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In inflammatory gastrointestinal disorders, healing of the damaged intestinal epithelial barrier involves a complex process of epithelial cell migration known as restitution. Babbin et al. (p.8112 ) observed that formylpeptide receptor-1 (FPR) was expressed along the lateral membrane of crypt epithelial cells in normal human colon and therefore investigated whether this G protein-coupled receptor could be involved in intestinal epithelial cell restitution. Using an in vitro model of this process, the authors found that treatment with the FPR ligand fMLF significantly accelerated wound closure, and this healing could be blocked by an FPR antagonist. FPR was found to be expressed along actin filaments in lamellipodial and filopodial extrusions in migrating cells, further supporting its direct involvement in epithelial cell migration. Investigation of the signaling mechanisms responsible for FPR action indicated that PI3K and the downstream Rho GTPases Rac1 and Cdc42 were required for FPR-mediated epithelial migration. All three signaling molecules were up-regulated upon fMLF treatment, and selective inhibitors of any of them eliminated the enhanced wound closure mediated by FPR. These data could lead to novel therapeutic strategies for the treatment of inflammatory colonic damage.

It has been suggested that intracellular pathogens protect themselves by interfering with host cell apoptosis. To assess how this might occur, Persson et al. (p.8357 ) analyzed the effects of death receptor ligation on Toxoplasma gondii-infected cells. Surprisingly, T. gondii-infected cells were not protected from cell death but instead rapidly underwent necrosis after Fas ligation. Necrotic cell death was accompanied by rapid egress of infectious parasites from the dying cells. Systematic analysis determined that early in the apoptotic pathway, caspase-induced calcium mobilization induced this parasite egress, which was an active process that caused host cell necrosis. Further studies indicated that activated primary T cells could induce T. gondii egress via either a Fas-mediated, caspase-dependent pathway or a perforin-mediated, caspase-independent pathway and that both pathways led to the release of parasites able to infect bystander cells or the effector cells themselves. These data were supported by in vivo experiments demonstrating that activated Ag-specific CD8⁺ T cells were infected by T. gondii after interacting with infected cells. This study presents the intriguing possibility that T cell effector mechanisms intended to eliminate intracellular pathogens may instead contribute to their spread.

Scurfy (Sf) mice, which bear a mutation in Foxp3 and thus lack regulatory T cells (Tregs), develop a severe multiorgan autoimmune syndrome and die before weaning. Intriguingly, mice lacking IL-2, which is thought to be important for Treg generation, survival, and function, have a significantly longer lifespan than Sf mice. To determine the roles IL-2 and Fas might play in the fatal autoimmunity seen in Sf mice, Zheng et al. (p.8035 ) developed doubly deficient mice lacking Foxp3 and either IL-2 or Fas. Current dogma would predict that these mice would demonstrate more severe inflammation than mice lacking Foxp3 alone; however, both sets of double-knockout mice lived significantly longer than Sf mice. Surprisingly, lymphoproliferation and FasL-mediated cytotoxicity were enhanced, not reduced, in Sf.Il2^−/− mice, indicating that a reduction in activation-induced cell death did not enhance survival of these mice. Despite their high level of lymphoproliferation, the Sf.Il2^−/− mice displayed greatly reduced autoimmune disease and very little inflammation in the skin and lung. In contrast, Sf.Fas^−/− mice demonstrated similar autoimmune inflammation to Sf mice yet still lived longer than these mice, suggesting that Fas and IL-2 employ distinct mechanisms for inflammatory regulation. These data indicate that IL-2 may regulate organ-specific autoimmune responses through mechanisms not involving Treg.

The non-classical MHC class I molecule HLA-G has been proposed to play an important role in maternal-fetal tolerance, but an appropriate animal model has not been available to explore this possibility. To address this problem, Bondarenko et al. (p.8042 ) analyzed the effect of interfering with Mamu-AG, a putative rhesus monkey homologue of HLA-G, during early pregnancy. Rhesus monkeys treated daily with an anti-Mamu-AG Ab (25D3) showed a broad spectrum of morphological changes in their placenta and decidua when compared with control animals. A delay in placental development was observed, involving delays in stromal growth and trophoblast proliferation, reduced villous vascularization, and reduced expression of chorionic gonadotropin. Although Ab treatment did not affect endometrial infiltration by NK cells, the numbers of DC-SIGN⁺ macrophages were reduced and CD3⁺ T cells were increased in the decidua of animals treated with 25D3 vs controls. Finally, decidual differentiation and maturation were impaired in 25D3-treated animals, further supporting a role for Mamu-AG in the support of pregnancy. The mechanisms responsible for these striking and generally unexpected effects of Mamu-AG interference remain to be determined; however, these in vivo data are some of the first to demonstrate the role of nonclassical MHC molecules in pregnancy.

Previously, prolonged Ag exposure was thought to be necessary to make an effective vaccine. However, Radcliffe et al. (p.8313 ) have shown that shorter Ag expression is better for eliciting an effector CD8⁺ T cell response. Using a DNA vaccine consisting of a mifepristone-responsive plasmid expressing SIINFEKL, the authors looked at CD8⁺ T cell effector responses to timed Ag doses. Surprisingly, they found that reducing Ag expression to a single burst led to significantly more peptide-specific cells than did prolonged Ag expression. IFN-γ production was impaired in T cells primed with persistent Ag expression, whereas T cells primed with a single burst of Ag produced IFN-γ when stimulated with 10-fold less peptide than those primed under prolonged Ag conditions. When memory responses were examined, more persistent Ag expression led to greater overall numbers of CD8⁺ T cells, but these could only expand ∼4-fold. This is in contrast with the transient burst of Ag, which produced memory cells that expanded ∼35-fold. These results help explain the CD8⁺ T cell impairment seen in chronic lymphocytic choriomeningitis virus infections where there is prolonged Ag exposure. The authors thus conclude that the most effective DNA vaccines directed at generating a CD8⁺ T cell response will be those that do not prolong Ag expression.

During the pathogenesis of multiple sclerosis and its animal model, experimental autoimmune encephalomyelitis (EAE), circulating leukocytes cross the blood-brain barrier (BBB). The involvement of E- and P-selectins in this process has been controversial, so Döring et al. (p.8470 ) reanalyzed the CNS expression and requirements for these adhesion molecules in EAE development in C57BL/6 and SJL mice. Using a unique polyclonal Ab, the authors observed expression of P-selectin in the meninges, but rarely the parenchyma, of healthy mice. They found, however, that P-selectin was up-regulated on endothelial cells and platelets in blood vessels in both the parenchyma and meninges during EAE. Surprisingly, although this adhesion molecule was up-regulated during disease, mice on either the C57BL/6 or SJL background lacking both E- and P-selectin developed EAE comparably to wild-type mice. T cells from these doubly deficient mice showed no alteration in myelin-specific proliferation or cytokine production, nor in the size or composition of the inflammatory CNS infiltrates during EAE. Finally, transgenic overexpression of E-selectin did not affect EAE development or recruitment across the BBB. These data, conducted in two different mouse strains, should help to resolve the controversy regarding selectin involvement in EAE pathogenesis.

The stimulatory receptor for advanced glycation end products (RAGE) has been broadly implicated in innate immunity, but its role in adaptive immunity is less clear. Moser et al. (p.8051 ) sought to clarify this receptor’s activity by evaluating the contribution of RAGE expression to the responses of T cells and dendritic cells (DCs) both in vitro and in vivo. Using an adoptive transfer system, the authors found that T cell priming was impaired if either the adoptively transferred T cells or the recipient cells lacked RAGE expression. In vitro experiments determined that CD3-mediated signaling was intact in RAGE-deficient T cells but that T:DC interactions and subsequent T cell proliferation were impaired if T cells, but not DCs, lacked RAGE. Additionally, RAGE-deficient T cells demonstrated impaired Th1 differentiation, leading to Th2 skewing in vitro. The authors next extensively analyzed the effects of RAGE deficiency in DCs to address previous data suggesting the importance RAGE in DC function. However, DCs expressing or lacking RAGE were found to be comparable in Ag processing and presentation, in vitro differentiation, activation by TLRs, migration, and T:DC adhesive interactions. Taken together, these data identify RAGE as an important player in adaptive immunity through T cell-intrinsic regulation of T cell priming and differentiation.

Lipid bodies are dynamic organelles important for lipid homeostasis that also participate in inflammatory processes. To better understand the role these organelles play in immune responses to infection, Pacheco et al. (p.8500 ) assessed the role of the chemokine MCP-1 in macrophage lipid body biogenesis. In in vivo models of peritonitis, macrophages from mice deficient in MCP-1 did not demonstrate the increase in lipid body biogenesis and production of leukotriene B₄ (LTB₄) observed in the macrophages of wild-type mice. In vitro studies were then employed to analyze the mechanism by which MCP-1 might induce the formation of lipid bodies synthesizing LTB₄. MCP-1 produced by the macrophages themselves was found to directly trigger lipid body biogenesis via signaling through its receptor CCR2, which required the activity of PI3K and ERK. Additionally, interference with microtubule networks inhibited MCP-1-induced lipid body biogenesis and LTB₄ synthesis, indicating that the proper regulation of microtubule dynamics is required for lipid body development and activity. This work enhances our knowledge of the activities of macrophages during inflammation by demonstrating that MCP-1 is required for the formation of lipid bodies and that these organelles can synthesize the inflammatory mediator LTB₄.

Summaries written by Jennifer Hartt Meyers, Ph.D.