Real-time imaging reveals the dynamics of leukocyte behaviour during experimental cerebral malaria pathogenesis

Saparna Pai; Jim Qin; Lois Cavanagh; Andrew Mitchell; Fatima El-Assaad; Rohit Jain; Valery Combes; Nicholas H Hunt; Georges E R Grau; Wolfgang Weninger

doi:10.1371/journal.ppat.1004236

Real-time imaging reveals the dynamics of leukocyte behaviour during experimental cerebral malaria pathogenesis

PLoS Pathog. 2014 Jul 17;10(7):e1004236. doi: 10.1371/journal.ppat.1004236. eCollection 2014 Jul.

Authors

Affiliations

¹ Immune Imaging Laboratory, The Centenary Institute, Newtown, Sydney, New South Wales, Australia; Sydney Medical School, University of Sydney, Sydney, New South Wales, Australia.
² Immune Imaging Laboratory, The Centenary Institute, Newtown, Sydney, New South Wales, Australia.
³ Vascular Immunology Unit, Discipline of Pathology, Sydney Medical School, University of Sydney, Camperdown, Sydney, New South Wales, Australia.
⁴ Sydney Medical School, University of Sydney, Sydney, New South Wales, Australia; Vascular Immunology Unit, Discipline of Pathology, Sydney Medical School, University of Sydney, Camperdown, Sydney, New South Wales, Australia.
⁵ Sydney Medical School, University of Sydney, Sydney, New South Wales, Australia; Molecular Immunopathology Unit, Discipline of Pathology, Sydney Medical School and Bosch Institute, University of Sydney, Camperdown, Sydney, New South Wales, Australia.
⁶ Immune Imaging Laboratory, The Centenary Institute, Newtown, Sydney, New South Wales, Australia; Discipline of Dermatology, University of Sydney, Sydney, New South Wales, Australia; Department of Dermatology, Royal Prince Alfred Hospital, Camperdown, Sydney, New South Wales, Australia.

Abstract

During experimental cerebral malaria (ECM) mice develop a lethal neuropathological syndrome associated with microcirculatory dysfunction and intravascular leukocyte sequestration. The precise spatio-temporal context in which the intravascular immune response unfolds is incompletely understood. We developed a 2-photon intravital microscopy (2P-IVM)-based brain-imaging model to monitor the real-time behaviour of leukocytes directly within the brain vasculature during ECM. Ly6C(hi) monocytes, but not neutrophils, started to accumulate in the blood vessels of Plasmodium berghei ANKA (PbA)-infected MacGreen mice, in which myeloid cells express GFP, one to two days prior to the onset of the neurological signs (NS). A decrease in the rolling speed of monocytes, a measure of endothelial cell activation, was associated with progressive worsening of clinical symptoms. Adoptive transfer experiments with defined immune cell subsets in recombinase activating gene (RAG)-1-deficient mice showed that these changes were mediated by Plasmodium-specific CD8(+) T lymphocytes. A critical number of CD8(+) T effectors was required to induce disease and monocyte adherence to the vasculature. Depletion of monocytes at the onset of disease symptoms resulted in decreased lymphocyte accumulation, suggesting reciprocal effects of monocytes and T cells on their recruitment within the brain. Together, our studies define the real-time kinetics of leukocyte behaviour in the central nervous system during ECM, and reveal a significant role for Plasmodium-specific CD8(+) T lymphocytes in regulating vascular pathology in this disease.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
CD8-Positive T-Lymphocytes* / metabolism
CD8-Positive T-Lymphocytes* / pathology
Endothelial Cells* / metabolism
Endothelial Cells* / pathology
Malaria, Cerebral* / metabolism
Malaria, Cerebral* / pathology
Malaria, Cerebral* / physiopathology
Mice
Mice, Knockout
Microscopy, Fluorescence
Monocytes* / metabolism
Monocytes* / pathology
Plasmodium berghei / metabolism*

Grants and funding

This study was supported by the Dermatology Research Foundation and Sydney Medical School, The University of Sydney. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.