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Rosetting Plasmodium falciparum-infected erythrocytes bind to human brain microvascular endothelial cells in vitro, demonstrating a dual adhesion phenotype mediated by distinct P. falciparum erythrocyte membrane protein 1 domains.

Adams Y, Kuhnrae P, Higgins MK, Ghumra A, Rowe JA - Infect. Immun. (2013)

Bottom Line: Antibodies to the N-terminal regions of the IT4var09 PfEMP1 variant (NTS-DBL1α and DBL2γ domains) specifically inhibited and reversed cytoadherence down to low concentrations (<10 μg/ml of total IgG).Surface plasmon resonance experiments showed that the NTS-DBLα and DBL2γ domains bind strongly to heparin, with half-maximal binding at a concentration of ∼0.5 μM in both cases.Therefore, cytoadherence of IT/R29 IE is distinct from rosetting, which is primarily mediated by NTS-DBL1α interactions with complement receptor 1.

View Article: PubMed Central - PubMed

Affiliation: Centre for Immunity, Infection and Evolution, Institute of Infection and Immunity, School of Biological Sciences, University of Edinburgh, Edinburgh, United Kingdom.

ABSTRACT
Adhesion interactions between Plasmodium falciparum-infected erythrocytes (IE) and human cells underlie the pathology of severe malaria. IE cytoadhere to microvascular endothelium or form rosettes with uninfected erythrocytes to survive in vivo by sequestering IE in the microvasculature and avoiding splenic clearance mechanisms. Both rosetting and cytoadherence are mediated by the parasite-derived IE surface protein family Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1). Rosetting and cytoadherence have been widely studied as separate entities; however, the ability of rosetting P. falciparum strains to cytoadhere has received little attention. Here, we show that IE of the IT/R29 strain expressing a rosette-mediating PfEMP1 variant (IT4var09) cytoadhere in vitro to a human brain microvascular endothelial cell line (HBEC-5i). Cytoadherence was inhibited by heparin and by treatment of HBEC-5i with heparinase III, suggesting that the endothelial receptors for IE binding are heparan sulfate proteoglycans. Antibodies to the N-terminal regions of the IT4var09 PfEMP1 variant (NTS-DBL1α and DBL2γ domains) specifically inhibited and reversed cytoadherence down to low concentrations (<10 μg/ml of total IgG). Surface plasmon resonance experiments showed that the NTS-DBLα and DBL2γ domains bind strongly to heparin, with half-maximal binding at a concentration of ∼0.5 μM in both cases. Therefore, cytoadherence of IT/R29 IE is distinct from rosetting, which is primarily mediated by NTS-DBL1α interactions with complement receptor 1. These data show that IT4var09-expressing parasites are capable of dual interactions with both endothelial cells and uninfected erythrocytes via distinct receptor-ligand interactions.

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Adhesion of IT/R29 IE to HBEC-5i is not CR1 dependent. Mouse monoclonal antibodies to CR1 were tested for their ability to inhibit adhesion of IT/R29 IE to HBEC-5i, and no significant inhibition was seen. Data shown are the means and standard errors from two independent experiments.
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Figure 6: Adhesion of IT/R29 IE to HBEC-5i is not CR1 dependent. Mouse monoclonal antibodies to CR1 were tested for their ability to inhibit adhesion of IT/R29 IE to HBEC-5i, and no significant inhibition was seen. Data shown are the means and standard errors from two independent experiments.

Mentions: Rosetting of IT/R29 parasites is dependent on CR1 on uninfected erythrocytes (15, 19). To explore whether CR1 also could play a role in endothelial cytoadherence, we tested the ability of a rosette-disrupting CR1 MAb, J3Bll, to inhibit adhesion of IT/R29 IE to HBEC-5i. J3B11, the control MAb J3D3 (a non-rosette-disrupting CR1 MAb), and an isotype control failed to significantly inhibit adhesion of IT/R29 IE to HBEC-5i (Fig. 6), indicating that CR1 does not play a role in IT/R29 cytoadherence. These results are consistent with previous data showing that CR1 could not be detected on HBEC-5i or HPMEC (34).


Rosetting Plasmodium falciparum-infected erythrocytes bind to human brain microvascular endothelial cells in vitro, demonstrating a dual adhesion phenotype mediated by distinct P. falciparum erythrocyte membrane protein 1 domains.

Adams Y, Kuhnrae P, Higgins MK, Ghumra A, Rowe JA - Infect. Immun. (2013)

Adhesion of IT/R29 IE to HBEC-5i is not CR1 dependent. Mouse monoclonal antibodies to CR1 were tested for their ability to inhibit adhesion of IT/R29 IE to HBEC-5i, and no significant inhibition was seen. Data shown are the means and standard errors from two independent experiments.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3958005&req=5

Figure 6: Adhesion of IT/R29 IE to HBEC-5i is not CR1 dependent. Mouse monoclonal antibodies to CR1 were tested for their ability to inhibit adhesion of IT/R29 IE to HBEC-5i, and no significant inhibition was seen. Data shown are the means and standard errors from two independent experiments.
Mentions: Rosetting of IT/R29 parasites is dependent on CR1 on uninfected erythrocytes (15, 19). To explore whether CR1 also could play a role in endothelial cytoadherence, we tested the ability of a rosette-disrupting CR1 MAb, J3Bll, to inhibit adhesion of IT/R29 IE to HBEC-5i. J3B11, the control MAb J3D3 (a non-rosette-disrupting CR1 MAb), and an isotype control failed to significantly inhibit adhesion of IT/R29 IE to HBEC-5i (Fig. 6), indicating that CR1 does not play a role in IT/R29 cytoadherence. These results are consistent with previous data showing that CR1 could not be detected on HBEC-5i or HPMEC (34).

Bottom Line: Antibodies to the N-terminal regions of the IT4var09 PfEMP1 variant (NTS-DBL1α and DBL2γ domains) specifically inhibited and reversed cytoadherence down to low concentrations (<10 μg/ml of total IgG).Surface plasmon resonance experiments showed that the NTS-DBLα and DBL2γ domains bind strongly to heparin, with half-maximal binding at a concentration of ∼0.5 μM in both cases.Therefore, cytoadherence of IT/R29 IE is distinct from rosetting, which is primarily mediated by NTS-DBL1α interactions with complement receptor 1.

View Article: PubMed Central - PubMed

Affiliation: Centre for Immunity, Infection and Evolution, Institute of Infection and Immunity, School of Biological Sciences, University of Edinburgh, Edinburgh, United Kingdom.

ABSTRACT
Adhesion interactions between Plasmodium falciparum-infected erythrocytes (IE) and human cells underlie the pathology of severe malaria. IE cytoadhere to microvascular endothelium or form rosettes with uninfected erythrocytes to survive in vivo by sequestering IE in the microvasculature and avoiding splenic clearance mechanisms. Both rosetting and cytoadherence are mediated by the parasite-derived IE surface protein family Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1). Rosetting and cytoadherence have been widely studied as separate entities; however, the ability of rosetting P. falciparum strains to cytoadhere has received little attention. Here, we show that IE of the IT/R29 strain expressing a rosette-mediating PfEMP1 variant (IT4var09) cytoadhere in vitro to a human brain microvascular endothelial cell line (HBEC-5i). Cytoadherence was inhibited by heparin and by treatment of HBEC-5i with heparinase III, suggesting that the endothelial receptors for IE binding are heparan sulfate proteoglycans. Antibodies to the N-terminal regions of the IT4var09 PfEMP1 variant (NTS-DBL1α and DBL2γ domains) specifically inhibited and reversed cytoadherence down to low concentrations (<10 μg/ml of total IgG). Surface plasmon resonance experiments showed that the NTS-DBLα and DBL2γ domains bind strongly to heparin, with half-maximal binding at a concentration of ∼0.5 μM in both cases. Therefore, cytoadherence of IT/R29 IE is distinct from rosetting, which is primarily mediated by NTS-DBL1α interactions with complement receptor 1. These data show that IT4var09-expressing parasites are capable of dual interactions with both endothelial cells and uninfected erythrocytes via distinct receptor-ligand interactions.

Show MeSH
Related in: MedlinePlus