Malaria is a major cause of mortality worldwide. The majority of deaths are due to infections with Plasmodium falciparum (P. falciparum). Cerebral malaria (CM) is a major cause of death in these patients. Despite its virulence, the pathophysiologic basis of CM is poorly understood. Recent investigations suggest that microvascular damage is a factor with associated dysfunction/ dysruption of the blood brain barrier. Endothelial cell damage is caused by several factors including: the sequestration of infected red blood cells, hypoxic damage and "cytokine storm". Repair of damaged microvasculature may occur either by the proliferation of local endothelial cells or the recruitment of bone-marrow derived circulating endothelial progenitor cells (cEPCs). Low levels and dysfunction of cEPCs have been associated with the development of symptoms and adverse events in diseases associated with microvascular damage including cardiovascular disease, diabetes and stroke. Circulating EPC ontogeny and migration from the bone marrow is mediated by chemokines/proteases such as the stromal derived growth factor 1 (SDF-1) and the matrix metalloprotein-9 (MMP-9). We hypothesize that P. falciparum infection results in an imbalance between microvascular damage and repair. Cerebral malaria occurs when cEPCs are diminished and damaged endothelial cells cannot be replaced. A R21 supported study supports this hypothesis. The mean percentage of cEPCs are significantly lower in Ghanaian children with CM as compared to those with uncomplicated malaria (UM), asymptomatic parasitemia (AP) or healthy controls (HC) (p<0.0001). In addition, mean plasma levels of SDF-1 are significantly increased in children with CM and UM as compared to those with AP or HC (p<0.001).
The Specific Aims of this proposal will further define the host response to microvascular damage in malaria and determine its association with the development of or recovery from CM. P. falciparum-infected children in Ghana with different degrees of disease severity (CM, UM and AP) will be compared with healthy controls.
Aim 1) To determine the time course of the host response to microvascular damage. Levels of cEPCs, SDF-1 and MMP-9 will be measured prospectively in the same patient. We predict that patients with UM who develop CM should have decreasing cEPC levels leading to the development of CM. Increases in cEPCs should be detected in patients who recover from CM. SDF-1 and MMP-9 levels should be elevated in patients who develop CM and decline with recovery.
Aim 2) To assess the function of circulating EPCs in malaria infections. Transwell migration assays and blood outgrowth endothelial cell culture coupled with DiLDL metabolic labeling will be performed. We predict that the cEPCs of patients with CM will have functional abnormalities.
Aim 3) To assess qRT-PCR assays for the determination of cEPC levels. Quantitative RT-PCR is potentially a facile tool for use in endemic areas for determining cEPC levels as well as assessing function. The utility of qRT-PCR will be assessed in comparison to flow cytometry analysis and functional assays.
Cerebral malaria is a major killer of young children worldwide for which there are limited treatment options. This proposal will establish a novel paradigm for the pathogenesis of the disease: that microvascular repair is an important component of recovery. The proposed studies therefore may provide evidence for using novel classes of chemotherapeutics, which are being actively developed by industry for the treatment of diseases such as coronary artery disease and stroke, for the treatment of cerebral malaria.