SummaryMultiple sclerosis (MS) is a demyelinating inflammatory disorder of the central nervous system (CNS), which involves autoimmune responses to myelin antigens. Studies in experimental autoimmune encephalomyelitis (EAE), an animal model for MS, have provided convincing evidence that T cells specific for self-antigens mediate pathology in these diseases. Until recently, T helper type 1 (Th1) cells were thought to be the main effector T cells responsible for the autoimmune inflammation. However more recent studies have highlighted an important pathogenic role for CD4 + T cells that secrete interleukin (IL)-17, termed Th17, but also IL-17-secreting gd T cells in EAE as well as other autoimmune and chronic inflammatory conditions. This has prompted intensive study of the induction, function and regulation of IL-17-producing T cells in MS and EAE. In this paper, we review the contribution of Th1, Th17, gd, CD8+ and regulatory T cells as well as the possible development of new therapeutic approaches for MS based on manipulating these T cell subtypes.

Despite the fact that CD4+CD25+Foxp3+ regulatory T cells (Treg cells) play a central role in maintaining self-tolerance and that IL-17-producing CD4+ T cells (Th17 cells) are pathogenic in many autoimmune diseases, evidence to date has indicated that Th17 cells are resistant to suppression by human Foxp3+ Treg cells. It was recently demonstrated that CD39, an ectonucleotidase which hydrolyzes ATP, is expressed on a subset of human natural Treg cells. We found that although both CD4+CD25highCD39+ and CD4+CD25highCD39− T cells suppressed proliferation and IFN-γ production by responder T cells, only the CD4+CD25highCD39+, which were predominantly FoxP3+, suppressed IL-17 production, whereas CD4+CD25highCD39− T cells produced IL-17. An examination of T cells from multiple sclerosis patients revealed a normal frequency of CD4+CD25+CD127lowFoxP3+, but interestingly a deficit in the relative frequency and the suppressive function of CD4+CD25+CD127lowFoxP3+CD39+ Treg cells. The mechanism of suppression by CD39+ Treg cells appears to require cell contact and can be duplicated by adenosine, which is produced from ATP by the ectonucleotidases CD39 and CD73. Our findings suggest that CD4+CD25+Foxp3+CD39+ Treg cells play an important role in constraining pathogenic Th17 cells and their reduction in multiple sclerosis patients might lead to an inability to control IL-17 mediated autoimmune inflammation.

Repeated antigenic encounter drives proliferation and differentiation of memory T cell pools. An important question is whether certain specific T cells may be driven eventually to exhaustion in elderly individuals since the human life expectancy is increasing. We found that CMV-specific CD4+ T cells were significantly expanded in healthy young and old carriers compared with purified protein derivative-, varicella zoster virus-, EBV-, and HSV-specific populations. These CMV-specific CD4+ T cells exhibited a late differentiated phenotype since they were largely CD27 and CD28 negative and had shorter telomeres. Interestingly, in elderly CMV-seropositive subjects, CD4+ T cells of different specificities were significantly more differentiated than the same cells in CMV-seronegative individuals. This suggested the involvement of bystander-secreted, differentiation-inducing factors during CMV infection. One candidate was IFN-α, which induced loss of costimulatory receptors and inhibited telomerase in activated CD4+ T cells and was secreted at high levels by CMV-stimulated plasmacytoid dendritic cells (PDC). The CMV-specific CD4+ T cells in elderly subjects had severely restricted replicative capacity. This is the first description of a human memory T cell population that is susceptible to being lost through end-stage differentiation due to the combined effects of lifelong virus reactivation in the presence of bystander differentiation-inducing factors.

IL-1β plays a critical role in promoting IL-17 production by γδ and CD4 T cells. However, IL-1–targeted drugs, although effective against autoinflammatory diseases, are less effective against autoimmune diseases. Conversely, gain-of-function mutations in the NLRP3 inflammasome complex are associated with enhanced IL-1β and IL-18 production and Th17 responses. In this study, we examined the role of caspase-1–processed cytokines in IL-17 production and in induction of experimental autoimmune encephalomyelitis (EAE). Killed Mycobacterium tuberculosis, the immunostimulatory component in CFA used for inducing EAE, stimulated IL-1β and IL-18 production by dendritic cells through activation of the inflammasome complex and caspase-1. Dendritic cells stimulated with M. tuberculosis and myelin oligodendrocyte glycoprotein promoted IL-17 production by T cells and induced EAE following transfer to naive mice, and this was suppressed by a caspase-1 inhibitor and reversed by administration of IL-1β or IL-18. Direct injection of the caspase-1 inhibitor suppressed IL-17 production by CD4 T cells and γδ T cells in vivo and attenuated the clinical signs of EAE. γδ T cells expressed high levels of IL-18R and the combination of IL-18 and IL-23, as with IL-1β and IL-23, stimulated IL-17 production by γδ T cells, but also from CD4 T cells, in the absence of TCR engagement. Our findings demonstrate that caspase-1–processed cytokines IL-1β and IL-18 not only promote autoimmunity by stimulating innate IL-17 production by T cells but also reveal redundancy in the functions of IL-1β and IL-18, suggesting that caspase-1 or the inflammasome may be an important drug target for autoimmune diseases.

While memory T cells are maintained by continuous turnover, it is not clear how human regulatory CD4 + CD45RO + CD25 hi Foxp3 + T lymphocyte populations persist throughout life. We therefore used deuterium labeling of cycling cells in vivo to determine whether these cells could be replenished by proliferation. We found that CD4 + CD45RO + Foxp3 + CD25 hi T lymphocytes were highly proliferative, with a doubling time of 8 days, compared with memory CD4 + CD45RO + Foxp3 -CD25 -(24 days) or naive CD4 + CD45RA + Foxp3 -CD25 -populations (199 days). However, the regulatory population was susceptible to apoptosis and had critically short telomeres and low telomerase activity. It was therefore unlikely to be self regenerating. These data are consistent with continuous production from another population source. We found extremely close TCR clonal homology between regulatory and memory CD4 + T cells. Furthermore, antigen-related expansions within certain TCR Vb families were associated with parallel numerical increases of CD4 + CD45RO + CD25 hi Foxp3 + Tregs with the same Vb usage. It is therefore unlikely that all human CD4 + CD25 + Foxp3 + Tregs are generated as a separate functional lineage in the thymus. Instead, our data suggest that a proportion of this regulatory population is generated from rapidly dividing, highly differentiated memory CD4 + T cells; this has considerable implications for the therapeutic manipulation of these cells in vivo. IntroductionBoth memory and regulatory populations of T cells must be maintained in tandem in order to generate controlled immunity for the lifetime of the organism. Since the thymus involutes early in life, memory T cells have to largely be maintained by lifelong turnover of preexisting populations of specific T cells in adults (1, 2). The corollary of this is that thymic involution during aging will also severely restrict the production of Tregs by this organ. The source of these cells in adult humans and the relative contributions of long-term survival and ongoing turnover to the maintenance of CD4 + CD25 hi Foxp3 + Treg populations remain unknown.The naturally occurring CD4 + CD25 hi Treg subset that expresses the lineage marker Foxp3 represents an important population of suppressive T cells that can prevent reactivity to both self and nonself antigens (3-5). These cells also downregulate immune responses as pathogen is cleared (3)(4)(5). Early studies demonstrated that in mice, CD4 + CD25 hi Tregs are generated as a distinct population in the thymus (6). Indeed, in mice, there is substantial overlap of TCR repertoires between thymic and peripheral CD4 + Foxp3 + Tregs, suggesting that the thymic regulatory pool makes a significant contribution to the peripheral regulatory cells (7). However, murine CD4 + CD25 hi Tregs, which are phenotypically and functionally identical to the thymus-derived population can also be

The enzyme telomerase is essential for maintaining the replicative capacity of memory T cells. Although CD28 costimulatory signals can up-regulate telomerase activity, human CD8+ T cells lose CD28 expression after repeated activation. Nevertheless, telomerase is still inducible in CD8+CD28− T cells. To identify alternative costimulatory pathways that may be involved, we introduced chimeric receptors containing the signaling domains of CD28, CD27, CD137, CD134, and ICOS in series with the CD3 zeta (ζ) chain into primary human CD8+ T cells. Although CD3 ζ-chain signals alone were ineffective, triggering of all the other constructs induced proliferation and telomerase activity. However, not all CD8+CD28− T cells could up-regulate this enzyme. The further fractionation of CD8+CD28− T cells into CD8+CD28− CD27+ and CD8+CD28−CD27− subsets showed that the latter had significantly shorter telomeres and extremely poor telomerase activity. The restoration of CD28 signaling in CD8+CD28−CD27− T cells could not reverse the low telomerase activity that was not due to decreased expression of human telomerase reverse transcriptase, the enzyme catalytic subunit. Instead, the defect was associated with decreased phosphorylation of the kinase Akt, that phosphorylates human telomerase reverse transcriptase to induce telomerase activity. Furthermore, the defective Akt phosphorylation in these cells was specific for the Ser473 but not the Thr308 phosphorylation site of this molecule. Telomerase down-regulation in highly differentiated CD8+CD28−CD27− T cells marks their inexorable progress toward a replicative end stage after activation. This limits the ability of memory CD8+ T cells to be maintained by continuous proliferation in vivo.

Anergic/suppressive CD4+CD25+ T cells have been proposed to play an important role in the maintenance of peripheral tolerance. Here we demonstrate that in humans these cells suppress proliferation to self antigens, but also to dietary and foreign antigens. The suppressive CD4+CD25+ T cells display a broad usage of the T cell receptor Vβ repertoire,suggesting that they recognize a wide variety of antigens. They reside in the primed/memory CD4+CD45RO+CD45RBlow subset and have short telomeres, indicating that these cells have the phenotype of highly differentiated CD4+ T cells that have experienced repeated episodes of antigen‐specific stimulation in vivo. This suggests that anergic/suppressiveCD4+CD25+ T cells may be generated in the periphery as a consequence of repeated antigenic encounter. This is supported by the observation that highly differentiated CD4+T cells can be induced to become anergic/suppressive when stimulated by antigen presented by non‐professional antigen‐presenting cells. We suggest that besides being generated in the thymus, CD4+CD25+ regulatory T cells may also be generated in the periphery. This would provide a mechanism for the generation of regulatory cells that induce tolerance to a wide array of antigens that may not be encountered in the thymus.

Infection of endothelial cells with an endothelial cell-tropic clinical isolate of cytomegalovirus (CMV), C1FE, induced enhanced production of the neutrophil chemoattractant C-X-C chemokines interleukin-8 and GROalpha. Infected endothelial cell supernatants induced neutrophil chemotaxis in a transendothelial migration assay. Neutrophils acquired the CMV structural protein pp65 following either coculture with infected endothelial cells or transmigration through infected endothelium. The lack of CMV p72 expression in the neutrophils indicated that viral replication had not occurred in these cells. Of importance, neutrophils acquired infectious CMV during transmigration across infected endothelium and were subsequently able to transmit infectious virus to fibroblasts. Thus, CMV-infected endothelial cells can recruit neutrophils by the secretion of C-X-C chemokines and can transmit the virus to them by direct cell-to-cell contact and during neutrophil transendothelial migration, suggesting that the neutrophil-endothelial cell interaction plays an important role in virus dissemination in vivo.