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Journal of Neuroinflammation
Published in: Journal of Neuroinflammation 1/2016

Open Access 01-12-2016 | Research

Clozapine metabolites protect dopaminergic neurons through inhibition of microglial NADPH oxidase

Authors: Lulu Jiang, Xuefei Wu, Shuo Wang, Shih-Heng Chen, Hui Zhou, Belinda Wilson, Chun-Yang Jin, Ru-Band Lu, Keqin Xie, Qingshan Wang, Jau-Shyong Hong

Published in: Journal of Neuroinflammation | Issue 1/2016

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Abstract

Background

Clozapine, an atypical antipsychotic medication, has been effectively used to treat refractory schizophrenia. However, the clinical usage of clozapine is limited due to a high incidence of neutropenia or agranulocytosis. We previously reported that clozapine protected dopaminergic neurons through inhibition of microglial activation. The purpose of this study was to explore the neuroprotective effects of clozapine metabolites clozapine N-oxide (CNO) and N-desmethylclozapine (NDC), as well as their propensity to cause neutropenia.

Methods

The primary midbrain neuron-glia culture was applied to detect the neuroprotective and anti-inflammatory effect of clozapine and its metabolites in lipopolysaccharide (LPS) and MPP+-induced toxicity. And the subsequent mechanism was demonstrated by gp91 phox mutant cell cultures as well as microgliosis cell lines. In vivo, to confirm the neuroprotective effect of clozapine and CNO, we measured the dopaminergic neuronal loss and rotarod motor deficits in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-generated mouse Parkinson’s disease (PD) model. The neutropenia or agranulocytosis of clozapine and its metabolites was illustrated by white blood cell count of the treated mice.

Results

We found that, in midbrain neuron-glia cultures, CNO and NDC were more potent than clozapine in protecting dopaminergic neurons against LPS and MPP+-induced toxicity. CNO and NDC-afforded neuroprotection was linked to inhibition of microglia-mediated neuroinflammation, as demonstrated by abolished neuroprotection in microglia-depleted cultures and their capacity of inhibiting LPS-induced release of proinflammatory factors from activated microglia. NADPH oxidase (NOX2) was subsequently recognized as the main target of CNO and NDC since genetic ablation of gp91 phox , the catalytic subunit of NOX2, abolished their neuroprotective effects. CNO and NDC inhibited NOX2 activation through interfering with the membrane translocation of the NOX2 cytosolic subunit, p47 phox . The neuroprotective effects of CNO were further verified in vivo as shown by attenuation of dopaminergic neurodegeneration, motor deficits, and reactive microgliosis in MPTP-generated mouse PD model. More importantly, unlike clozapine, CNO did not lower the white blood cell count.

Conclusions

Altogether, our results show that clozapine metabolites elicited neuroprotection through inactivation of microglia by inhibiting NOX2. The robust neuroprotective effects and lack of neutropenia suggest that clozapine metabolites may be promising candidates for potential therapy for neurodegenerative diseases.
Appendix
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Metadata
Title
Clozapine metabolites protect dopaminergic neurons through inhibition of microglial NADPH oxidase
Authors
Lulu Jiang
Xuefei Wu
Shuo Wang
Shih-Heng Chen
Hui Zhou
Belinda Wilson
Chun-Yang Jin
Ru-Band Lu
Keqin Xie
Qingshan Wang
Jau-Shyong Hong
Publication date
01-12-2016
Publisher
BioMed Central
Published in
Journal of Neuroinflammation / Issue 1/2016
Electronic ISSN: 1742-2094
DOI
https://doi.org/10.1186/s12974-016-0573-z

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