Inhibition of Plasmodium falciparum Field Isolates-Mediated Endothelial Cell Apoptosis by Fasudil: Therapeutic Implications for Severe Malaria

Plasmodium falciparum infection can abruptly progress to severe malaria, a life-threatening complication resulting from sequestration of parasitized red blood cells (PRBC) in the microvasculature of various organs such as the brain and lungs. PRBC adhesion can induce endothelial cell (EC) activation and apoptosis, thereby disrupting the blood-brain barrier. Moreover, hemozoin, the malarial pigment, induces the erythroid precursor apoptosis. Despite the current efficiency of antimalarial drugs in killing parasites, severe malaria still causes up to one million deaths every year. A new strategy targeting both parasite elimination and EC protection is urgently needed in the field. Recently, a rho-kinase inhibitior Fasudil, a drug already in clinical use in humans for cardio- and neuro-vascular diseases, was successfully tested on laboratory strains of P. falciparum to protect and to reverse damages of the endothelium. We therefore assessed herein whether Fasudil would have a similar efficiency on P. falciparum taken directly from malaria patients using contact and non-contact experiments. Seven (23.3%) of 30 PRBC preparations from different patients were apoptogenic, four (13.3%) acting by cytoadherence and three (10%) via soluble factors. None of the apoptogenic PRBC preparations used both mechanisms indicating a possible mutual exclusion of signal transduction ligand. Three PRBC preparations (42.9%) induced EC apoptosis by cytoadherence after 4 h of coculture (“rapid transducers”), and four (57.1%) after a minimum of 24 h (“slow transducers”). The intensity of apoptosis increased with time. Interestingly, Fasudil inhibited EC apoptosis mediated both by cell-cell contact and by soluble factors but did not affect PRBC cytoadherence. Fasudil was found to be able to prevent endothelium apoptosis from all the P. falciparum isolates tested. Our data provide evidence of the strong anti-apoptogenic effect of Fasudil and show that endothelial cell-P. falciparum interactions are more complicated than previously thought. These findings may warrant clinical trials of Fasudil in severe malaria management

Hepatitis C Virus Infection May Lead to Slower Emergence of P. falciparum in Blood

International audienceBACKGROUND: Areas endemic for Plasmodium falciparum, hepatitis B virus (HBV) and hepatitis C virus (HCV) overlap in many parts of sub-Saharan Africa. HBV and HCV infections develop in the liver, where takes place the first development stage of P. falciparum before its further spread in blood. The complex mechanisms involved in the development of hepatitis may potentially influence the development of the liver stage of malaria parasites. Understanding the molecular mechanisms of these interactions could provide new pathophysiological insights for treatment strategies in Malaria. METHODOLOGY: We studied a cohort of 319 individuals living in a village where the three infections are prevalent. The patients were initially given a curative antimalarial treatment and were then monitored for the emergence of asexual P. falciparum forms in blood, fortnightly for one year, by microscopy and polymerase chain reaction. PRINCIPAL FINDINGS: At inclusion, 65 (20.4%) subjects had detectable malaria parasites in blood, 36 (11.3%) were HBV chronic carriers, and 61 (18.9%) were HCV chronic carriers. During follow-up, asexual P. falciparum forms were detected in the blood of 203 patients. The median time to P. falciparum emergence in blood was respectively 140 and 120 days in HBV- and HBV+ individuals, and 135 and 224 days in HCV- and HCV+ individuals. HCV carriage was associated with delayed emergence of asexual P. falciparum forms in blood relative to patients without HCV infection. CONCLUSIONS: This pilot study represents first tentative evidence of a potential epidemiological interaction between HBV, HCV and P. falciparum infections. Age is an important confounding factor in this setting however multivariate analysis points to an interaction between P. falciparum and HCV at the hepatic level with a slower emergence of P. falciparum in HCV chronic carriers. More in depth analysis are necessary to unravel the basis of hepatic interactions between these two pathogens, which could help in identifying new therapeutic approaches against malaria

Comparison of apoptosis in human primary pulmonary endothelial cells and a brain microvascular endothelial cell line co-cultured with Plasmodium falciparum field isolates

Abstract Background Plasmodium falciparum infection can progress unpredictably to severe forms including respiratory distress and cerebral malaria. The mechanisms underlying the variable natural course of malaria remain elusive. Methods The cerebral microvascular endothelial cells-D3 and lung endothelial cells both from human were cultured separately and challenged with P. falciparum field isolates taken directly from malaria patients or 3D7 strain (in vitro maintained culture). The capacity of these P. falciparum isolates to induce endothelial cell apoptosis via cytoadherence or not was then assessed. Results Overall, 27 P. falciparum isolates were collected from patients with uncomplicated malaria (n = 25) or severe malaria (n = 2). About half the isolates (n = 17) were able to bind brain endothelial cells (12 isolates, 44%) or lung endothelial cells (17 isolates, 63%) or both (12 isolates, 44%). Sixteen (59%) of the 27 isolates were apoptogenic for brain and/or lung endothelial cells. The apoptosis stimulus could be cytoadherence, direct cell-cell contact without cytoadherence, or diffusible soluble factors. While some of the apoptogenic isolates used two stimuli (direct contact with or without cytoadherence, plus soluble factors) to induce apoptosis, others used only one. Among the 16 apoptogenic isolates, eight specifically targeted brain endothelial cells, one lung endothelial cells, and seven both. Conclusion These results indicate that the brain microvascular cell line was more susceptible to apoptosis triggered by P. falciparum than the primary pulmonary endothelial cells and may have relevance to host-parasite interaction

<i>Plasmodium falciparum</i> parasitized red blood cells (PRBC)-mediated human lung endothelial cells (HLEC) apoptosis induction and inhibition by Fasudil: Seventeen PRBC from F747 to F625 were cocultured in contact conditions (cytoadherence).

<p>Para., parasitemia; Nb, number; schiz. Schizont;</p><p>*(8×10⁶); Cyto., cytoadherence; OD., optical density. In red colour OD values of apoptosis positive and inhibition by Fasudil.</p><p>**Hyperparasitemia (Severe malaria case).</p

<i>Plasmodium falciparum</i> parasitized red blood cells (PRBC)-mediated human lung endothelial cells (HLEC) apoptosis induction and inhibition by Fasudil: Ten PRBC from F528 to F625 were cocultured in non contact experiments.

<p>Pf, <i>Plasmodium falciparum</i>; Para., parasitemia; Nb, number; schiz. Schizont;</p><p>*(24×10⁶); Cyto., cytoadherence; OD, optical density. In red colour OD values of apoptosis positive and inhibition by Fasudil.</p><p>**Hyperparasitemia (Severe malaria case).</p

PRBC soluble factors-mediated HLEC apoptosis and inhibition by Fasudil.

<p>PRBC soluble factors-mediated HLEC apoptosis and inhibition by Fasudil.</p

Marked Rise in the Prevalence of Asymptomatic <i>Plasmodium falciparum</i> Infection in Rural Gabon

<div><p>Control strategies implemented a decade ago led to a marked reduction in the prevalence of malaria in many countries. In Dienga, southeastern Gabon, the prevalence of microscopic <i>P</i>. <i>falciparum</i> infection was 7% in 2003, close to the pre-elimination threshold of 5%. The aim of this work was to determine the prevalence of <i>P</i>. <i>falciparum</i> infection in the same community a decade later. A cohort of 370 individuals aged from 3 to 85 years living in Dienga was investigated for <i>P</i>. <i>falciparum</i> infection; during six passages (P) in 15-month period. Demographic data were collected, along with behaviors and attitudes towards malaria. <i>Plasmodium</i> infection was diagnosed by microscopy (ME), followed by PCR to detect submicroscopic infection. The prevalence of <i>P</i>. <i>falciparum</i> infection in P1, P2, P3, P4, P5 and P6 was respectively 43.5% (25.1% ME+, 18.4% PCR+); 40.9% (27.0% ME+, 13.9% PCR+), 52.7% (26.1% ME+, 26.6% PCR+); 34.1% (14.1% ME+, 20% PCR+), 57.7% (25.4.% ME+, 32.3% PCR+); and 46.2% (21.4% ME+, 24.8% PCR+) with an overall average of 45.9% (95%CI [37.0–54.7], 23.2% ME+ and 22.7% PCR+). P4 and P5 prevalences were statically different throughout the six passages. Microscopic prevalence was significantly higher than that observed ten years ago (23% [n = 370] vs 7% [n = 323], p < 0.001). Asymptomatic infections were the most frequent (96%). Gametocytes were detected in levels ranging from 5.9% to 13.9%. Insecticide-treated nets, indoor residual insecticides, and self-medication were used by respectively 33.2% (95%CI [29.0–37.4]), 17.7% (95%CI [15.5–19.9]) and 12.1% (95%CI [10.6–13.6]) of the study population. A near-threefold increase in <i>P</i>. <i>falciparum</i> infection has been observed in a rural area of southeastern Gabon during a 10-year period. Most infections were asymptomatic, but these subjects likely represent a parasite reservoir. These findings call for urgent reinforcement of preventive measures.</p></div

Attitudes and behaviors concerning malaria.

<p>Attitudes and behaviors concerning malaria.</p