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Extracellular Methemoglobin Mediated Early ROS Spike Triggers Osmotic Fragility and RBC Destruction: An Insight into the Enhanced Hemolysis During Malaria

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Abstract

Malaria infection is known to cause severe hemolysis due to production of abnormal RBCs and enhanced RBC destruction through apoptosis. Infected RBC lysis exposes uninfected RBC to the large amount of pro-oxidant molecules such as methemoglobin. Methemoglobin (MetHb) exposure dose dependently makes RBCs susceptible to osmotic stress and causes hemolysis. MetHb mediated oxidative stress in RBC correlated well with osmotic fragility and hemolysis. Interestingly, a reactive oxygen species (ROS) spike at 15 min was responsible for the observed effects on RBC cells. Two natural antioxidants N-acetyl cysteine and mannitol protected the RBC from MetHb-mediated defects, which clearly indicated involvement of oxidative stress in the process. MetHb due to its pseudo-peroxidase activity produces ROS in the external microenvironment. Therefore, classical peroxidase inhibitors were tested to probe peroxidase activity mediated ROS production with defects in RBCs. Clotrimazole (CLT), which irreversibly inactivates the MetHb (CLT-MetHb) and abolishes peroxidase activity, did not produce significant ROS outside RBC and was inefficient to cause osmotic fragility and hemolysis. Hence, initiating a chain reaction, MetHb released from ruptured RBC produces significant ROS in the external microenvironment to make RBC membrane leaky and enhanced hemolysis. Together data presented in the current work explored the role of MetHb in accelerated humorless during malaria which could be responsible for severe outcomes of pathological disorders.

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Abbreviations

GSH:

Reduced glutathione

NAC:

N-acetyl cysteine

ROS:

Reactive oxygen species

MetHb:

Methemoglobin

CLT-MetHb:

Clotrimazole modified methemoglobin

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Acknowledgments

This work has partially been supported by the Department of Atomic energy (DAE) young scientist award and IITG startup grant to VT. Both Authors acknowledge the infrastructure facility provided by the Department of Biotechnology, IIT-Guwahati, Guwahati, Assam (India). We are thankful to Dr. Amogh A Sahasrabuddhe, Scientist, Central Drug Research Institute (CDRI), Lucknow for valuable suggestions and for critically reviewing the manuscript.

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No competing financial interests exist.

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Authors and Affiliations

  1. Malaria Research Group, Department of Biotechnology, Indian Institute of Technology-Guwahati, Guwahati, 781039, Assam, India

    S. N. Balaji & Vishal Trivedi

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  1. S. N. Balaji
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  2. Vishal Trivedi
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Correspondence to Vishal Trivedi.

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Balaji, S.N., Trivedi, V. Extracellular Methemoglobin Mediated Early ROS Spike Triggers Osmotic Fragility and RBC Destruction: An Insight into the Enhanced Hemolysis During Malaria. Ind J Clin Biochem 27, 178–185 (2012). https://doi.org/10.1007/s12291-011-0176-5

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  • DOI: https://doi.org/10.1007/s12291-011-0176-5

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