Origin of CD8^＋ Effector and Memory T Cell Subsets

It is well accepted that CD8＋ T cells play a pivotal role in providing protection against infection with intracellular pathogens and some tumors. In many cases protective immunity is maintained for long periods of time （immunological memory）. Over the past years, it has become evident that in order to fulfill these multiple tasks, distinct subsets of effector and memory T cells have to be generated. Until today, however, little is known about the underlying mechanisms of subset differentiation and the timing of lineage fate decisions. In this context, it is of special importance to determine at which level of clonal expansion functional and phenotypical heterogeneity is achieved. Different models for T cell subset diversification have been proposed; these differ mainly in the time point during priming and clonal expansion （prior, during, or beyond the first cell division） when differentiation programs are induced. Recently developed single-cell adoptive transfer technology has allowed us to demonstrate that individual precursor cell still bears the full plasticity to develop into a plethora different T cell subsets. This observation targets the shaping of T cell subset differentiation towards factors that are still operative beyond the first cell division. These findings have important implications for vaccine development, as the modulation of differentiation patterns towards distinct subsets could become a powerful strategy to enhance the efficacy and quality of vaccines. Cellular ＆ Molecular Immunology.     

Defect of CD8^＋ Memory T Cells Developed in Absence of IL-12 Priming for Secondary Expansion

IL-12 priming plays an important role in stimulation of CD8^＋ effector T cells and development of CD8^＋ memory T （Tm） cells. However, the functional alteration of CD8^＋ Tm cells developed in the absence of IL-12 priming is elusive. In this study, we investigated the capacity of secondary expansion of CD8~ Tm cells developed from transgenic OT I CD8^＋ T cells. The latter cells were in vitro and in vivo stimulated by ovalbumin （OVA）-pulsed dendritic cells [DCovA and （IL-12^-/-）DCovA] derived from wild-type C57BL/6 and IL-12 gene knockout mice, respectively. We demonstrated that IL-12 priming is important not only in CD8^＋ T cell clonal expansion, but also in generation of CD8^＋ Tm cells with the capacity of secondary expansion upon antigen re-encounter. However, IL-12 signaling is not involved in CD8^＋ Tm cell survival and recall responses. Therefore, this study provides useful information for vaccine design and development. Cellular ＆ Molecular Immunology.     

Generation and Regulation of CD8＋ Regulatory T Cells

Research into the suppressive activity of CD4＋FoxP3＋ T regulatory cells （Treg） has defined a sublineage of CD4＋ cells that contribute to self-tolerance and resistance to autoimmune disease. Much less attention has been given to the potential contribution of regulatory sublineages of CD8＋ cells. Analysis of a small fraction of CD8＋ cells that target autoreactive CD4＋ cells through recognition of the MHC class Ib molecule Qa-1 in mouse and HLA-E in human has revitalized interest in CD8＋ Treg. Here we summarize recent progress and future directions of research into the role of this CD8＋ sublineage in resistance to autoimmune disease. Cellular ＆ Molecular Immunology. 2008; 5（6）：401-406.     

PEG10 Activation by Co-Stimulation of CXCR5 and CCR7 Essentially Contributes to Resistance to Apoptosis in CD19^＋CD34^＋ B Cells from Patients with B Cell Lineage Acute and Chronic Lymphocytic Leukemia

We investigated CD19~+CD34~+ and CD19~+CD34 B cells from cord blood (CB) and typical patients with B celllineage acute and chronic lymphocytic leukemia (B-ALL and B-CLL) in terms of expression and functions ofCXCR5/CXCL13 and CCR7/CCL19.CXCR5 and CCR7 were selectively frequent expressed on B-ALL,B-CLLand CB CD19~+CD34~+ B cells,but not on CD19~+CD34 B cells.Instead of induction of impressive chemotacticresponsiveness,CXCL13 and CCL19 together induced significant resistance to TNF-α-mediated apoptosis inB-ALL and B-CLL but not CB CD19~+CD34~+ B cells.B-ALL and B-CLL CD19~+CD34~+ B cells expressed elevatedlevel of Paternally Expressed Gene 10 (PEG10),and CXCL13 and CCL19 together significantly up-regulatedPEG10 expression in the cells.We found that CXCL13 and CCL19 together by means of activation of CXCR5and CCR7 up-regulated PEG10 expression and function,subsequent stabilized caspase-3 and caspase-8 inB-ALL and B-CLL CD19~+CD34~+ B cells,and rescued the cells from TNF-α-mediated apoptosis.We suggestedthat normal lymphocytes,especially naive B and T cells,utilized CXCR5/CXCL13 and CCR7/CCL19 formigration,homing,maturation,and cell homeostasis as well as secondary lymphoid tissues organogenesis.Meanwhile certain malignant cells took advantages of CXCR5/CXCL13 and CCR7/CCL19 for infiltration,resistance to apoptosis,and inappropriate proliferation.Cellular & Molecular Immunology.2004;1(4):280-294.     

Global Gene Expression Profiling in Interleukin-12-Induced Activation of CD8^＋ Cytotoxic T Lymphocytes against MouseMammary Carcinoma

Interleukin-12(IL-12) is a critical cytokine representing the link between the cellular and humoral branches ofhost immune defense apparatus.IL-12-induced cytotoxic lymphocyte(CTL) development is a centralmechanism in immune responses against intracellular infectious agents as well as malignant growth.However,the molecular basis of tumor-specific CTL responses mediated by IL-12 remains poorly defined.In this study,we addressed this issue in a comprehensive manner to probe into IL-12-induced anti-tumor responses by globalgene expression profiling of mRNA expression in CD8~+ T cells in a transplantable syngeneic mouse mammarycarcinoma model treated or not with recombinant IL-12.A strong tumor regression was induced by the IL-12treatment.An introspection of differential gene expression at an early stage of the IL-12-initiated CTLactivation reveals interesting genes and molecular pathways that may account for the marked tumor regression,and is likely to provide a rich source of potential targets for further research and development of effectivetherapeutic modalities.Cellular & Molecular Immunology.2004;1(5):357-366.     

Type 1 CD8^＋ T Cells are Superior to Type 2 CD8^＋ T Cells in Tumor Immunotherapy due to Their Efficient Cytotoxicity, Prolonged Survival and Type 1 Immune Modulation

CD8^＋ cytotoxic T （Tc） cells play a crucial role in host immune responses to cancer, and in this context, adoptive CD8^＋ Tc cell therapy has been studied in numerous animal tumor models. Its antitumor efficacy is, to a large extent, determined by the ability of Tc cells to survive and infiltrate tumors. In clinical trials, such in vitro-activated T cells often die within hours to days, and this greatly limits their therapeutic efficacy. CD8^＋ Tc cells fall into two subpopulations based upon their differential cytokine secretion. In this study, we in vitro generated that ovalbumin （OVA）-pulsed dendritic cell （DCovA）-activated CD8^＋ type 1 Tc （Tcl） cells secreting IFN-T, and CD8^＋ type 2 Tc （Tc2） cells secreting IL-4, IL-5 and IL-10, which were derived from OVA-specific T cell receptor （TCR） transgenic OT I mice. We then systemically investigated the in vitro and in vivo effector function and survival of Tcl and Tc2 cells, and then assessed their survival kinetics after adoptively transferred into C57BL/6 mice, respectively. We demonstrated that, when compared to CD8^＋ Tc2, Tcl cells were significantly more effective in perforin-mediated cytotoxicity to tumor cells, had a significantly higher capacity for in vivo survival after the adoptive T cell transfer, and had a significantly stronger therapeutic effect on eradication of well-established tumors expressing OVA in animal models. In addition, CD8^＋ Tcl and Tc2 cells skewed the phenotype of CD4^＋ T cells toward Thl and Th2 type, respectively. Therefore, the information regarding the differential effector function, survival and immune modulation of CD8^＋ Tcl and Tc2 cells may provide useful information when preparing in vitro DC-activated CD8^＋ T cells for adoptive T cell therapy of cancer.     

Wharton’s Jelly Mesenchymal Stromal Cells as a Feeder Layer for the Ex Vivo Expansion of Hematopoietic Stem and Progenitor Cells: a Review

In recent years, umbilical cord blood (UCB) has been widely used as an alternative source to bone marrow (BM) for transplantation of hematopoietic stem and progenitor cells (HSPCs) in a variety of hematological and non-hematological disorders. Nevertheless, the insufficient number of UCB-HSPCs for graft represents a major challenge. HSPCs ex vivo expansion prior to transplantation is a valid strategy to overcome this limit. Several attempts to optimize the expansion conditions have been reported, including the use of mesenchymal stromal cells (MSCs) as feeder layer. Wharton’s Jelly (WJ), the main component of umbilical cord (UC) matrix, is especially rich in MSCs, which are considered ideal candidates for feeder layer in co-culture systems. In fact, they can be easily harvested and grow robustly in culture, producing a confluent monolayer in a short time. Similarly to bone marrow-mesenchymal stromal cells (BM-MSCs), WJ-derived MSCs (WJ-MSCs) have been used to support hematopoiesis in vitro and in vivo. Here, we review the rationale for using MSCs, particularly WJ-MSCs, as a feeder layer for UCB-HSPCs ex vivo expansion. In addition, we report the main findings attesting the use of these MSCs as a support in hematopoiesis.     

EBV-Induced Human CD8^＋ NKT Cells Synergise CD4^＋ NKT Cells Suppressing EBV-Associated Tumours upon Induction of Thl-Bias

CD8^＋ natural killer T （NKT） cells from EBV-associated tumour patients are quantitatively and functionally impaired. EBV-induced CD8^＋ NKT cells drive syngeneic T cells into a Thl-bias response to suppress EBV-associated malignancies. IL-4-biased CD4^＋ NKT cells do not affect either syngeneic T cell cytotoxicity or Th cytokine secretion. Circulating mDC1 cells from patients with EBV-associated malignancies impair the production of IFN-T by CD8^＋ NKT cells. In this study, we have established a human-thymus-SCID chimaera model to further investigate the underlying mechanism of EBV-induced CD8^＋ NKT cells in suppressing EBV-associated malignancies. In the human-thymus-SCID chimera, EBV-induced CD8^＋ NKT cells suppress EBV-associated malignancies in a manner dependent on the Thl-bias response and syngeneic CD3^＋ T cells. However, adoptive transfer with CD4^＋ NKT cells alone inhibits T cell immunity. Interestingly, CD4^＋ NKT cells themselves secrete high levels of IL-2, enhancing the persistence of adoptively transferred CD8^＋ NKT cells and T cells, thereby leading to a more pronounced T cell anti-tumour response in chimaeras co-transferred with CD4^＋ and CD8^＋ NKT cells. Thus, immune reconstitution with EBV-induced CD4^＋ and CD8^＋ NKT cells synergistically enhances T cell tumour immunity, providing a potential prophylactic and therapeutic treatment for EBV-associated malignancies.     

Analysis of the Conservation of T Cell Receptor Alpha and Beta Chain Variable Regions Gene in pp65 Peptide-Specific HLA-A＊0201-Restricted CD8^＋ T Cells

Many viral epitope specific T cell receptors （TCRs） in MHC-matched individuals have been demonstrated to involve conserved amino acid motifs in β chain complementarity-determining region 3 （CDR3）. However, it is not sure whether the conserved motifs can also be found in TCR β chain. In previous studies, we developed a modified method to enlarge the percentage of cytomegalovirus （CMV） pp65 peptide-specific CD8^＋ T cells in PBMC by continuous peptide stimulation in vitro, which provides sufficient number of specific T cells for detection. In this study, we further analyzed the restrictive usage of TCR Vα and Vβ gene families and investigated the CDR3 gene sequence of pp65 peptide-specific CD8β T cells. Analysis of CDR3 spectratypes suggested a restricted usage of TCR α chain AV8, AV12, AV21, AV31 families and TCR βchain BV3, BV14, BV21, BV23, BVll families in donor CD8^＋ T cells stimulated by pp65 peptide. The sequences of these T cells involved similar sequence （TX） G （X） A in CDR3 region of TCR α chain and L （XT） G （X） A in TCR β chain.     

Establishment and Characterization of a Cell Based Artificial Antigen-Presenting Cell for Expansion and Activation of CD8+ T Cells Ex Vivo

Atificial antigen-presenting cells are expected to stimulate the expansion and acquisition of optimal therapeutic features of T cells before infusion. Here CD32 that binds to a crystallizable fragment of IgG monoclonal antibody was genetically expressed on human K562 leukemia cells to provide a ligand for T-cell receptor. CD86 and 4-1BBL, which are ligands of CO. stimulating receptors of CD28 and 4-1BB. respectively, were also expressed on K562 cells. Then we accomplished the artificial antigen-presenting cells by coupling K32, CD86/4-IBBL cell with OKT3 monoclonal antibody against CD3.named K32/CD86/4-lBBL/OKT3 cells. These artificial modified cells had the abilities of inducing CD8+ T cell activation. promoting CD8+ T cell proliferation, division, and long-term growth, inhibiting CD8+ T cell apoptosis, and enhancing CD8+ T cell secretion of IFN-Y and perforin. Furthermore, antigen. secific cytotoxic T lymphocytes could be retained in the culture stimulated with K32/CD86/4-1BBL/OKT3 cells at least within 28 day This approach was robust, simple, reproducible and economical for expansion and activation of CD8+ T cells and may have important therapeutic implications for adoptive immunotherapy. Cellular & Molecular Immunology.2007;5(1):47-53.     

人NK细胞体外高效扩增的实验研究

目的:建立人NK细胞体外大量扩增的方法。方法:采用基因工程方法,在K562细胞上同时表达IL-15、IL-18、4-1BBL3种基因,构建特定的K562工程细胞作为刺激细胞。IL-15、IL-18基因分别与一段特殊的跨膜蛋白基因融合,4-1BBL直接跨膜表达,使这3种蛋白在K562细胞中表达后锚定于细胞膜表面。其次,以照射致死的该K562工程细胞作为刺激细胞,以人外周血单个核细胞(PBMC)为扩增培养对象,通过与IL-2的共刺激作用,使NK细胞在体外培养条件下得到大量的扩增。结果:经过21d的刺激培养后,CD56 CD3-细胞数量扩增了(520±75)倍。CD56 CD3-细胞的纯度从培养前占PBMC的7%±4%到扩增后占总细胞比例的93%±3%。PBMC中的T细胞基本上没有得到扩增,扩增后的细胞中CD3 细胞只占2%±1.2%。扩增的细胞具备了NK细胞的基本特征和生物学特性,除了CD56 CD3-外,还对扩增的NK细胞上NKG2D、NKp46、NKp44、NKp30、CD94、CD158b、CD158a、NKB1、NKAT2等标记进行了验证。细胞毒实验表明,在效应细胞∶靶细胞为5∶1时,扩增的NK细胞的杀伤率达到了95%±4%。结论:建立的NK细胞体外扩增方法,达到了较高的扩增水平,且扩增的细胞活性较好。本方法以PBMC为原始材料,能够实现NK细胞体外的大规模制备,这将为抗病毒与抗肿瘤的NK细胞免疫治疗奠定基础。     

Artificial antigen-presenting cells expressing CD80, CD70, and 4-1BB ligand efficiently expand functional T cells specific to tumor-associated antigens

Professional antigen-presenting cells (APCs), notably dendritic cells (DCs), are the most potent for expanding antigen-specific T cells ex vivo. However, the labor-intensive and expensive procedure for customized preparation of autologous APCs has hampered their broad clinical application. Artificial APC (aAPC) systems, which can be readily prepared from off-the-shelf components, have been proposed as a promising alternative to custom-made professional APCs. Here, in order to develop a novel aAPC system, we established K562 erythroleukemia cells expressing different combinations of co-stimulatory molecule ligands, CD80, CD70, and/or 4-1BB ligand (4-1BBL). When nucleofected with in vitro-generated mRNA encoding a tumor-associated antigen, MART-1, the K562 cells expressing all of CD80, CD70, and 4-1BBL were the most efficient for expansion of functional T cells specific to an HLA-A2-restricted immunodominant epitope, MART-1_26–35. In addition, only the K562 cells expressing all three of these co-stimulatory molecule ligands could clearly expand T cells specific to other less immunogenic antigen epitopes, gp100_154–162 and Cyp1B1_239–247, through transfection with in vitro generated gp100 and Cyp1B1 mRNA, respectively. These results indicated that non-redundant and synergistic effects of co-stimulation via CD28/CD80, CD27/CD70, and 4-1BB/4-1BBL might be critical for eliciting efficient expansion of T cells; co-stimulation via the 4-1BB/4-1BBL interaction might expand antigen-specific T cells by preventing apoptotic cell death triggered by specific antigens in the presence of the CD28/CD80 and CD27/CD70 signaling. Taken together, our findings suggested that this K562-based aAPC system expressing CD80, CD70, and 4-1BBL would be useful for efficiently stimulating functional antigen-specific T cells ex vivo, in particular when detailed information on the epitope specificities is unavailable.     

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.     

Dispensable role for 4-1BB and 4-1BBL in development of vaccinia virus-specific CD8 T cells

CD8 T cells are strongly induced in response to certain strains of vaccinia virus (VACV) and the generation of this population is tightly regulated by two Tumor Necrosis Factor (TNF)/TNFR superfamily members, OX40 (CD134) and CD27. In this study, we examined the role of another member of the TNFR superfamily, 4-1BB (CD137, TNFRSF9), and its ligand (4-1BBL, CD137L, TNFSF9), that have been described to control the generation of memory CD8 T cell populations elicited by other viruses such as influenza. Expression of 4-1BB and 4-1BBL was observed in wild-type mice during the primary infection, but we found that both 4-1BB and 4-1BBL deficient mice generated normal numbers of VACV-specific effector CD8 T cells that produced IFN-γ and TNF. Additionally, CD8 T cells deficient in 4-1BB were able to expand and persist comparably to wild-type T cells in response to VACV infection. Furthermore, the knockout mice also showed no defect in development of VACV-specific CD8 memory T cell populations. Lastly, showing alternate control mechanisms were not active in the gene-deficient environments that masked any activity, blocking 4-1BB/4-1BBL interactions using neutralizing antibody also had no effect on the number of VACV-specific memory CD8 T cells induced. Thus, our data demonstrate that 4-1BB and 4-1BBL do not play a strong or dominant role in driving the generation of high frequencies of VACV-specific CD8 T cells.     

CD5 blockade enhances ex vivo CD8+ T cell activation and tumour cell cytotoxicity

CD5 is expressed on T cells and a subset of B cells (B1a). It can attenuate TCR signalling and impair CTL activation and is a therapeutic targetable tumour antigen expressed on leukemic T and B cells. However, the potential therapeutic effect of functionally blocking CD5 to increase T cell anti-tumour activity against tumours (including solid tumours) has not been explored. CD5 knockout mice show increased anti-tumour immunity: reducing CD5 on CTLs may be therapeutically beneficial to enhance the anti-tumour response. Here, we show that ex vivo administration of a function-blocking anti-CD5 MAb to primary mouse CTLs of both tumour-naïve mice and mice bearing murine 4T1 breast tumour homografts enhanced their capacity to respond to activation by treatment with anti-CD3/anti-CD28 MAbs or 4T1 tumour cell lysates. Furthermore, it enhanced TCR signalling (ERK activation) and increased markers of T cell activation, including proliferation, CD69 levels, IFN-γ production, apoptosis and Fas receptor and Fas ligand levels. Finally, CD5 function-blocking MAb treatment enhanced the capacity of CD8⁺ T cells to kill 4T1-mouse tumour cells in an ex vivo assay. These data support the potential of blockade of CD5 function to enhance T cell-mediated anti-tumour immunity.     

Determination of cell expansion and surface molecule expression on anti‐CD3/28 expanded CD4+ T cells

CD4⁺ T cell immunotherapy has potential for treatment in HIV‐infected patients. A large number of expanded CD4⁺ T cells and confirmation of functional‐related phenotypes are required for ensuring the successful outcomes of treatment. Freshly isolated CD4⁺ T cells from healthy donors were activated with anti‐CD3/28‐coated magnetic beads at different bead‐to‐cell ratios and cultured in the absence and presence of IL‐2 supplementation for 3 weeks. Fold expansion, cell viability, growth kinetic and lymphocyte subset identities were determined. Data demonstrated that a 1:1 bead‐to‐cell ratio rendered the highest expansion of 1044‐fold with 88% viability and 99.5% purity followed by the 2:1 and 0.5:1 ratios. No significant difference in proliferation and phenotypes was found between non–IL‐2 and IL‐2 supplementation groups. Several specific surface molecule expressions of the expanded cells including chemokine receptors, adhesion molecules, co‐stimulatory molecules, activation molecules, maturation markers, cytokine receptors and other molecules were altered when compared to the unexpanded cells. This optimized expansion protocol using the 1:1 bead‐to‐cell ratio of anti‐CD3/28‐coated magnetic beads and culture condition without IL‐2 supplementation provided the satisfactory yield with good reproducibility. Specific surface molecule expressions of the expanded cells presented potential roles in proliferation, differentiation, homeostasis, apoptosis and organ homing.     

High-frequency activation of single CD4+ and CD8+ T cells to proliferate and secrete cytokines using anti-receptor antibodies and IL-2(1).

A high cloning efficiency single-cell culture system was developed to define the activation requirements of isolated CD4+ and CD8+ T cells to proliferate and secrete cytokines. T cells were triggered using solid-phase anti-CD3 and anti-CD4 or anti-CD8 antibodies plus rIL-2. Activation was measured by microscopic scoring of proliferation and by measurement of cytokine production using the cytokine-responsive cell lines FDC-P1, which responds to GM-CSF, IL-3, IFN-gamma and IL-4, and 32D clone 3 which responds to IL-3 only. Whilst anti-CD3 plus rIL-2 triggered only 4% of peripheral T cells to proliferate, anti-CD3 plus anti-CD8 mAb triggered about 40% of CD8+ T cells; 80% of the resultant clones secreted cytokine and 90% of these were IL-3+. Anti-CD3 plus anti-CD4 mAb triggered proliferation in about 20% of CD4+ T cells, of which 34% formed cytokine-producing clones with 47% of these secreting IL-3. In addition to responding at higher frequency, CD8+ T cells formed larger clones which produced higher levels of cytokines than CD4+ cells. Cell separation on the basis of Pgp-1 expression suggested that this culture system did not select for previously activated cells. Whereas Pgp-1+ T cells from keyhole limpet haemocyanin (KLH)-primed mice were enriched in KLH-specific cells, no significant differences were observed in the clonogenicity or cytokine-secreting capacity of Pgp-1+ and Pgp-1- T cells from normal mice.     

CD86对CD8+T细胞分化的影响

目的：探讨CD86(B7-2)对CD8^ T细胞分化的影响。方法：用限制性内切酶Xho Ⅰ酶切质粒pCDM8得到CD86基因，并将其插入pCDNA3，用BamH Ⅰ酶切鉴定。用脂质体法介导pCDNA3-CD86真核表达载体转染人肝癌细胞株HMCT／21，600μg/ml G418筛选，稳定且高表达CD86的抗性克隆用流式细胞仪进行鉴定。从健康志愿者血中分离外周血单个核细胞(PB-MC)，使PBMC与靶细胞之比为20：1，共同培养48h后，用流式细胞仪检测CD3^ T细胞内IL-4和IFN-γ的表达率。结果：成功构建pCDNA3-CD86真核表达载体；CD86在HMC7721-CD86细胞中的表达率为30．8％，而在HMC7721细胞中的表达率为0．98％；健康志愿者CD3^ T细胞内IL-4和IFN-γ的表达率分别为1．92％和24．4％；PBMC与靶细胞共同培养48h后，无论是否用IFN-α刺激，IL-4，IFN-γ的阳性比值在HMC7721-CD86转染组均大于1，而在HMC7721未转染组均小于1。结论：在细胞培养中，CD86可诱导CD8^ T细胞活化，并向Tc2表型转化。