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Interferon Gamma Potentiates the Injury Caused by MPP(+) on SH-SY5Y Cells, Which is Attenuated by the Nitric Oxide Synthases Inhibition

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Abstract

This study examined whether the cytokine interferon (IFN) gamma plays a role in the injury of SH-SY5Y cells caused by MPP(+) (1-methyl-4-phenylpyridinium). First of all, IFN-gamma sensitized cells to the neurotoxin MPP(+), as determined by MTT (3-(4,5-dimethylthiazol-2-y1)-2,5-diphenyltetrazolium bromide) assay. MPP(+)-injured cells showed higher reactive oxygen species (ROS) levels, which was reinforced by IFN-gamma. The injury triggered a marked expression of the neuronal NOS (nNOS) enzyme. L-NAME [N(ω)-nitro-l-arginine methyl ester, a non-specific NOS inhibitor] reestablished the cell viability after IFN-gamma challenging, and recovered cells from MPP(+) injury (95.0 vs. 84.7 %; P < 0.05). Seven-NI (7-nitroindazole, a nNOS inhibitor) protected cells against the injury by MPP(+) co-administered with IFN-gamma. Both inhibitors restrained the apoptosis of SH-SY5Y cells caused by MPP(+)/IFN-gamma. Regarding oxidative stress, L-NAME and 7-NI attenuated the increase in ROS levels caused by MPP(+) (45.3 or 48.4 vs. 87.9 %, P < 0.05). Indeed, L-NAME was more effective than 7-NI for reducing oxidative stress caused by MPP(+) under IFN-gamma exposition. The nNOS gene silencing by small-interfering RNAs recovered cells challenged by IFN-gamma (24 h), or MPP(+) (8 h). In conclusion, IFN-gamma sensitizes cells to MPP(+)-induced injury, also causing an increase in ROS levels. Pretreating cells with L-NAME or 7-NI reverts both the oxidative stress and apoptosis triggered by the neurotoxin MPP(+). Taking together, our data reinforce that IFN-gamma and NOS enzymes play a role in oxidative stress and dopaminergic cell death triggered by MPP(+).

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Acknowledgments

The present research received financial support from the following Brazilian institutions: Coordenação de Aperfeiçoamento de Pessoal de Nível Superior—CAPES, Conselho Nacional de Desenvolvimento Científico e Tecnológico—CNPq, and Fundação de Apoio à Pesquisa do Distrito Federal—FAP-DF.

Conflict of interest

Data regarding nNOS sequences used for designing the structures siRNAnNOShum_3987 (SEQ ID NO: 38/39) and siRNAnNOShum_4400 (SEQ ID NO: 41/42) are under patent registration at the Brazilian Institute for Industrial Property—“Instituto Nacional da Propriedade Industrial”—INPI, BR 10 2012 032844 5.

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  1. Laboratório de Tecnologias para Terapia Gênica, ASS 128, ICC Sul. Universidade de Brasília - UnB, Campus Darcy Ribeiro, FAV, Brasília, DF, 70910-970, Brazil

    Simoneide S. Titze-de-Almeida, Cátia Faria Lustosa, Camila Hillesheim Horst & Ricardo Titze-de-Almeida

  2. Laboratório de Neurofisiologia e Biologia Molecular, Dept. Morfologia Fisiologia e Patologia Básica, FORP, Universidade de São Paulo - USP, Ribeirão Prêto, SP, 14040-904, Brazil

    Elaine Del Bel

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Titze-de-Almeida, S.S., Lustosa, C.F., Horst, C.H. et al. Interferon Gamma Potentiates the Injury Caused by MPP(+) on SH-SY5Y Cells, Which is Attenuated by the Nitric Oxide Synthases Inhibition. Neurochem Res 39, 2452–2464 (2014). https://doi.org/10.1007/s11064-014-1449-1

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