实验性自身免疫性脑脊髓炎小鼠模型视神经炎发病机制：与辅助性T细胞亚群的关系

背景：视神经炎的病因多为自身免疫引起的炎性脱髓鞘,很多的视神经炎为多发性硬化的早期表现。在外周CD4+T细胞中,有5%-10%为调节性T细胞,其免疫抑制能力很强。自身免疫性脑脊髓炎所引起的视神经炎发病机制是否与辅助性T细胞亚群的有关呢？目的：分析实验性自身免疫性脑脊髓炎小鼠模型视神经炎发病机制与辅助性T细胞亚群的关系。方法：将小鼠腹腔注射百日咳菌液建立实验性自身免疫性脑脊髓炎模型,分别免疫11,15,19 d,并设腹腔注射生理盐水的佐剂组小鼠作对照。
　　结果与结论：酶联免疫吸附检测显示,与佐剂组相比,免疫后19 d组视神经白细胞介素4蛋白含量降低(P<0.05);免疫后11,15 d组视神经白细胞介素17蛋白含量升高(P<0.05);免疫后15,19 d组视神经干扰素γ蛋白含量升高(P <0.05);免疫后11,15,19 d组视神经Foxp3蛋白含量显著降低(P <0.05)。实时PCR检测显示,与佐剂组相比,免疫后11,15,19 d组视神经中干扰素γ、Foxp3 mRNA表达降低(P<0.05), RORt mRNA表达升高;免疫后15,19 d组视神经中白细胞介素4,白细胞介素17,T-beat mRNA表达升高(P<0.05)。免疫后19 d组GATA3 mRNA表达降低(P<0.05)。结果证实,实验性自身免疫性脑脊髓炎小鼠模型视神经炎的发生发展可能受到Foxp3和调节性T细胞表达减少的影响,免疫后早期白细胞介素17可能介导对炎症损伤,发病高峰期干扰素γ可能使炎症损伤程度加重。     

A pathogenic role for myelin-specific CD8(+) T cells in a model for multiple sclerosis

Multiple sclerosis (MS) is a demyelinating disease of the central nervous system (CNS) characterized by plaques of infiltrating CD4(+) and CD8(+) T cells. Studies of MS and experimental autoimmune encephalomyelitis (EAE), an animal model of MS, focus on the contribution of CD4(+) myelin-specific T cells. The role of CD8(+) myelin-specific T cells in mediating EAE or MS has not been described previously. Here, we demonstrate that myelin-specific CD8(+) T cells induce severe CNS autoimmunity in mice. The pathology and clinical symptoms in CD8(+) T cell-mediated CNS autoimmunity demonstrate similarities to MS not seen in myelin-specific CD4(+) T cell-mediated EAE. These data suggest that myelin-specific CD8(+) T cells could function as effector cells in the pathogenesis of MS.     

Andrographolide interferes with T cell activation and reduces experimental autoimmune encephalomyelitis in the mouse

Andrographolide is a bicyclic diterpenoid lactone derived from extracts of Andrographis paniculata, a plant indigenous to South Asian countries that shows anti-inflammatory properties. The molecular and cellular bases for this immunomodulatory capacity remain unknown. Here, we show that andrographolide is able to down-modulate both humoral and cellular adaptive immune responses. In vitro, this molecule was able to interfere with T cell proliferation and cytokine release in response to allogenic stimulation. These results were consistent with the observation that T cell activation by dendritic cells (DCs) was completely abolished by exposing DCs to andrographolide during antigen pulse. This molecule was able to interfere with maturation of DCs and with their ability to present antigens to T cells. Furthermore, in vivo immune responses such as antibody response to a thymus-dependent antigen and delayed-type hypersensitivity were drastically diminished in mice by andrographolide treatment. Finally, the ability of andrographolide to inhibit T cell activation was applied to interfere with the onset of experimental autoimmune encephalomyelitis (EAE), an inflammatory demyelinating disease of the central nervous system that is primarily mediated by CD4(+) T cells and serves as an animal model for human multiple sclerosis. Treatment with andrographolide was able to significantly reduce EAE symptoms in mice by inhibiting T cell and antibody responses directed to myelin antigens. Our data suggest that andrographolide is able to efficiently block T cell activation in vitro, as well as in vivo, a feature that could be useful for interfering with detrimental T cell responses.     

Spontaneous opticospinal encephalomyelitis in a double-transgenic mouse model of autoimmune T cell/B cell cooperation

We describe a double-transgenic mouse strain (opticospinal EAE [OSE] mouse) that spontaneously develops an EAE-like neurological syndrome closely resembling a human variant of multiple sclerosis, Devic disease (also called neuromyelitis optica). Like in Devic disease, the inflammatory, demyelinating lesions were located in the optic nerve and spinal cord, sparing brain and cerebellum, and the murine lesions showed histological similarity with their human correlates. OSE mice have recombination-competent immune cells expressing a TCR-alphabeta specific for myelin oligodendrocyte glycoprotein (MOG) aa 35-55 peptide in the context of I-Ab along with an Ig J region replaced by the recombined heavy chain of a monoclonal antibody binding to a conformational epitope on MOG. OSE mouse B cells bound even high dilutions of recombinant MOG, but not MOG peptide, and processed and presented it to autologous T cells. In addition, in OSE mice, but not in single-transgenic parental mice, anti-MOG antibodies were switched from IgM to IgG1.     

Proteinase-activated receptor 2 modulates neuroinflammation in experimental autoimmune encephalomyelitis and multiple sclerosis

The proteinase-activated receptors (PARs) are widely recognized for their modulatory properties of inflammation and neurodegeneration. We investigated the role of PAR2 in the pathogenesis of multiple sclerosis (MS) in humans and experimental autoimmune encephalomyelitis (EAE) in mice. PAR2 expression was increased on astrocytes and infiltrating macrophages in human MS and murine EAE central nervous system (CNS) white matter (P < 0.05). Macrophages and astrocytes from PAR2 wild-type (WT) and knockout (KO) mice exhibited differential immune gene expression with PAR2 KO macrophages showing significantly higher interleukin 10 production after lipopolysaccharide stimulation (P < 0.001). PAR2 activation in macrophages resulted in the release of soluble oligodendrocyte cytotoxins (P < 0.01). Myelin oligodendrocyte glycoprotein-induced EAE caused more severe inflammatory gene expression in the CNS of PAR2 WT animals (P < 0.05), together with enhanced T cell proliferation and interferon gamma production (P < 0.05), compared with KO littermates. Indeed, PAR2 WT animals showed markedly greater microglial activation and T lymphocyte infiltration accompanied by worsened demyelination and axonal injury in the CNS compared with their PAR2 KO littermates. Enhanced neuropathological changes were associated with a more severe progressive relapsing disease phenotype (P < 0.001) in WT animals. These findings reveal previously unreported pathogenic interactions between CNS PAR2 expression and neuroinflammation with ensuing demyelination and axonal injury.     

Inactivation of T cell receptor peptide-specific CD4 regulatory T cells induces chronic experimental autoimmune encephalomyelitis (EAE)

T cell receptor (TCR)-recognizing regulatory cells, induced after vaccination with self-reactive T cells or TCR peptides, have been shown to prevent autoimmunity. We have asked whether this regulation is involved in the maintenance of peripheral tolerance to myelin basic protein (MBP) in an autoimmune disease model, experimental autoimmune encephalomyelitis (EAE). Antigen-induced EAE in (SJL x B10.PL)F1 mice is transient in that most animals recover permanently from the disease. Most of the initial encephalitogenic T cells recognize MBP Ac1-9 and predominantly use the TCR V beta 8.2 gene segment. In mice recovering from MBP-induced EAE, regulatory CD4+ T cells (Treg) specific for a single immunodominant TCR peptide B5 (76-101) from framework region 3 of the V beta 8.2 chain, become primed. We have earlier shown that cloned B5-reactive Treg can specifically downregulate responses to Ac1- 9 and also protect mice from EAE. These CD4 Treg clones predominantly use the TCR V beta 14 or V beta 3 gene segments. Here we have directly tested whether deletion/blocking of the Treg from the peripheral repertoire affects the spontaneous recovery from EAE. Treatment of F1 mice with appropriate V beta-specific monoclonal antibodies resulted in an increase in the severity and duration of the disease; even relapses were seen in one-third to one-half of the Treg-deleted mice. Interestingly, chronic disease in treated mice appears to be due to the presence of Ac1-9-specific T cells. Thus, once self-tolerance to MBP is broken by immunization with the antigen in strong adjuvant, TCR peptide- specific CD4 Treg cells participate in reestablishing peripheral tolerance. Thus, a failure to generate Treg may be implicated in chronic autoimmune conditions.     

Glatiramer acetate (Copaxone) therapy induces CD8(+) T cell responses in patients with multiple sclerosis

Glatiramer acetate (GA; Copaxone) is a random copolymer of glutamic acid, lysine, alanine, and tyrosine that is used therapeutically in patients with multiple sclerosis (MS). To investigate the mechanism of the drug's immunomodulatory effect, we used immunophenotypic approaches to characterize the precise nature of GA-induced T cell responses. We demonstrate here that healthy individuals and untreated MS patients exhibit prominent T cell proliferative responses to GA. However, these responses are different in distinct subsets of T cells. Whereas GA-induced CD4(+) T cell responses are comparable in healthy individuals and MS patients, CD8(+) T cell responses are significantly lower in untreated MS patients. Treatment with GA results in upregulation of these CD8(+) responses with restoration to levels observed in healthy individuals. Both CD4(+) and CD8(+) GA-specific responses are HLA-restricted. GA therapy also induces a change in the cytokine profile of GA-specific CD4(+) and CD8(+) T cells. This study provides the first direct immunophenotypic evidence, to our knowledge, of GA-specific CD8(+) T cell responses and their upregulation during the course of therapy, which may suggest a role for these responses in the immunomodulatory effects of the drug.     

CD4+T细胞及其亚群与多发性硬化

目的:总结近年来国内外有关多发性硬化的免疫学研究进展,分析CD4 T淋巴细胞,Th1/Th2,CD4 CD25 T淋巴细胞与多发性硬化之间的关系。资料来源:应用计算机检索Medline1992-12/2004-12与CD4 T淋巴细胞,Th1/Th2,CD4 CD25 T淋巴细胞和多发性硬化相关文章,检索词“CD4 Tcell,Th1/Th2cell,CD4 CD25 TLymphocytes,Multiplesclerosis”,并限定文章语言种类为“English”。资料选择:就检索到的120余篇文献进行筛选,选择以CD4 Tcell,Th1/Th2,CD4 CD25 T淋巴细胞为主要研究内容的文献27篇,无论研究对象为动物还是患者全部纳入,其中研究内容相似的,以近3年且发表在较权威杂志者优先。资料提炼:将筛选到的27篇文献按CD4 T淋巴细胞,Th1/Th2,CD4 CD25 T淋巴细胞与多发性硬化的关系进行分类:其中6篇与CD4 T淋巴细胞,3篇与Th1/Th2相关,18篇与CD4 CD25 T淋巴细胞有关。资料综合:6篇文献,说明了CD4 T淋巴细胞在多发性硬化的发病过程中居于中心环节,3篇文献提示Th1/Th2细胞的失衡与多发性硬化的疾病进程有关,18篇文献提到CD4 CD25 T淋巴细胞的基础研究及其与多发性硬化的关系。结论:病灶部位CD4 T淋巴细胞的增加,Th1/Th2细胞的失衡会导致多发性硬化发病。而CD4 CD25 T淋巴细胞功能的减弱与多发性硬化的发病有关,但数量是否有关还需要进一步研究。     

An electrophysiological model of spinal transmission deficits in mouse experimental autoimmune encephalomyelitis

Chronic relapsing/remitting experimental autoimmune encephalomyelitis (EAE) can be induced in 8-week-old female SJL/J(H-2) mice via inoculation with the p139-151 peptide of myelin proteolipid protein (PLP), Mycobacterium tuberculosis (MT), complete Freund's adjuvant (CFA), and Bordatella pertussis. EAE is a relevant preclinical model of MS that incorporates several aspects of the clinical disease. Chief among these are the inflammatory mediated neurological deficits. While the impact of localized spinal cord demyelination on neurotransmission has been modeled successfully, relatively little work has been done with spinal cord from animals with EAE. The goal of this study was to assess the utility of a grease-gap tissue bath methodology in the detection of transmission deficits in EAE spinal cord tissue. Spinal cords removed from EAE mice at different phases of the neurological deficit were assessed for their response to both lumbar and sacral application of one of several depolarizing agents (veratridine, potassium chloride [KCl], (+/-)-alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid [AMPA]). The main finding of this study is that transmission deficits were detected in EAE mice at the onset of the neurological deficits. They were sustained for a period of approximately 2 to 3 weeks post disease onset followed by a gradual recovery of group function. The other finding is that there is a decrease in the latency to achieve AMPA-mediated depolarization in sacral spinal cord that is independent of the magnitude of the depolarization response. These results suggest that this methodology can be utilized to assess sensory and motor deficits in spinal cord from EAE animals.     

CD4(+) T cell vaccination overcomes defective cross-presentation of fungal antigens in a mouse model of chronic granulomatous disease

Aspergillus fumigatus is a model fungal pathogen and a common cause of infection in individuals with the primary immunodeficiency chronic granulomatous disease (CGD). Although primarily considered a deficiency of innate immunity, CGD is also linked to dysfunctional T cell reactivity. Both CD4(+) and CD8(+) T cells mediate vaccine-induced protection from experimental aspergillosis, but the molecular mechanisms leading to the generation of protective immunity and whether these mechanisms are dysregulated in individuals with CGD have not been determined. Here, we show that activation of either T cell subset in a mouse model of CGD is contingent upon the nature of the fungal vaccine, the involvement of distinct innate receptor signaling pathways, and the mode of antigen routing and presentation in DCs. Aspergillus conidia activated CD8(+) T cells upon sorting to the Rab14(+) endosomal compartment required for alternative MHC class I presentation. Cross-priming of CD8(+) T cells failed to occur in mice with CGD due to defective DC endosomal alkalinization and autophagy. However, long-lasting antifungal protection and disease control were successfully achieved upon vaccination with purified fungal antigens that activated CD4(+) T cells through the endosome/lysosome pathway. Our study thus indicates that distinct intracellular pathways are exploited for the priming of CD4(+) and CD8(+) T cells to A. fumigatus and suggests that CD4(+) T cell vaccination may be able to overcome defective antifungal CD8(+) T cell memory in individuals with CGD.     

Clonal expansions of CD8(+) T cells dominate the T cell infiltrate in active multiple sclerosis lesions as shown by micromanipulation and single cell polymerase chain reaction

Clonal composition and T cell receptor (TCR) repertoire of CD4(+) and CD8(+) T cells infiltrating actively demyelinating multiple sclerosis (MS) lesions were determined with unprecedented resolution at the level of single cells. Individual CD4(+) or CD8(+) T cells were isolated from frozen sections of lesional tissue by micromanipulation and subjected to single target amplification of TCR-beta gene rearrangements. This strategy allows the assignment of a TCR variable region (V region) sequence to the particular T cell from which it was amplified. Sequence analysis revealed that in both cases investigated, the majority of CD8(+) T cells belonged to few clones. One of these clones accounted for 35% of CD8(+) T cells in case 1. V region sequence comparison revealed signs of selection for common peptide specificities for some of the CD8(+) T cells in case 1. In both cases, the CD4(+) T cell population was more heterogeneous. Most CD4(+) and CD8(+) clones were represented in perivascular infiltrates as well as among parenchymal T cells. In case 2, two of the CD8(+) clones identified in brain tissue were also detected in peripheral blood. Investigation of the antigenic specificities of expanded clones may help to elucidate their functional properties.     

Spontaneous regression of primary autoreactivity during chronic progression of experimental autoimmune encephalomyelitis and multiple sclerosis

Experimental autoimmune encephalomyelitis (EAE) is a widely used animal model for multiple sclerosis (MS). EAE is typically initiated by CD4(+) T helper cell type 1 (Th1) autoreactivity directed against a single priming immunodominant myelin peptide determinant. Recent studies have shown that clinical progression of EAE involves the accumulation of neo-autoreactivity, commonly referred to as epitope spreading, directed against peptide determinants not involved in the priming process. This study directly addresses the relative roles of primary autoreactivity and secondary epitope spreading in the progression of both EAE and MS. To this end we serially evaluated the development of several epitope-spreading cascades in SWXJ mice primed with distinctly different encephalitogenic determinants of myelin proteolipid protein. In a series of analogous experiments, we examined the development of epitope spreading in patients with isolated monosymptomatic demyelinating syndrome as their disease progressed to clinically definite MS. Our results indicate that in both EAE and MS, primary proliferative autoreactivity associated with onset of clinical disease invariably regresses with time and is often undetectable during periods of disease progression. In contrast, the emergence of sustained secondary autoreactivity to spreading determinants is consistently associated with disease progression in both EAE and MS. Our results indicate that chronic progression of EAE and MS involves a shifting of autoreactivity from primary initiating self-determinants to defined cascades of secondary determinants that sustain the self-recognition process during disease progression.     

Homologies between T cell receptor junctional sequences unique to multiple sclerosis and T cells mediating experimental allergic encephalomyelitis.

The selection of T cell clones with mutations in the hypoxanthine guanine phosphoribosyltransferase (hprt) gene has been used to isolate T cells reactive to myelin basic protein (MBP) in patients with multiple sclerosis (MS). These T cell clones are activated in vivo, and are not found in healthy individuals. The third complementarity determining regions (CDR3) of the T cell receptor (TCR) alpha and beta chains are the putative contact sites for peptide fragments of MBP bound in the groove of the HLA molecule. The TCR V gene usage and CDR3s of these MBP-reactive hprt-T cell clones are homologous to TCRs from other T cells relevant to MS, including T cells causing experimental allergic encephalomyelitis (EAE) and T cells found in brain lesions and in the cerebrospinal fluid (CSF) of MS patients. In vivo activated MBP-reactive T cells in MS patients may be critical in the pathogenesis of MS.     

Failure to suppress the expansion of the activated CD4 T cell population in interferon gamma-deficient mice leads to exacerbation of experimental autoimmune encephalomyelitis

Mice deficient in interferon (IFN)-gamma or IFN-gamma receptor develop progressive and fatal experimental autoimmune encephalomyelitis (EAE). We demonstrate that CD4 T cells lacking IFN-gamma production were required to passively transfer EAE, indicating that they were disease-mediating cells in IFN-gamma knockout (KO) mice. IFN-gamma KO mice accumulated 10-16-fold more activated CD4 T cells (CD4(+)CD44(hi)) than wild-type mice in the central nervous system during EAE. CD4(+)CD44(hi) T cells in the spleen and central nervous system of IFN-gamma KO mice during EAE showed markedly increased in vivo proliferation and significantly decreased ex vivo apoptosis compared with those of wild-type mice. IFN-gamma KO CD4(+)CD44(hi) T cells proliferated extensively to antigen restimulation in vitro and accumulated larger numbers of live CD4(+) CD44(hi) T cells. IFN-gamma completely suppressed proliferation and significantly induced apoptosis of CD4(+)CD44(hi) T cells responding to antigen and hence inhibited accumulation of live, activated CD4 T cells. We thus present novel in vivo and in vitro evidence that IFN-gamma may limit the extent of EAE by suppressing expansion of activated CD4 T cells.     

Simultaneous isolation of CD8(+) and CD4(+) T cells specific for multiple viruses for broad antiviral immune reconstitution after allogeneic stem cell transplantation

Opportunistic viral infections can cause serious morbidity and mortality in immunocompromised patients after allogeneic stem cell transplantation. Clinical studies have shown that adoptive transfer of donor-derived T cells specific for cytomegalovirus (CMV), Epstein-Barr virus (EBV), or human adenovirus (HAdV) can be a safe and effective treatment of infections with these major viral pathogens. The aim of this study was to develop a method for the simultaneous isolation of coordinated CD8(+) and CD4(+) memory T-cell responses against a broad repertoire of viral epitopes. To ensure that the method was applicable to a wide variety of virus-specific T cells that may differ in phenotypic and functional properties, we focused on T cells specific for the persistent viruses, CMV and EBV, and T cells specific for HAdV and influenza (FLU), which are not repetitively activated in vivo after initial viral clearance. Following in vitro activation, nearly all T cells specific for these viruses produced interferon γ (IFN-γ) and tumor necrosis factor α, and expressed CD137, whereas the populations varied in the production of interleukin-2, degranulation, and expression of phenotypic markers. Different kinetics of IFN-γ production were observed in CMV/EBV-specific T cells and HAdV/FLU-specific T cells. However, after the stimulation of peripheral blood from seropositive donors with viral protein-spanning peptide pools, the activated virus-specific CD8(+) and CD4(+) T cells could be simultaneously isolated by either IFN-γ-based or CD137-based enrichment. This study provides an efficient and widely applicable strategy for the isolation of virus-specific T cells, which may be used for the reconstitution of virus-specific immunity in allogeneic stem cell transplantation recipients.     

One naive T cell,multiple fates in CD8+ T cell differentiation

The mechanism by which the immune system produces effector and memory T cells is largely unclear. To allow a large-scale assessment of the development of single naive T cells into different subsets, we have developed a technology that introduces unique genetic tags (barcodes) into naive T cells. By comparing the barcodes present in antigen-specific effector and memory T cell populations in systemic and local infection models, at different anatomical sites, and for TCR–pMHC interactions of different avidities, we demonstrate that under all conditions tested, individual naive T cells yield both effector and memory CD8⁺ T cell progeny. This indicates that effector and memory fate decisions are not determined by the nature of the priming antigen-presenting cell or the time of T cell priming. Instead, for both low and high avidity T cells, individual naive T cells have multiple fates and can differentiate into effector and memory T cell subsets.Activation of naive antigen-specific T cells is characterized by a vigorous proliferative burst, resulting in the formation of a large pool of effector T cells. After pathogen clearance, ∼95% of activated T cells die, leaving behind a stable pool of long-lived memory cells (Williams and Bevan, 2007). Two fundamentally different mechanisms could give rise to the production of effector and memory T cells during an immune response. First, single naive T cells may be destined to produce either effector T cells or memory T cells, but not both (“one naive cell, one fate”). As an alternative, effector and memory T cells could derive from the same clonal precursors within the naive T cell pool (“one naive cell, multiple fates”). As the fate decisions that control T cell differentiation could either be taken during initial T cell priming (i.e., before the first cell division) or at later stages, at least four conceptually different models describing effector and memory T cell differentiation can be formulated (Fig. S1).A first model predicts a separate origin of effector and memory T cells as a result of differential T cell priming by APCs. In this scenario, fate decisions would be taken before the first cell division, and even though cells destined to become memory cells may transiently display traits associated with effector T cells (e.g., expression of granzyme B or IFN-γ; see the following paragraphs), their ability for long-term survival would be predetermined. In line with this model, several studies have provided evidence that the fate of CD8⁺ T cells may, to some extent, be programmed during initial activation (Kaech and Ahmed, 2001; van Stipdonk et al., 2003; Masopust et al., 2004; Williams and Bevan, 2007; Bannard et al., 2009).A second model, which relies on recent data from Chang et al. (2007), likewise suggests that the priming APC plays the crucial role in determining effector or memory T cell fate, but by a strikingly different mechanism and with an opposite prediction concerning the lineage relationship of effector and memory T cells. Specifically, analysis of T cell–APC conjugates has shown that the first division of activated T cells can be asymmetric, with the daughter T cell that is formed proximal to the APC being more likely to contribute to the effector T cell subset and the distal daughter T cell being more likely to generate memory T cells (Chang et al., 2007). Assuming that all primary daughter cells survive and yield further progeny, these data would predict that single naive T cells contribute to both the effector and the memory subset.In contrast to these two models that are based on a determining role of the priming APC, two other models predict that T cell fate is determined by the cumulative effect of signals that not only naive T cells but also their descendants receive. The first of these models, termed the “decreasing potential model,” argues that T cell progeny that receive additional stimulation after priming undergo terminal differentiation toward the effector subset, whereas descendants that do not encounter these signals may transiently display certain effector functions but will ultimately become memory T cells (Ahmed and Gray, 1996). In support of this model, it has been demonstrated that continued inflammatory signals (Badovinac et al., 2004; Joshi et al., 2007) and prolonged antigenic stimulation (Sarkar et al., 2008) can lead descendant CD8⁺ T cells to preferentially develop into effector cells.If the descendants of all individual naive T cells have an equal chance of receiving signals for terminal differentiation, the standard decreasing potential model predicts that memory and effector T cells will be derived from the same population of naive T cells. However, there is evidence that the environmental factors that promote either terminal differentiation or memory T cell development may alter over the course of infection (Sarkar et al., 2008). A fourth model therefore argues that the progeny of T cells that are activated early or late during infection will receive distinct signals and, hence, assume (partially) different fates (van Faassen et al., 2005; D’Souza and Hedrick, 2006; Quigley et al., 2007; Stemberger et al., 2007a).A large number of studies in which cell differentiation was analyzed at the population level have been informative in revealing which effector properties can be displayed by T cells that subsequently differentiate into memory T cells (for review see Jameson and Masopust, 2009). In particular, two recent studies using IFN-γ or granzyme B reporter mice have shown that memory T cells can arise from cells that have previously transcribed IFN-γ or granzyme B genes (Harrington et al., 2008; Bannard et al., 2009). However, it is important to realize that these studies reveal little with regard to the developmental potential of individual naive T cells. Specifically, the fact that T cells that have a particular effector capacity can become memory T cells does not indicate that all naive T cells yield such effector cells, nor does it indicate that all memory T cells have gone through an effector phase.To determine the developmental potential of naive T cells, it is essential to develop technologies in which T cell responses can be analyzed at the single naive T cell level. In early work that aimed to follow T cell responses at the clonal level, TCR repertoire analysis has been used to assess the kinship of T cell populations (Maryanski et al., 1996; Kedzierska et al., 2004). However, as several naive T cell clones can share the same TCR, it has been argued that such analyses do not necessarily monitor T cell fate at the single T cell level (Stemberger et al., 2007b; Obar et al., 2008). Recently, Stemberger et al. (2007a) have reported on a more elegant approach to address naive T cell potency. Using the transfer of single naive CD8⁺ T cells into mice, this study provides direct evidence that a single naive CD8⁺ T cell can form both effector and memory cell subsets. However, the statistical power of single-cell transfer studies obviously has limitations. In addition, if homeostatic proliferation would occur before antigen-driven proliferation in this system, this would limit the conclusions that can be drawn with regard to the pluripotency of a single naive T cell.In this study, we have developed a technology that allows the generation of naive T cells that carry unique genetic tags (barcodes), and we describe how this technology can be used for the large-scale assessment of the developmental potential of single naive T cells. Using physiological frequencies of barcode-labeled naive CD8⁺ T cells of different functional avidities, we demonstrate that in both systemic and local infection models, effector and memory CD8⁺ T cell subsets share the same precursors in the naive T cell pool. These data demonstrate that under all conditions analyzed, single naive T cells do not selectively yield effector or memory T cells. Rather, T cell differentiation into effector and memory T cell subsets occurs by a one naive cell, multiple fates principle.     

Cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) can regulate dendritic cell-induced activation and cytotoxicity of CD8(+) T cells independently of CD4(+) T cell help

The mechanisms that regulate the strength and duration of CD8(+) cytotoxic T cell activity determine the effectiveness of an antitumor immune response. To better understand the antitumor effects of anti-cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) antibody treatment, we analyzed the effect of CTLA-4 signaling on CD8(+) T cells in vitro and in vivo. In vitro, cross-linking of CTLA-4 on purified CD8(+) T cells caused decreased proliferative responses to anti-CD3 stimulation and rapid loss of activation marker expression. In vivo, blockade of CTLA-4 by neutralizing anti-CTLA-4 mAb greatly enhanced the accumulation, activation, and cytotoxic activity of CD8(+) T cells induced by immunization with Ag on dendritic cells (DC). This enhanced response did not require the expression of MHC class II molecules on DC or the presence of CD4(+) T cells. These results demonstrate that CTLA-4 blockade is able to directly enhance the proliferation and activation of specific CD8(+) T cells, indicating its potential for tumor immunotherapy even in situations in which CD4(+) T cell help is limited or absent.