重组人可溶性CD40配体对树突状细胞体外分化、成熟及功能的影响

用Ficoll密度离心及贴壁法获得外周血单个核细胞 (PBMC ) ,PBMC经细胞因子组合诱导分化成树突状细胞 (DC ) :GM CSF (10 0ng/ml)与IL 4 (5 0ng/ml)诱导 5d后 ,分别加入TNF α (10ng/ml)或rhsCD4 0L (2 μg/ml)继续培养 4d ;倒置显微镜下观察DC形态 ,免疫荧光标记和流式细胞术分析DC表型 (CD1a、CD80、CD83、HLA DR、CD14、CD16、CD19)及摄取FITC Dextran抗原的能力 ;3 H TdR掺入法检测DC刺激自体混合淋巴细胞体外增殖反应 (MLR )能力 ;ELISA法分析DC培养上清中IL 12的水平 ;Trans well细胞趋化实验检测DC对自体外周T淋巴细胞的趋化能力。发现经rhsCD4 0L刺激的DC表面分子 (CD1a、CD80、CD83、HLA DR )的表达水平高于经典的细胞因子组合组 (GM CSF +IL 4 +TNF α ) ,同时rhsCD4 0L刺激后的DC摄取FITC Dextran的能力下降而刺激自体MLR和分泌IL 12的能力明显提高 ;而且rhsCD4 0L诱导的DC表面趋化因子受体CXCR4的表达水平及对自体外周T淋巴细胞的趋化能力均强于TNF α或FL激发的DC。rhsCD4 0L在体外不仅具有显著的诱导DC分化 ,促进DC成熟的功能 ,而且经rhsCD4 0L作用的DC能更有效地激发T淋巴细胞     

树突状细胞介导的肿瘤免疫治疗研究

树突状细胞是抗原递呈能力最强的抗原递呈细胞。树突状细胞可通过交叉致敏途径呈递外源性抗原如肿瘤抗原,诱导CD8+的CTL应答。应用不同形式的肿瘤抗原负载DC,将致敏DC作为疫苗回输体内可诱导机体产生有效的抗肿瘤免疫应答。     

树突状细胞介导的肿瘤免疫治疗研究

树突状细胞是抗原递呈能力最强的抗原递呈细胞。树突状细胞可通过交叉致敏途径呈递外源性抗原如肿瘤抗原，诱导CD8 的CTL应答。应用不同形式的肿瘤抗原负载DC，将致敏DC作为疫苗回输体内可诱导机体产生有效的抗肿瘤免疫应答。     

IL-3对小鼠骨髓来源树突状细胞分化发育的影响

比较 GM- CSF IL- 4与 IL- 3 IL- 4两种方法培养制备的树突状细胞 (Dendritic cell,DC)在形态、产量、免疫表型和抗原摄取能力方面的差异。用 GM- CSF IL- 4和 IL- 3 IL- 4分别诱导小鼠骨髓来源的前体细胞分化为成熟树突状细胞 ,通过相差显微镜、扫描电镜、激光共聚焦扫描显微镜进行形态观察 ,流式细胞术分析细胞免疫表型和摄取抗原能力。结果表明 ,IL- 3 IL- 4诱导的 DC产量高 ,细胞纯度好 ,形态上与 GM- CSF IL- 4诱导的DC类似 ;细胞免疫表型显示高表达 MHC 类分子 ,但低表达或不表达 CD80、CD86 ;IL- 3 IL- 4诱导的 DC具有强大的摄取抗原能力。 IL- 3替代 GM- CSF可诱导低表达共刺激分子的耐受型 DC     

诱导实体瘤细胞生成具有树突状细胞样的抗原提呈细胞

目的　探讨体外用细胞因子诱导实体瘤细胞生成具有树突状细胞样的抗原提呈功能。方法　以人肝母细胞瘤为研究模型 ,用人胰岛素样生长因子 (IGF 1)、人重组粒细胞单核细胞集落刺激因子 (GM CSF)、人重组白细胞介素 4 (IL 4 )及肿瘤坏死因子α(TNF α)诱导培养瘤细胞 2周 ,用流式细胞仪及MTT法检测诱导前后瘤细胞表型及抗原提呈功能。结果　用细胞因子处理瘤细胞后 ,树突状细胞样的特异性标志CD1a和CD83表达阳性率显著上调 ;MHCⅡ、B7 1、B7 2、ICAM 1和CD4 0表达水平显著增加。同时 ,瘤细胞能显著刺激同种异体淋巴细胞增殖 ,促进IL 12的产生 ,明显提高细胞毒性T淋巴细胞 (CTL)对瘤细胞的杀伤活性。结论　初步实验结果提示 ,体外用IGF 1、GM CSF、IL 4及TNF α等细胞因子能诱导肝母细胞瘤细胞生成具有树突状细胞样特征的抗原提呈细胞。     

HBsAg体外冲击的慢性乙肝患者树突状细胞的生物学特性及其对HBV特异性CTL的诱导作用

目的 :研究HBsAg冲击的慢性乙肝患者单核细胞来源的树突状细胞 (DCs)的功能状况及体外对HBV特异性CTL的诱导作用 ,初步探讨诱导特异性抗HBV细胞免疫的途径。方法 :分离慢性乙肝患者外周血单核细胞 ,以GM CSF +IL 4 +TNF α培养诱导DCs,加入HBsAg冲击以诱导HBV特异性DCs。采用FCM测定细胞表面免疫分子CD1a、CD83、CD86、CD80、CD4 0以及HLA DR的表达水平 ,ELISA法检测培养上清中细胞因子IL 6、IL 12的分泌含量 ,MTT法测定DC刺激同种异体淋巴细胞增殖的能力 ,LDH法检测DC诱导的患者外周血T细胞对HepG2 2 2 15 (转染HBVDNA)、HepG2肝癌细胞株及K5 6 2白血病细胞株的细胞毒作用。结果 :HBsAg冲击的DC其表达CD1a、CD83、CD86、CD80、CD4 0、HLA DR表面分子明显高于对照组 (P <0 0 1,P <0 0 5 ) ,分泌IL 12的水平也高于对照组 (P <0 0 1) ,而分泌IL 6的水平则较对照组显著降低(P <0 0 1) ;HBsAg冲击的DC刺激同种异体淋巴细胞增殖的能力明显增强 (P <0 0 5 ) ,并可有效地诱导自体CTL对转HBV基因的HepG2 2 2 15细胞高效特异性杀伤作用 (P <0 0 1)。结论 :慢性乙型肝炎患者单核细胞来源的DCs经HBsAg抗原冲击后 ,生物学活性增强 ,并且能有效地诱导对HBV特异性反应的CTL。     

凋亡肿瘤细胞负载的树突状细胞对抗原特异性CTL的激活

为探讨凋亡Daudi细胞负载的树突状细胞 (DC )激发诱导肿瘤抗原特异性CTL及其生物学特性 ,采用常规方法从人外周血单个核细胞 (PBMC )诱导DC ,激发型CD40mAb联合 50Gyγ射线辐照诱导Daudi凋亡后负载DC ,然后与自体T细胞共育 ,并联合IL 2激发诱导肿瘤特异性CTL ;采用免疫荧光标记和流式细胞仪测定分析膜分子的表达 ;ELISA检测细胞因子的产生 ;Dextran FITC内吞实验分析DC的抗原摄取能力 ;3H掺入试验和51 Cr释放试验分别测定CTL的增殖和细胞毒效应。结果 :(1 )细胞因子序贯体外诱导 7d的DC ,对Dextran吞噬能力最强 ;(2 )CD40mAb诱导联合γ射线辐照 ,能有效介导Daudi细胞的凋亡 ;(3 )抗原负载联合CD40mAb激发可使DC上调表达CD1a、CD80、CD86和HLA DR ,并能促进IL 1 2的分泌 ;(4)凋亡Daudi负载后的DC在激发型CD40mAb作用下 ,能激发和扩增对Daudi细胞具有高效和特异杀伤作用的细胞毒性T细胞。因而认为凋亡肿瘤细胞负载联合CD40mAb刺激的DC可有效激活和扩增肿瘤特异性CTL。     

冻融抗原负载树突状细胞诱导细胞毒性T淋巴细胞特异性杀伤急性白血病细胞的实验研究

我们应用基因重组人白介素 4 (rhIL 4 ) ,基因重组人粒 /单细胞集落刺激因子 (rhGM CSF)联合培养体系自健康志愿者骨髓单个核细胞诱导产生DC ,使其负载急性髓系白血病 (AML)细胞冻融抗原 ,体外诱导细胞毒性T细胞 (CTL) ,观察负载AML细胞冻融抗原后DC的状态对所激发的T淋巴细胞表型及功能的影响及CTL对AML细胞特异性杀伤活性。我们发现负载AML冻融抗原后DC的CD1a、CD83、CD86、CD11C、HLA DR表达率为较培养前明显增高 (P <0 .0 1) ,且负载AML冻融抗原的DC诱导的CTL中CD3 CD8 T细胞比例 ,较诱导前明显增高 (P <0 .0 1) ,而负载AML细胞冻融抗原的DC诱导CTL对AML细胞有较强的杀伤作用 ,明显强于加或不加IL 2培养的T细胞对照组 (P <0 .0 1) ,而对K5 6 2细胞无明显的杀伤活性。通过以上实验我们认为经rhGM CSF ,rhIL 4培养产生的DC为CD14CD1a DC ,能诱导CTL对AML细胞产生明显的特异性杀伤作用 ,另外 ,负载AML细胞冻融抗原DC诱导的CTL中CD3 CD8 T细胞比例明显增高 ,提示CD8 T细胞在抗肿瘤免疫中具有极其重要的作用。     

Gp130分子的活化在树突状细胞分化和成熟中的意义

目的 :研究激发型抗gp130分子单抗B S12对树突状细胞 (DC)分化成熟和功能的影响。方法 :人外周血单核细胞在GM CSF和IL 4作用下分化为未成熟DC后 ,加入B S12单抗 ,观察它对DC表型、摄取抗原的能力以及DC分泌IL 12、进行混合淋巴细胞反应及趋化T细胞能力的影响 ;同时比较B S12和激发型抗CD4 0单抗 5C11对DC的作用。结果 :激发型抗gp130分子单抗B S12作用于未成熟DC ,可以上调DC上CD1a和共刺激分子CD80、CD86以及成熟DC的标志CD83分子的表达 ,下调CD14的表达 ,降低DC摄取抗原的能力 ,增强DC分泌IL 12、进行混合淋巴细胞反应及趋化T细胞的能力 ,但这些作用都稍弱于 5C11对DC的作用。结论 :未成熟DC上gp130分子的直接活化可以诱导DC的分化和功能成熟。     

多发性骨髓瘤患者树突状细胞介导的特异性抗瘤活性的体外诱导

目的 :观察U2 6 6 细胞及其可溶性抗原激发的多发性骨髓瘤 (MM)患者树突状细胞 (DC)体外诱导U2 6 6 特异性CTL的作用。方法 :将MM患者外周血来源的单核细胞在rhGM CSF 80 0U ml与IFNα 6 0 0U ml条件下利用无血清技术培养生成DC ,应用丝裂霉素C处理的U2 6 6 细胞及用U2 6 6 细胞制备的可溶性抗原预刺激DC ,然后与自体淋巴细胞共同孵育 5～ 7d以诱生特异性CTL ,采用MTT法检测对U2 6 6 细胞的特异性杀伤效果。结果 :MM患者外周血单核细胞在GM CSF IFNα条件下培养 8d后生成具有典型特征的DC ,高度表达CD86、CD5 4及MHCII类分子HLA DR。应用MTT法检测U2 6 6 细胞及其可溶性抗原激发的DC诱导特异性CTL对靶细胞U2 6 6 的杀伤率分别为 2 1 2 %± 5 4 %和 2 8 0 %± 7 6 % ,对照组未用抗原刺激组都为 11 7%±4 3%。而以抗原直接刺激自体淋巴细胞组为 15 6 %± 4 8%和 13 1%± 5 5 % (P <0 0 1)。结论 :U2 6 6 细胞及其可溶性抗原激发的DC与自体淋巴细胞孵育能诱导抗U2 6 6 特异性CTL。     

Influence of CD4 T cells and the source of major histocompatibility complex class II-restricted peptides on cytotoxic T-cell priming by dendritic cells

We have previously reported that bone marrow derived dendritic cells (DC) pulsed with major histocompatibility complex (MHC) class I-restricted peptide efficiently prime a cytotoxic T lymphocyte (CTL) response in vivo. Here we assess the involvement of CD4(+) T cells in the induction of CD8(+) CTL by DC by testing the ability of class II-deficient (C2D) DC, class II mutant (Alpha beta mut) DC and autologous serum generated DC (AS DC) to present class I-restricted antigens in vitro and in vivo. DC generated from the bone marrow of class II knockout mice and transgenic mice expressing a mutant class II that can not bind CD4 were phenotypically similar to wild type (wt) DC, except with regard to MHC class II expression. The C2D and Alpha beta mut DC, though fully capable of presenting the class I-restricted ovalbumin (OVA) peptide to a T-cell hybridoma in vitro, failed to prime a CTL response in vivo. Restoration of class II expression on C2D DC allowed priming of a CTL response; thus, the defect in CTL priming was indeed caused by the absence of class II expression. Likewise, DC generated in autologous serum were unable to prime a CTL response as these DC only express 'self' class II epitopes and therefore would not activate syngeneic CD4(+) T cells. Addition of exogenous class II epitopes rescued the ability of AS DC to prime a CTL response. These observations provide convincing evidence that efficient CTL induction by DC in vivo requires concomitant presentation of class II epitopes for CD4(+) T-cell induction.     

IL—3、GM—CSF扩增骨髓树突状细胞及促进MHC Class—I途径抗原提呈

目的：研究 IL-3、GM-CSF能否扩增小鼠骨髓树突状细胞，并通过 MHC Class-I途径提呈外源性抗原。方法：以C57BL/6小鼠骨髓2 h粘附细胞作为树突状细胞来源，在 IL-3（10 ng/ml）及 GM-CSF（1000 U/ml）条件下培养5 d，观察细胞形态、数量和免疫表型的变化，以及对外源性抗原的摄取能力和Class-I途径提呈能力。结果：培养后细胞绝对计数显著增加，MHC分子及共刺激分子表达显著增加，对颗粒性抗原beads-OVA具有高效摄取能力，Class-I途径提呈beads-OVA及SIIFEKL表位的能力显著增强。结论：IL-3、GM-CSF能有效扩增具有高效抗原摄取和提呈能力的树突状细胞，提示在树突状细胞和肿瘤免疫治疗研究中具有重要价值。     

Interleukin-3 and granulocyte-macrophage colony-stimulating factor enhance the generation and function of dendritic cells.

Dendritic cells, well-known for their potent antigen-presenting activity, are generally present at very low frequency in the spleens of naive mice. We examined the ability of mice to generate functional dendritic cells (DC) following exposure to the cytokines interleukin-3 (IL-3) and granulocyte-macrophage colony-stimulating factor (GM-CSF). Tumours secreting these cytokines provided a continuous stimulus resulting in a greatly increased number and frequency of DC in the spleen. These cells were purified by conventional DC isolation techniques and were found to exhibit many of the characteristics of DC from unmanipulated mice, including high allo-stimulatory activity in mixed lymphocyte reactions and expression of many similar cell surface markers. Using ovalbumin-peptide specific class I- and class II-restricted hybridomas containing the lacZ reporter gene, we found that these cytokine-generated DC had a greatly increased efficacy in the uptake and processing of particulate antigen. These cells appear to have retained the ability to ingest antigen that is generally associated with immature DC, but also exhibit the peptide/major histocompatibility complex (MHC)-presenting capabilities of mature DC. Development of an assay to measure the activity of a single DC revealed that these dual activities were the properties of the majority of the cytokine-generated DC. These findings indicate that exposure in vivo to the cytokines IL-3 and GM-CSF can result in the generation of large numbers of DC with increased capability of stimulating T cells. Thus, these cells may be important in vivo in the process of cross-priming and the subsequent generation of tumour-reactive cytotoxic T lymphocytes (CTL).     

Coupling of antigen to cholera toxin for dendritic cell vaccination promotes the induction of MHC class I-restricted cytotoxic T cells and the rejection of a cognate antigen-expressing model tumor

We previously demonstrated that cholera toxin (CT) is highly efficient as a combined carrier and adjuvant for dendritic cell (DC) vaccination, inducing strong Th1-dominated B cell and CD4(+) T cell responses. In this study we show that vaccination with DC pre-pulsed ex vivo with CT-conjugated OVA (OVA-CT) gives rise to OVA-specific CD8(+) T cells that produce IFN-gamma and are cytotoxic for OVA-expressing E.G7 tumor cells both in vitro and in vivo. The induction of specific CD8(+) CTL by OVA-CT-treated DC was associated with enhanced presentation of OVA peptide (SIINFEKL) on MHC class I in combination with an overall activation of the pulsed DC. Vaccination of mice with OVA-CT-pulsed DC resulted in rejection of already established MHC class I-positive, MHC class II-negative, OVA-expressing E.G7 tumors in an antigen-specific, CD8(+) T cell-dependent fashion and was associated with high numbers of tumor-infiltrating CD8(+) T cells. Conjugation of antigen to CT facilitated DC uptake of the linked antigen through the GM1 receptor-binding B subunit and induced strong activation-maturation signals through the biologically active A subunit. These results have interesting implications for DC vaccination aimed at inducing CTL immune responses.     

Mechanisms of cytokine synergy essential for vaccine protection against viral challenge.

The ability of cytokines to steer CD4(+) T(h) cell responses toward a T(h)1 or T(h)2 phenotype and enhance the magnitude of both CD8(+) cytotoxic T lymphocytes (CTL) and antibody responses has clearly been demonstrated by our lab and others, but the influence of cytokines on protective immune responses is much less clear. Here we show an essential role for CD4(+) T(h)1 helper cell induction and IFN-gamma production in protection from viral challenge with a recombinant vaccinia virus expressing HIV-1MN viral envelope glycoprotein gp160. Complete protection from viral challenge is achieved only when the triple combination of exogenous cytokines granulocyte macrophage colony stimulating factor (GM-CSF), IL-12 and tumor necrosis factor (TNF)-alpha are co-administered with the peptide vaccine. In vivo depletion of CD4(+) cells or immunization of IFN-gamma-deficient mice abrogates protection. GM-CSF, IL-12 and TNF-alpha also synergize for the enhanced induction of CTL; however, adoptive transfer of a CD8(+) CTL line afforded only partial protection in this viral challenge model. As a possible mechanism of in vivo protection we show that GM-CSF increases the percentage and activity of antigen-presenting dendritic cells in draining lymph nodes where the immune response is initiated. We further demonstrate synergy between IL-12 and the proinflammatory cytokine TNF-alpha in driving IFN-gamma production. Thus, a combination of IL-12 and TNF-alpha is essential for the optimal development of T(h)1 responses and help for CTL induction in BALB/c mice, and is complemented by a third cytokine, GM-CSF, which enhances antigen presentation.     

Vaccination with dendritic cells pulsed in vitro with tumor antigen conjugated to cholera toxin efficiently induces specific tumoricidal CD8+ cytotoxic lymphocytes dependent on cyclic AMP activation of dendritic cells

We investigated the development of CD8+ tumor-specific cytotoxic lymphocytes (CTL) and protection against tumor growth after vaccination with bone marrow-derived dendritic cells (DC) pulsed with a model protein ovalbumin conjugated to cholera toxin (OVA-CT) in B6 mice using E.G7 tumor cells expressing OVA(257-264) peptide (SIINFEKL) as target cells in vitro and in vivo. Vaccination with OVA-CT-pulsed DC concurrently induced strong CTL in vitro activity and anti-E.G7 tumor protection in vivo in WT, NK-depleted and CD4-deficient mice as well as in IL-12-/- and IFN-gamma-/- mice but not in CD8-deficient mice. Importantly, activation of CTL by OVA-CT-pulsed DC was dependent on CT-induced activation of adenylate cyclase and increased cAMP production by DC associated with increased expression of MHC class I and co-stimulatory molecules (CD80, CD86 and CD40). These results show that vaccination with DC pulsed with antigens (Ag) conjugated to CT induces a strong CTL response and suggest that conjugation of tumor Ag to CT for DC vaccination represents a promising approach for tumor vaccination and immunotherapy.     

Effective therapeutic anticancer vaccines based on precision guiding of cytolytic T lymphocytes

Summary: In natural immune responses CD4⁺ T helper (Th) cells, reactive with peptide antigens presented by major histocompatibility complex (MHC) class II molecules on dendritic cells (DC), can drive the maturation of DC that is required for induction of CD8⁺ cytolytic T‐lymphocyte (CTL) immunity. Proper induction, expansion and maintenance of CTL responses are achieved through delicate interactions between CD4⁺ T cells, DC and CD8⁺ T cells involving several ligand–receptor pairs. Th cells to a large extent operate through up‐regulation of CD40L, which then interacts with CD40 on DC to cause DC maturation. Subsequent CTL induction by activated DC requires CD80/CD86 on the DC to interact with the CD28 costimulatory receptor on CD8⁺ T cells. For maintenance and full expansion of CTL, interaction of the DC‐expressed 4–1BB ligand with its receptor 4–1BB on CTL is also important. Alternative molecular triggers of DC activation that can support induction of powerful CTL responses include agonistic anti‐CD40 antibody or ligands of Toll‐like receptors (TLR) such as LPS (TLR4 ligand) or oligodeoxynucleotides containing CpG‐motifs (TLR9 ligand). The combination of CpG adjuvant with a 35 amino acid long synthetic peptide comprising both tumor‐specific CTL and Th epitopes proved to be a highly effective vaccine formulation capable of inducing therapeutic immunity against human papillomavirus‐induced mouse tumors. The recently acquired insights into antigen presentation and costimulatory signals have made possible the development of a new generation of therapeutic anticancer vaccines.     

Dexamethasone inhibits the antigen presentation of dendritic cells in MHC class II pathway

Glucocorticoids (GC) are physiological inhibitors of inflammatory responses and are widely used as anti-inflammatory and immunosuppressive agents in treatment of many autoimmune and allergic diseases. In the present study, we demonstrated that one of the mechanisms by which GC can suppress the immune responses is to inhibit the differentiation and antigen presentation of dendritic cells (DC). DC were differentiated from murine bone marrow hematopoietic progenitor cells by culture with GM-CSF and IL-4 with or without dexamethasone (Dex). Our data showed that Dex, in a dose dependent manner, down-regulated surface expression of CD86, CD40, CD54 and MHC class II molecules by DC, but the expression of MHC class I, CD80, CD95 and CD95L were not affected. In addition, Dex-treated DC showed an impaired function to activate alloreactive T cells and to secrete IL-Ibeta and IL-12p70. Moreover, Dex inhibited DC to present antigen by MHC class II pathway. However, the endocytotic activity of DC was not affected. The inhibitory effect of Dex on the expression of costimulatory molecules and the antigen-presenting capacity of DC could be blocked by the addition of RU486, a potent steroid hormone antagonist, suggesting the requirement of binding to cytosolic receptors in the above-described action of Dex. Since DC have the unique property to present antigen to responding naive T cells and are required in the induction of a primary response, the functional suppression of DC by Dex may be one of the mechanisms by which GC regulate immune responses in vivo.     

Mechanisms of induction of primary virus-specific cytotoxic T lymphocyte responses.

We have investigated the ability of various antigen-presenting cell (APC) types to induce primary anti-viral cytotoxic T lymphocyte (CTL) responses by single in vitro stimulation. Of these APC types, only dendritic cells (DC) and RMA-S lymphoma cells could induce primary CTL responses, but by divergent mechanisms. DC were capable of generating primary virus-specific CTL, either by presenting viral peptide or processed infectious virus. In contrast, RMA-S cells could not present endogenous antigen, e.g. after virus infection, but this cell line very efficiently presented exogenous viral peptides to induce primary virus-specific CTL in vitro. Spleen cells, lipopolysaccharide-induced B cell blasts or the non-mutated RMA cells did not have the ability to trigger unprimed T cells by single in vitro stimulation. We have investigated several characteristics important for primary CTL response induction by DC and RMA-S cells (summarized in Fig. 6). Primary CTL response induction by DC or RMA-S cells was blocked by anti-LFA-1 or anti-CD8 monoclonal antibodies (mAb). DC rapidly aggregated with unprimed T cells, which was independent of LFA-1 and CD8 molecules. RMA-S cells did not form conjugates with unprimed T cells. Despite their abundant major histocompatibility complex (MHC) class I cell-surface expression, DC did not bind much exogenously added viral peptide. In contrast, the MHC class I molecules on RMA-S cells bound a large quantity of exogenously administered peptide. Powerful adhesion by DC and high expression of relevant MHC/peptide complexes on RMA-S cells are important features in the initial contact with unprimed T lymphocytes. In a later stage of contact, both DC and RMA-S cells activate LFA-1 (and CD8) molecules at the T cell surface to strengthen and maintain the contact between T cell and APC.