EpOMEs and DiHOMEs Activate TRPA1 and TRPV1 Channels in Airway Smooth Muscle
Bibliographic record
Abstract
Abstract RATIONALE: Oxylipins, bioactive lipid mediators found throughout the lung, are derived from polyunsaturated fatty acids. Classical oxylipins such as the prostaglandins, thromboxanes, and leukotrienes are capable of promoting Airway Smooth Muscle (ASM) contraction or relaxation. Most oxylipins in the lungs remain unexplored in the context of ASM contractility. We have previously shown epoxy-12-octadecenoic acids (EpOMEs) and dihydroxy-12-octadecenoic acids (DiHOMEs), which are elevated in asthmatic lungs following pollen exposure, initiate a rise in ASM intracellular calcium, dependent on the presence of extracellular calcium. Previous research suggests these oxylipins activate the calcium channels ‘TRPA1’ and ‘TRPV1’ in neuronal tissue. These channels are abundant in ASM and may be activated in a similar manner. We therefore hypothesize that EpOMEs and DiHOMEs use TRPA1 and TRPV1 channels to induce transient calcium changes in ASM. METHODS: hTERT immortalized human ASM were incubated with calcium binding indicator dye (Fluo-4) and fluorescence was measured using ‘Cytation 5’. ASM were pre-incubated with either DMSO (vehicle), HC-030031 (TRPA1-inhibitor), AMG 517 (TRPV1-inhibitor) or Gadolinium (cation channel blocker) for 1 hour. 10µM of 12,13-EpOME, 9,10-DiHOME, or 12,13-DiHOME were injected, and resulting fluorescence was measured in triplicate wells. Technical triplicates were averaged, and the final sample size was n=5-8 individual replicates. Data is presented as ΔF/F0 and analyzed via One-Way ANOVA with Dunnet's post-test versus vehicle, significance set at p<0.05. RESULTS: 12,13-EpOME increased fluorescence intensity in ASM pre-incubated with DMSO (0.22±0.09), this increase was significantly ablated by the cation channel inhibitor ‘Gadolinium’ (0.01±0.02). Pre-incubation with the TRPV1 inhibitor ‘AMG 517’ also significantly diminished the response (-0.02±0.19) but pre-incubation with the TRPA1 inhibitor ‘HC-030031’ did not (1.95±0.28). 9,10-DiHOME increased fluorescence intensity with DMSO (0.38±0.34), which was significantly abolished by Gadolinium (0.04±0.04) and AMG 517 (0.05±0.08). HC030031 significantly reduced 9,10-DiHOME response by ∼65% (0.14±0.14). 12,13-DiHOME increased fluorescence intensity with DMSO (0.50±0.28) and was significantly reduced by HC030031 (0.08±0.05), Gadolinium (0.15±0.07), and AMG 517 (0.21±0.25). CONCLUSION: Our data provides mechanistic understanding for the action of EpOMEs and DiHOMEs in ASM. Blocking cation channels reduced the activity of all oxylipins, indicating requirement of an extracellular calcium channel. All oxylipins appear to activate TRPV1 to mediate calcium entry, while only the DiHOMEs appear to activate TRPA1. Interestingly, inhibiting TRPA1 appears to augment the calcium response to 12,13-EpOME, suggesting crosstalk between TRPA1 and TRPV1 in ASM. More work is needed to understand this crosstalk, and to understand whether these pathways govern EpOME and DiHOME induced airway narrowing.
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.001 |
| Insufficient payload (model declined to judge) | 0.004 | 0.001 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one source (direct Gemma or distilled Codex), not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".