Protease activated receptor signaling is required for African trypanosome traversal of human brain microvascular endothelial cells
Using human brain microvascular endothelial cells (HBMECs) as an in vitro model for how African trypanosomes cross the human blood-brain barrier (BBB) we recently reported that the parasites cross the BBB by generating calcium activation signals in HBMECs through the activity of parasite cysteine pr...
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Published in | PLoS neglected tropical diseases Vol. 3; no. 7; p. e479 |
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Main Authors | , , , , , , , , , , , |
Format | Journal Article |
Language | English |
Published |
United States
Public Library of Science
21.07.2009
Public Library of Science (PLoS) |
Subjects | |
Online Access | Get full text |
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Summary: | Using human brain microvascular endothelial cells (HBMECs) as an in vitro model for how African trypanosomes cross the human blood-brain barrier (BBB) we recently reported that the parasites cross the BBB by generating calcium activation signals in HBMECs through the activity of parasite cysteine proteases, particularly cathepsin L (brucipain). In the current study, we examined the possible role of a class of protease stimulated HBMEC G protein coupled receptors (GPCRs) known as protease activated receptors (PARs) that might be implicated in calcium signaling by African trypanosomes.
Using RNA interference (RNAi) we found that in vitro PAR-2 gene (F2RL1) expression in HBMEC monolayers could be reduced by over 95%. We also found that the ability of Trypanosoma brucei rhodesiense to cross F2RL1-silenced HBMEC monolayers was reduced (39%-49%) and that HBMECs silenced for F2RL1 maintained control levels of barrier function in the presence of the parasite. Consistent with the role of PAR-2, we found that HBMEC barrier function was also maintained after blockade of Galpha(q) with Pasteurella multocida toxin (PMT). PAR-2 signaling has been shown in other systems to have neuroinflammatory and neuroprotective roles and our data implicate a role for proteases (i.e. brucipain) and PAR-2 in African trypanosome/HBMEC interactions. Using gene-profiling methods to interrogate candidate HBMEC pathways specifically triggered by brucipain, several pathways that potentially link some pathophysiologic processes associated with CNS HAT were identified.
Together, the data support a role, in part, for GPCRs as molecular targets for parasite proteases that lead to the activation of Galpha(q)-mediated calcium signaling. The consequence of these events is predicted to be increased permeability of the BBB to parasite transmigration and the initiation of neuroinflammation, events precursory to CNS disease. |
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Bibliography: | Conceived and designed the experiments: DJG JCGG YVK CAP KGB BAW APCdAL JS JSD. Performed the experiments: DJG JCGG OVN AB. Analyzed the data: DJG JCGG OVN YZ KGB JSD. Contributed reagents/materials/analysis tools: DJG BAW APCdAL. Wrote the paper: DJG JCGG JSD. Current address: Institute of Human Virology, School of Medicine University of Maryland, Baltimore, Maryland, United States of America. |
ISSN: | 1935-2735 1935-2727 1935-2735 |
DOI: | 10.1371/journal.pntd.0000479 |