CD4+ regulatory T cells are spared from deletion by antilymphocyte serum, a polyclonal anti-T cell antibody

Broad T cell depletion has been used as an integral part of treatment in transplantation and autoimmune diseases. Following depletion, residual T cells undergo homeostatic proliferation and convert to memory-like T cells. In this study, we investigated the effect of T cell depletion by antilymphocyte serum (ALS), a polyclonal anti-T cell Ab, on CD4(+) regulatory T cells. After ALS treatment, CD4(+)CD25(+) T cells underwent proliferation and expressed a memory T cell marker, CD44. One week after ALS treatment, both CD25(+) and CD25(-) T cells exhibited increased suppression of alloresponses in vitro, which waned thereafter to the levels mediated by naive CD25(+) and CD25(-) T cells. By real-time PCR analyses, ALS treatment of CD4-deficient mice adoptively transferred with Thy1.2(+)CD4(+)CD25(+)Foxp3(+) and Thy1.1(+)CD4(+)CD25(-)Foxp3(-) T cells resulted in the appearance of Thy1.2(+)CD4(+)CD25(-)Foxp3(+) and Thy1.1(+)CD4(+)CD25(+)Foxp3(+) T cells, suggesting the conversion between CD25(+) and CD25(-) T cells. Naive CD25(+) T cells expressed a higher level of intracellular Bcl-x(L) than CD25(-) T cells. Up-regulation of the Bcl-x(L) molecule during ALS-induced homeostatic expansion further promoted survival of CD25(+) and, to a lessor degree, CD25(-) cells. These results indicate that CD25(+) T cells are spared from ALS-mediated deletion, with some CD25(+) T cells converting to CD25(-) T cells, and continue to exhibit regulatory activity. The concomitant presence of T cell deletion and continuous regulatory T cell activity may underlie the therapeutic effect of ALS, particularly in treatment of autoimmune diseases.     

Cutting edge: control of CD8+ T cell activation by CD4+CD25+ immunoregulatory cells

CD4(+)CD25(+) regulatory T cells inhibit organ-specific autoimmune diseases induced by CD4(+)CD25(-) T cells and are potent suppressors of CD4(+)CD25(-) T cell activation in vitro. We demonstrate that CD4(+)CD25(+) T cells also suppress both proliferation and IFN-gamma production by CD8(+) T cells induced either by polyclonal or Ag-specific stimuli. CD4(+)CD25(+) T cells inhibit the activation of CD8(+) responders by inhibiting both IL-2 production and up-regulation of IL-2Ralpha-chain (CD25) expression. Suppression is mediated via a T-T interaction as activated CD4(+)CD25(+) T cells suppress the responses of TCR-transgenic CD8(+) T cells stimulated with soluble peptide-MHC class I tetramers in the complete absence of APC. These results broaden the immunoregulatory role played by CD4(+)CD25(+) T cells in the prevention of autoimmune diseases, but also raise the possibility that they may hinder the induction of effector CD8(+) T cells to tumor or foreign Ags.     

Segregated regulatory CD39+CD4+ T cell function: TGF-β-producing Foxp3- and IL-10-producing Foxp3+ cells are interdependent for protection against collagen-induced arthritis

Oral immunization with a Salmonella vaccine vector expressing enterotoxigenic Escherichia coli colonization factor Ag I (CFA/I) can protect against collagen-induced arthritis (CIA) by dampening IL-17 and IFN-γ via enhanced IL-4, IL-10, and TGF-β. To identify the responsible regulatory CD4(+) T cells making the host refractory to CIA, Salmonella-CFA/I induced CD39(+)CD4(+) T cells with enhanced apyrase activity relative to Salmonella vector-immunized mice. Adoptive transfer of vaccine-induced CD39(+)CD4(+) T cells into CIA mice conferred complete protection, whereas CD39(-)CD4(+) T cells did not. Subsequent analysis of vaccinated Foxp3-GFP mice revealed the CD39(+) T cells were composed of Foxp3-GFP(-) and Foxp3-GFP(+) subpopulations. Although each adoptively transferred Salmonella-CFA/I-induced Foxp3(-) and Foxp3(+)CD39(+)CD4(+) T cells could protect against CIA, each subset was not as efficacious as total CD39(+)CD4(+) T cells, suggesting their interdependence for optimal protection. Cytokine analysis revealed Foxp3(-) CD39(+)CD4(+) T cells produced TGF-β, and Foxp3(+)CD39(+)CD4(+) T cells produced IL-10, showing a segregation of function. Moreover, donor Foxp3-GFP(-) CD4(+) T cells converted to Foxp3-GFP(+) CD39(+)CD4(+) T cells in the recipients, showing plasticity of these regulatory T cells. TGF-β was found to be essential for protection because in vivo TGF-β neutralization reversed activation of CREB and reduced the development of CD39(+)CD4(+) T cells. Thus, CD39 apyrase-expressing CD4(+) T cells stimulated by Salmonella-CFA/I are composed of TGF-β-producing Foxp3(-) CD39(+)CD4(+) T cells and support the stimulation of IL-10-producing Foxp3(+) CD39(+)CD4(+) T cells.     

IL-15 expands unconventional CD8alphaalphaNK1.1+ T cells but not Valpha14Jalpha18+ NKT cells

Despite recent gains in knowledge regarding CD1d-restricted NKT cells, very little is understood of non-CD1d-restricted NKT cells such as CD8(+)NK1.1(+) T cells, in part because of the very small proportion of these cells in the periphery. In this study we took advantage of the high number of CD8(+)NK1.1(+) T cells in IL-15-transgenic mice to characterize this T cell population. In the IL-15-transgenic mice, the absolute number of CD1d-tetramer(+) NKT cells did not increase, although IL-15 has been shown to play a critical role in the development and expansion of these cells. The CD8(+)NK1.1(+) T cells in the IL-15-transgenic mice did not react with CD1d-tetramer. Approximately 50% of CD8(+)NK1.1(+) T cells were CD8alphaalpha. In contrast to CD4(+)NK1.1(+) T cells, which were mostly CD1d-restricted NKT cells and of which approximately 70% were CD69(+)CD44(+), approximately 70% of CD8(+)NK1.1(+) T cells were CD69(-)CD44(+). We could also expand similar CD8alphaalphaNK1.1(+) T cells but not CD4(+) NKT cells from CD8alpha(+)beta(-) bone marrow cells cultured ex vivo with IL-15. These results indicate that the increased CD8alphaalphaNK1.1(+) T cells are not activated conventional CD8(+) T cells and do not arise from conventional CD8alphabeta precursors. CD8alphaalphaNK1.1(+) T cells produced very large amounts of IFN-gamma and degranulated upon TCR activation. These results suggest that high levels of IL-15 induce expansion or differentiation of a novel NK1.1(+) T cell subset, CD8alphaalphaNK1.1(+) T cells, and that IL-15-transgenic mice may be a useful resource for studying the functional relevance of CD8(+)NK1.1(+) T cells.     

Human CD4+CD25+ regulatory T cells share equally complex and comparable repertoires with CD4+CD25- counterparts

CD4(+)CD25(+) T cells are critical mediators of peripheral immune tolerance. However, many developmental and functional characteristics of these cells are unknown, and knowledge of human regulatory T cells is particularly limited. To better understand how human CD4(+)CD25(+) T cells develop and function, we examined the diversity of CD4(+)CD25(+) and CD4(+)CD25(-) T cell repertoires in both thymus and peripheral blood. Levels of T receptor excision circles (TREC) were comparable in purified CD4(+)CD25(+) and CD4(+)CD25(-) thymic populations, but were significantly higher than those in samples derived from peripheral blood, consistent with murine studies demonstrating thymic development of CD4(+)CD25(+) regulatory T cells. Surprisingly, CD4(+)CD25(-) T cells isolated from peripheral blood had greater TREC quantities than their CD4(+)CD25(+) counterparts, supporting the possibility of extrathymic expansion as well. CD4(+)CD25(+) and CD4(+)CD25(-) T cells from a given individual showed overlapping profiles with respect to diversity by Vbeta staining and spectratyping. Interestingly, CD4(+)CD25(+) T cells have lower quantities of CD3 than CD4(+)CD25(-) T cells. Collectively, these data suggest that human CD4(+)CD25(+) T cells recognize a similar array of Ags as CD4(+)CD25(-) T cells. However, reduced levels of TCR on regulatory T cells suggest different requirements for activation and may contribute to how the immune system regulates whether a particular response is suppressed or augmented.     

Maturation of adult peripheral blood CD38(+)CD4(+) T cells demonstrated by cytokine production in response to a superantigen,TSST-1

This study looks at immunoincompetent CD4(+) T cells in adult peripheral blood (APB) using cytokine production in response to a superantigen as a measure of function. We compared the function of APB CD38(+)CD4(+) and CD38(-/low)CD4(+) T cells to that of cord blood (CB) CD4(+) T cells. APB CD4(+) T cell blasts produce substantial amounts of IL-2 in response to TSST-1 restimulation, while CB CD4(+) T cell blasts produce less. APB CD38(+)CD4(+) T cells produce low levels of IL-4 and IFN-gamma in response to TSST-1, even after activation, while APB CD38(-/low)CD4(+) T cells retain their ability to produce high levels of these cytokines despite high CD38 expression. These results suggest that the developmental stage of APB CD38(+)CD4(+) T cells lies between that of CB CD4(+) T cells and APB CD38(-/low)CD4(+) T cells and that APB CD38(+)CD45RO(-)CD4(+) T cells gradually cease to express CD38 as they acquire full function. We reconsider CD4(+) cell maturation and response to TSST-1 and discuss the implications of T cell maturity on infectious diseases.     

CD27 instructs CD4+ T cells to provide help for the memory CD8+ T cell response after protein immunization

For optimal quality, memory CD8(+) T cells require CD4(+) T cell help. We have examined whether CD4(+) T cells require CD27 to deliver this help, in a model of intranasal OVA protein immunization. CD27 deficiency reduced the capacity of CD4(+) T cells to support Ag-specific CD8(+) T cell accumulation at the tissue site after primary and secondary immunization. CD27-dependent CD4(+) T cell help for the memory CD8(+) T cell response was delivered during priming. It did not detectably affect formation of CD8(+) memory T cells, but promoted their secondary expansion. CD27 improved survival of primed CD4(+) T cells, but its contribution to the memory CD8(+) T cell response relied on altered CD4(+) T cell quality rather than quantity. CD27 induced a Th1-diagnostic gene expression profile in CD4(+) T cells, which included the membrane molecule MS4A4B. Accordingly, CD27 increased the frequency of IFN-gamma- and IL-2-producing CD4(+) T cells. It did not affect CD40L expression. Strikingly, MS4A4B was also identified as a unique marker of CD8(+) memory T cells that had received CD27-proficient CD4(+) T cell help during the primary response. This apparent imprinting effect suggests a role for MS4A4B as a downstream effector in CD27-dependent help for CD8(+) T cell memory.     

IL-21 counteracts the regulatory T cell-mediated suppression of human CD4+ T lymphocytes

High expression of IL-21 and/or IL-21R has been described in T cell-mediated inflammatory diseases characterized by defects of counterregulatory mechanisms. CD4(+)CD25(+) regulatory T cells (Treg) are a T cell subset involved in the control of the immune responses. A diminished ability of these cells to inhibit T cell activation has been documented in immune-inflammatory diseases, raising the possibility that inflammatory stimuli can block the regulatory properties of Treg. We therefore examined whether IL-21 controls CD4(+)CD25(+) T cell function. We demonstrate in this study that IL-21 markedly enhances the proliferation of human CD4(+)CD25(-) T cells and counteracts the suppressive activities of CD4(+)CD25(+) T cells on CD4(+)CD25(-) T cells without affecting the percentage of Foxp3(+) cells or survival of Treg. Additionally, CD4(+)CD25(+) T cells induced in the presence of IL-21 maintain the ability to suppress alloresponses. Notably, IL-21 enhances the growth of CD8(+)CD25(-) T cells but does not revert the CD4(+)CD25(+) T cell-mediated suppression of this cell type, indicating that IL-21 makes CD4(+) T cells resistant to suppression rather than inhibiting CD4(+)CD25(+) T cell activity. Finally, we show that IL-2, IL-7, and IL-15, but not IL-21, reverse the anergic phenotype of CD4(+)CD25(+) T cells. Data indicate that IL-21 renders human CD4(+)CD25(-) T cells resistant to Treg-mediated suppression and suggest a novel mechanism by which IL-21 could augment T cell-activated responses in human immune-inflammatory diseases.     

Cross-linking of CD45 on suppressive/regulatory T cells leads to the abrogation of their suppressive activity in vitro

CD4(+)CD25(+) T cells have immunoregulatory and suppressive functions and are responsible for suppressing self-reactive cells and maintaining self-tolerance. In addition to CD4(+)CD25(+) T cells, there is some evidence that a fraction of CD4(+)CD25(-) T cells exhibit suppressive activity in vitro or in vivo. We have shown, using aged mice, that aging not only leads to a decline in the ability to mount CD4(+)CD25(-) T cell responses, but, at the same time, renders aged CD4(+)CD25(-) T cells suppressive. In this study we report two newly established mAbs that could abrogate the suppressive function of aged CD4(+)CD25(-) T cells. These mAbs recognized the same protein, the transmembrane phosphatase CD45. Cross-linking of CD45 on aged CD4(+)CD25(-) T cells was required for the disruption of their suppressive activity. Surprisingly, these mAbs also abrogated the suppressive action of CD4(+)CD25(+) T cells in vitro. Our results demonstrate an unexpected function of CD45 as a negative regulator neutralizing the suppressive activity of aged CD4(+)CD25(-) and young CD4(+)CD25(+) T cells.     

Regulation of murine inflammatory bowel disease by CD25+ and CD25- CD4+ glucocorticoid-induced TNF receptor family-related gene+ regulatory T cells

CD4(+)CD25(+) regulatory T cells in normal animals are engaged in the maintenance of immunological self-tolerance and prevention of autoimmune disease. However, accumulating evidence suggests that a fraction of the peripheral CD4(+)CD25(-) T cell population also possesses regulatory activity in vivo. Recently, it has been shown glucocorticoid-induced TNFR family-related gene (GITR) is predominantly expressed on CD4(+)CD25(+) regulatory T cells. In this study, we show evidence that CD4(+)GITR(+) T cells, regardless of the CD25 expression, regulate the mucosal immune responses and intestinal inflammation. SCID mice restored with the CD4(+)GITR(-) T cell population developed wasting disease and severe chronic colitis. Cotransfer of CD4(+)GITR(+) population prevented the development of CD4(+)CD45RB(high) T cell-transferred colitis. Administration of anti-GITR mAb-induced chronic colitis in mice restored both CD45RB(high) and CD45RB(low) CD4(+) T cells. Interestingly, both CD4(+)CD25(+) and CD4(+)CD25(-) GITR(+) T cells prevented wasting disease and colitis. Furthermore, in vitro studies revealed that CD4(+)CD25(-)GITR(+) T cells as well as CD4(+)CD25(+)GITR(+) T cells expressed CTLA-4 intracellularly, showed anergic, suppressed T cell proliferation, and produced IL-10 and TGF-beta. These data suggest that GITR can be used as a specific marker for regulatory T cells controlling mucosal inflammation and also as a target for treatment of inflammatory bowel disease.     

Small intestine CD11c+ CD8+ T cells suppress CD4+ T cell-induced immune colitis

The large (LI) and small intestine (SI) differ in patterns of susceptibility to chronic mucosal inflammation. In this study, we evaluated whether this might, in part, reflect differences in resident mucosal CD11c(+) T cells. These cells comprised 39-48% (SI) and 12-17% (LI) of the intraepithelial compartment, most of which were T-cell receptor-αβ(+). In the SI, the majority of these cells were CD103(+) CD8(+) NK1.1(-), whereas the opposite phenotype prevailed in the LI. In transfer models of CD4(+) T cell-induced colitis, small numbers (2.5 × 10(5)) of SI CD11c(+) CD8(+) T cells suppressed proinflammatory cytokine-producing CD4(+) T cells in mesenteric lymph nodes and mucosa-associated lymphoid compartments (SI and LI) and protected mice from chronic inflammation. On a per-cell basis, the regulatory function of SI CD11c(+) T cells in CD4(+) T cell colitis was potent compared with other reported regulatory CD4(+) or CD8(+) T cells. In contrast, neither LI CD11c(+) T cells nor SI CD11c(-) T cells were effective in such immunoregulation. SI CD11c(+) CD8(+) T cells were similarly effective in suppressing CD4(+)CD45RB(hi) T cell colitis, as evidenced by inhibition of intracellular proinflammatory cytokine expression and histological inflammation. These findings indicate that SI CD11c(+) CD8(+) T cells are a distinct intestinal T cell population that plays an immunoregulatory role in control of proinflammatory CD4(+) T cells and maintenance of intestinal mucosal homeostasis.     

Homeostasis of peripheral CD4+ T cells: IL-2R alpha and IL-2 shape a population of regulatory cells that controls CD4+ T cell numbers

We show that the lymphoid hyperplasia observed in IL-2Ralpha- and IL-2-deficient mice is due to the lack of a population of regulatory cells essential for CD4 T cell homeostasis. In chimeras reconstituted with bone marrow cells from IL-2Ralpha-deficient donors, restitution of a population of CD25(+)CD4(+) T cells prevents the chaotic accumulation of lymphoid cells, and rescues the mice from autoimmune disease and death. The reintroduction of IL-2-producing cells in IL-2-deficient chimeras establishes a population of CD25(+)CD4(+) T cells, and restores the peripheral lymphoid compartments to normal. The CD25(+)CD4(+) T cells regulated selectively the number of naive CD4(+) T cells transferred into T cell-deficient hosts. The CD25(+)CD4(+)/naive CD4 T cell ratio and the sequence of cell transfer determines the homeostatic plateau of CD4(+) T cells. Overall, our findings demonstrate that IL-2Ralpha is an absolute requirement for the development of the regulatory CD25(+)CD4(+) T cells that control peripheral CD4 T cell homeostasis, while IL-2 is required for establishing a sizeable population of these cells in the peripheral pools.     

Activation of CD25(+)CD4(+) regulatory T cells by oral antigen administration

CD25(+)CD4(+) T cells are naturally occurring regulatory T cells that are anergic and have suppressive properties. Although they can be isolated from the spleens of normal mice, there are limited studies on how they can be activated or expanded in vivo. We found that oral administration of OVA to OVA TCR transgenic mice resulted in a modification of the ratio of CD25(+)CD4(+) to CD25(-)CD4(+) cells with an increase of CD25(+)CD4(+) T cells accompanied by a decrease of CD25(-)CD4(+) T cells. The relative increase in CD25(+)CD4(+) T cells persisted for as long as 4 wk post feeding. We also found that CTLA-4 was dominantly expressed in CD25(+)CD4(+) T cells and there was an increase in the percentage of CD25(+)CD4(+) T cells expressing CTLA-4 in OVA-fed mice. In contrast to CD25(-)CD4(+) cells, CD25(+)CD4(+) cells from fed mice proliferated only minimally to OVA or anti-CD3 and secreted IL-10 and elevated levels of TGF-beta(1) following anti-CD3 stimulation. CD25(+)CD4(+) cells from fed mice suppressed the proliferation of CD25(-)CD4(+) T cells in vitro more potently than CD25(+)CD4(+) T cells isolated from unfed mice, and this suppression was partially reversible by IL-10 soluble receptor or TGF-beta soluble receptor and high concentration of anti-CTLA-4. With anti-CD3 stimulation, CD25(+)CD4(+) cells from unfed mice secreted IFN-gamma, whereas CD25(+)CD4(+) cells from fed mice did not. Adoptive transfer of CD25(+)CD4(+) T cells from fed mice suppressed in vivo delayed-type hypersensitivity responses in BALB/c mice. These results demonstrate an Ag-specific in vivo method to activate CD25(+)CD4(+) regulatory T cells and suggest that they may be involved in oral tolerance.     

CD4+/CD25+ regulatory cells inhibit activation of tumor-primed CD4+ T cells with IFN-gamma-dependent antiangiogenic activity, as well as long-lasting tumor immunity elicited by peptide vaccination

CD25(+) regulatory T (T reg) cells suppress the activation/proliferation of other CD4(+) or CD8(+) T cells in vitro. Also, down-regulation of CD25(+) T reg cells enhance antitumor immune responses. In this study, we show that depletion of CD25(+) T reg cells allows the host to induce both CD4(+) and CD8(+) antitumoral responses following tumor challenge. Simultaneous depletion of CD25(+) and CD8(+) cells, as well as adoptive transfer experiments, revealed that tumor-specific CD4(+) T cells, which emerged in the absence of CD25(+) T reg cells, were able to reject CT26 colon cancer cells, a MHC class II-negative tumor. The antitumoral effect mediated by CD4(+) T cells was dependent on IFN-gamma production, which exerted a potent antiangiogenic activity. The capacity of the host to mount this antitumor response is lost once the number of CD25(+) T reg cells is restored over time. However, CD25(+) T reg cell depletion before immunization with AH1 (a cytotoxic T cell determinant from CT26 tumor cells) permits the induction of a long-lasting antitumoral immune response, not observed if immunization is conducted in the presence of regulatory cells. A study of the effect of different levels of depletion of CD25(+) T reg cells before immunization with the peptide AH1 alone, or in combination with a Th determinant, unraveled that Th cells play an important role in overcoming the suppressive effect of CD25(+) T reg on the induction of long-lasting cellular immune responses.     

CD4(+) T cells are required for the development of cytotoxic CD8(+) T cells during Mycobacterium tuberculosis infection.

The control of acute and chronic Mycobacterium tuberculosis infection is dependent on CD4(+) T cells. In a variety of systems CD8(+) T cell effector responses are dependent on CD4(+) T cell help. The development of CD8(+) T cell-mediated immune responses in the absence of CD4(+) T cells was investigated in a murine model of acute tuberculosis. In vitro and in vivo, priming of mycobacteria-specific CD8(+) T cells was unaffected by the absence of CD4(+) T cells. Infiltration of CD8(+) T cells into infected lungs of CD4(-/-) or wild-type mice was similar. IFN-gamma production by lung CD8(+) T cells in CD4(-/-) and wild-type mice was also comparable, suggesting that emergence of IFN-gamma-producing mycobacteria-specific CD8(+) T cells in the lungs was independent of CD4(+) T cell help. In contrast, cytotoxic activity of CD8(+) T cells from lungs of M. tuberculosis-infected mice was impaired in CD4(-/-) mice. Expression of mRNA for IL-2 and IL-15, cytokines critical for the development of cytotoxic effector cells, was diminished in the lungs of M. tuberculosis-infected CD4(-/-) mice. As tuberculosis is frequently associated with HIV infection and a subsequent loss of CD4(+) T cells, understanding the interaction between CD4(+) and CD8(+) T cell subsets during the immune response to M. tuberculosis is imperative for the design of successful vaccination strategies.     

CD4+ Th1 and CD8+ type 1 cytotoxic T cells both play a crucial role in the full development of contact hypersensitivity

The role of CD4(+) vs CD8(+) T cells in contact hypersensitivity (CHS) remains controversial. In this study, we used gene knockout (KO) mice deficient in CD4(+) or CD8(+) T cells to directly address this issue. Mice lacking either CD4(+) or CD8(+) T cells demonstrated depressed CHS responses to dinitrofluorobenzene and oxazolone compared with wild-type C57BL/6 mice. The depression of CHS was more significant in CD8 KO mice than in CD4 KO mice. Furthermore, in vivo depletion of either CD8(+) T cells from CD4 KO mice or CD4(+) T cells from CD8 KO mice virtually abolished CHS responses. Lymph node cells (LNCs) from hapten-sensitized CD4 and CD8 KO mice showed a decreased capacity for transferring CHS. In vitro depletion of either CD4(+) T cells from CD8 KO LNCs or CD8(+) T cells from CD4 KO LNCs resulted in a complete loss of CHS transfer. LNCs from CD4 and CD8 KO mice produced significant amounts of IFN-gamma, indicating that both CD4(+) and CD8(+) T cells are able to secrete IFN-gamma. LNCs from CD8, but not CD4, KO mice were able to produce IL-4 and IL-10, suggesting that IL-4 and IL-10 are mainly derived from CD4(+) T cells. Intracellular cytokine staining of LNCs confirmed that IFN-gamma-positive cells consisted of CD4(+) (Th1) and CD8(+) (type 1 cytotoxic T) T cells, whereas IL-10-positive cells were exclusively CD4(+) (Th2) T cells. Collectively, these results suggest that both CD4(+) Th1 and CD8(+) type 1 cytotoxic T cells are crucial effector cells in CHS responses to dinitrofluorobenzene and oxazolone in C57BL/6 mice.     

Enhanced neointima formation following arterial injury in immune deficient Rag-1-/- mice is attenuated by adoptive transfer of CD8 T cells

T cells modulate neointima formation after arterial injury but the specific T cell population that is activated in response to arterial injury remains unknown. The objective of the study was to identify the T cell populations that are activated and modulate neointimal thickening after arterial injury in mice. Arterial injury in wild type C57Bl6 mice resulted in T cell activation characterized by increased CD4(+)CD44(hi) and CD8(+)CD44(hi) T cells in the lymph nodes and spleens. Splenic CD8(+)CD25(+) T cells and CD8(+)CD28(+) T cells, but not CD4(+)CD25(+) and CD4(+)CD28(+) T cells, were also significantly increased. Adoptive cell transfer of CD4(+) or CD8(+) T cells from donor CD8-/- or CD4-/- mice, respectively, to immune-deficient Rag-1-/- mice was performed to determine the T cell subtype that inhibits neointima formation after arterial injury. Rag-1-/- mice that received CD8(+) T cells had significantly reduced neointima formation compared with Rag-1-/- mice without cell transfer. CD4(+) T cell transfer did not reduce neointima formation. CD8(+) T cells from CD4-/- mice had cytotoxic activity against syngeneic smooth muscle cells in vitro. The study shows that although both CD8(+) T cells and CD4(+) T cells are activated in response to arterial injury, adoptive cell transfer identifies CD8(+) T cells as the specific and selective cell type involved in inhibiting neointima formation.     

T cells with a CD4+CD25+ regulatory phenotype suppress in vitro proliferation of virus-specific CD8+ T cells during chronic hepatitis C virus infection

Chronic hepatitis C virus (HCV) infection is associated with impaired proliferative, cytokine, and cytotoxic effector functions of HCV-specific CD8(+) T cells that probably contribute significantly to viral persistence. Here, we investigated the potential role of T cells with a CD4(+)CD25(+) regulatory phenotype in suppressing virus-specific CD8(+) T-cell proliferation during chronic HCV infection. In vitro depletion studies and coculture experiments revealed that peptide specific proliferation as well as gamma interferon production of HCV-specific CD8(+) T cells were inhibited by CD4(+)CD25(+) T cells. This inhibition was dose dependent, required direct cell-cell contact, and was independent of interleukin-10 and transforming growth factor beta. Interestingly, the T-cell-mediated suppression in chronically HCV-infected patients was not restricted to HCV-specific CD8(+) T cells but also to influenza virus-specific CD8(+) T cells. Importantly, CD4(+)CD25(+) T cells from persons recovered from HCV infection and from healthy blood donors exhibited significantly less suppressor activity. Thus, the inhibition of virus-specific CD8(+) T-cell proliferation was enhanced in chronically HCV-infected patients. This was associated with a higher frequency of circulating CD4(+)CD25(+) cells observed in this patient group. Taken together, our results suggest that chronic HCV infection leads to the expansion of CD4(+)CD25(+) T cells that are able to suppress CD8(+) T-cell responses to different viral antigens. Our results further suggest that CD4(+)CD25(+) T cells may contribute to viral persistence in chronically HCV-infected patients and may be a target for immunotherapy of chronic hepatitis C.     

Functional characterization of MHC class II-restricted CD8+CD4- and CD8-CD4- T cell responses to infection in CD4-/- mice

Classical CD4(+) and CD8(+) T cells recognize Ag presented by MHC class II (MHCII) and MHC class I (MHCI), respectively. However, our results show that CD4(-/-) mice mount a strong, readily detectable CD8(+) T cell response to MHCII-restricted epitopes after a primary bacterial or viral infection. These MHCII-restricted CD8(+)CD4(-) T cells are more similar to classical CD8(+) T cells than to CD4(+) T cells in their expression of effector functions during a primary infection, yet they also differ from MHCI-restricted CD8(+) T cells by their inability to produce high levels of the cytolytic molecule granzyme B. After resolution of a primary infection, epitope-specific MHCII-restricted T cells in CD4(-/-) mice persist for a long period of time as memory T cells. Surprisingly, upon reinfection the secondary MHCII-restricted response in CD4(-/-) mice consists mainly of CD8(-)CD4(-) T cells. In contrast to CD8(+) T cells, MHCII-restricted CD8(-)CD4(-) T cells are capable of producing IL-2 in addition to IFN-gamma and thus appear to have attributes characteristic of CD4(+) T cells rather than CD8(+) T cells. Therefore, MHCII-restricted T cells in CD4(-/-) mice do not share all phenotypic and functional characteristics with MHCI-restricted CD8(+) T cells or with MHCII-restricted CD4(+) T cells, but, rather, adopt attributes from each of these subsets. These results have implications for understanding thymic T cell selection and for elucidating the mechanisms regulating the peripheral immune response and memory differentiation.     

Cognate memory CD4+ T cells generated with dendritic cell priming influence the expansion, trafficking, and differentiation of secondary CD8+ T cells and enhance tumor control

CD4(+) T cells are known to provide support for the activation and expansion of primary CD8(+) T cells, their subsequent differentiation, and ultimately their survival as memory cells. However, the importance of cognate memory CD4(+) T cells in the expansion of memory CD8(+) T cells after re-exposure to Ag has been not been examined in detail. Using bone marrow-derived dendritic cells pulsed with cognate or noncognate MHC class I- and class II-restricted peptides, we examined whether the presence of memory CD4(+) T cells with the same Ag specificity as memory CD8(+) T cells influenced the quantity and quality of the secondary CD8(+) T cell response. After recombinant vaccinia virus-mediated challenge, we demonstrate that, although cognate memory CD4(+) T cells are not required for activation of secondary CD8(+) T cells, their presence enhances the expansion of cognate memory CD8(+) T cells. Cognate CD4(+) T cell help results in an approximate 2-fold increase in the frequency of secondary CD8(+) T cells in secondary lymphoid tissues, and can be accounted for by enhanced proliferation in the secondary CD8(+) T cell population. In addition, cognate memory CD4(+) T cells further selectively enhance secondary CD8(+) T cell infiltration of tumor-associated peripheral tissue, and this is accompanied by increased differentiation into effector phenotype within the secondary CD8(+) T cell population. The consequence of these improvements to the magnitude and phenotype of the secondary CD8(+) T cell response is substantial increase in control of tumor outgrowth.