Journal: Physiological Reports

Article Title: Effects of cAMP and CFTR modulation on apical fluid pH in human airway Calu‐3 cells

doi: 10.14814/phy2.70747

Figure Lengend Snippet: Concentration‐response analysis of cAMP elevating agents on apical fluid pH. Human airway epithelial (Calu‐3) cells were exposed to cAMP elevating agent (a and b) forskolin and (c and d) isoproterenol in the basolateral (bottom) compartment for 3 h at various concentrations. (a and c) A dose–response curve for measured apical fluid pH is depicted and (b and d) comparisons between treatment groups were performed. Data presented as means ± SD ( n = 4–5, FSK; n = 5, ISO). A one‐way ANOVA with subsequent multiple comparisons was used for statistical analysis. **** p ≤ 0.0001.

Article Snippet: The human airway epithelial cell line Calu‐3 (male, age 25) (ATCC, HTB‐55), derived from lung adenocarcinoma tissue, was cultured in Alpha‐Minimum Essential medium (α‐MEM) (Corning, 10‐022‐CV) supplemented with 10% fetal bovine serum (VWR, 080‐450), 1% HEPES (Corning, 25‐060‐CI), and 1% penicillin–streptomycin (VWR, 97063‐708) at 37°C and 5% CO 2 for both submerged and air‐liquid interface (ALI) culture conditions.

Techniques: Concentration Assay

Journal: Physiological Reports

Article Title: Effects of cAMP and CFTR modulation on apical fluid pH in human airway Calu‐3 cells

doi: 10.14814/phy2.70747

Figure Lengend Snippet: Control experiment in Ringer's solution to validate drug‐induced pH changes in the absence of cells. Various combinations of cAMP elevating agents forskolin (0.1 μM) and isoproterenol (0.01 μM), CFTR potentiator VX‐770 (1 μM), PDE‐4 inhibitor roflumilast (1 μM), and ABCC4 inhibitor MK‐571 (10 μM) were administered to the basolateral (bottom) compartment for 3 h to evaluate the change in pH in the absence of human airway epithelial (Calu‐3) cells. Data presented as means ± SD ( n = 3). A one‐way ANOVA with subsequent multiple comparisons was used for statistical analysis.

Article Snippet: The human airway epithelial cell line Calu‐3 (male, age 25) (ATCC, HTB‐55), derived from lung adenocarcinoma tissue, was cultured in Alpha‐Minimum Essential medium (α‐MEM) (Corning, 10‐022‐CV) supplemented with 10% fetal bovine serum (VWR, 080‐450), 1% HEPES (Corning, 25‐060‐CI), and 1% penicillin–streptomycin (VWR, 97063‐708) at 37°C and 5% CO 2 for both submerged and air‐liquid interface (ALI) culture conditions.

Techniques: Control

Journal: Physiological Reports

Article Title: Effects of cAMP and CFTR modulation on apical fluid pH in human airway Calu‐3 cells

doi: 10.14814/phy2.70747

Figure Lengend Snippet: Effect of various pharmacological interventions on apical fluid pH. Human airway epithelial (Calu‐3) cells were exposed to various combinations of cAMP elevating agents forskolin (0.1 μM) and isoproterenol (0.01 μM), CFTR potentiator VX‐770 (1 μM), PDE‐4 inhibitor roflumilast (1 μM), and ABCC4 inhibitor MK‐571 (10 μM) to the basolateral (bottom) compartment for 3 h. Measured apical fluid pH is depicted and comparisons between treatment groups were performed. Data presented as means ± SD ( n = 7). A two‐way ANOVA with subsequent multiple comparisons was used for statistical analysis. **** p ≤ 0.0001; #### p ≤ 0.0001.

Article Snippet: The human airway epithelial cell line Calu‐3 (male, age 25) (ATCC, HTB‐55), derived from lung adenocarcinoma tissue, was cultured in Alpha‐Minimum Essential medium (α‐MEM) (Corning, 10‐022‐CV) supplemented with 10% fetal bovine serum (VWR, 080‐450), 1% HEPES (Corning, 25‐060‐CI), and 1% penicillin–streptomycin (VWR, 97063‐708) at 37°C and 5% CO 2 for both submerged and air‐liquid interface (ALI) culture conditions.

Techniques:

Journal: Physiological Reports

Article Title: Effects of cAMP and CFTR modulation on apical fluid pH in human airway Calu‐3 cells

doi: 10.14814/phy2.70747

Figure Lengend Snippet: CFTR inhibitors administered in the apical or basolateral compartment do not impact apical fluid pH. Human airway epithelial (Calu‐3) cells were exposed to CFTR inhibitors (a and c) CFTRinh‐172 and (b and d) GlyH‐101 in the apical (a and b, top) or basolateral (c and d, bottom) compartment for 3 h at various concentrations. cAMP elevating agent forskolin (0.1 μM) was used as a positive control and comparisons between treatment groups were performed. Data presented as means ± SD ( n = 5). A one‐way ANOVA with subsequent multiple comparisons was used for statistical analysis.

Article Snippet: The human airway epithelial cell line Calu‐3 (male, age 25) (ATCC, HTB‐55), derived from lung adenocarcinoma tissue, was cultured in Alpha‐Minimum Essential medium (α‐MEM) (Corning, 10‐022‐CV) supplemented with 10% fetal bovine serum (VWR, 080‐450), 1% HEPES (Corning, 25‐060‐CI), and 1% penicillin–streptomycin (VWR, 97063‐708) at 37°C and 5% CO 2 for both submerged and air‐liquid interface (ALI) culture conditions.

Techniques: Positive Control

Journal: Physiological Reports

Article Title: Effects of cAMP and CFTR modulation on apical fluid pH in human airway Calu‐3 cells

doi: 10.14814/phy2.70747

Figure Lengend Snippet: Effect of cAMP elevating agents FSK and ISO post‐CFTR inhibition on apical fluid pH. Human airway epithelial (Calu‐3) cells were pre‐treated with CFTR inhibitors (a) CFTRinh‐172 (10 μM) and (b) GlyH‐101 (10 μM) to the apical (top) compartment for 30 min prior to treatment with cAMP elevating agents forskolin (0.1 μM) and isoproterenol (0.01 μM) to the basolateral (bottom) compartment for 3 h. Measured apical fluid pH is depicted and comparisons between treatment groups were performed. Data presented as means ± SD ( n = 4). A one‐way ANOVA with subsequent multiple comparisons was used for statistical analysis.

Article Snippet: The human airway epithelial cell line Calu‐3 (male, age 25) (ATCC, HTB‐55), derived from lung adenocarcinoma tissue, was cultured in Alpha‐Minimum Essential medium (α‐MEM) (Corning, 10‐022‐CV) supplemented with 10% fetal bovine serum (VWR, 080‐450), 1% HEPES (Corning, 25‐060‐CI), and 1% penicillin–streptomycin (VWR, 97063‐708) at 37°C and 5% CO 2 for both submerged and air‐liquid interface (ALI) culture conditions.

Techniques: Inhibition

Journal: Physiological Reports

Article Title: Effects of cAMP and CFTR modulation on apical fluid pH in human airway Calu‐3 cells

doi: 10.14814/phy2.70747

Figure Lengend Snippet: Effect of CFTR potentiator VX‐770 post‐CFTR inhibition on apical fluid pH. Human airway epithelial (Calu‐3) cells were pre‐treated with CFTR inhibitors (a) CFTRinh‐172 (10 μM) and (b) GlyH‐101 (10 μM) to the apical (top) compartment for 30 min prior to treatment with CFTR potentiator VX‐770 (1 μM) to the basolateral (bottom) compartment for 3 h. Measured apical fluid pH is depicted and comparisons between treatment groups were performed. Data presented as means ± SD ( n = 4). A one‐way ANOVA with subsequent multiple comparisons was used for statistical analysis.

Article Snippet: The human airway epithelial cell line Calu‐3 (male, age 25) (ATCC, HTB‐55), derived from lung adenocarcinoma tissue, was cultured in Alpha‐Minimum Essential medium (α‐MEM) (Corning, 10‐022‐CV) supplemented with 10% fetal bovine serum (VWR, 080‐450), 1% HEPES (Corning, 25‐060‐CI), and 1% penicillin–streptomycin (VWR, 97063‐708) at 37°C and 5% CO 2 for both submerged and air‐liquid interface (ALI) culture conditions.

Techniques: Inhibition