Journal: JCI Insight

Article Title: CFTR negatively reprograms Th2 cell responses, and CFTR potentiation restrains allergic airway inflammation

doi: 10.1172/jci.insight.191098

Figure Lengend Snippet: ( A ) Schematic diagram showing isolation and stimulation of naive CD4 + T cells. ( B and C ) IL-5 and IL-13 by ELISA in cellular supernatant from Cftr +/+ and Cftr −/− CD4 + T cells grown in culture stimulated with mouse IL-4 ( n = 5 mice per genotype). ( D ) Representative gating strategy for IL-13 expression in cultured Cftr +/+ and Cftr −/− CD4 + T cell populations gated on live lymphoid cells. ( E ) IL-13 median fluorescence intensity (MFI) of cultured Cftr +/+ and Cftr −/− CD4 + T cells ( n = 5 mice per genotype). ( F ) IL-13 by ELISA in cellular supernatant from Cftr +/+ CD4 + T cells grown in culture with the CFTR inhibitor, GlyH-101, or control vehicle (DMSO) stimulated with mouse IL-4. ( G and H ) IL-5 and IL-13 by ELISA in cellular supernatant from mouse CD4 + T cells expressing either wild-type human CFTR or hCFTR ΔF508 grown in culture with Elexacaftor/Tezacaftor/ Ivacaftor (ETI) or DMSO control ( n = 5 mice per genotype per condition). Data are shown as mean ± SD. Statistical analysis were performed using unpaired Student’s t test ( B , C , E , and F ) and by 1-way ANOVA ( G and H ) followed by Tukey’s honestly significant difference (HSD) post hoc test for multiple comparisons. * P < 0.05, ** P < 0.01, and **** P < 0.0001.

Article Snippet: Murine IL-5, murine IL-13, and murine IgE were assayed using Quantikine ELISA kits following manufacturer’s instructions (R&D Systems).

Techniques: Isolation, Enzyme-linked Immunosorbent Assay, Expressing, Cell Culture, Fluorescence, Control

Journal: JCI Insight

Article Title: CFTR negatively reprograms Th2 cell responses, and CFTR potentiation restrains allergic airway inflammation

doi: 10.1172/jci.insight.191098

Figure Lengend Snippet: ( A ) Schematic diagram showing adaptive model of intranasal AE -induced airway inflammation in CD4 Cre− Cftr fl/fl and CD4 Cre+ Cftr fl/fl mice. ( B ) IgE concentrations by ELISA in serum from treated mice ( n = 4–5 depending on genotype and condition). ( C – F ) The number of macrophages ( C ), neutrophils ( D ), eosinophils ( E ), and lymphocytes ( F ) in the BALF of PBS- or AE -challenged mice ( n = 4–5 per genotype and condition). ( G and H ) IL-5 and IL-13 by ELISA in BAL from CD4 Cre− Cftr fl/fl and CD4 Cre+ Cftr fl/fl mice treated with either AE or PBS control ( n = 4–5 per genotype and condition). Open circles represent AE sensitized and challenged mice, and closed circles denote PBS control mice. Black circles denote CD4 Cre− Cftr fl/fl and red circles indicate CD4 Cre+ Cftr fl/fl mice. Statistical analysis in B – H were done by 1-way ANOVA followed by Tukey’s honestly significant difference (HSD) post hoc test for multiple comparisons. * P < 0.05, ** P < 0.01, *** P < 0.001, and **** P < 0.0001.

Article Snippet: Murine IL-5, murine IL-13, and murine IgE were assayed using Quantikine ELISA kits following manufacturer’s instructions (R&D Systems).

Techniques: Enzyme-linked Immunosorbent Assay, Control

Journal: JCI Insight

Article Title: CFTR negatively reprograms Th2 cell responses, and CFTR potentiation restrains allergic airway inflammation

doi: 10.1172/jci.insight.191098

Figure Lengend Snippet: ( A ) Schematic diagram showing an adaptive model of intranasal AE -induced inflammation and i.p. ivacaftor (IVA, 1 μM) or DMSO administration schedule in Cftr −/− hCFTR +/+ mice. ( B ) IgE concentrations by ELISA in serum from sensitized/challenged mice treated with IVA ( n = 17) or DMSO ( n = 16). ( C – F ) The number of macrophages ( C ), neutrophils ( D ), eosinophils ( E ), and lymphocytes ( F ), in the BALF of AE -challenged mice treated with either IVA ( n = 17) or DMSO ( n = 16). ( G and H ) IL-5 and IL-13 by ELISA in BALF from AE- sensitized and challenged mice treated with either IVA ( n = 17) or DMSO control ( n = 16). Open circles represent IVA treated mice, and closed circles denote DMSO-treated control mice. Statistical analysis in B – H performed using unpaired Student’s t test. * P < 0.05, ** P < 0.01, *** P < 0.001, and **** P < 0.0001.

Article Snippet: Murine IL-5, murine IL-13, and murine IgE were assayed using Quantikine ELISA kits following manufacturer’s instructions (R&D Systems).

Techniques: Enzyme-linked Immunosorbent Assay, Control

Journal: JCI Insight

Article Title: CFTR negatively reprograms Th2 cell responses, and CFTR potentiation restrains allergic airway inflammation

doi: 10.1172/jci.insight.191098

Figure Lengend Snippet: ( A ) Schematic diagram detailing the isolation and Th2 polarization of naive human CD4 + T cells used for flow cytometry and cytokine analysis. ( B ) Viability of ivacaftor (IVA) or DMSO (control) cultured human Th2 cells at 7 days. ( C ) Representative flow cytometry histogram showing the median fluorescence intensity (MFI) of GATA3 at 7 days for DMSO- and IVA-treated Th2 cells. ( D ) The quantified MFI of GATA3 in DMSO- and IVA-treated Th2 cells at 72 hours ( n = 6 paired human samples). ( E ) Representative flow cytometry histogram showing the median fluorescence intensity (MFI) of IL-13 at 7 days for DMSO- and IVA-treated Th2 cells. ( F ) The quantified MFI of IL-13 in DMSO- and IVA-treated Th2 cells at 72 hours ( n = 6 paired human samples). ( G ) IL-13 by ELISA in cellular supernatant from cultured DMSO- and IVA-treated Th2 cells. Statistical analysis in B , D , F , and G was performed using paired Student’s t test. * P < 0.05.

Article Snippet: Murine IL-5, murine IL-13, and murine IgE were assayed using Quantikine ELISA kits following manufacturer’s instructions (R&D Systems).

Techniques: Isolation, Flow Cytometry, Control, Cell Culture, Fluorescence, Enzyme-linked Immunosorbent Assay

Journal: Communications biology

Article Title: Time-dependent cell-state selection identifies transiently expressed genes regulating ILC2 activation.

doi: 10.1038/s42003-023-05297-w

Figure Lengend Snippet: Fig. 4 Identification of genes expressed in different activation steps from human specimens. a Representative time-lapse images of secretion signal and cell morphology (left) and heatmap of IL-13 secretion signal (right) of individual hILC2s. Scale bar, 25 µm. b Heatmap showing the average mRNA levels in each activation state of each specimen in standard scores (z-score). Details of each class are provided in Supplementary Data 6. c Box plot illustrating the expression levels of representative TIGs; IL13, IL4 and MIR155HG in each activation state. Each colour represents a different specimen. The centre line, box limits, whiskers, and dots indicate the median, upper and lower quartiles, 1.5x interquartile range and individual cells, respectively. ND: not detected. d Comparison of IL-13 production in cultured human ILC2 with and without miR-155 inhibitor by ELISA of bulk culture supernatants. Inhibition was performed by electroporation of a synthetic miRNA inhibitor with miR-155 specific or control sequence. e Comparison of temporal changes in the estimated proportion of IL-13-secreting cells in cultured human ILC2s with and without miR-155 inhibitor using the Nelson-Aalen estimator with their respective 95% confidence intervals (coloured bands). The number of cells in negative control and inhibition were 259 and 153, respectively. The difference between them was compared with the log-rank test (observed p value = 0.0013). f Secretion signal traces and images of IL-4 producing individual hILC2s. Three secretion-positive traces are shown in magenta and other traces are shown in grey (388 traces).

Article Snippet: For the measurement of IL-13 in culture supernatant, we used enzyme-linked immuno-sorbent assay (ELISA) kit (Human IL-13 Quantikine ELISA Kit, R&D Systems).

Techniques: Activation Assay, Expressing, Comparison, Cell Culture, Enzyme-linked Immunosorbent Assay, Inhibition, Electroporation, Control, Sequencing, Negative Control