups), p < 0.05 (*), p < 0.01 (**), p < 0.001 (***) One-Way ANOVA with Bonferonni’s Multiple Comparison test. NOD islets were stimulated with IL-17F or IL-17A in the presence of TNFα + IFNγ (Cyto) for 24 h and ( E ) Nfkbiz , ( F ) Cxcl1 , ( G ) Cxcl2 , ( H ) Ccl20 expression measured by Taqman quantitative RT-PCR (n = 6 for all groups), p < 0.05 (*), p < 0.001 (***) One-Way ANOVA with Bonferonni’s Multiple Comparison test.

Article Snippet: Mouse cytokines were used at the following concentrations: mTNFα (In Vitro Technologies) at 100 ng/ml, mIFNγ at 10 ng/ml (Australian Biosearch), hIL-1β at 50 ng/ml (Peprotech), mIL-17A at 25 ng/ml (R&D Systems) and mIL-17F at 100 ng/ml (R&D Systems).

Techniques: Expressing, Quantitative RT-PCR, Comparison

Journal: Scientific Reports

Article Title: IL-17F induces inflammation, dysfunction and cell death in mouse islets

doi: 10.1038/s41598-020-69805-2

Figure Lengend Snippet: IL-17F suppresses expression of β-cell identity genes and glucose stimulated insulin secretion in mouse islets. Min6 cells were stimulated with IL-17F or IL-17A in the presence of TNFα + IFNγ (Cyto) for 24 h and ( A ) Ins1 , ( B ) Ins2 , ( C ) Glut2 , ( D ) Pdx1 and ( E ) Foxo1 expression measured by Taqman quantitative RT-PCR (n = 4 for all groups). NOD islets were stimulated with IL-17F or IL-17A in the presence of TNFα + IFNγ (Cyto) for 24 h and ( F ) Ins1 , ( G ) Ins2 , ( H ) Glut2 , ( I ) Pdx1 and ( J ) Foxo1 expression measured by Taqman quantitative RT-PCR (n = 6 for all groups), p < 0.05 (*), p < 0.01 (**), p < 0.001 (***) One-Way ANOVA with Bonferonni’s Multiple Comparison test. ( K ) NOD islets were stimulated with IL-17F or IL-17A in the presence of TNFα + IFNγ (Cyto) for 48 h and glucose stimulated insulin secretion assay performed (3 independent experiments, n = 4 for all groups), p < 0.05 (*), p < 0.01 (**), p < 0.001 (***) Two-Way ANOVA with Bonferonni’s Multiple Comparison test.

Article Snippet: Mouse cytokines were used at the following concentrations: mTNFα (In Vitro Technologies) at 100 ng/ml, mIFNγ at 10 ng/ml (Australian Biosearch), hIL-1β at 50 ng/ml (Peprotech), mIL-17A at 25 ng/ml (R&D Systems) and mIL-17F at 100 ng/ml (R&D Systems).

Techniques: Expressing, Quantitative RT-PCR, Comparison

Journal: Scientific Reports

Article Title: IL-17F induces inflammation, dysfunction and cell death in mouse islets

doi: 10.1038/s41598-020-69805-2

Figure Lengend Snippet: IL-17F induces cell death in mouse islets. NOD islets were stimulated with IL-17F or IL-17A in the presence of ( A , C ) TNFα + IFNγ or ( B , D ) IL-1β + IFNγ for 4 days and cell death quantified by DNA fragmentation (n = 5 for TNFα + IFNγ and n = 2 for IL-1β + IFNγ), p < 0.05 (*), p < 0.001 (***) One-Way ANOVA with Bonferonni’s Multiple Comparison test. ( C ) C57Bl/6 islets were stimulated with IL-17F or IL-17A in the presence of ( C ) TNFα + IFNγ or ( D ) IL-1β + IFNγ for 4 days and cell death quantified by DNA fragmentation (n = 3 for all groups), p < 0.05 (*), p < 0.001 (***) One-Way ANOVA with Bonferonni’s Multiple Comparison test.

Article Snippet: Mouse cytokines were used at the following concentrations: mTNFα (In Vitro Technologies) at 100 ng/ml, mIFNγ at 10 ng/ml (Australian Biosearch), hIL-1β at 50 ng/ml (Peprotech), mIL-17A at 25 ng/ml (R&D Systems) and mIL-17F at 100 ng/ml (R&D Systems).

Techniques: Comparison

Journal: Scientific Reports

Article Title: IL-17F induces inflammation, dysfunction and cell death in mouse islets

doi: 10.1038/s41598-020-69805-2

Figure Lengend Snippet: IL-17F induced cell death is iNOS dependent in mouse islets. NOD islets were stimulated with IL-17F or IL-17A in the presence of ( A ) IFNγ or ( B ) TNFα for 4 h and Nos2 expression measured by Taqman quantitative RT-PCR (n = 5 for all groups), p < 0.05 (*), p < 0.01 (**) One-Way ANOVA with Bonferonni’s Multiple Comparison test. ( C ) Min6 cells, ( D ) NOD islets and ( E ) C57Bl/6 islets were stimulated with IL-17F or IL-17A in the presence of TNFα + IFNγ (Cyto) for 24 h and Nos2 expression measured by Taqman quantitative RT-PCR (n = 4 for Min6 cells and C57Bl/6 islets, n = 6 for NOD islets), p < 0.05 (*), p < 0.001 (***) One-Way ANOVA with Bonferonni’s Multiple Comparison test. ( F ) NOD islets were stimulated with IL-17F or IL-17A and TNFα + IFNγ (Cyto) +/− Jnk inhibitor (SP600125) at 50 μM for 24 h and Nos2 expression measured by Taqman quantitative RT-PCR (n = 2 for all groups), p < 0.001 (***) Two-Way ANOVA with Bonferonni’s Multiple Comparison test. ( G ) NOD islets were stimulated with IL-17F or IL-17A and TNFα + IFNγ (Cyto) ± p38 inhibitor (SB203580) at 20 μM for 24 h and Nos2 expression measured by Taqman quantitative RT-PCR (n = 2 for all groups), p < 0.001 (***) Two-Way ANOVA with Bonferonni’s Multiple Comparison test. ( H ) NOD islets were stimulated with IL-17F or IL-17A and TNFα + IFNγ (Cyto) ± NF-κB inhibitor (BAY11-7082) at 10 μM for 24 h and Nos2 expression measured by Taqman quantitative RT-PCR (n = 2 for all groups), p < 0.01 (**) Two-Way ANOVA with Bonferonni’s Multiple Comparison test. ( I ) NOD islets were stimulated with IL-17F or IL-17A in the presence of TNFα + IFNγ (Cyto) for 4 days ± NMMA (1 mM) and cell death quantified by DNA fragmentation (n = 2 for all groups), p < 0.01 (**) One-Way ANOVA with Bonferonni’s Multiple Comparison test.

Article Snippet: Mouse cytokines were used at the following concentrations: mTNFα (In Vitro Technologies) at 100 ng/ml, mIFNγ at 10 ng/ml (Australian Biosearch), hIL-1β at 50 ng/ml (Peprotech), mIL-17A at 25 ng/ml (R&D Systems) and mIL-17F at 100 ng/ml (R&D Systems).

Techniques: Expressing, Quantitative RT-PCR, Comparison