Tempol, an intracellular antioxidant, inhibits tissue factor expression, attenuates dendritic cell function, and is partially protective in a murine model of cerebral malaria

PLoS One. 2014 Feb 28;9(2):e87140. doi: 10.1371/journal.pone.0087140. eCollection 2014.

Abstract

Background: The role of intracellular radical oxygen species (ROS) in pathogenesis of cerebral malaria (CM) remains incompletely understood.

Methods and findings: We undertook testing Tempol--a superoxide dismutase (SOD) mimetic and pleiotropic intracellular antioxidant--in cells relevant to malaria pathogenesis in the context of coagulation and inflammation. Tempol was also tested in a murine model of CM induced by Plasmodium berghei Anka infection. Tempol was found to prevent transcription and functional expression of procoagulant tissue factor in endothelial cells (ECs) stimulated by lipopolysaccharide (LPS). This effect was accompanied by inhibition of IL-6, IL-8, and monocyte chemoattractant protein (MCP-1) production. Tempol also attenuated platelet aggregation and human promyelocytic leukemia HL60 cells oxidative burst. In dendritic cells, Tempol inhibited LPS-induced production of TNF-α, IL-6, and IL-12p70, downregulated expression of co-stimulatory molecules, and prevented antigen-dependent lymphocyte proliferation. Notably, Tempol (20 mg/kg) partially increased the survival of mice with CM. Mechanistically, treated mice had lowered plasma levels of MCP-1, suggesting that Tempol downmodulates EC function and vascular inflammation. Tempol also diminished blood brain barrier permeability associated with CM when started at day 4 post infection but not at day 1, suggesting that ROS production is tightly regulated. Other antioxidants-such as α-phenyl N-tertiary-butyl nitrone (PBN; a spin trap), MnTe-2-PyP and MnTBAP (Mn-phorphyrin), Mitoquinone (MitoQ) and Mitotempo (mitochondrial antioxidants), M30 (an iron chelator), and epigallocatechin gallate (EGCG; polyphenol from green tea) did not improve survival. By contrast, these compounds (except PBN) inhibited Plasmodium falciparum growth in culture with different IC50s. Knockout mice for SOD1 or phagocyte nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (gp91(phox-/-)) or mice treated with inhibitors of SOD (diethyldithiocarbamate) or NADPH oxidase (diphenyleneiodonium) did not show protection or exacerbation for CM.

Conclusion: Results with Tempol suggest that intracellular ROS contribute, in part, to CM pathogenesis. Therapeutic targeting of intracellular ROS in CM is discussed.

MeSH terms

  • Animals
  • Antioxidants / pharmacology*
  • Antioxidants / therapeutic use
  • Cells, Cultured
  • Chemokine CCL2 / metabolism
  • Cyclic N-Oxides / pharmacology*
  • Cyclic N-Oxides / therapeutic use
  • Dendritic Cells / drug effects*
  • Dendritic Cells / metabolism*
  • Enzyme-Linked Immunosorbent Assay
  • Humans
  • Interleukin-6 / metabolism
  • Interleukin-8 / metabolism
  • Malaria, Cerebral / drug therapy*
  • Malaria, Cerebral / metabolism
  • Mice
  • Reactive Oxygen Species / metabolism
  • Real-Time Polymerase Chain Reaction
  • Spin Labels
  • Thromboplastin / metabolism*

Substances

  • Antioxidants
  • Chemokine CCL2
  • Cyclic N-Oxides
  • Interleukin-6
  • Interleukin-8
  • Reactive Oxygen Species
  • Spin Labels
  • Thromboplastin
  • tempol

Grant support

This work was supported by the Intramural Research Program of the Division of Intramural Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health. Dr. Anderson Sá-Nunes is recipient of grants from Brazilian Malaria Network (MCT/CNPq/MS/SCTIE/DECIT/PRONEX - 555648/2009-5) and from the Research Network on Bioactive Molecules from Arthropod Vectors (NAP-MOBIARVE, University of São Paulo). GIA Reference Center is supported by the PATH/Malaria Vaccine Initiative. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.