Journal: American journal of physiology. Cell physiology

Article Title: (R)-vanzacaftor potentiates BK Ca channels in the absence of CFTR correction or potentiation

doi: 10.1152/ajpcell.00654.2025

Figure Lengend Snippet: Following overnight incubation in S-VX-121 ( A; 1 μM) or R-VX-121 ( B; 1 μM), forskolin (10 μM) and VX-770 (1 μM) induced an increase in I Cl that was inhibited by CFTR inh172 (3 μM). C. Average responses to forskolin + VX-770 for experiments in A and B. DMSO (0.1%) was used as a control. D. Immunoblot of CFTR showing correction of F508del CFTR by S-VX-121, but not R-VX-121. E. Average results for 3 experiments. F. Cl − currents in response to forskolin following overnight incubation in S-VX-121 (1 μM; blue trace) or S-VX-121 (1 μM) plus R-VX-121 (3 μM) (red trace). G. Concentration-response relationship for inhibition of S-VX-121-induced correction of I Cl by R-VX-121. The data were fit to the Hill equation with an apparent IC 50 of 3.5 μM. H. Cl − currents in response to forskolin following overnight incubation in VX-445 (1 μM; blue trace) or VX-445 (1 μM) plus R-VX-121 (6 μM) (red trace). I. Average response to forskolin for the experiments shown in H (n=23). J, K, L. F508del CFTR was corrected with VX-445 (1 μM) plus VX-661 (3 μM). Forskolin was used to stimulate I Cl and the effect of VX-770 (1 μM; J ), S-VX-121 (10 μM) followed by VX-770 ( K ) or R-VX-121 (10 μM) followed by VX-770 ( L ) determined. CFTR inh172 (3 μM) was used to confirm I Cl was due to CFTR. M. Chemical structures of VX-770, S-VX-121 and R-VX-121. #, P<0.01.

Article Snippet: Paxilline (HY-N6778), VX-445 (HY-111772), S-VX-121 (HY-145603), R-VX-121 (HY-145603A) and CFTR inh172 (HY-16671) were obtained from MedChemExpress.

Techniques: Incubation, Control, Western Blot, Concentration Assay, Inhibition

Journal: American journal of physiology. Cell physiology

Article Title: (R)-vanzacaftor potentiates BK Ca channels in the absence of CFTR correction or potentiation

doi: 10.1152/ajpcell.00654.2025

Figure Lengend Snippet: A. Concentration-dependent vasodilation of mesenteric artery by R-VX-121 from a male mouse. B. Average responses in male mice to S- (blue line, n=7) and R- (red line, n=7) VX-121. C and D . Average responses in male mice to R-VX-121 ( C ) and S-VX-121 ( D ) under control conditions (solid lines) and in the presence of 10 μM paxilline (Pax; dashed lines). Paxilline inhibited the response to both R- and S-VX-121 (n=8 for all conditions, P<0.01). E. Average responses in female mice to S- (blue line, n=7) and R- (red line, n=7) VX-121. F and G. Average responses in male mice to S-VX-121 ( F ) and R-VX-121 ( G ) under control conditions (solid lines) and in the presence of 3 μM CFTR inh172 (dashed lines). CFTR inh172 had no effect (n=8 for all conditions).

Article Snippet: Paxilline (HY-N6778), VX-445 (HY-111772), S-VX-121 (HY-145603), R-VX-121 (HY-145603A) and CFTR inh172 (HY-16671) were obtained from MedChemExpress.

Techniques: Concentration Assay, Control

Journal: International Journal of Molecular Sciences

Article Title: Identification of Actionable Gene Variants in Pulmonary Large-Cell Neuroendocrine Carcinoma: A Real-World Analysis of a Polish Cohort

doi: 10.3390/ijms27072939

Figure Lengend Snippet: The graphic representation of TMEM79::NTRK1 fusion, including exonic (TMEM79 NM_032323.3 exon 3; NTRK1 NM_002529.3 exon 2) and chromosomal (chr1:156256264, chr1:156834146) breakpoint positions based on the output data from the Archer Analysis software version 7.2 (ArcherDX, Inc., Boulder, CO, USA; ( A )) and on the variant visualization by the Alamut Visual Plus v1.12 software (SOPHiA GENETICS, Inc., Boston, MA, USA; ( B )).

Article Snippet: Gene fusion variants were further evaluated using Alamut Visual Plus (v1.12; SOPHiA GENETICS, Inc., Boston, MA, USA).

Techniques: Software, Variant Assay

Journal: F1000Research

Article Title: An unexpected effect of TNF-α on F508del-CFTR maturation and function

doi: 10.12688/f1000research.6683.2

Figure Lengend Snippet: Differentiated primary HBE cell cultures grown at an air-liquid interface were incubated with 50ng/ml TNFα for 10–30min, 3–6h and 24h. CFTR immunodetection was performed with 24.1 anti-CFTR antibody and analyzed with confocal microscopy. Green staining represents CFTR (Alexa Fluor 488), red color staining represents ZO-1 protein of tight junctions (Alexa Fluor 594) and blue DAPI staining visualizes nuclei. Independent TNFα treatments and CFTR immunodetection were performed on HBE cells from three different F508del/F508del CF patients. Representative images of one experiment are demonstrated (Scale bars = 20µm).

Article Snippet: Anti-CFTR antibodies (abs): MM13-4 mouse monoclonal ab against N-terminus of CFTR, (Millipore, France, 05-581); 24-1 mouse monoclonal ab against C-terminus of CFTR (R&D Systems, MAB, 25031).

Techniques: Incubation, Immunodetection, Confocal Microscopy, Staining

Journal: Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie

Article Title: Activation of constitutive androstane receptor inhibits intestinal CFTR-mediated chloride transport.

doi: 10.1016/j.biopha.2019.01.015

Figure Lengend Snippet: Fig. 4. Effects of CAR agonists on CFTR-mediated Cl−

Article Snippet: After centrifugation of the cell and tissue homogenates at 12,000 rpm for 20min at 4 °C, equal amounts of protein (50 μg) were separated by SDS-PAGE and transferred onto a nitrocellulose membrane (Amersham Biosciences, Buckinghamshire, UK).The membrane was then incubated for 1 h with 5% non-fat milk and probed overnight at 4 °C with anti-human CFTR, anti-β actin (Cell signaling), anti-mouse CFTR (Abcam) and anti-CAR (R&D systems) (all at 1000-fold dilution) followed by a further incubation with HRP-conjugated secondary antibody for 1 h at room temperature.

Techniques:

Journal: Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie

Article Title: Activation of constitutive androstane receptor inhibits intestinal CFTR-mediated chloride transport.

doi: 10.1016/j.biopha.2019.01.015

Figure Lengend Snippet: Fig. 5. Effects of CAR agonist on mRNA and protein expres- sion of CFTR in T84 cells (A): Quantitative real time PCR ana- lysis of CFTR mRNA expression in T84 cells after treatment with vehicle (0.01% DMSO), CITCO, or phenytoin at the indicated concentrations for 24 h. The results were normalized to GAPDH. (B) Representative blot and densitometric quantification of CFTR protein expression were normalized to β-actin levels in T84 cells treated with vehicle (0.01% DMSO), CITCO, or phenytoin at the indicated concentrations for 24 h. The data are expressed as means ± S.E. (n = 5–7). *, P < 0.05; **, P < 0.01; ***, P < 0.001 compared to control.

Article Snippet: After centrifugation of the cell and tissue homogenates at 12,000 rpm for 20min at 4 °C, equal amounts of protein (50 μg) were separated by SDS-PAGE and transferred onto a nitrocellulose membrane (Amersham Biosciences, Buckinghamshire, UK).The membrane was then incubated for 1 h with 5% non-fat milk and probed overnight at 4 °C with anti-human CFTR, anti-β actin (Cell signaling), anti-mouse CFTR (Abcam) and anti-CAR (R&D systems) (all at 1000-fold dilution) followed by a further incubation with HRP-conjugated secondary antibody for 1 h at room temperature.

Techniques: Real-time Polymerase Chain Reaction, Lysis, Expressing, Control

Journal: Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie

Article Title: Activation of constitutive androstane receptor inhibits intestinal CFTR-mediated chloride transport.

doi: 10.1016/j.biopha.2019.01.015

Figure Lengend Snippet: Fig. 6. Effects of CAR antagonist on CITCO- induced inhibition of Cl−secretion and CFTR protein expression in T84 cells (A). Effects of CAR antagonist on CITCO-induced inhibition of Cl−secretion in T84 cells. T84 cells were treated with vehicle (0.1% DMSO), CITCO (1 μM), CAR antagonist [CINPA1 (1 μM)], or CITCO plus CINPA1 for 24 h, then the forskolin-stimulated Isc was measured. (left) Representative Isc tracings and (right) summary of the data are shown. Data are ex- pressed as means ± S.E. of percentages of forskolin-stimulated Isc (n = 5). (B) Representative blot and densitometric quanti- fication of CFTR protein expression in T84 cells treated with vehicle alone or together with CITCO (1 μM) and/or CINPA1 (1 μM) for 24 h. The data are expressed as means ± S.E. (n = 5). *, P < 0.05 compared to the vehicle- treated control. NS, not significantly different from control. ##, P < 0.05; ##, P < 0.01 compared to CITCO-treatment group.

Article Snippet: After centrifugation of the cell and tissue homogenates at 12,000 rpm for 20min at 4 °C, equal amounts of protein (50 μg) were separated by SDS-PAGE and transferred onto a nitrocellulose membrane (Amersham Biosciences, Buckinghamshire, UK).The membrane was then incubated for 1 h with 5% non-fat milk and probed overnight at 4 °C with anti-human CFTR, anti-β actin (Cell signaling), anti-mouse CFTR (Abcam) and anti-CAR (R&D systems) (all at 1000-fold dilution) followed by a further incubation with HRP-conjugated secondary antibody for 1 h at room temperature.

Techniques: Inhibition, Expressing, Control

Journal: Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie

Article Title: Activation of constitutive androstane receptor inhibits intestinal CFTR-mediated chloride transport.

doi: 10.1016/j.biopha.2019.01.015

Figure Lengend Snippet: Fig. 7. Effect of CAR activation on CFTR-mediated Cl−transport during inhibition of mRNA synthesis in T84 cells. T84 cells were treated with ve- hicle (0.1% DMSO), CITCO (1 μM), or actinomycin D (Act D) (5 μg/ml), or CITCO plus Act D for 24 h, after that CFTR expression was measured by real- time PCR analysis. The results were normalized to GAPDH. The data are ex- pressed as means ± S.E. (n = 5). *, P < 0.05; ***, P < 0.001 compared to the vehicle-treated control. NS, not significantly different.

Article Snippet: After centrifugation of the cell and tissue homogenates at 12,000 rpm for 20min at 4 °C, equal amounts of protein (50 μg) were separated by SDS-PAGE and transferred onto a nitrocellulose membrane (Amersham Biosciences, Buckinghamshire, UK).The membrane was then incubated for 1 h with 5% non-fat milk and probed overnight at 4 °C with anti-human CFTR, anti-β actin (Cell signaling), anti-mouse CFTR (Abcam) and anti-CAR (R&D systems) (all at 1000-fold dilution) followed by a further incubation with HRP-conjugated secondary antibody for 1 h at room temperature.

Techniques: Activation Assay, Inhibition, Expressing, Real-time Polymerase Chain Reaction, Control

Journal: Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie

Article Title: Activation of constitutive androstane receptor inhibits intestinal CFTR-mediated chloride transport.

doi: 10.1016/j.biopha.2019.01.015

Figure Lengend Snippet: Fig. 9. Effect of CAR activation on CFTR expression and function in mouse intestine. (A): Activation of CAR target gene (MDR1) in mouse intestine. B: Quantitative PCR analysis of CFTR mRNA expression in mouse intestinal tissues after intraperitoneally injected the mice with 3 mg/kgBW TCPOBOP (CAR agonist) or vehicle (corn oil) for 7 days. The results were normalized to GAPDH. C: Representative blot and densitometric quantification of CFTR pro- tein expression in mouse intestinal tissues after treated the mice with TCPOBOP or vehicle (corn oil) for 7 days. The results were nor- malized to β-actin levels. The data are pre- sented as mean ± SE (n = 8). *, P < 0.05; **, P < 0.01 compared to control. D. Antidiarrheal application of CAR activation on cholera toxin-induced intestinal fluid accumu- lation in mouse closed-loop model. ICR mice were treated with 3 mg/kgBW TCPOBOP or vehicle (corn oil) for 7 days. After treatment, PBS or PBS containing cholera toxin (CT) (1 μg/loop) were instilled into ileal loops. Four hours later, ileal loops were removed for de- termination of loop weight/length ratio. Representative photograph of ileal loops and summary of the data are shown. Data are ex- pressed as means ± S.E. of loop weight/ length ratio (n = 8). **, P < 0.01 compared with CT-treated control. NS, not significantly different compared with PBS-treated control.

Article Snippet: After centrifugation of the cell and tissue homogenates at 12,000 rpm for 20min at 4 °C, equal amounts of protein (50 μg) were separated by SDS-PAGE and transferred onto a nitrocellulose membrane (Amersham Biosciences, Buckinghamshire, UK).The membrane was then incubated for 1 h with 5% non-fat milk and probed overnight at 4 °C with anti-human CFTR, anti-β actin (Cell signaling), anti-mouse CFTR (Abcam) and anti-CAR (R&D systems) (all at 1000-fold dilution) followed by a further incubation with HRP-conjugated secondary antibody for 1 h at room temperature.

Techniques: Activation Assay, Expressing, Real-time Polymerase Chain Reaction, Injection, Control

Journal: PLoS ONE

Article Title: Heterogeneous expression of CFTR in insulin-secreting β-cells of the normal human islet

doi: 10.1371/journal.pone.0242749

Figure Lengend Snippet: List of primary antibodies raised against the indicated antigens are identified by their clone/catalogue number from vendors/suppliers.

Article Snippet: Cftr (20738-1-ap) , Proteintech , 1:100 , IF , No signal.

Techniques: Staining

Journal: Journal of Biological Chemistry

Article Title: Probing Conformational Rescue Induced by a Chemical Corrector of F508del-Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) Mutant

doi: 10.1074/jbc.m111.239699

Figure Lengend Snippet: FIGURE 5. The effect of chemical correction and ablation of the R553XR555 sequence on conformational correction of F508del-CFTR probed using limited proteolysis. Limited proteolysis of control or chemically corrected F508del-CFTR, F508del-KXK-CFTR, or WT-CFTR with increasing levels of trypsin (0, 1.56, 3.13, 6.25, 12.5, 25, 50, 100 g/ml) was performed. Digests were analyzed by SDS-PAGE, and immunoreactive bands were detected using the Odyssey infrared imaging system from LI-COR Biosciences. The colored signal was converted to grayscale for the preparation of this figure. A, samples were immuno- blotted with anti-CFTR monoclonal antibody, L12B4 (NBD1-specific). Subsequent analyses focused on the larger NBD1 fragments (34–36 kDa) and the smaller NBD1 fragments (lower than 28 kDa). Both sets of fragments were indicated using brackets on the right-hand side of each image. B, samples were immuno- blotted with anti-CFTR monoclonal antibody, M3A7 (NBD2-specific). NBD2 fragments of 28 kDa were analyzed subsequently (indicated using brackets on the right-hand side of each image).

Article Snippet: L12B4 anti-CFTR antibody (1:1000) was used for immunodetection conjugated to IRDye800 (1:10 000) (Rockland Immunochemical Inc).

Techniques: Sequencing, Control, SDS Page, Imaging

Journal: The Journal of pharmacology and experimental therapeutics

Article Title: Differential regulation of cystic fibrosis transmembrane conductance regulator by interferon gamma in mast cells and epithelial cells.

doi: 10.1124/jpet.105.087528

Figure Lengend Snippet: Fig. 1. Western blot analysis of CFTR and STAT1 expression in un- treated (lane 1), PMA-treated (lane 2), TNF-treated (lane 3), and IFN- treated (lane 4) T84 (A) and RCMC (B). T84 cells were treated with PMA (10 ng/ml), human recombinant TNF (10 ng/ml), or IFN (10 ng/ml) for 24 h. RCMC were treated with PMA (10 ng/ml) or rat recombinant TNF (10 ng/ml) or IFN (10 ng/ml) for 24 h. Cell lysates were resolved on an 4 to 12% SDS-PAGE and blotted with anti-CFTR, anti-STAT1, or anti-actin antibody. Western blots shown represent three independent experiments. The effect of 24 h IFN treatment (80 ng/ml) on CFTR expression was characterized in T84 cells (C) and rat peritoneal mast cells (D), using confocal laser scanning microscopy (bar on the figure represents 10 m).

Article Snippet: The membranes were blocked with 3% milk in Tris-buffered saline-0.05% Tween for 1 h and then probed with primary antibodies against CFTR (clone H-182) and STAT1 (Santa Cruz Biotechnology Inc., Santa Cruz, CA), phosphoSTAT1 (BD Transduction Laboratories, Chicago, IL), phospho-stress-activated MAPK/c-Jun NH2-terminal kinase (JNK) (Thr183/Tyr185; Cell Signaling Technology Inc.), phospho-p38 MAPK (Thr180/Tyr182; Cell Signaling Technology Inc.), and phospho ERK1/2 (Thr202/Tyr204; Cell Signaling Technology Inc.), or anti-actin (Sigma-Aldrich) in 4% BSA/PBS for 1 h at room temperature.

Techniques: Western Blot, Expressing, Recombinant, SDS Page, Confocal Laser Scanning Microscopy

Journal: The Journal of pharmacology and experimental therapeutics

Article Title: Differential regulation of cystic fibrosis transmembrane conductance regulator by interferon gamma in mast cells and epithelial cells.

doi: 10.1124/jpet.105.087528

Figure Lengend Snippet: Fig. 4. A, IFN-mediated up-regulation of mast cell CFTR protein requires activation of JAK2 and/or p38 and ERK. RCMC, LAD2, and T84 were treated with IFN (10 ng/ml) with or without JAK2 inhibitor (AG-490; 30 mg/ml), p38 inhibitor (SB202190; 30 mg/ml), or ERK MAPK inhibitor (U0126; 30 mg/ml) for 24 h. Representative of three inde- pendent experiments. B, densitometry summary of data in Fig. 4. RCMC were treated with IFN (10 ng/ml) with or without JAK2 inhibitor (AG-490; 30 mg/ml), p38 inhibitor (SP202190; 30 mg/ml), or ERK inhibitor (U0126; 30 mg/ml) for 24 h, and expression of CFTR (black columns) and STAT1 (gray columns) was analyzed by Western blot (n 3 independent experiments; error bars represent S.E.M.). As- terisks represent statistical significance as determined by Student’s t test (p 0.05).

Article Snippet: The membranes were blocked with 3% milk in Tris-buffered saline-0.05% Tween for 1 h and then probed with primary antibodies against CFTR (clone H-182) and STAT1 (Santa Cruz Biotechnology Inc., Santa Cruz, CA), phosphoSTAT1 (BD Transduction Laboratories, Chicago, IL), phospho-stress-activated MAPK/c-Jun NH2-terminal kinase (JNK) (Thr183/Tyr185; Cell Signaling Technology Inc.), phospho-p38 MAPK (Thr180/Tyr182; Cell Signaling Technology Inc.), and phospho ERK1/2 (Thr202/Tyr204; Cell Signaling Technology Inc.), or anti-actin (Sigma-Aldrich) in 4% BSA/PBS for 1 h at room temperature.

Techniques: Activation Assay, Expressing, Western Blot

Journal: The Journal of Physiology

Article Title: The apical mucus layer alters the pharmacological properties of the airway epithelium

doi: 10.1113/JP287891

Figure Lengend Snippet: Representative traces (left) and summary graphs (right) showing short‐circuit current results obtained in epithelia untreated ( A ) or treated ( B ) with IL‐4 (10 ng/ml) for 72 h. During short‐circuit current recordings, epithelia were exposed sequentially to: amiloride (10 µM, apical), CPT‐cAMP (100 µM, apical and basolateral), CFTR inh ‐172 (I‐172, 10 µM, apical), PPQ‐102 (30 µM, apical), UTP (100 µM, apical) and Ani9 (10 µM, apical). Dotted line indicates the baseline that was used to measure cAMP‐activated current before and after I‐172. Dashed line indicates absolute zero current level. The graphs report, for each experiment, the value of the cAMP‐activated current before and after CFTR inh ‐172 addition. Data in the figure ( n = 11–19 per condition) are from separate preparations of cells from three non‐cystic fibrosis donors.

Article Snippet: CFTR inh ‐172 (HY‐16 671), PPQ‐102 (HY‐14 179), bumetanide (HY‐17 468), VX‐445 (HY‐111 772) and VX‐661 (HY‐15 448) were from MedChemExpress.

Techniques:

Journal: The Journal of Physiology

Article Title: The apical mucus layer alters the pharmacological properties of the airway epithelium

doi: 10.1113/JP287891

Figure Lengend Snippet: Representative traces (left) and summary graphs (right) showing short‐circuit current results obtained in epithelia untreated ( A ) or treated ( B ) with IL‐4 (10 ng/ml) for 72 h. Before experiments, the apical surface of epithelia was left unwashed or washed with dithiothreitol (DTT, 5 mM). Compounds added during short‐circuit current recordings were as listed for Fig. except for PPQ‐102, which was omitted. Dotted line indicates the baseline used to measure cAMP‐activated current before and after I‐172. Dashed line indicates the absolute zero current level. The graphs report the fraction of the cAMP‐activated current that remains after addition of CFTR inh ‐172. *** P < 0.001 vs . no wash; ### P < 0.001 IL‐4 no wash vs . untreated no wash (ANOVA with Tukey's post hoc test). Data shown ( n = 10–17 per condition) are from separate preparations of cells from three non‐cystic fibrosis donors.

Article Snippet: CFTR inh ‐172 (HY‐16 671), PPQ‐102 (HY‐14 179), bumetanide (HY‐17 468), VX‐445 (HY‐111 772) and VX‐661 (HY‐15 448) were from MedChemExpress.

Techniques:

Journal: The Journal of Physiology

Article Title: The apical mucus layer alters the pharmacological properties of the airway epithelium

doi: 10.1113/JP287891

Figure Lengend Snippet: Representative short‐circuit current traces (left) and summary graphs (right) from epithelia kept in control conditions ( A ), treated with IL‐4 ( B ) or treated with IL‐17A/TNF‐α ( C ). Where indicated, epithelia were washed apically with dithiothreitol (DTT) to remove mucus. During recordings, after CFTR stimulation with CPT‐cAMP, currents were inhibited with apical PPQ‐102 (30 µM) followed by apical CFTR inh ‐172 (I‐172, 10 µM). Epithelia were then stimulated with apical UTP (100 µM) followed by apical Ani9 (10 µM). Dotted and dashed lines indicate the baseline for cAMP‐activated current and zero current level, as in previous legends. The graphs report the fraction of the cAMP‐activated current that remains after addition of PPQ‐102, with/without DTT treatment. * P = 0.0220; *** P < 0.001 (unpaired Student's t ‐test). Data ( n = 4 or 5 per condition) are from cells of one non‐CF donor.

Article Snippet: CFTR inh ‐172 (HY‐16 671), PPQ‐102 (HY‐14 179), bumetanide (HY‐17 468), VX‐445 (HY‐111 772) and VX‐661 (HY‐15 448) were from MedChemExpress.

Techniques: Control