Lipid moiety of emozoin promotes chemokines release and MMP-9 activation by human microvascular endothelial cells

Background Cerebral malaria (CM) is a fatal complication of P.falciparum infection. There is growing evidence that the combination of both parasite and host factors could be involved in the blood-brain barrier (BBB) breakdown. In the present work we studied the effects of hemozoin (HZ), the malaria lipid-enriched ferriprotoporphyrin IX pigment, on matrix metalloproteinases (MMPs) regulation and chemokines release by human microvascular endothelial cells (HMEC-1). Methods Cells were treated with native (n)HZ, delipidized (d)HZ or lipid-free synthetic (s)HZ. Cell morphology was evaluated by optical microscopy, chemokines (CXCL-8, RANTES) production by ELISA, total gelatinolytic activity by in situ gelatin zymography, MMP-2 and MMP-9 activity by direct gelatin zymography, proMMP-9, MMP-1 and MMP-3 protein expression by western blotting and microvessel-like structures formation by Matrigel Results Usually, cultured HMEC-1 show a polygonal morphology. The treatment with nHZ modified HMEC-1 in elongated cells with a reduced planar surface. Moreover, nHZ stimulated the production of the chemokines CXCL-8 and RANTES in a dose and time-dependent manner. Additionally, nHZ enhanced total gelatinolytic activity by inducing ex novo MMP-9 without affecting basal MMP-2 activity; nHZ promoted proMMP-9 protein expression, as well as MMP-1 and MMP-3, enzymes involved in the proMMP-9 proteolytic activation cascade. Since an increase in MMPs activity is required for the degradation of the basement membrane and endothelial cell migration, HMEC-1 ability to migrate and self-organize in microvessel-like structures was investigated. In the presence of nHZ, HMEC-1 migrated and got organized in microvessel-like structures more quickly than control cells. Interestingly, neither sHZ nor dHZ were able to induce the same effects observed with nHZ on HMEC-1. Conclusions Collectively, the present data suggest that the lipid moiety of HZ promotes MMP-9 activity and chemokines release by HMEC-1. As a consequence, cell invasiveness, tight junctions disruption and leukocytes recruitment might be promoted, concurring to BBB damage in CM. Acknowledgments The financial support of the FP6-IP18834 ANTIMAL, Regione Piemonte, Ricerca Sanitaria Finalizzata, Compagnia di San Paolo (Italian Malaria Network), Università di Milano (PUR 2009) is acknowledged