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Human red blood cells at work: identification and visualization of erythrocytic eNOS activity in health and disease

Cortese-Krott, Miriam M., Rodriguez-Mateos, Ana, Sansone, Roberto, Kuhnle, Gunter G.C., Thasian-Sivarajah, Sivatharsini, Krenz, Thomas, Horn, Patrick, Krisp, Christoph, Wolters, Dirk, Heiß, Christian, Kroncke, Klaus-Dietrich, Hogg, Neil, Feelisch, Martin and Kelm, Malte (2012) Human red blood cells at work: identification and visualization of erythrocytic eNOS activity in health and disease Blood, 120, pp. 4229-4237. (doi:10.1182/blood-2012-07-442277). (PMID:23007404).

Record type: Article

Abstract

A nitric oxide synthase (NOS)-like activity has been demonstrated in human red blood cells (RBCs), but doubts about its functional significance, isoform identity and disease relevance remain. Using flow cytometry in combination with the NO-imaging probe DAF-FM we find that all blood cells form NO intracellularly, with a rank order of monocytes > neutrophils > lymphocytes > RBCs > platelets. The observation of a NO-related fluorescence within RBCs was unexpected given the abundance of the NO-scavenger oxyhemoglobin. Constitutive normoxic NO formation was abolished by NOS inhibition and intracellular NO scavenging, confirmed by laser-scanning microscopy and unequivocally validated by detection of the DAF-FM reaction product with NO using HPLC and LC-MS/MS. Employing immunoprecipitation, ESI-MS/MS-based peptide sequencing and enzymatic assay we further demonstrate that human RBCs contain an endothelial NOS (eNOS) that converts L-3H-Arginine to L-3H-Citrulline in a Ca2+/Calmodulin-dependent fashion. Moreover, in patients with coronary artery disease, red cell eNOS expression and activity are both lower than in age-matched healthy individuals and correlate with the degree of endothelial dysfunction. Thus, human RBCs constitutively produce NO under normoxic conditions via an active eNOS isoform the activity of which is compromised in patients with coronary artery disease.

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e-pub ahead of print date: 24 September 2012
Organisations: Clinical & Experimental Sciences

Identifiers

Local EPrints ID: 343579
URI: http://eprints.soton.ac.uk/id/eprint/343579
ISSN: 0006-4971
PURE UUID: 06fee4b7-1ac3-44ff-9529-4aa1273cfb61
ORCID for Martin Feelisch: ORCID iD orcid.org/0000-0003-2320-1158

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Date deposited: 04 Oct 2012 16:05
Last modified: 18 Jul 2017 05:21

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Contributors

Author: Miriam M. Cortese-Krott
Author: Ana Rodriguez-Mateos
Author: Roberto Sansone
Author: Gunter G.C. Kuhnle
Author: Sivatharsini Thasian-Sivarajah
Author: Thomas Krenz
Author: Patrick Horn
Author: Christoph Krisp
Author: Dirk Wolters
Author: Christian Heiß
Author: Klaus-Dietrich Kroncke
Author: Neil Hogg
Author: Martin Feelisch ORCID iD
Author: Malte Kelm

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