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Mechanisms associated to impaired activity of cardiac P-type ATPases in endothelial nitric oxide synthase knockout mice

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Abstract

The effect of long-lasting in vivo restriction of nitric oxide (NO) bioavailability on cardiac and renal P-type ATPases critical for intracellular ion homeostasis is controversial. Previous work has shown in eNOS knockout (eNOS−/−) mice hearts that Na+/K+- and Ca2+-ATPase activities were depressed but the underlying mechanisms are still unclear. The goal of this study was to characterize potential alterations responsible for impaired enzyme activity in eNOS−/− mice. Na+/K+-ATPase activity from crude preparations of adult male eNOS−/− mice hearts and kidneys was reduced compared with wild-type animals (32 %, p < 0.05 and 16 %, p < 0.0001, respectively). Immunoblot analysis showed that although the expression of the predominant (or exclusive, for the kidney) Na+/K+-ATPase α1 isoform was not significantly changed, there was an important downregulation of the less abundant α2 isoform in the heart (57 %, p < 0.0001). In addition, although cardiac Ca2+-ATPase activity was unaltered, the expression of sarco/endoplasmic reticulum Ca2+-ATPase 2 protein in eNOS−/− mice was very high (290 % compared with wild-type animals, p < 0.0001) without any significant change in phospholamban expression. Consistent with these findings, the content of cardiac and renal free sulfhydryl groups, essential for the catalytic function of such ATPases, was decreased (23 %, p < 0.01 and 35 %, p < 0.05, respectively). Altogether, the present results suggest that the absence of eNOS promotes a compartmentalized altered redox balance that affects the activity and expression of ion transport ATPases.

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Abbreviations

DTNB:

5,50-dithiobis-(2-nitrobenzoic acid)

eNOS:

Endothelial nitric oxide synthase

eNOS−/− :

eNOS knockout

l-NAME:

N G-nitro-l-arginine methyl ester

NO:

Nitric oxide

PMCA:

Plasma membrane Ca2+-ATPase

PMSF:

Phenylmethylsulfonyl fluoride

ROS:

Reactive oxygen species

SERCA:

Sarco/endoplasmic reticulum Ca2+-ATPase

WT:

Wild type

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Acknowledgments

Research supported by FAPERJ, FAPESP, and CNPq-PRONEX. A. Caricati-Neto, A. Jurkiewicz, and F. Noël are recipients of CNPq research fellowships.

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Authors and Affiliations

  1. Laboratório de Farmacologia Bioquímica e Molecular, Instituto de Ciências Biomédicas, Universidade Federal do Rio de Janeiro, Av. Carlos Chagas Filho, 373, CCS, Bloco J, sala J01-017, Ilha do Fundão, 21941-902, Rio de Janeiro, RJ, Brazil

    Daniele C. Rezende, François Noël & Luis E. M. Quintas

  2. Insitituto Federal de Educação, Ciência e Tecnologia do Rio de Janeiro, Rio de Janeiro, RJ, Brazil

    Elisa S. C. Pôças

  3. Laboratório de Farmacologia de Órgãos e Sistemas, Instituto de Ciências Biomédicas, Universidade Federal do Rio de Janeiro, Rio de Janeiro, RJ, Brazil

    Humberto Muzi-Filho & Valéria M. N. Cunha

  4. Departamento de Farmacologia, Universidade Federal de São Paulo, São Paulo, SP, Brazil

    Afonso Caricati-Neto & Aron Jurkiewicz

Authors
  1. Daniele C. Rezende
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  2. Elisa S. C. Pôças
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  3. Humberto Muzi-Filho
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  8. Luis E. M. Quintas
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Correspondence to Luis E. M. Quintas.

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Rezende, D.C., Pôças, E.S.C., Muzi-Filho, H. et al. Mechanisms associated to impaired activity of cardiac P-type ATPases in endothelial nitric oxide synthase knockout mice. J Physiol Biochem 69, 207–214 (2013). https://doi.org/10.1007/s13105-012-0203-x

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  • DOI: https://doi.org/10.1007/s13105-012-0203-x

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