Further improvements of the P. falciparum humanized mouse model.

BACKGROUND: It has been shown previously that it is possible to obtain growth of Plasmodium falciparum in human erythrocytes grafted in mice lacking adaptive immune responses by controlling, to a certain extent, innate defences with liposomes containing clodronate (clo-lip). However, the reproducibility of those models is limited, with only a proportion of animals supporting longstanding parasitemia, due to strong inflammation induced by P. falciparum. Optimisation of the model is much needed for the study of new anti-malarial drugs, drug combinations, and candidate vaccines. MATERIALS/METHODS: We investigated the possibility of improving previous models by employing the intravenous route (IV) for delivery of both human erythrocytes (huRBC) and P. falciparum, instead of the intraperitoneal route (IP), by testing various immunosuppressive drugs that might help to control innate mouse defences, and by exploring the potential benefits of using immunodeficient mice with additional genetic defects, such as those with IL-2Rγ deficiency (NSG mice). RESULTS: We demonstrate here the role of aging, of inosine and of the IL-2 receptor γ mutation in controlling P. falciparum induced inflammation. IV delivery of huRBC and P. falciparum in clo-lip treated NSG mice led to successful infection in 100% of inoculated mice, rapid rise of parasitemia to high levels (up to 40%), long-lasting parasitemia, and consistent results from mouse-to-mouse. Characteristics were closer to human infection than in previous models, with evidence of synchronisation, partial sequestration, and receptivity to various P. falciparum strains without preliminary adaptation. However, results show that a major IL-12p70 inflammatory response remains prevalent. CONCLUSION: The combination of the NSG mouse, clodronate loaded liposomes, and IV delivery of huRBC has produced a reliable and more relevant model that better meets the needs of Malaria research.

The expression and function of cathepsin E in dendritic cells.

Cathepsin E is an aspartic proteinase that has been implicated in Ag processing within the class II MHC pathway. In this study, we document the presence of cathepsin E message and protein in human myeloid dendritic cells, the preeminent APCs of the immune system. Cathepsin E is found in a perinuclear compartment, which is likely to form part of the endoplasmic reticulum, and also a peripheral compartment just beneath the cell membrane, with a similar distribution to that of Texas Red-dextran within 2 min of endocytosis. To investigate the function of cathepsin E in processing, a new soluble targeted inhibitor was synthesized by linking the microbial aspartic proteinase inhibitor pepstatin to mannosylated BSA via a cleavable disulfide linker. This inhibitor was shown to block cathepsin D/E activity in cell-free assays and within dendritic cells. The inhibitor blocked the ability of dendritic cells from wild-type as well as cathepsin D-deficient mice to present intact OVA, but not an OVA-derived peptide, to cognate T cells. The data therefore support the hypothesis that cathepsin E has an important nonredundant role in the class II MHC Ag processing pathway within dendritic cells.

Extension, retraction and contraction in the formation of a dendritic cell dendrite: distinct roles for Rho GTPases.

The morphology of antigen-presenting dendritic cells (DC) is characterized by the possession of numerous long arborizing processes known as dendrites. The formation of these processes by DC, both in the periphery and in lymphoid organs, is believed to contribute to the remarkable efficiency with which they take up, process and present antigen to T cells. However, the process of dendrite formation and the signaling pathways that lead to the formation of these dendrites remain obscure. In this study we describe an in vitro model in which human immature DC form long processes similar to those formed in vivo. The formation of these processes involves initial attachment of a cell protrusion to the extracellular matrix substrate, and subsequent movement of the cell body away from the adhesion site, leaving behind a long slender dendrite. Dendrite formation, but not their maintenance, was found to be dependent on the activity of Rho GTPases. More specifically, Cdc42 and Rac1 were both required for the migration step of process formation, promoting cell spreading and extension. In contrast, Rho, and its downstream effector p160ROCK, regulated the release of adhesions to the substratum, and associated cellular contraction. Consequently, inhibition of Rho/p160ROCK leads to the formation of longer dendrites. DC therefore coordinate adhesion and protrusion to perform a specialized process of cellular morphogenesis, which differentiates these cells from all other cells of the immune system and may contribute to their distinctive function.