Internalization and down-regulation of the prostacyclin receptor in human platelets
| Autor: | Ornella Letari, Simonetta Nicosia, S. Giovanazzi, Maria Rosa Accomazzo, D. Oliva |
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| Rok vydání: | 1997 |
| Předmět: |
Blood Platelets
Platelet Aggregation media_common.quotation_subject Receptors Prostaglandin Down-Regulation Prostacyclin In Vitro Techniques Receptors Epoprostenol Binding Competitive Biochemistry medicine Humans Platelet Iloprost Binding site Internalization Receptor Molecular Biology Prostacyclin receptor media_common Chemistry Cell Membrane Infant Newborn Cell Biology Epoprostenol Kinetics Dinitrophenol Biophysics lipids (amino acids peptides and proteins) Adenylyl Cyclases Subcellular Fractions Research Article medicine.drug |
| Zdroj: | Biochemical Journal. 325:71-77 |
| ISSN: | 1470-8728 0264-6021 |
| Popis: | The internalization of [3H]iloprost, a prostacyclin analogue, was studied in human platelets by binding studies. After incubation with [3H]iloprost at 37 degrees C, addition of unlabelled ligand at either 37 degrees C or 4 degrees C caused dissociation of 74% and 52% of the bound ligand respectively, suggesting that a portion had been internalized. The percentage of [3H]iloprost bound at equilibrium to the surface (evaluated by acid treatment) at either 37 degrees C or 4 degrees C was markedly different (80% versus 25%). Internalization was dependent on time and on the ligand nature and concentration. Energy-depleting agents (dinitrophenol and 2-deoxyglucose) completely inhibited internalization, whereas probenecid (inhibitor of organic anion transporters) did not affect it significantly. Subcellular fractionation indicated that, at 4 degrees C or in the absence of ligand, most of the receptor was present in membrane fractions (pellet at 27000 or 105000 g), whereas, when platelets were preincubated at 37 degrees C with iloprost, the receptor was found mainly in the cytosolic fraction. In platelets preincubated with iloprost at 4 degrees C, two classes of binding sites were present, whereas after preincubation at 37 degrees C only the lower-affinity sites were detected. After exposure to the agonist, iloprost-induced inhibition of platelet aggregation and activation of adenylate cyclase and cAMP production were significantly lower. Taken together, these data demonstrate that human platelets can internalize a high-affinity binding site for iloprost, presumably the prostacyclin receptor. |
| Databáze: | OpenAIRE |
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