Journal article

Publication year: 2016

Pages: 75-83

Journal: Molecular and Biochemical Parasitology

Volume number: 206

Issue number: 1-2

ISSN: 0166-6851

DOI Link: https://doi.org/10.1016/j.molbiopara.2015.11.002

Publisher: Elsevier

Abstract: 
Glutathione plays a crucial role in the redox regulation of the malaria parasite Plasmodium falciparum and is linked to drug resistance mechanisms, especially in resistance against the antimalarial drug chloroquine (CQ). The determination of the glutathione-dependent redox potential was recently established in living parasites using a cytosolically expressed biosensor comprising redox-sensitive green fluorescent protein coupled to human glutaredoxin 1 (hGrx1-roGFP2). In order to further elucidate redox changes induced by antimalarial drugs and to consolidate the application spectrum of the ratiometric biosensor we systematically compared it to other methods probing thiol and redox metabolism. Among these methods were cell disruptive and non -disruptive approaches including spectrophotometric assays with Ellman's reagent and naphthalene dicarboxyaldehyde as well as molecular probes such as ThiolTracker (TM) Violet and the dichlorofluorescein-based probe CM-H2DCFDA. To directly compare the methods, blood stages of the CQ-sensitive P. falciparum 3D7 strain were challenged with the oxidative agent diamide and the antimalarial drugs artemisinin and CQ for 1 h, 4 h, and 24 h. For all conditions, dose -dependent changes in the different redox parameters could be monitored which are compared and discussed. We furthermore detected slight differences in thiol status of parasites transiently transfected with hGrxlroGFP2 in comparison with control 3D7 cells. In conclusion, ThiolTrackerTM Violet and, even more so, the hGrxl-roGFP2 probe reacted reliably and sensitively to drug induced changes in intracellular redox metabolism. These results were substantiated by classical cell disruptive methods. (C) 2015 Elsevier B.V. All rights reserved.

Harvard Citation style: Mohring, F., Jortzik, E. and Becker, K. (2016) Comparison of methods probing the intracellular redox milieu in Plasmodium falciparum, Molecular and Biochemical Parasitology, 206(1-2), pp. 75-83. https://doi.org/10.1016/j.molbiopara.2015.11.002

APA Citation style: Mohring, F., Jortzik, E., & Becker, K. (2016). Comparison of methods probing the intracellular redox milieu in Plasmodium falciparum. Molecular and Biochemical Parasitology. 206(1-2), 75-83. https://doi.org/10.1016/j.molbiopara.2015.11.002