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

Cortese-Krott, M. M., Rodriguez-Mateos, Z, Sansone, R., Kuhnle, G. G. C., Thasian-Sivarajah, S., Krenz, T., Horn, P., Krisp, C., Wolters, D., Heiss, C. , Kroncke, K.-D., Hogg, N., Feelisch, M. and Kelm, M. (2012) Human red blood cells at work: identification and visualization of erythrocytic eNOS activity in health and disease BLOOD, 120 (20). pp. 4229-4237.

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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.

Item Type: Article
Divisions : Faculty of Health and Medical Sciences > School of Biosciences and Medicine > Department of Clinical and Experimental Medicine
Faculty of Health and Medical Sciences > School of Biosciences and Medicine
Faculty of Health and Medical Sciences
Authors :
NameEmailORCID
Cortese-Krott, M. M.
Rodriguez-Mateos, Z
Sansone, R.
Kuhnle, G. G. C.
Thasian-Sivarajah, S.
Krenz, T.
Horn, P.
Krisp, C.
Wolters, D.
Heiss, C.c.heiss@surrey.ac.uk
Kroncke, K.-D.
Hogg, N.
Feelisch, M.
Kelm, M.
Date : 24 September 2012
DOI : 10.1182/blood-2012-07-442277
Copyright Disclaimer : Copyright © 2012 American Society of Hematology
Depositing User : Diane Maxfield
Date Deposited : 16 Aug 2018 14:14
Last Modified : 16 Aug 2018 14:14
URI: http://epubs.surrey.ac.uk/id/eprint/848989

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