Marked anti-tumor effects of CD8+CD62L+ T cells from melanoma-bearing mice

CD8⁺CD62L⁺ T cells have been shown to play pivotal roles in anti-viral immunity, chronic myeloid leukemia and renal cell carcinoma. Recently, CD8⁺CD62L⁺ T cells from naïve mice (nCD8⁺CD62L⁺ T cells) have shown superior anti-tumor properties in melanoma-bearing mice. Considering that antigen-specific memory T cells have shown to possess more potent immunity than non-specific memory T cells, we hypothesized that CD8⁺CD62L⁺ T cells from tumor-bearing individuals (mCD8⁺CD62L⁺ T cells) might have superior anti-tumor effect than nCD8⁺CD62L⁺ T cells. Therefore, we investigated phenotypes, functions and the in vivo distribution of mCD8⁺CD62L⁺ T cells in tumor-bearing mice. We found that, while keeping the features of central memory T cells, the frequency of mCD8⁺CD62L⁺ T cell in the spleen of tumor-bearing mice was significantly higher than that the one of nCD8⁺CD62L⁺ T cell in naive mice. Moreover, we demonstrated that mCD8⁺CD62L⁺ T cells had higher proliferation rate and IFN-γ production than nCD8⁺CD62L⁺ T cells, in vitro. We performed adoptive transfer of mCD8⁺CD62L⁺ T cells into melanoma-bearing mice and tracked them in spleen, lymph nodes and in melanoma tissues. Our results show that mCD8⁺CD62L⁺ T cells had stronger in vivo anti-tumoral activity than nCD8⁺CD62L⁺ T cells. This study highlights the therapeutic potential of mCD8⁺CD62L⁺ T cells in the immunotherapy of melanoma and possibly other tumors.     

Impact of mTOR signaling pathway on CD8+ T cell immunity through Eomesodermin in response to invasive candidiasis

BackgroundWe investigated the effect of the mammalian target of rapamycin (mTOR) pathway on CD8+ T cell immunity through Eomesodermin (Eomes) in intensive care unit (ICU) patients with invasive candidiasis (IC) and in a mouse model.MethodsWe evaluated quantitative changes in parameters of the mTOR/phosphorylated ribosomal S6 kinase (pS6K) pathway and immune system at the onset of infection in ICU patients. The study was registered on 28 February 2017 at chictr.org.cn (ChiCTR-ROC-17010750). We also used a mouse model of Candida infection and constructed T-cell-specific mTOR and T-cell-specific tuberous sclerosis complex (TSC) 1 conditional knockout mice to elucidate the molecular mechanisms.ResultsWe enrolled 88 patients, including 8 with IC. The IC group had lower CD8+ T cell counts, higher serum levels of mTOR, pS6K, Eomes and interleukin (IL)-6. The mouse model with IC showed results consistent in the clinical study. The CD8+ T cell immune response to IC seemed to be weakened in TSC1 knockout mice compared with wild-type IC mice, demonstrating that mTOR activation resulted in the impaired CD8+ T cell immunity in IC.ConclusionsIn IC, the mTOR activation may play a vital role in impaired CD8+ T cell immunity through enhancing expression of Eomes.The study was registered on 28 February 2017 at chictr.org.cn (identifier ChiCTR-ROC-17010750).     

Anomalies of the CD8+ T cell pool in haemochromatosis: HLA-A3-linked expansions of CD8+CD28− T cells

The present study consists of a phenotypic and functional characterization of peripheral blood T lymphocytes in a group of 21 patients with hereditary haemochromatosis (HH), an MHC class I-linked genetic disease resulting in iron overload, and a group of 30 healthy individuals, both HLA-phenotyped. The HH patients studied showed an increased percentage of CD8⁺ CD28⁻ T cells with a corresponding reduction in the percentage of CD8⁺ CD28⁺ T cells in peripheral blood relative to healthy blood donors. No anomalies of CD28 expression were found in the CD4⁺ subset. The presence of the HLA-A3 antigen but not age accounted for these imbalances. Thus, an apparent failure of the CD8⁺ CD28⁺ T cell population ‘to expand’, coinciding with an ‘expansion’ of CD8⁺ CD28⁻ T cells in peripheral blood of HLA-A3⁺ but not HLA-A3⁻ HH patients was observed when compared with the respective HLA-A3-matched control group. A significantly higher percentage of HLA-DR⁺ but not CD45RO⁺ cells was also found within the peripheral CD8⁺ T cell subset in HH patients relative to controls. Phytohaemagglutinin (PHA) stimulation of peripheral blood mononuclear cells (PBMC) for 5 days showed: (i) that CD8⁺ CD28⁺ T cells both in controls and HH were able to expand in vitro; (ii) that CD8⁺ CD28⁻ T cells decreased markedly after activation in controls but not in HH patients. Moreover, functional studies showed that CD8⁺ cytotoxic T lymphocytes (CTL) from HH patients exhibited a diminished cytotoxic activity (approx. two-fold) in standard ⁵¹Cr-release assays when compared with CD8⁺ CTL from healthy controls. The present results provide additional evidence for the existence of phenotypic and functional anomalies of the peripheral CD8⁺ T cell pool that may underlie the clinical heterogeneity of this iron overload disease. They are of particular relevance given the recent discovery of a novel mutated MHC class I-like gene in HH.     

High-avidity CD8+ T cells

The primary goal of vaccination is the establishment of protective immunity. Thus there has been significant effort put toward the identification of attributes of the immune response that are associated with optimal protection. Although the number of virus-specific cells elicited is unquestionably important, recent studies have identified an additional parameter, functional avidity, as critical in determining the efficiency of viral clearance. T-cell avidity is a measure of the sensitivity of a cell to peptide antigen. High-avidity cells are those that can recognize antigen-presenting cells (APC) bearing very low levels of peptide antigen, whereas low-avidity cells require much higher numbers of peptide major histocompatibility complex (MHC) complexes in order to become activated or exert effector function. We are only now beginning to gain insights into the molecular control of avidity and the signals required for the optimal activation, expansion, and retention of high-avidity cells in vivo. This review summarizes the current knowledge regarding CD8⁺ T-cell avidity and explores some of the important issues that are, as of yet, unresolved.     

Thymic stromal lymphopoietin plays an adjuvant role in BCG-mediated CD8 cytotoxic T cell responses through dendritic cell activation

Although Bacillus Calmette–Guérin (BCG) has historically emerged as a potent adjuvant in cancer immunization through dendritic cell (DC) activation, the efficacy of its antitumor effect has been limited. Therefore, the strategy of adjuvant therapy using BCG needs to be improved by adding enhancers. Here we found that thymic stromal lymphopoietin (TSLP) acts as an enhancer for the BCG-mediated antitumor effect. While BCG-stimulated DCs induced CD8⁺ T cell production of IFN-γ without strong cell expansion, TSLP-stimulated DCs induced robust CD8⁺ T cell expansion without high quantities of IFN-γ production. Notably, DCs stimulated with both BCG and TSLP induced robust expansion of CD8⁺ T cells that produced a large amount of IFN-γ with a potent cytolytic activity related to granzyme B expression. Our data suggest that TSLP is a good adjuvant to enhance the BCG-mediated cytotoxic T cell effect through DC activation, and provide a functional basis for a novel strategy for antitumor immune-based therapy.     

CD8+ T cells and not CD4+ T cells are hyporesponsive to CD28- and CD40L-mediated activation in HIV-infected subjects

T cell dysfunction in HIV-infected subjects could be the consequence of altered sensitivity of CD4⁺ or CD8⁺ T cells to various costimulatory signals. Therefore, we studied proliferation and cytokine production in highly purified CD8⁺ and CD4⁺ T cells from HIV-infected and HIV⁻ subjects, induced by co-activation via cell-bound CD80, CD86 and CD40 or by allo-activation. Regardless of the nature of the first and the costimulatory signal, CD8⁺ T cells from patients proliferated consistently less than controls, while responses from CD4⁺ T cells were similar in patients and controls. This phenomenon was observed after ligation of CD28 combined with anti-CD3 or phorbol myristate acetate (PMA), but also after allogeneic stimulation and after activation by CD40 and anti-CD3. Anti-CD3 combined with CD80 or CD86 induced a mixed Th1/Th2-type cytokine profile in both CD4⁺ and CD8⁺ T cells from controls, whereas anti-CD3 plus CD40 induced only low levels of Th2-type cytokines and no interferon-gamma (IFN-γ) in CD4⁺ T cells. Compared with controls, CD4⁺ T cells from patients produced slightly lower levels of IL-10 but equal amounts of IFN-γ, IL-4 and IL-5, while CD8⁺ T cells from patients produced less of all cytokines tested. In conclusion, responses of purified CD4⁺ T cells from HIV⁺ subjects to various costimulatory pathways are relatively intact, whereas CD8⁺ T cells are hyporesponsive at the level of proliferation and cytokine production. A generalized intrinsic CD8⁺ T cell failure might contribute to viral and neoplastic complications of HIV infection.     

CD4+ T cell-independent maintenance and expansion of memory CD8+ T cells derived from in vitro dendritic cell activation

CD4+ T cells are essential for the maintenance of CD8+ memory T (Tm) cells following acute infection, but the importance of CD4+ T cells for the maintenance and expansion of CD8+ Tm cells to non-infectious antigens remains mostly unknown. Here, we showed that ovalbumin (OVA)-specific CD8+ Tm cell precursors derived from in vitro stimulation of TCR transgenic OT I CD8+ T cells with OVA protein-pulsed bone marrow-derived dendritic cells (DCOVA) can give rise to functional CD8+ Tm cells after adoptively transferred into mice. These CD8+ Tm cells can be maintained and remain fully functional in CD4+ T cell-absent environments in vivo. Furthermore, CD4+ T cells are not essential for the expansion of these CD8+ Tm cells. Finally, these in vitro DCOVA-activated CD8+ Tm cells maintained in CD4-deficient mice are also able to confer fully protective immunity against a later challenge of OVA-expressing tumor cells. Collectively, these findings demonstrate that in contrast to acute infections, maintenance and expansion of CD8+ Tm cells after priming with OVA protein-pulsed dendritic cells are independent of CD4+ T cells.     

CD4+CD25+ T regulatory cells inhibit cytotoxic activity of T CD8+ and NK lymphocytes in the direct cell-to-cell interaction

There are reports suggesting an influence of CD4(+)CD25(+) T regulatory cells (Treg) on cytotoxic lymphocytes. The aim of the study was to evaluate such an influence. Cytotoxic activity was examined in the cultures of peripheral blood mononuclear cells (PBMC) as well as in the cultures of separate T CD8(+) or NK cells mixed with Treg and other subpopulations of PBMC. We found that the production of IFNgamma, perforin and cytotoxic activity of T CD8(+) or NK cells were decreased in the presence of Treg, however, the percentage of conjugates formed by cytotoxic cells with target cells during cytotoxic reaction was decreased only in the cultures of T CD8(+) cells. Inhibition of the cytotoxic reactions in the presence of Treg cells was found to be associated with the generation of conglomerates formed by CD4(+)CD25(+) and the cytotoxic cells, as observed under the fluorescence microscope. Treg produced IL10 when mixed with the cytotoxic lymphocytes, however, an addition of anti-IL10 mAb into the cultures did not affect the results. It is concluded that Treg were able to inhibit both T CD8+ and NK lymphocyte cytotoxic activities in a direct cell-to-cell interaction. Treg decreased the number of T CD8+ cells attached to the target cells, while the mechanism underlying a decrease in NK cytotoxicity remained unclear.     

A role for dendritic cells in the priming of antigen-specific CD4+ and CD8+ T lymphocytes by immune-stimulating complexes in vivo

Immune-stimulating complexes (ISCOMS) are adjuvant vectors which are unusual in being able to prime both CD4(+) and CD8(+) T cells by parenteral and mucosal routes. However, their mode of action is unclear and to define better the cellular interactions involved we have studied the ability of ISCOMS containing ovalbumin (OVA) to prime TCR transgenic CD4(+) or CD8(+) T cells in vivo. Immunization with OVA ISCOMS caused activation and clonal expansion of CD4(+) and CD8(+) T cells in the T cell areas of the draining lymph nodes, followed by the migration of both CD4(+) and CD8(+) T cells into the B cell follicle. The T cells were primed to proliferate and secrete IFN-gamma after re-stimulation in vitro with the appropriate OVA peptide and CD8(+) T cell priming occurred in the absence of CD4(+) T cells. Increasing the number of dendritic cells (DC) in vivo with flt3 ligand augmented the expansion and activation of the OVA-specific T cells, particularly CD8(+) T cells. These studies indicate DC play a central role in the priming of both CD4(+) and CD8(+) T cells in vivo, and suggest that an ability to target DC may allow ISCOMS to be powerful vaccine vectors for stimulating protective immunity.     

LIGHT-deficiency impairs CD8+ T cell expansion,but not effector function

LIGHT, a newly identified member of the tumor necrosis factor (TNF) family, is expressed on activated T lymphocytes. To evaluate how LIGHT contributes to T cell functions, we generated LIGHT-deficient (LIGHT(-/-)) mice using gene targeting. Disruption of LIGHT significantly reduced CD8(+) T cell-cycle progression, leading to reduced proliferation to anti-CD3, anti-CD3/anti-CD28 or allogeneic stimulation, whereas proliferation of CD4(+) T cells remained unchanged. In contrast to the observed proliferative defects, isolated CD8(+) T cells from LIGHT(-/-) mice displayed normal cytotoxic effector function development when compared to wild-type CD8(+) T cells. Underlying a potential mechanism of reduced CD8(+) T cell proliferation, LIGHT(-/-) CD8(+) T cells displayed reduced surface levels of CD25 and a diminished ability to proliferate in response to exogenous IL-2. Furthermore, addition of IL-12 to LIGHT(-/-) CD8(+) T cell cultures could not ameliorate this proliferative defect. These results reveal a potential mechanism of action for LIGHT as a positive regulator of CD8(+) T cell expansion, but not lytic effector function development.     

Impaired CD4 and CD8 T cell phenotype and reduced chemokine secretion in recent-onset type 1 diabetic children

Although the role of the T cell-mediated autoimmune reaction in type 1 diabetes (T1D) is conclusive, studies including data from human circulating CD4(+) and CD8(+) lymphocytes subsets during the disease onset and posterior development are scarce. Further, chemokines and chemokine receptors are key players in the migration of pathogenic T cells into the islets of non-obese diabetic mice developing T1D, but few studies have investigated these markers in human T1D patients. We studied the expression of T helper 1 (Th1)- and Th2-associated chemokine receptors, and the two isoforms of CD45 leucocyte antigen on CD4(+) and CD8(+) lymphocytes from T1D and healthy children, as well as the secretion of chemokines in cell supernatants in peripheral blood mononuclear cells. Our results showed increased expression of CCR7 and CD45RA and reduced CD45RO on CD8(+) cells among recent-onset T1D patients. The percentages of CD4(+) cells expressing CXC chemokine receptor 3 (CXCR3), CXCR6 and CCR5, and the secretion of interferon-gamma-induced protein-10, monocyte chemoattractant protein-1, macrophage inflammatory protein (MIP)-1alpha and MIP-1beta was lower among diabetics. Low expression of Th1-associated receptors and secretion of chemokines, together with an increased amount of CD8(+) cells expressing CD45RA and CCR7 in T1D patients therefore might represent suboptimal Th function in T1D, leading to impaired T cytotoxic responses or alternatively reflect a selective recruitment of Th1 cells into the pancreas.     

4-1BB co-stimulation enhances human CD8(+) T cell priming by augmenting the proliferation and survival of effector CD8(+) T cells

Interactions between 4-1BB and its ligand, 4-1BBL, enhance CD8(+) T cell-mediated antiviral and antitumor immunity in vivo. However, mechanisms regulating the priming of CD8(+) T cell responses by 4-1BB remain unclear, particularly in humans. The 4-1BB receptor was undetectable on naive or resting human CD8(+) T cells and induced in vitro by TCR triggering. Naive cord blood cells were therefore primed in vitro against peptides or cellular antigens and then co-stimulated with 4-1BBL or agonistic antibodies. Co-stimulation enhanced effector function such as IFN-gamma production and cytotoxicity by augmenting numbers of antigen-specific and effector CD8(+) T cells. OKT3 responses also showed reduced cell death and revealed that the proliferation of CD8(+) T cells required two independently regulated events. One, the induction of IL-2 production, could be directly triggered by 4-1BB engagement on CD8(+) T cells in the absence of accessory cells. The other, expression of CD25, was induced with variable efficacy by accessory cells. Thus, suboptimal accessory cells and 4-1BB co-stimulation combined their effects to enhance IL-2 production and proliferation. Reduced apoptosis observed after co-stimulation in the presence of accessory cells correlated with increased levels of Bcl-X(L) in CD8(+) T cells, while Bcl-2 expression remained unchanged. Altogether, 4-1BB enhanced expansion, survival and effector functions of newly primed CD8(+) T cells, acting in part directly on these cells. As 4-1BB triggering could be protracted from the TCR signal, 4-1BB agonists may function through these mechanisms to enhance or rescue suboptimal immune responses.     

Relative importance of CD4+ and CD8+ T cells in the resolution of Chlamydophila abortus primary infection in mice

The role of the specific cellular immune response is well established in Chlamydiaceae infections, but the importance of each T-cell subset seems to be species-dependent. This study was designed to clarify the role of T-cell subsets in the response to Chlamydophila abortus primary infection. C57BL/6 mice were depleted of CD4+ or CD8+, or both, by monoclonal antibody injections and subsequently infected with C. abortus. Mice were killed at intervals and samples were collected for bacteriological and histopathological analysis. Also carried out were spleen cell culture, cytokine quantification, immunolabelling for C. abortus antigen, and a TUNEL assay for apoptosis. CD8+ T cell-depleted mice all died within 12 days of C. abortus infection, while no mortality was observed in the other groups; surprisingly, CD4+ T cell-depleted mice showed lower morbidity (expressed as weight loss) than did a non-depleted (control) group. CD8+ T cell-depleted mice also differed from the other groups in showing a significantly higher chlamydial burden in the liver. CD8+ T cell-depleted mice also had a higher number of apoptotic cells in hepatic inflammatory foci and showed exacerbated IFN-gamma production by spleen cells after specific stimulation. Simultaneous depletion of both T-cell subpopulations led to a chronic infection, but not to early mortality. It is concluded that CD8+ T cells may play a role in the regulatory control of the CD4+ T-cell response and may have a direct cytotoxic or IFN-gamma-mediated effect on infected cells.     

Analysis of the immune response of Hantaan virus nucleocapsid protein-specific CD8+ T cells in mice

The major histocompatibility complex (MHC) class-I restricted epitope of Hantaan virus nucleocapsid protein (N) was identified using overlapping peptides and BALB/c mice. Using the MHC tetramer derived from the epitope, we found that the level of N-specific CD8(+) T cells increased to approximately 20% of all antigen-specific CD8(+) T cells in a mouse model of transient infection. However, N-specific CD8(+) T cells were undetectable in a mouse model of persistent infection, both in the persistently infected phase and in the convalescent phase. Levels of CD8(+) T cells producing interferon-gamma were weak in both the acute and convalescent phases in the persistently infected model. These results indicate that hantavirus strongly suppresses the production of N-specific CD8(+) T cells throughout the course of infection in persistently infected mice. Moreover, N-specific CD8(+) T cells were not effective in recovering persistently infected mice, despite the existence of abundant N antigen in vivo.     

Attenuation of Peripheral Regulatory T-Cell Suppression of Skin-Homing CD8+T Cells in Atopic Dermatitis

Purpose

Cutaneous lymphocyte-associated antigen (CLA)-expressing CD8⁺T cells have been known to play an important role in the pathogenesis of atopic dermatitis (AD). However, the mechanisms underlying the loss of self-tolerance remain unclear. Regulatory T cells (Tregs) play a key role in the development of homeostasis in the immune system. We, therefore, hypothesized that a reduced ability of Tregs to inhibit autologous CD8⁺CLA⁺T cells might be underlying mechanism in AD.

Materials and Methods

CD8⁺CLA⁺T cells and Tregs were obtained from the peripheral blood of AD patients and control volunteers. The frequencies of CD8⁺CLA⁺T cells were evaluated. The proliferative responses of CD8⁺CLA⁺T cells were assessed by flow cytometry, and the levels of transforming growth factor-β1 (TGF-β1) and interleukin-10 (IL-10) in culture supernatants were detected by enzyme-linked immunosorbent assay.

Results

Our results revealed higher frequency and increased expression of perforin and granzyme-B in peripheral CD8⁺CLA⁺T cells in AD, and lower inhibitory ability of Tregs on proliferation of CD8⁺CLA⁺T cells in AD. Meanwhile, the levels of TGF-β1 produced by Tregs were significantly lower in AD, and anti-TGF-β1 abolished such suppression.

Conclusion

The attenuated inhibitory ability of Tregs on hyper-activated autologous CD8⁺CLA⁺T cells, mediated by TGF-β1, plays an important role in the pathogenesis of AD.     

Epigallocatechin gallate attenuates adhesion and migration of CD8+ T cells by binding to CD11b

BACKGROUND: Although green tea polyphenol catechin has been reported to have antiallergic and anti-inflammatory activities, the precise mechanisms of its effect on the immune system have been poorly investigated. OBJECTIVE: In this study, we aimed to elucidate the mechanisms of the anti-inflammatory effect of catechin. For this purpose, we studied the effect of 2 kinds of catechin, epigallocatechin gallate (EGCG) and epicatechin gallate, on peripheral blood CD8+ T cells, which play the key role in immune responses. METHODS: Isolated peripheral blood mononuclear cells or CD8+ T cells were incubated without or with catechin, and the changes in the surface expression of integrin molecules were investigated by flow cytometry and the direct binding of catechin to CD11b molecule by competitive ELISA. Also, the effect of catechin on the ability of CD8+ T cells to bind intracellular adhesion molecule 1 and to migrate in response to chemokines was evaluated by using the adhesion and migration assays. RESULTS: The 2 catechins directly bound to CD11b expressed on CD8+ T cells, which caused a consequent decrease of flow-cytometric CD11b expression. The effect was more prominent with EGCG than epicatechin gallate, and the impaired expression of CD11b induced by EGCG resulted in decreased ability of CD8+ T cells to adhere intercellular adhesion molecule 1, and consequently decreased migration in response to chemokines. CONCLUSION: We concluded that catechin, especially EGCG, by downregulating CD11b expression on CD8+ T cells and, in consequence, inhibiting infiltration of these cells into the sites of inflammation, is a promising new potent anti-inflammatory agent.     

Human TSLP directly enhances expansion of CD8+ T cells

Human thymic stromal lymphopoietin (TSLP) promotes CD4(+) T-cell proliferation both directly and indirectly through dendritic cell (DC) activation. Although human TSLP-activated DCs induce CD8(+) T-cell proliferation, it is not clear whether TSLP acts directly on CD8(+) T cells. In this study, we show that human CD8(+) T cells activated by T-cell receptor stimulation expressed TSLP receptor (TSLPR), and that TSLP directly enhanced proliferation of activated CD8(+) T cells. Although non-stimulated human CD8(+) T cells from peripheral blood did not express TSLPR, CD8(+) T cells activated by anti-CD3 plus anti-CD28 did express TSLPR. After T-cell receptor stimulation, TSLP directly enhanced the expansion of activated CD8(+) T cells. Interestingly, using monocyte-derived DCs pulsed with a cytomegalovirus (CMV)-specific pp65 peptide, we found that although interleukin-2 allowed expansion of both CMV-specific and non-specific CD8(+) T cells, TSLP induced expansion of only CMV-specific CD8(+) T cells. These results suggest that human TSLP directly enhances expansion of CD8(+) T cells and that the direct and indirect action of TSLP on expansion of target antigen-specific CD8(+) T cells may be beneficial to adoptive cell transfer immunotherapy.