Infection of Endothelial Cells with Trypanosoma cruzi Activates NF-κB and Induces Vascular Adhesion Molecule Expression

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Abstract
Transcriptional activation of vascular adhesion molecule expression, a major component of an inflammatory response, is regulated, in part, by the nuclear factor-κB/Rel (NF-κB) family of transcription factors. We therefore determined whether Trypanosoma cruzi infection of endothelial cells resulted in the activation of NF-κB and the induction or increased expression of adhesion molecules. Human umbilical vein endothelial cells (HUVEC) were infected with trypomastigotes of the Tulahuen strain of T. cruzi . Electrophoretic mobility shift assays with an NF-κB-specific oligonucleotide and nuclear extracts from T. cruzi -infected HUVEC (6 to 48 h postinfection) detected two major shifted complexes. Pretreatment with 50× cold NF-κB consensus sequence abolished both gel-shifted complexes while excess SP-1 consensus sequence had no effect. These data indicate that nuclear extracts from T. cruzi -infected HUVEC specifically bound to the NF-κB consensus DNA sequence. Supershift analysis revealed that the gel-shifted complexes were comprised of p65 (RelA) and p50 (NF-κB1). Northern blot analyses demonstrated both the induction of vascular cell adhesion molecule 1 and E-selectin and the upregulation of intercellular adhesion molecule 1 mRNA in HUVEC infected with T. cruzi . Immunocytochemical staining confirmed adhesion molecule expression in response to T. cruzi infection. These findings are consistent with the hypothesis that the activation of the NF-κB pathway in endothelial cells associated with T. cruzi infection may be an important factor in the inflammatory response and subsequent vascular injury and endothelial dysfunction that lead to chronic cardiomyopathy.