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Erythrocytes l-Arginine y+ Transporter Inhibition by N-Ethylmaleimide in Ice-bath

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Abstract

Erythrocytes l-arginine uptake is conveyed by y+ and y+L membrane transport systems. Pre-incubation with N-ethylmaleimide for 10 min at 37°C inhibits the y+ system. The aim of this study was to determine the ideal pre-incubation temperature in evaluating y+ and y+L systems. Cells were pre-incubated with or without N-ethylmaleimide for 10 min at 4°C and 37°C. l-Arginine uptake was quantified by radioisotope and standard erythrocytes membrane flux methodology. Results demonstrate that erythrocytes l-arginine content is depleted by pre-incubation at 37°C for 10 min, thus changing the V max measurement. The inhibitory effect of N-ethylmaleimide pre-incubation was temperature independent and already complete after 1 min of incubation. No significant difference in kinetic parameters was detected between cells pre-incubated at 37°C or 4°C, under zero-trans conditions. In conclusion, we suggest that measurement of erythrocytes l-arginine uptake by y+ and y+L systems could be carried out without N-ethylmaleimide pre-incubation at 37°C.

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Acknowledgments

We thank the São Lucas Hospital Blood Bank for providing human blood samples, and Dr Célia Carlini for critical review of the manuscript. The Nephrology Laboratory has the support of Secretaria de Ciência e Tecnologia do Estado do Rio Grande do Sul (SCT-RS), Fundação de Amparo à Pesquisa do Estado do Rio Grande do Sul (FAPERGS), Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES), Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) and PUCRS, Brazil. Poli-de-Figueiredo is a CNPq researcher.

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  1. Programa de Pós-graduação em Medicina e Ciências da Saúde (Nefrologia), Faculdade de Medicina/Instituto de Pesquisas Biomédicas/Hospital São Lucas, Pontifícia Universidade Católica do Rio Grande do Sul, 2° Andar - Laboratório de Nefrologia, Instituto de Pesquisas Biomédicas/HSL, Av Ipiranga 6690, Porto Alegre, RS, 90610-000, Brazil

    Bartira Ercília Pinheiro da Costa, Priscilla Barcellos de Almeida, Ioná Rosine Conceição, Ivan Carlos Ferreira Antonello, Domingos O. d’Avila & Carlos Eduardo Poli-de-Figueiredo

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  1. Bartira Ercília Pinheiro da Costa
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  2. Priscilla Barcellos de Almeida
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  6. Carlos Eduardo Poli-de-Figueiredo
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Correspondence to Bartira Ercília Pinheiro da Costa.

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Pinheiro da Costa, B.E., de Almeida, P.B., Conceição, I.R. et al. Erythrocytes l-Arginine y+ Transporter Inhibition by N-Ethylmaleimide in Ice-bath. Cell Biochem Biophys 58, 69–73 (2010). https://doi.org/10.1007/s12013-010-9089-9

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