Severe malaria: what's new on the pathogenesis front?

Plasmodium falciparum causes the most severe and fatal form of malaria in humans with over half a million deaths each year. Cerebral malaria (CM), a complex neurological syndrome of severe falciparum malaria, is often fatal and represents a major public health burden. Despite vigorous efforts, the pathophysiology of CM remains to be elucidated, thereby hindering the development of adjunctive therapies. In recent years, multidisciplinary and collaborative approaches have led to groundbreaking progress both in the laboratory and in the field. Here we review the latest breakthroughs in severe malaria pathogenesis, with a specific focus on new pathogenetic mechanisms leading to CM. The most recent findings point towards specific parasite phenotypes targeting brain microvasculature, endothelial dysfunction and subsequent oedema-induced brain swelling

Inhibition of endothelial activation: A new way to treat cerebral malaria?

Background: Malaria is still a major public health problem, partly because the pathogenesis of its major complication, cerebral malaria (CM), remains incompletely understood. However tumor necrosis factor (TNF) is thought to play a key role in the development of this neurological syndrome, as well as lymphotoxin α (LT). Methods and Findings: Using an in vitro model of CM based on human brain-derived endothelial cells (HBEC-5i), we demonstrate the anti-inflammatory effect of LMP-420, a 2-NH2-6-Cl-9-[(5- dihydroxyboryl)-pentyl] purine that is a transcriptional inhibitor of TNF. When added before or concomitantly to TNF, LMP-420 inhibits endothelial cell (EC) activation, i.e., the up-regulation of both ICAM-1 and VCAM-1 on HBEC-5i surfaces. Subsequently, LMP-420 abolishes the cytoadherence of ICAM-1-specific Plasmodium falciparum-parasitized red blood cells on these EC. Identical but weaker effects are observed when LMP-420 is added with LT. LMP-420 also causes a dramatic reduction of HBEC-5i vesiculation induced by TNF or LT stimulation, as assessed by microparticle release. Conclusion: These data provide evidence for a strong in vitro anti-inflammatory effect of LMP-420 and suggest that targeting host cell pathogenic mechanisms might provide a new therapeutic approach to improving the outcome of CM patients. © 2005 Wassmer et al

Platelet-Induced Clumping of Plasmodium falciparum–Infected Erythrocytes from Malawian Patients with Cerebral Malaria—Possible Modulation In Vivo by Thrombocytopenia

Platelets may play a role in the pathogenesis of human cerebral malaria (CM), and they have been shown to induce clumping of Plasmodium falciparum–parasitized red blood cells (PRBCs) in vitro. Both thrombocytopenia and platelet-inducedPRBCclumping are associated with severe malaria and, especially, withCM.In the present study, we investigated the occurrence of the clumping phenomenon in patients with CM by isolating and coincubating their plasma and PRBCs ex vivo. Malawian children with CM all had low platelet counts, with the degree of thrombocytopenia directly proportional to the density of parasitemia. Plasma samples obtained from these patients subsequently induced weak PRBC clumping. When the assays were repeated, with the plasma platelet concentrations adjusted to within the physiological range considered to be normal, massive clumping occurred. The results of this study suggest that thrombocytopenia may, through reduction of platelet-mediated clumping of PRBCs, provide a protective mechanism for the host during CM

Experimental conditions affect the outcome of Plasmodium falciparum platelet-mediated clumping assays

<p>Abstract</p> <p>Background</p> <p>Platelet-mediated clumping of <it>Plasmodium falciparum</it>-infected erythrocytes (IE) is a parasite adhesion phenotype that has been associated with severe malaria in some, but not all, field isolate studies. A variety of experimental conditions have been used to study clumping <it>in vitro</it>, with substantial differences in parasitaemia (Pt), haematocrit (Ht), and time of reaction between studies. It is unknown whether these experimental variables affect the outcome of parasite clumping assays.</p> <p>Methods</p> <p>The effects of Pt (1, 4 and 12%), Ht (2, 5 and 10%) and time (15 min, 30 min, 1 h, 2 h) on the clumping of <it>P. falciparum </it>clone HB3 were examined. The effects of platelet freshness and parasite maturity were also studied.</p> <p>Results</p> <p>At low Ht (2%), the Pt of the culture has a large effect on clumping, with significantly higher clumping occurring at 12% Pt (mean 47% of IE in clumps) compared to 4% Pt (mean 26% IE in clumps) or 1% Pt (mean 7% IE in clumps) (ANOVA, p = 0.0004). Similarly, at low Pt (1%), the Ht of the culture has a large effect on clumping, with significantly higher clumping occurring at 10% Ht (mean 62% IE in clumps) compared to 5% Ht (mean 25% IE in clumps) or 2% Ht (mean 10% IE in clumps) (ANOVA, p = 0.0004). Combinations of high Ht and high Pt were impractical because of the difficulty assessing clumping in densely packed IE and the rapid formation of enormous clumps that could not be counted accurately. There was no significant difference in clumping when fresh platelets were used compared to platelets stored at 4°C for 10 days. Clumping was a property of mature pigmented-trophozoites and schizonts but not ring stage parasites.</p> <p>Conclusion</p> <p>The Pt and Ht at which <it>in vitro </it>clumping assays are set up have a profound effect on the outcome. All previous field isolate studies on clumping and malaria severity suffer from potential problems in experimental design and methodology. Future studies of clumping should use standardized conditions and control for Pt, and should take into account the limitations and variability inherent in the assay.</p

Plasmodium vivax in Hematopoietic Niches: Hidden and Dangerous

This is the author accepted manuscript. The final version is available from Elsevier via the DOI in this recordEstimation of Plasmodium vivax biomass based on circulating biomarkers indicates the existence of a predominant biomass outside of the circulation that is not captured by peripheral parasitemia, in particular in patients with complicated outcomes. A series of recent studies have suggested that the hematopoietic niche of the bone marrow (BM) is a major reservoir for parasite replication and the development of transmission stages. However, significant knowledge gaps remain in our understanding of host–parasite interactions, pathophysiology, and the implications for treatment and diagnosis of such a reservoir. Here, we discuss the current status of this emerging research field in the context of P. vivax.National Institute of Allergy and Infectious Diseases of the National Institutes of HealthMedical Research Council (MRC)Royal SocietySao Paulo Research Foundatio

Platelets Alter Gene Expression Profile in Human Brain Endothelial Cells in an In Vitro Model of Cerebral Malaria

Platelet adhesion to the brain microvasculature has been associated with cerebral malaria (CM) in humans, suggesting that platelets play a role in the pathogenesis of this syndrome. In vitro co-cultures have shown that platelets can act as a bridge between Plasmodium falciparum-infected red blood cells (pRBC) and human brain microvascular endothelial cells (HBEC) and potentiate HBEC apoptosis. Using cDNA microarray technology, we analyzed transcriptional changes of HBEC in response to platelets in the presence or the absence of tumor necrosis factor (TNF) and pRBC, which have been reported to alter gene expression in endothelial cells. Using a rigorous statistical approach with multiple test corrections, we showed a significant effect of platelets on gene expression in HBEC. We also detected a strong effect of TNF, whereas there was no transcriptional change induced specifically by pRBC. Nevertheless, a global ANOVA and a two-way ANOVA suggested that pRBC acted in interaction with platelets and TNF to alter gene expression in HBEC. The expression of selected genes was validated by RT-qPCR. The analysis of gene functional annotation indicated that platelets induce the expression of genes involved in inflammation and apoptosis, such as genes involved in chemokine-, TREM1-, cytokine-, IL10-, TGFβ-, death-receptor-, and apoptosis-signaling. Overall, our results support the hypothesis that platelets play a pathogenic role in CM

Competitive endothelial adhesion between Plasmodium falciparum isolates under physiological flow conditions

<p>Abstract</p> <p>Background</p> <p>Sequestration of parasitized red blood cells in the microvasculature of major organs involves a sequence of events that is believed to contribute to the pathogenesis of severe falciparum malaria. <it>Plasmodium falciparum </it>infections are commonly composed of multiple subpopulations of parasites with varied adhesive properties. A key question is: do these subpopulations compete for adhesion to endothelium? This study investigated whether, in a laboratory model of cytoadherence, there is competition in binding to endothelium between pRBC infected with <it>P. falciparum </it>of variant adhesive phenotypes, particularly under flow conditions.</p> <p>Methods</p> <p>Four different <it>P. falciparum </it>isolates, of known adherence phenotypes, were matched in pairs, mixed in different proportions and allowed to bind to cultured human endothelium. Using <it>in vitro </it>competitive static and flow-based adhesion assays, that allow simultaneous testing of the adhesive properties of two different parasite lines, adherence levels of paired <it>P. falciparum </it>isolates were quantified and analysed using either non-parametric Wilcoxon's paired signed rank test or Student paired test.</p> <p>Results</p> <p>Study findings show that <it>P. falciparum </it>parasite lines show marked differences in the efficiency of adhesion to endothelium.</p> <p>Conclusion</p> <p><it>Plasmodium falciparum </it>variants will compete for adhesion to endothelia and variants can be ranked by their efficiency of binding. These findings suggest that variants from a mixed infection will not show uniform cytoadherence and so may vary in their ability to cause disease.</p

Differentially expressed microRNAs in experimental cerebral malaria and their involvement in endocytosis, adherens junctions, FoxO and TGF-β signalling pathways

Cerebral malaria (CM) is the most severe manifestation of infection with Plasmodium, however its pathogenesis is still not completely understood. microRNA (miRNA) have been an area of focus in infectious disease research, due to their ability to affect normal biological processes, and have been shown to play roles in various viral, bacterial and parasitic infections, including malaria. The expression of miRNA was studied following infection of CBA mice with either Plasmodium berghei ANKA (causing CM), or Plasmodium yoelii (causing severe but non-cerebral malaria (NCM)). Using microarray analysis, miRNA expression was compared in the brains of non-infected (NI), NCM and CM mice. Six miRNA were significantly dysregulated between NCM and CM mice, and four of these, miR-19a-3p, miR-19b-3p, miR-142-3p and miR-223-3p, were further validated by qPCR assays. These miRNA are significantly involved in several pathways relevant to CM, including the TGF-β and endocytosis pathways. Dysregulation of these miRNA during CM specifically compared with NCM suggests that these miRNA, through their regulation of downstream targets, may be vitally involved in the neurological syndrome. Our data implies that, at least in the mouse model, miRNA may play a regulatory role in CM pathogenesis.This work was funded by the National Health and Medical Research Council (#1099920 for GEG). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.S

Infectious Diseases of the Nervous System and Their Impact in Developing Countries

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Cerebral malaria: insights from host-parasite protein-protein interactions

<p>Abstract</p> <p>Background</p> <p>Cerebral malaria is a form of human malaria wherein <it>Plasmodium falciparum</it>-infected red blood cells adhere to the blood capillaries in the brain, potentially leading to coma and death. Interactions between parasite and host proteins are important in understanding the pathogenesis of this deadly form of malaria. It is, therefore, necessary to study available protein-protein interactions to identify lesser known interactions that could throw light on key events of cerebral malaria.</p> <p>Methods</p> <p>Sequestration, haemostasis dysfunction, systemic inflammation and neuronal damage are key processes of cerebral malaria. Key events were identified from literature as being crucial to these processes. An integrated interactome was created using available experimental and predicted datasets as well as from literature. Interactions from this interactome were filtered based on Gene Ontology and tissue-specific annotations, and further analysed for relevance to the key events.</p> <p>Results</p> <p>PfEMP1 presentation, platelet activation and astrocyte dysfunction were identified as the key events influencing the disease. 48896 host-parasite along with other host-parasite, host-host and parasite-parasite protein-protein interactions obtained from a disease-specific corpus were combined to form an integrated interactome. Filtering of the interactome resulted in five host-parasite PPI, six parasite-parasite and two host-host PPI. The analysis of these interactions revealed the potential significance of apolipoproteins and temperature/Hsp expression on efficient PfEMP1 presentation; role of MSP-1 in platelet activation; effect of parasite proteins in TGF-β regulation and the role of albumin in astrocyte dysfunction.</p> <p>Conclusions</p> <p>This work links key host-parasite, parasite-parasite and host-host protein-protein interactions to key processes of cerebral malaria and generates hypotheses for disease pathogenesis based on a filtered interaction dataset. These hypotheses provide novel and significant insights to cerebral malaria.</p