Malaria starts with Plasmodium sporozoites infection of the host's liver, where development into blood stage parasites occurs. It is not clear why natural infections do not induce protection against the initial liver stage and generate low CD8+ T cell responses. Using a rodent malaria model, we show that Plasmodium blood stage infection suppresses CD8+ T cell immune responses that were induced against the initial liver stage. Blood stage Plasmodium affects dendritic cell (DC) functions, inhibiting maturation and the capacity to initiate immune responses and inverting the interleukin (IL)-12/IL-10 secretion pattern. The interaction of blood stage parasites with DCs induces the secretion of soluble factors that inhibit the activation of CD8+ T cells in vitro and the suppression of protective CD8+ T cell responses against the liver stage in vivo. We propose that blood stage infection induces DCs to suppress CD8+ T cell responses in natural malaria infections. This evasion mechanism leaves the host unprotected against reinfection by inhibiting the immune response against the initial liver stage of the disease.

IntroductionDendritic cells (DCs) are antigen-presenting cells (APCs) that play a central role in both innate and adaptive immune responses. Immature DCs take up and process antigens in the periphery. After detecting microbial products such as LPS, peptidoglycans, bacterial DNA or viral RNA, or proinflammatory cytokines, DCs mature and migrate to lymphoid organs to initiate immune responses by priming naive T cells. 1 Maturation of DCs augments their antigenpresentation capacity, since peptide loading, half-life, and delivery of MHC molecules to the cell surface are increased, as is surface expression of T-cell costimulatory molecules. 2 Peptides derived from exogenous antigens are internalized by endocytosis, processed, and presented on MHCII molecules. 3 Loading of exogenous peptides onto the MHCII occurs in the endocytic pathway in MHC class II-containing vesicular compartments. 4 Transport of peptide-loaded MHCII molecules to the plasma membrane is regulated by mechanisms controlling the traffic of endosomal compartments, for example, by the Rho subfamily of small GTPases through their regulation of the F-actin cytoskeleton. 5,6 In activated DCs, surface localization of peptide-loaded MHCII molecules is achieved by direct fusion of MHCII-containing vesicles with the plasma membrane, supported by tubular structures extending from these compartments to the cell membrane. [7][8][9] The mechanisms controlling the movement of MHCII-containing compartments to the plasma membrane are not entirely clear, and probably require Rho GTPases, 10,11 actin-based motor proteins, 12 and actin cytoskeleton rearrangement. 13 In the plasma membrane of activated DCs, MHCII is localized in glycolipid-enriched membrane microdomains, commonly referred to as lipid rafts. This localization is important in intracellular signaling and T-cell activation. 14,15 Regulatory mechanisms of the pathways to properly localize functional MHCII within the cell membrane remain to be fully understood.Murine SWAP-70 was originally isolated from activated, mature B lymphocytes, 16 and is also strongly expressed in mast cells. 17,18 SWAP-70 is expressed in DCs obtained from bone marrow precursors (BMDCs) and has been shown to localize to DC membranes at sites of cell-cell contact and of micropinosomes. 19 A pleckstrin homology (PH) domain exists in the center region of SWAP-70, binds PIP3, the second messenger product generated by PI3K, and likely mediates membrane localization of the protein.Binding of PIP3 is necessary for SWAP-70 to localize to membrane F-actin structures called membrane ruffles, 20 sites of membrane F-actin remodeling. SWAP-70 directly binds F-actin, but not G-actin. 21 The Rho GTPase family (Cdc42, Rac, and Rho) plays important roles in regulating DC functions, 22,23 for example, macropinocytosis/endocytosis, 24,25 effective migration, 23 efficient T-cell priming, 26 and up-regulation of surface MHCII molecules. 10,11 Molecularly, SWAP-70 loosely resembles proteins of the Dbl-family of guanine nucleotide exchange factors for smal...

The phospholipid mediator sphingosine 1-phosphate (S1P) enhances motility and endocytosis of mature dendritic cells (DCs). We show that in vitro migration of Swap-70−/− bone marrow-derived DCs (BMDCs) in response to S1P and S1P-induced upregulation of endocytosis are significantly reduced. S1P-stimulated movement of Swap-70−/− BMDCs, specifically retraction of their trailing edge, in a collagen three-dimensional environment is impaired. These in vitro observations correlate with delayed entry into lymphatic vessels and migration to lymph nodes of skin DCs in Swap-70−/− mice. Expression of S1P receptors (S1P1–3) by wild-type and Swap-70−/− BMDCs is similar, but Swap-70−/− BMDCs fail to activate RhoA and to localize Rac1 and RhoA into areas of actin polymerization after S1P stimulus. The Rho-activating G protein Gαi interacts with SWAP-70, which also supports the localization of Gα13 to membrane rafts in BMDCs. LPS-matured Swap-70−/− BMDCs contain significantly more active RhoA than wild-type DCs. Preinhibition of Rho activation restored migration to S1P, S1P-induced upregulation of endocytosis in mature Swap-70−/− BMDCs, and localization of Gα13 to membrane rafts. These data demonstrate SWAP-70 as a novel regulator of S1P signaling necessary for DC motility and endocytosis.

During an acute blood-stage malaria infection, T cell responses to malaria and other bystander antigens are inhibited. Plasmodium infection induces strong cytokine responses that facilitate parasite clearance but may interfere with T cell functions, as some of the soluble immune mediators induced are also general inhibitors of T cell responses. Using a malaria mouse model, we have analyzed the cytokines produced by dendritic cells in response to P. yoelii infection that have potential T cell inhibitory activity. We found that during acute infection DC migrate to the spleen and secrete TGF-b, prostaglandin E 2 (PGE 2 ) and IL-10. We have analyzed the role of these general T cell inhibitors in a particular T cell response of evident importance in malaria infections: the CD8 + T cells generated against the liver-stage of the disease. During blood-stage infection, inhibition of the activity of TGF-b and PGE 2 restores the CD8 + T cell responses generated by sporozoites, increasing protection against re-infection. Our findings suggest that the strong cytokine response induced by blood-stage P. yoelii infection affects host T cell responses, inhibiting protective CD8 + T cells against the liver-stage of the disease.