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  • Selective blockade of phosphodiesterase types 2, 5 and 9 results in cyclic 3′5′ guanosine monophosphate accumulation in retinal pigment epithelium cells

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Laboratory science - Extended reports
Selective blockade of phosphodiesterase types 2, 5 and 9 results in cyclic 3′5′ guanosine monophosphate accumulation in retinal pigment epithelium cells
  1. R M H Diederen1,
  2. E C La Heij2,
  3. M Markerink-van Ittersum1,
  4. A Kijlstra2,
  5. F Hendrikse2,
  6. J de Vente1
  1. 1Department of Psychiatry and Neuropsychology, European Graduate School of Neuroscience, University of Maastricht, Maastricht, The Netherlands
  2. 2Department of Ophthalmology, Eye Research Institute Maastricht, University Hospital Maastricht, Maastricht, The Netherlands
  1. Correspondence to: R Diederen Department of Ophthalmology, University Hospital Maastricht, PO Box 5800, P Debyelaan 25, 6202 AZ Maastricht, The Netherlands; r.diederen{at}NP.unimaas.nl

Abstract

Aim: To investigate which phosphodiesterase (PDE) is involved in regulating cyclic 3′5′ guanosine monophosphate breakdown in retinal pigment epithelium (RPE) cells.

Methods: cGMP content in the cultured RPE cells (D407 cell line) was evaluated by immunocytochemistry in the presence of non-selective or isoform-selective PDE inhibitors in combination with the particulate guanylyl cyclase stimulator atrial natriuretic peptide (ANP) or the soluble guanylyl cyclase stimulator sodium nitroprusside (SNP). mRNA expression of PDE2, PDE5 and PDE9 was studied in cultured human RPE cells and rat RPE cell layers using non-radioactive in situ hybridisation.

Results: In the absence of PDE inhibitors, cGMP levels in cultured RPE cells are very low. cGMP accumulation was readily detected in cultured human RPE cells after incubation with Bay60–7550 as a selective PDE2 inhibitor, sildenafil as a selective PDE5 inhibitor or Sch51866 as a selective PDE9 inhibitor. In the presence of PDE inhibition, cGMP content increased markedly after stimulation of the particulate guanylyl cyclase. mRNA of PDE2,PDE5 and PDE9 was detected in all cultured human RPE cells and also in rat RPE cell layers.

Conclusions: PDE2, PDE5 and PDE9 have a role in cGMP metabolism in RPE cells.

  • ANP, atrial natriuretic peptide
  • cAMP, cyclic adenosine monophosphate
  • cGMP, cyclic 3′,5′guanosine monophosphate
  • DIG, digoxigenin
  • IBMX, 3-isobutyl-1-methylxanthine
  • PDE, phosphodiesterase
  • RPE, retinal pigment epithelium
  • SNP, sodium nitroprusside
  • SSC, TBS, TRIS-buffered saline
  • TBS-T, TRIS-buffered saline Triton X

https://doi.org/10.1136/bjo.2006.100628

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    Footnotes

    • Published Online First 30 August 2006

    • Funding: This study was supported by the Algemene Nederlandse Vereniging ter Voorkoming van Blindheid.

    • Competing interests: None.