MRP4 Influences cAMP Levels in Ciliary Epithelial cells of the Human Eye
Bibliographic record
Abstract
The ciliary epithelium is important for aqueous humor inflow and maintenance of intraocular pressure. Treatment of ciliary epithelium with β‐adrenergic agonists leads to increased extracellular cAMP and adenosine and stimulation of aqueous humor inflow. The mechanism by which cAMP gets out ciliary epithelial cells is unknown. The purpose was to determine if the multidrug resistance‐associated protein 4 (MRP4) in ciliary epithelial cells influences intracellular and extracellular cAMP levels. Intracellular and extracellular cAMP levels were monitored in cultured ciliary epithelial cells using a FRET‐based assay. Also examined by RT‐PCR in cultured cells and native human ciliary body was the expression of MRP4, and the expression of receptors and enzymes associated with the cAMP‐adenosine signaling pathway. MRP4 mRNA and protein (immunoblotting) were detected in cultured cells and native human ciliary body. There was a dose‐dependent increase in both intracellular and extracellular cAMP levels following treatment of the cells with isoproterenol (ISO, 0–1 mM). Intracellular cAMP levels were ~10‐fold higher in the presence vs. absence of ISO (1 mM), and extracellular cAMP went from being undetectable to detectable. The β2‐adrenergic receptor was detected in the cultured cells and native ciliary body, but β1‐ and β3‐adrenergic receptors were undetectable. MRP4 mRNA was undetectable by qPCR and RT‐PCR in cultured cells following treatment with siRNA against MRP4. There was no difference in basal intracellular cAMP levels between cells treated with negative control siRNA (control) and MRP4 siRNA. Following treatment with ISO (1 mM) intracellular cAMP levels were 2.7‐fold significantly higher in cells treated with MRP4 siRNA compared to control. There was no difference in extracellular cAMP levels between the two treatments. Also present in both cultured cells and native ciliary body were message for enzymes necessary for conversion of cAMP to adenosine, such as ecto‐5‐nucleotidase (CD73). These data show that MRP4 likely contributes to efflux of cAMP from ciliary epithelial cells where it is then converted to adenosine. The cultured cells should prove useful for further study of the cAMP‐adenosine signaling pathway in the ciliary epithelium of the human eye. Support or Funding Information Natural Sciences and Engineering Research Council of Canada
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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 | 0.001 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.001 |
| Insufficient payload (model declined to judge) | 0.003 | 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".