Severe malaria is associated with sequestration of Plasmodium falciparum-infected red blood cells (PRBC) in the microvasculature and elevation of intercellular adhesion molecule-1 (ICAM-1) and TNF. In vitro co-culture of human umbilical vein endothelial cells (HUVEC), with either PRBC or uninfected RBC, required the presence of low level TNF (5pg/ml) for significant up-regulation of ICAM-1, which may contribute to increased cytoadhesion in vivo. These effects were independent of P. falciparum erythrocyte membrane protein-1 (PfEMP-1)-mediated adhesion but critically dependent on cell-cell contact. Further changes included increases in IL8 release and soluble TNF receptor shedding. Microarray analysis of HUVEC transcriptome following co-culture, using a human Affymetrix microarray chip, showed significant differential regulation of genes which defined gene ontologies such as cell communication, cell adhesion, signal transduction and immune response. Our data demonstrate that endothelial cells have the ability to mobilise immune and pro-adhesive responses when exposed to both PRBC and TNF. In addition, there is also a previously un-described positive regulation by RBC and TNF and a concurrent negative regulation of a range of genes involved in inflammation and cell-death, by PRBC and TNF. We propose that the balance between positive and negative regulation demonstrated in our study will determine endothelial pathology during a malaria infection.