Plasmodium falciparum and TNF-α Differentially Regulate Inflammatory and Barrier Integrity Pathways in Human Brain Endothelial Cells

Abstract

Cerebral malaria is a severe complication of Plasmodium falciparum infection that causes the loss of blood-brain barrier integrity and frequently results in death. Here, we compared the effect of P. falciparum -infected red blood cells and inflammatory cytokines, like TNF-α, in the loss of BBB integrity.

Transcriptional responses of brain endothelium toPlasmodium falciparumpatient-derived isolatesin vitro

“…Nonetheless, this model allowed for the study of the HBMEC transcriptional response to patient-derived parasites and to assess if these responses were dependent on IE cytoadhesion or clinical syndrome. The ratio of 110 IE/HBMEC in our experiments is similar to the range employed in other studies 23,55 , which was calculated in Zuniga et al to be equivalent to approximately two layers of IE on top of the HBMEC. We tested whether the Fluidigm panel would reflect endothelial gene expression changes by exposing HBMEC overnight to IL-1beta and TNF and comparing the gene expression with a media control (Supplementary Table 2).…”

“…In this model, HBMEC were stimulated with TNF overnight before the cytokine stimuli was removed and the IE introduced for 6-hours co-culture. The ratio of 110 IE/HBMEC in our experiments is similar to the range employed in other studies 23,51 , which was calculated in Zuniga et al to be equivalent to approximately two layers of IE on top of the HBMEC. We validated whether the Fluidigm panel would reflect gene expression changes in the co-culture model by exposing HBMEC overnight to IL-1beta and TNF and comparing the gene expression with a media control (Table 1).…”

“…The multiplex was performed at an early stage of the study and was not performed with all the isolates or SBP1-KO, and only performed with one experiment, thus, the results are only an indication of the effects of IE co-culture. Stimulation with TNF, as positive control, did increase the concentrations of multiple cytokines, including IL-6, IL-8, IP-10 and MCP-1, also reported in other studies 23,51,61 . Indeed, Zuniga et al 23 noted they observed transcriptionally similar but translationally different levels of cytokines between their treatment groups even when total protein synthesis levels were similar, suggesting a role for post-translational regulation in HBMEC after incubation with IE.…”

“…It has been reported that IE affect the endothelial responses to thrombin 23,51 , which reduces barrier function rapidly and recovers within 2 hours. The presence of RBC or IE affected the response of HBMEC to thrombin with a reduced maximum response and AUC, compared to medium (Supplementary Figures 3E and F).…”

“…Although IE derived from malaria patients have exhibited different binding phenotypes, most studies employ well characterized PfEMP1 laboratory strains to investigate cytoadherence and interaction of different PfEMP1 variants to brain endothelial cells 19,[21][22][23][24] . Separate investigations have highlighted the divergent effect of both TNF stimulation and IE cytoadhesion on endothelial transcriptional responses and the pathways they induce 23,25 . IE isolated from peripheral blood of UM and CM patients have been used to study cytoadherence to human brain microvascular endothelial cells (HBMEC).…”