Characterization of a constitutive type III nitric oxide synthase in human U937 monocytic cells: stimulation by soluble CD23
| Autor: | J P Kolb, B Dugas, A Abadie, C Amirand, V. Roman, Haixia Zhao, N Dugas |
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| Rok vydání: | 1997 |
| Předmět: |
Calmodulin
Immunology chemistry.chemical_element Gene Expression Calcium Polymerase Chain Reaction Monocytes Nitric oxide chemistry.chemical_compound Tumor Cells Cultured Immunology and Allergy Humans RNA Messenger Cyclic GMP Calcium metabolism U937 cell biology Receptors IgE CD23 Molecular biology Nitric oxide synthase EGTA chemistry Biochemistry Solubility biology.protein Nitric Oxide Synthase Research Article |
| Zdroj: | Scopus-Elsevier |
| ISSN: | 0019-2805 |
| Popis: | The soluble cleavage fragment of the low-affinity immunoglobulin E (IgE) receptor/CD23 (sCD23 25000 MW) and antibodies directed against their receptors on monocytes, CD11b and CD11c, stimulate the production of nitric oxide (NO) by these cells and we have suggested that the enzyme involved could be related to the endothelial constitutive type III nitric oxide synthase (ecNOS). In the present work, we have analysed the characteristic properties of this NOS isoform in the model of the human promonocytic cells U937 By reverse-transcription polymerase chain reaction (RT-PCR), the presence of an mRNA coding for type III NOS was found in U937 cells and the corresponding protein was detected by immunofluorescence in permeabilized cells with a specific anti-ecNOS monoclonal antibody (mAb). Membrane extracts displayed a NOS activity dependent on the presence of calcium and calmodulin in the reaction medium and that was abrogated in the presence of EGTA. Recombinant soluble CD23 (25000 MW) was found to trigger an NO-dependent cGMP accumulation in these cells, which was abrogated by calcium chelators and inhibitors of the calcium/calmodulin complex. Moreover, sCD23 elicited a transient augmentation of intracytoplasmic free calcium concentration [Ca2+]i that was dependent on the presence of calcium in the external buffer and was prevented in the presence of EGTA, indicating that it was due to a calcium influx. In conclusion, human promonocytic cells such as U937 exhibit a functional type III NOS that can be stimulated by calcium-raising agents, such as sCD23. |
| Databáze: | OpenAIRE |
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