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Relationship: 2440
Title
CFTR Function, Decreased leads to ASL Height, Decreased
Upstream event
Downstream event
AOPs Referencing Relationship
| AOP Name | Adjacency | Weight of Evidence | Quantitative Understanding | Point of Contact | Author Status | OECD Status |
|---|---|---|---|---|---|---|
| Oxidative stress Leading to Decreased Lung Function via CFTR dysfunction | adjacent | High | Moderate | Arthur Author (send email) | Open for comment. Do not cite |
Taxonomic Applicability
| Term | Scientific Term | Evidence | Link |
|---|---|---|---|
| Homo sapiens | Homo sapiens | High | NCBI |
Sex Applicability
| Sex | Evidence |
|---|---|
| Mixed | High |
Life Stage Applicability
| Term | Evidence |
|---|---|
| All life stages | High |
Serous and glandular secretions of the airway epithelium contribute to the ASL, and epithelial ion channel (e.g. CFTR, ENaC, CaCC, BK) function is critical to normal ASL homeostasis. Should the PCL decrease in depth, liquid will be absorbed from the mucus layer until the necessary depth is restored. Conversely, the mucus layer will absorb surplus PCL to reduce any increase in its depth. The regulation of these reabsorption processes is complex and not fully elucidated (Boucher, 2004). Experimental evidence suggests that the balance between Na+ absorption and Cl− secretion mediated by ENaC and CFTR plays a major role, with the ion channels affecting each other’s activity (increased CFTR activity leads to decreased ENaC activity and vice versa) (Boucher, 2003; Boucher, 2004; Schmid et al., 2011). Mechanistic studies with selective CFTR and ENaC inhibitors suggest that the sensors for regulating ASL height lie within the ASL itself (Boucher, 2003; Hobbs et al., 2013). Additionally, ATP, adenosine and other purinergic receptor agonists, adenylate cyclase and cyclic adenosine monophosphate (cAMP)-dependent protein kinases acting on CFTR and/or ENaC ensure that the ASL height is adjusted to the appropriate height, resulting in maintenance of PCL depth at approximately the length of cilia (Antunes and Cohen, 2007). If the CFTR-ENaC interaction is perturbed, the airways become “dehydrated” (i.e., the ASL height decreases), resulting in slowing or inhibition of cilia movement and impaired MCC (Munkholm and Mortensen, 2014).
| ID | Experimental Design | Species | Upstream Observation | Downstream Observation | Citation (first author, year) | Notes |
|---|
| Title | First Author | Biological Plausibility |
Dose Concordance |
Temporal Concordance |
Incidence Concordance |
|---|
Biological Plausibility
Dose Concordance Evidence
Temporal Concordance Evidence
Incidence Concordance Evidence
Uncertainties and Inconsistencies
The process of reabsorption of excess liquid to regulate ASL height is also known as “isosmotic volume hypothesis” or “isotonic volume transport/mucus clearance hypothesis” and implies that CFTR assumes a critical role in regulating ASL height by inhibiting ENaC activity (Ganesan et al., 2013; Matsui et al., 1998). However, an alternative, opposing hypothesis exists, the “hypotonic hypothesis” which states “that normal airway epithelia are covered by an ASL with a [NaCl] sufficiently low (≤ 50 mM NaCl) to activate defensins and create an antimicrobial “shield” on airway surfaces”, and there is evidence to both support and refute it (Cowley et al., 1997; Goldman et al., 1997; Jayaraman et al., 2001; Knowles et al., 1997; Landry and Eidelman, 2001; Matsui et al., 1998; Tarran et al., 2001a; Tarran et al., 2001b; Verkman et al., 2003). Other studies suggest the involvement of additional ion channels such as alternative chloride channels (Grasemann et al., 2007) and cyclic nucleotide-gated cation channels, particularly in the alveolar epithelium (Schwiebert et al., 1997; Wilkinson et al., 2011) in the regulation of ASL height. In addition, one study showing that instillation of Pseudomonas aeruginosa-laden agarose beads into excised swine tracheas significantly increased ASL height, and that this increase could be blocked by pre-incubation with the CFTR inhibitor CFTRinh172 (100 μM, 30 minutes) (Luan et al., 2014) presents an inconsistency with the available evidence presented here.
Unknown
While there are convincing quantitative data in support of this KER, it becomes clear from the review of the evidence that the downstream KE can only be modulated to a certain extent (e.g., maximal decrease in ASL height did not exceed 50% in the majority of studies), independent of the extent of change in the upstream KE. In addition, the available temporal data indicates that acute exposures predominantly cause a transient, rather than a lasting change in the downstream KE. Since chronic treatment data are not available, we judge our quantitative understanding as being moderate.
Response-response Relationship
Treatment of fully differentiated primary human bronchial epithelial cells (HBECs) with 2% cigarette smoke extract (CSE; bubbling 10 puffs of smoke from one 3R4F reference into 1 mL DMSO, at 2 s/10 mL puff, 10 puffs over 3 min; defined as 100%) for 20 minutes reduced CFTR channel activity by 50% and ASL by approx. 2-fold (Raju et al., 2016).
Apical treatment of primary HBECs grown in monolayers with 2% CSE (not further described) for 24 h decreased forskolin-induced Cl− currents by ca. 20% and ASL height by approx. 25%, and this could be counteracted by co-treatment with 10 µM ivacaftor, a CFTR potentiator known to significantly augment cAMP-mediated ion transport activity (Sloane et al., 2012).
Exposure of primary HBECs to cigarette smoke (5 min, ca. 12 puffs at 1 puff every 30 s; generated according to ISO standards) resulted in efficient removal of CFTR from the plasma membrane and a ca. 2-fold reduction in ASL height (Xu et al., 2015).
Exposure of primary HBECs, differentiated at the air-liquid interface, to cigarette smoke from 1 cigarette (ten 35-mL puffs, 2R4F reference cigarette) nearly abolished responses of the transepithelial electric potential difference Vt to ADO (i.e., blocking the ADO-A2b-cAMP-CFTR- active ion transport) and significantly decreased ASL volume/height by approx. 2-fold after 30 min (Clunes et al., 2012).
Exposure of fully differentiated primary HBECs to 30 puffs of whole smoke from 2 cigarettes (generated according to ISO standards) every day for 5 days (120 h) resulted in a ca. 40% reduction in CFTR expression and approx. 50% reduction in ASL height (Hassan et al., 2014).
Exposure of primary human airway epithelial cells grown in monolayers to whole smoke (3R4F reference cigarette; inExpose exposure system; 3L/min) resulted in significant reduction of CFTR Cl− currents (ca. 40% for a 30-min exposure) and significantly decreased ASL depth from 11.4± 4.1 to 5.6± 2.0 µm (Lambert et al., 2014).
Exposure of fully differentiated primary HBECs to smoke from 1 cigarette or little cigar every day for 5 days (1 × 35 ml puff per 30 second, up to a butt length of 36 mm) significantly reduced CFTR protein expression by ca. 2- to 4-fold and ASL height by 10 to 20% (Ghosh et al., 2017).
Cell surface CFTR protein expression was reduced by ca. 70% following exposure of baby hamster kidney cells expressing human CFTR (BHKCFTR) to cigarette smoke for 10 min (3R4F reference cigarette, 1 puff per min according to ISO standards). This was accompanied by a significant reduction in ASL height by approx. 50% (Rasmussen et al., 2014).
Treatment of primary HBECs from a cystic fibrosis patient with the ΔF508 mutation, grown as monolayer at the air-liquid interface, with the CFTR corrector VX-809 for 48 h increased CFTR maturation by ca. 8-fold and enhanced Cl− transport by approx. 4-fold, from 1.9±0.4 to 7.8±1.3 μA/cm2. VX-809 treatment for 5 days increased ASL height from 4.5±0.2 to 6.7±0.5 μm. Addition of 3 μM VX-770 further increased the ASL height to 9.2±0.2 μm (Van Goor et al., 2011). Treatment of primary HBECs from a G551D/ΔF508 cystic fibrosis patient, grown as monolayer at the air-liquid interface, with the CFTR potentiator VX-770 (10 µM) for 72 h dose-dependently increased forskolin-mediated Cl− currents by ca. 10-fold to 27±2 μA/cm2 and ASL volume to 125% that of controls (Van Goor et al., 2009).
ASL depth in the excised tracheas of rats without functional CFTR expression was approx. half that of wild-type animals (Tuggle et al., 2014).
Knockdown of mRNAs for the α- and β-ENaC subunits resulted in a ca. 70% decrease in amiloride-sensitive currents and a significant increase in ASL height from 6.8±0.5 and 7.4±0.5 µm to 9.8±0.6 and 9.6±0.8 µm in non-CF and CF epithelia, respectively (Gianotti et al., 2013).
Knockdown of α-ENaC mRNA in BMI1-transduced cystic fibrosis bronchial epithelial cells resulted in ca. 50% reduction in protein expression, reduction in amiloride-sensitive short-circuit current from 11.5 (siRNA control) to 6.4 μA/cm2 and increase in ASL height from 7.9 (siRNA control) to 12.1 μm (Tagalakis et al., 2018).
Overexpression of the β-ENaC subunit in mouse airways increased basal and amiloride-sensitive short-circuit currents approx. 2-fold (excised tracheas; compared to wild-type) and significantly reduced ASL height in bronchi and tracheas (by approx. 2 µm) (Mall et al., 2004).
Time-scale
Treatment of fully differentiated primary HBECs with 2% CSE (bubbling 10 puffs of smoke from one 3R4F reference into 1 mL DMSO, at 2 s/10 mL puff, 10 puffs over 3 minutes; defined as 100%) for 24 h decreased total CFTR expression and cell surface CFTR expression by approx. 20 and 25%, respectively, but treatment for 20 min did not. A 50% reduction in CFTR channel activity occurred immediately after addition of CSE and lasted for at least 20 minutes. A 2-fold reduction in ASL height was seen after 20 minutes, and ASL height was only partially restored at 1 h after CSE treatment (Raju et al., 2016a).
Following exposure of primary HBECs, differentiated at the air-liquid interface, to cigarette smoke from 1 cigarette (ten 35-mL puffs, 2R4F reference cigarette), ASL volume/height was significantly decreased by approx. 2-fold after 30 min. This decrease lasted for >2.5 h, and ASL height was restored at 4 h post-exposure (Clunes et al., 2012). Exposure of BHKCFTR cells to cigarette smoke for 10 min (3R4F reference cigarette, 1 puff per min according to ISO standards) resulted in a reduction in ASL height by approx. 50% within 30 min; the decrease lasted for up to 1 h post-exposure (Rasmussen et al., 2014).
Exposure of fully differentiated primary human HBECs to 30 puffs of whole smoke from 2 cigarettes (generated according to ISO standards) was sufficient to decrease ASL height by approx. 50% within 1 h of exposure, and following daily exposure for another 4 days, ASL height remained at around this level (Hassan et al., 2014).
Exposure of fully differentiated primary HBECs to whole smoke from four 3R4F reference cigarettes (generated according to ISO standard 3308; Vitrocell VC10 exposure system) resulted in a small, non-significant increase in ASL volume 4 h post-exposure. ASL volume decreased to baseline levels 7 h post-exposure and continued to drop below baseline levels until 24 h post-exposure (Schmid et al., 2015).
ASL height of primary HBECs dropped within 30 min of exposure to cigarette smoke (5 min, ca. 12 puffs at 1 puff every 30 seconds). ASL height stayed at that reduced level up to until 70 min post-exposure (Xu et al., 2015).
The maximum effect of VX-809 treatment on Cl− currents of primary human bronchial epithelial cells, grown as monolayer at the air-liquid interface, occurred following 24 h, and Cl– transport returned to uncorrected levels within 48 h of compound washout (concurrent ASL data not available) (Van Goor et al., 2011).
Known Feedforward/Feedback loops influencing this KER
Unknown
Phylogenetic analysis of CFTR DNA sequences across multiple species suggests a close evolutionary relationship between human and primate CFTR, followed by rabbit, guinea pig, equine, ovine, and bovine CFTR, whereas rodent CFTR DNA largely diverges from the human DNA (Chen et al., 2001). Of note, CFTR ion permeability differs from species to species (Higgins, 1992). For example, murine CFTR displays reduced channel activity compared with its human counterpart, while ovine CFTR exhibits higher ATP sensitivity, greater single-channel conductance and larger open probability than human CFTR. Moreover, sensitivity to pharmacological agents able to potentiate or block CFTR gating varies greatly from species to species (Bose et al., 2015). Therefore, results from animal studies are not directly transferable to human.
To date, ASL has been investigated in several species including mice, rats, guinea pigs, ferrets, cats, dogs, cows, monkeys, and humans. Although most studies provide data on its composition rather than its height, it is reasonable to assume that regulation of ASL height is equally critical to MCC across these species.
CFTR dysfunction as a consequence of inherited CFTR gene defects is studied in pediatric as well as adult cystic fibrosis patients. Acquired CFTR dysfunction following inhalation exposures (e.g. to cigarette smoke) may also apply to both pediatric and adult populations, depending on the setting and type of exposure, and this also applies to decreased ASL height.
To our knowledge, the role of gender has not been systematically evaluated in acquired CFTR dysfunction and its impact on ASL height. It is thought that the observed suppression of CFTR expression and impairment of CFTR function in cigarette smokers is a contributing factor to the pathogenesis of chronic obstructive pulmonary disease (COPD)(Dransfield et al., 2013; Raju et al., 2016). The main risk factor for COPD is cigarette smoking, and COPD is more common in men than in women, which may be directly related to the higher prevalence of smoking in men, although this gender gap is closing (Hitchman and Fong, 2011; Ntritsos et al., 2018; Syamlal et al., 2014). Nevertheless, the available clinical evidence in support of this AOP suggests that there is no remarkable gender difference.