Cystic fibrosis transmembrane conductance regulator activation by roflumilast contributes to therapeutic benefit in chronic bronchitis.
Study subjects
Primary human bronchial epithelial (HBE) monolayers
Mouse, ligated intestinal segment
Ligated murine intestinal segments monitored ex vivo; the abstract does not state strain, sex, or age.
CFTR assays
CFTR-dependent anion transport - roflumilast potency in normal HBE cells
Roflumilast activated CFTR-dependent anion transport in normal (non-smoke-exposed) primary HBE cells with a half maximal effective concentration of 2.9 nM, measured in modified Ussing chambers. The abstract reports the EC50 only; no short-circuit current value is given for this experiment.
Exposure conditions
| Condition | Agent | Concentration | Duration |
|---|---|---|---|
| Roflumilast at its half maximal effective concentration (2.9 nM) EXPOSURE:lambert-roflumilast-ec50-hbe |
roflumilast CHEBI:47657 |
2.9 nanomolar | — |
Evidence
Roflumilast activated CFTR-dependent anion transport in normal HBE cells with a half maximal effective concentration of 2.9 nM.
The abstract states the potency of roflumilast as an activator of CFTR-dependent anion transport in normal HBE cells, which is the EC50 this assay records on its exposure condition.
CFTR activity - WCS-exposed HBE cells with and without roflumilast
Roflumilast partially restored CFTR activity in WCS-exposed primary HBE cells: 5.3 +/- 1.1 microampere/cm2 with roflumilast vs. 1.2 +/- 0.2 microampere/cm2 in the WCS-exposed control, P < 0.05. The abstract does not state whether the +/- terms are SD or SEM, nor the sample size, so neither is recorded. Roflumilast was additive with ivacaftor, a specific CFTR potentiator approved for the treatment of CF; the abstract gives no value for the combination.
Exposure conditions
| Condition | Agent | Concentration | Duration |
|---|---|---|---|
| Whole cigarette smoke (WCS), no roflumilast (control arm) EXPOSURE:lambert-wcs-hbe |
smoke ENVO:01000838 |
— | — |
| Whole cigarette smoke (WCS) with roflumilast treatment EXPOSURE:lambert-wcs-roflumilast-hbe |
roflumilast CHEBI:47657 |
— | — |
Outputs
| Group | Output | Description |
|---|---|---|
| control | CFTR activity in WCS-exposed HBE cells (control arm) CFTR:lambert-hbe-wcs-roflumilast-output-control · under EXPOSURE:lambert-wcs-hbe |
cftr chloride secretion: 1.2 microampere per square centimeter (mean)
|
| treated | CFTR activity in WCS-exposed HBE cells treated with roflumilast CFTR:lambert-hbe-wcs-roflumilast-output-roflumilast · under EXPOSURE:lambert-wcs-roflumilast-hbe |
cftr chloride secretion: 5.3 microampere per square centimeter (mean)
|
Evidence
Roflumilast partially restored CFTR activity in WCS-exposed HBE cells (5.3 ± 1.1 μA/cm(2) vs. 1.2 ± 0.2 μA/cm(2) ... P < 0.05) and was additive with ivacaftor, a specific CFTR potentiator approved for the treatment of CF.
The abstract gives both short-circuit current values recorded by this assay - the WCS-exposed control and the roflumilast-treated arm - together with the significance of the difference. The elision marks the abstract's own '[control]' label, which the quote checker strips from bracketed spans.
Primary human bronchial epithelial (HBE) cells, Calu-3, and T84 monolayers were exposed to whole cigarette smoke (WCS) or air with or without roflumilast treatment.
The abstract describes the model system and exposure design behind this assay: HBE monolayers exposed to whole cigarette smoke or air, with or without roflumilast.
CFTR-dependent fluid secretion - ligated murine intestine, ex vivo
CFTR activation by roflumilast induced CFTR-dependent fluid secretion in ligated murine intestinal segments monitored ex vivo, increasing the wet:dry ratio and the segment diameter. The abstract reports the direction of both changes but no numeric values, so no measurement is recorded. This is the observation the authors link to the noninfectious diarrhea seen with roflumilast.
Exposure conditions
| Condition | Agent | Concentration | Duration |
|---|---|---|---|
| Roflumilast applied to ligated murine intestinal segments ex vivo EXPOSURE:lambert-roflumilast-murine-intestine |
roflumilast CHEBI:47657 |
— | — |
Evidence
CFTR activation by roflumilast also induced CFTR-dependent fluid secretion in murine intestine, increasing the wet:dry ratio and the diameter of ligated murine segments.
The abstract states the ex vivo murine intestinal result this assay records: roflumilast-driven CFTR activation induced CFTR-dependent fluid secretion, raising the wet:dry ratio and the diameter of the ligated segments.
ASL assays
ASL depth - WCS-exposed HBE monolayers with and without roflumilast
Roflumilast improved the depleted ASL depth of HBE monolayers exposed to WCS: 9.0 +/- 3.1 micrometer with roflumilast vs. 5.6 +/- 2.0 micrometer in the WCS-exposed control, P < 0.05, which the authors report as 79% of the depth observed in air controls. The abstract does not state whether the +/- terms are SD or SEM, nor the sample size, nor the absolute air-control depth, so none of those is recorded.
Exposure conditions
| Condition | Agent | Concentration | Duration |
|---|---|---|---|
| Whole cigarette smoke (WCS), no roflumilast (control arm) EXPOSURE:lambert-wcs-hbe |
smoke ENVO:01000838 |
— | — |
| Whole cigarette smoke (WCS) with roflumilast treatment EXPOSURE:lambert-wcs-roflumilast-hbe |
roflumilast CHEBI:47657 |
— | — |
Outputs
| Group | Output | Description |
|---|---|---|
| control | ASL depth in WCS-exposed HBE monolayers (control arm) ASL:lambert-hbe-wcs-roflumilast-output-control · under EXPOSURE:lambert-wcs-hbe |
asl height um: 5.6 micrometer (mean)
|
| treated | ASL depth in WCS-exposed HBE monolayers treated with roflumilast ASL:lambert-hbe-wcs-roflumilast-output-roflumilast · under EXPOSURE:lambert-wcs-roflumilast-hbe |
asl height um: 9.0 micrometer (mean)
|
Evidence
Roflumilast improved the depleted ASL depth of HBE monolayers exposed to WCS (9.0 ± 3.1 μm vs. 5.6 ± 2.0 μm ... P < 0.05), achieving 79% of that observed in air controls.
The abstract gives both ASL depth values recorded by this assay - the WCS-exposed control and the roflumilast-treated arm - and states that the treated depth reaches 79% of the air control. The elision marks the abstract's own '[control]' label, which the quote checker strips from bracketed spans.
Response comparisons
CFTR activity in WCS-exposed HBE cells, roflumilast vs. untreated control
Roflumilast partially restored CFTR activity in WCS-exposed HBE cells (5.3 vs. 1.2 microampere/cm2). The paper reports the significance threshold P < 0.05 rather than an exact p-value, and states no delta, percent change, or fold change, so change_value is not recorded.
ASL depth in WCS-exposed HBE monolayers, roflumilast vs. untreated control
Roflumilast improved the depleted ASL depth of WCS-exposed HBE monolayers (9.0 vs. 5.6 micrometer), reaching 79% of the air control. The paper reports the significance threshold P < 0.05 rather than an exact p-value, and states no delta, percent change, or fold change between these two arms, so change_value is not recorded.
Key-event relationships
CFTR function leads to airway surface liquid depth
Lambert 2014 concludes that roflumilast activates CFTR-mediated anion transport via a cyclic AMP-dependent pathway and thereby partially reverses the deleterious effects of whole cigarette smoke, resulting in augmented ASL depth - i.e. CFTR function determines ASL depth in this model.
Protocols
Modified Ussing chamber measurement of CFTR-dependent ion transport
CFTR-dependent ion transport of airway and intestinal epithelial monolayers measured in modified Ussing chambers. Short-circuit current reported in microamperes per square centimetre.
Equipment: Modified Ussing chamber
Confocal microscopy measurement of airway surface liquid depth
Airway surface liquid depth of HBE monolayers determined by confocal microscopy.
Equipment: Confocal microscope
Whole cigarette smoke exposure of epithelial monolayers
Primary HBE, Calu-3 and T84 monolayers exposed to whole cigarette smoke (WCS) or to air, with or without roflumilast treatment.
Ex vivo ligated murine intestinal segment fluid secretion
Intestinal fluid secretion of ligated murine intestine monitored ex vivo; readouts are the wet:dry ratio and the diameter of the ligated segments.