胃肿瘤抗原致敏的脐血 DC 联合 CIK 对胃癌细胞株 SGC-7901杀伤活性的研究

目的：探讨体外胃肿瘤抗原致敏脐血 DC 联合细胞因子诱导的杀伤细胞（CIK）对胃癌细胞株 SGC-7901的杀伤作用。方法分离脐血单个核细胞培养 DC 细胞和 CIK 细胞。流式细胞仪检测成熟 DC 细胞表面抗原 CD83、CD86、CD11c 及 CIK 细胞表面抗原 CD3、CD56、CD4、CD8、CD16表达。致敏 DC-CIK、非致敏 DC-CIK、CIK 作为效应细胞,SGC-7901作为靶细胞,利用乳酸脱氢酶（LDH）释放法检测致敏 DC-CIK、脐血 DC-CIK、CIK 分别在效靶比10：1、20：1、40：1时对胃癌细胞的杀伤活性。结果成熟脐血 DC 表面抗原 CD83＋ CD86＋、CD11c ＋ CD83＋、CD86＋ CD11c ＋表达率分别为（75.4±2.1）％、（79.3±1.4）％、（80.2±2.6）％。致敏脐血 DC 表面表达率分别为（77.7±1.5）％、（82.6±1.9）％、（76.9±2.6）％,二者差异无统计学意义（t ＝1.526,P ﹥0.05;t ＝0.958,P ﹥0.05;t ＝1.049,P ﹥0.05）。成熟 CIK 中 CD4＋细胞占（22.8±1.3）％,CD8＋细胞占（77.3±1.8）％,CD3＋ CD56＋ CD16＋细胞占（24.5±2.1）％。致敏 DC-CIK、DC-CIK、CIK 都对胃癌细胞有杀伤作用,致敏 DC-CIK 在效靶比为10：1、20：1、40：1时杀瘤活性分别为（37.68±1.49）％、（41.67±0.90）％、（42.71±0.98）％,在效靶比为40：1时杀瘤活性最强,DC-CIK 组分别为（36.77±0.46）％、（38.94±0.95）％、（41.15±0.89）％,CIK组分别为（34.74±1.01）％、（37.76±0.43）％、（39.65±0.79）％,三组间差异有统计学意义（F ＝5.92, P ﹤0.05;F ＝19.13,P ﹤0.05;F ＝8.88,P ﹤0.05）。结论胃肿瘤抗原致敏脐血 DC 可明显增强 DC-CIK的杀瘤活性,致敏脐血 DC-CIK 在效靶比为40：1时杀瘤活性最强。     

Bcl-2反义寡核苷酸诱导胃癌细胞凋亡的研究

目的　探讨反义寡核苷酸（ＡＳＯＤＮ） 对ｂｃｌ２ 基因的表达调控及诱导胃癌细胞凋亡的作用。方法　应用ＭＴＴ方法比较两条人工合成的ＡＳＯＤＮ 直接作用及其以脂质体（ＤＯＴＡＰ） 为载体对胃癌细胞的抑制效果。应用流式细胞术和ＲＴＰＣＲ 方法检测ＡＳＯＤＮ 对ｂｃｌ２ 蛋白和ｍＲＮＡ 表达量的调控。应用相差显微镜、电子显微镜、流式细胞术等方法,观察ｂｃｌ２ ＡＳＯＤＮ 诱导ＢＧＣ８２３ 胃癌细胞凋亡的效果。结果　ＭＴＴ实验显示两条ＡＳＯＤＮ 抑制胃癌细胞增殖的作用呈浓度和时间依赖性,且靶位点于ｍＲＮＡ蛋白编码区（ＡＳＯＤＮ２） 和以ＤＯＴＡＰ 为载体的ＡＳＯＤＮ 对胃癌细胞的增殖抑制效果为佳。ＡＳＯＤＮ作用于胃癌细胞,降低ｂｃｌ２ 蛋白和ｍＲＮＡ 表达。ｂｃｌ２ ＡＳＯＤＮ处理胃癌细胞,观察到肿瘤细胞缩小、凋亡小体出现、染色质浓缩、凋亡峰等凋亡特征性改变。结论　ｂｃｌ２ ＡＳＯＤＮ可降低ｂｃｌ２ ｍＲＮＡ和蛋白表达,诱导胃癌细胞凋亡     

Apoptosis induced by the sodium butyrate in human gastric cancer TMK-1 cells

The effects of sodium butyrate on cell proliferation, gene expression, and apoptosis were investigated. Upon exposure to sodium butyrate the cells exhibited marked morphological changes, reduced cell proliferation and most cells died through apoptosis within 48 hours. In the presence of dexamethasone, however, the sodium butyrate-triggered apoptosis was markedly reduced. Studies using the glucocorticoid receptor antagonist RU486 indicated that the protective effect of dexamethasone was mediated through glucocorticoid receptor. Sodium butyrate markedly induced the c-jun proteins level, whereas the c-Myc protein was down-regulated rapidly. c-Jun protein may play an important role in the action of sodium butyrate since its induction preceded the onset of DNA fragmentation. In addition, preincubation of the cells with dexamethasone markedly delayed the induction of c-jun levels by sodium butyrate. Analysis of the expression of bel-2-related genes indicated that the Bcl-xS protein level was increased in the presence of sodium butyrate and the up-regulation of Bcl-xS by sodium butyrate was also blocked by dexamethasone. Taken together, these results indicate that c-myc, c-jun and Bcl-xS proteins may be involved in the mechanism of sodium butyrate-triggered apoptosis in these cells.     

12-Lipoxygenase inhibition induced apoptosis in human gastric cancer cells

Arachidonic acid release from membrane phospholipids is essential for tumour cell proliferation. Lipoxygenases constitute a pathway for arachidonate metabolism. The present study investigated the expression of 12-lipoxygenase and its effect on cell proliferation as well as survival in two human gastric cancer cell lines (AGS and MKN-28). RT-PCR and western blots, respectively, showed 12-LOX mRNA and protein expression in both AGS and MKN-28 cell lines. Treatment with a 12-LOX inhibitor, baicalein, significantly inhibited cancer cell proliferation, but a metabolite of 12-LOX activity, 12 hydroxyeicosatetraenoic acid (12-HETE) reversed baicalein-induced growth inhibition. Furthermore, the blockade of the 12-LOX pathway through a 12-LOX inhibitor and antisense induced apoptosis of gastric cancer cell lines. The biochemical characteristics of apoptosis were p53-independent combined with a decrease in bcl-2 expression. Caspase-7 was proteolytically activated and responsible for the apoptosis execution.     

K-ras突变多肽与全细胞抗原致敏DCs诱导CTLs对胰腺癌的杀伤活性研究

Objective  

We studied the role of specific cytotoxic T lymphocytes (CTLs) activated by dendritic cells (DCs) presenting cationic nanoparticles with the K-ras (12-Val) mutant peptide and whole tumor antigen in the killing of different pancreatic cancer cell lines in vitro and in vitro.     

核黄素光化学反应诱导人胃癌细胞凋亡

背景与目的：核黄素在光照下具有抑癌活性。并且被认为与核黄素光化学反应产生的活性氧自由基有关。本研究探讨核黄素在光照处理下,体外诱导人胃癌MGC80-3细胞凋亡效应。方法：以台盼蓝拒染法计数细胞存活率,Giemas染色观察形态改变,DNA琼脂糖电泳分析DNA片段化,测定断裂及未断裂DNA含量定量凋亡效率。流式细胞术及Western blot分别检测细胞周期及凋亡相关基因表达变化。结果：细胞存活率依浓度及处理时间而下降;光照核黄素处理24h,细胞呈典型的凋亡形态;以10,10,30和40μmol/L的核黄素光照处理MGC80-3细胞,DNA电泳显示20μmol/L以上处理组均出现典型的凋亡DNA梯形带;且相应凋亡效率分别为35.4%,54.1%,70.6%和86.8%。与核黄素的浓度呈正相关;经10及20μmol/L核黄素光照处理后细胞周期主要阻滞于G2/M期,Western blot结果显示凋亡相关基因p53,c-myc及Bax表达上调,而Bcl-2表达下调。结论：核黄素光化学反应抑制胃癌MGC80-3细胞生长的作用是以诱导细胞凋亡为主要途径。并与诱导细胞阻滞于G2/M期有关。     

Leukemia-specific T-cell reactivity induced by leukemic dendritic cells is augmented by 4-1BB targeting.

PURPOSE: Acute myelogenous leukemia (AML) blasts are able to differentiate into leukemia-derived dendritic cells (AML-DC), thereby enabling efficient presentation of known and unknown leukemic antigens. Advances in culture techniques and AML-DC characterization justify clinical application. However, additional measures are likely needed to potentiate vaccines and overcome the intrinsic tolerant state of the patients' immune system. Engagement of the costimulatory molecule 4-1BB can break immunologic tolerance and increase CTL responses. In this study, we examined the role of the 4-1BB ligand (4-1BBL) on T-cell responses induced by AML-DC. EXPERIMENTAL DESIGN: In allogeneic and autologous cocultures of T cells and AML-DC, the effect of the addition of 4-1BBL on T-cell proliferation, T-cell subpopulations, and T-cell function was determined. RESULTS: Addition of 4-1BBL to cocultures of AML-DC and T cells induced a preferential increase in the proliferation of CD8(+) T cells. Increased differentiation into effector and central memory populations was observed in both CD4(+) and CD8(+) T cells in the presence of 4-1BBL. AML-DC induce a T helper 1 response, characterized by high IFN-gamma production, which is significantly increased by targeting 4-1BB. T cells primed in the presence of 4-1BBL show specificity for the leukemia-associated antigen Wilms' tumor 1, whereas cytotoxicity assays with leukemic blast targets showed the cytolytic potential of T cells primed in the presence of 4-1BBL. CONCLUSION: We conclude that 4-1BBL is an effective adjuvant to enhance T-cell responses elicited by AML-DC.     

细胞因子诱导杀伤细胞联合树突状细胞生物治疗辅助化疗对胃癌术后患者的治疗效果

目的观察细胞因子诱导杀伤细胞联合树突状细胞(DC-CIK)生物治疗辅助化疗对胃癌术后患者的治疗效果。方法将180例胃癌患者根据治疗方式不同分为观察组和对照组,观察组患者应用DC-CIK联合胃癌根治术及TP方案(紫杉醇+顺铂)化疗,对照组患者仅采用胃癌根治术及TP方案化疗,观察并比较两组患者的不良反应发生情况和3年生存率。结果两组患者治疗后干扰素γ(INF-γ)水平均高于治疗前(均P<0.05)。两组患者不良反应发生率差异无统计学意义(P>0.05);两组患者3年总生存率差异无统计学意义(P>0.05)。观察组患者的进展期生存率明显高于对照组,差异有统计学意义(P<0.05)。结论 DC-CIK生物治疗辅助胃癌术后化疗能够提高细胞因子水平,并且可有效地提高胃癌术后患者进展期生存率。     

白细胞介素24对人树突状细胞活化和成熟的诱导作用

目的: 〖HT5"SS〗研究白细胞介素24（interleukin24,IL24）对人外周血单核细胞来源树突状细胞（dendritic cell, DC）表型及抗原提呈功能的影响。〖HT5W〗方法： 〖HT5"SS〗利用Western blot检测DC培养上清中IL24的分泌水平;利用半定量RTPCR方法分析不同条件下DC表达IL24受体及趋化因子受体的情况;通过流式细胞术分析检测IL24共培养后DC细胞表面CD80、CD86、MHCⅡ类分子的表达水平;采用体外混合淋巴细胞实验分析经IL24共培养对DC抗原提呈功能的影响。〖HT5W〗结果：〖HT5"SS〗 体外培养过程中,用LPS刺激DC可诱导其分泌IL24;同时,LPS还能上调DC表达IL24受体亚单位IL22R1。IL24作用于DC可上调DC细胞表面CD80、CD86及MHCⅡ类分子的表达,并增强DC对T细胞的抗原提呈功能。〖HT5W〗结论： 〖HT5"SS〗IL24这一新型的细胞因子在体外实验中能促进DC的表型成熟,增强DC的抗原提呈功能。     

细胞因子诱导的杀伤细胞治疗胃癌的研究现状

细胞因子诱导的杀伤（cytokine induced killer,CIK）细胞免疫治疗作为一种毒副作用较轻、前景良好的过继性细胞免疫治疗方法,其联合化疗对胃癌的治疗已进行了一定的研究。虽然,目前所报道的临床研究已经证实CIK细胞治疗的安全性,但尚不足以充分证明CIK细胞联合化疗治疗胃癌的有效性,因为这些试验设计方案均存在一些缺陷,如为非随机对照的临床研究、样本量较小、未明确胃癌类型、未标准化的CIK细胞制备方法和治疗方案、无用于预测CIK细胞治疗效果的特异性生物标志物等。因此,迫切需要解决这些问题方能促进CIK细胞用于胃癌临床研究的发展。     

姜黄素诱导人胃癌细胞SGC-7901凋亡的作用机制

 目的 探讨姜黄素诱导人胃癌细胞SGC-7901凋亡的作用及其相关机制。方法 以不同浓度的姜黄素作用于胃癌SGC-7901细胞，利用倒置相差显微镜观察细胞生长形态变化，通过MTT检测细胞生长抑制率，流式细胞术检测细胞凋亡，Western blot 检测胃癌细胞中Fas及survivin的表达情况。结果 姜黄素能显著抑制体外培养的SGC-7901细胞的生长并呈量-效和时-效关系，流式细胞术检测到亚二倍体凋亡峰，细胞凋亡率增加，Western blot结果提示经姜黄素作用后Fas表达率上升，survivin表达率下降。结论 姜黄素能抑制胃癌细胞SGC-7901的生长并促进其凋亡，姜黄素可能通过上调Fas及下调survivin的表达而诱导凋亡。     

Infiltration of dendritic cells in relation to tumor invasion and lymph node metastasis in human gastric cancer

The infiltration of dendritic cells determined in 210 patients with gastric carcinoma was investigated from the standpoint of tumor invasion, lymph node metastasis, and prognosis. Dendritic cell infiltration was graded as "slight" and "marked." The 39% frequency in the marked infiltration group at the mucosal stage did not change in proportion to invasion into the deeper layers. The 5-year survival rate was 60.4% in patients with marked infiltration and 38.8% in those with slight infiltration, which was statistically different (P less than 0.01). The difference in survival rates was only statistically significant in those with cancer emerging from the serosa (P less than 0.001). There was a similar incidence of lymph node metastasis between the marked and slight infiltration groups in each grade of tumor invasion. However, marked infiltration of dendritic cells prevented widespread nodal involvement beyond the primary node in cases of advanced carcinoma (P less than 0.05). These findings indicate that infiltrating dendritic cells do not prevent the spread of tumor invasion but do prevent nodal involvement; therefore, for patients with a gastric cancer emerging from the serosa, the prognosis will be good.     

基因腺病毒感染树突状细胞诱导CTL对胶质瘤细胞的杀伤作用

目的:研究EPHA2基因修饰的树突状细胞(dendritic cell,DC)疫苗诱导细胞毒性T淋巴细胞(cytotoxic T lympho-cyte,CTL)对U251胶质瘤细胞的杀伤效应,为胶质瘤的免疫治疗提供新的方法。方法:将重组EPHA2腺病毒rAd-EPHA2感染HLA-A2阳性的人外周血来源的DC,制备EPHA2基因修饰的DC疫苗,Western blotting和FACS方法检测感染后DC的EPHA表达。以DC疫苗体外刺激HLA-A2阳性的单个核细胞,酶联免疫斑点实验(enzyme-linked immunospot assay,ELISPOT)和标准51Cr释放实验分别检测DC疫苗所诱导的CTL活性和对HLA-A2阳性的U251细胞的杀伤作用(另设rAd-Lac Z感染的DC组和PBS组作为对照)。结果:成功制备了EPHA2基因修饰的DC疫苗,其可有效表达EPHA2蛋白。与感染rAd-LacZ的DC和PBS组相比,感染rAd-EPHA2的DC疫苗能有效激发CTL活性[(187±21)vs(12±4)、(18±5)个,P<0.01];所诱导的CTL对胶质瘤U251细胞有明显的杀伤效应[(45.7±6.8)%vs(7,1±4.5)%,P<0.01],对自身淋巴细胞没有杀伤效应。结论:EPHA2基因修饰的DC疫苗能有效激发CTL活性,并对胶质瘤U251细胞有明显的杀伤活性。     

Induction of cytotoxic T lymphocytes against human cancer cell lines using dendritic cell-tumor cell hybrids generated by a newly developed electrofusion technique

Recently, dendritic cells (DCs) and DC-tumor cell hybrids (DC-tumor hybrids) have been used for cancer vaccine therapy in a clinical trial. DC-tumor hybrids combine the potent antigen-presenting capacity of DCs with the ability to present all tumor antigens expressed on tumor cells to T cells. We used DC-tumor hybrids as stimulator cells to induce tumor-specific cytotoxic T lymphocytes (CTLs) in vitro. DC-tumor hybrids were generated from human monocyte-derived DCs and human cancer-cell lines (GT3TKB, lung cancer; GCIY, gastric cancer) by our newly developed electrofusion technique, established and refined with the use of mouse cells. To evaluate the capacity of DC-tumor hybrids generated by our method to induce tumor antigen-specific CTLs, we performed a cytotoxic assay and an interferon-gamma release assay using CD8-dominant effector lymphocytes induced by them. DC-tumor hybrids more effectively induced tumor-specific primary T-cell response than did stimulation with DCs co-cultured with irradiated tumor cells overnight, irradiated tumor cells alone, or a mixture of DCs and irradiated tumor cells. DC-tumor hybrids were generated at a high fusion rate by our electrofusion technique. When CTLs were induced by DC-tumor hybrids in vitro, the high fusion rate did not contribute to the induction of CTLs with increased tumor-specific cytotoxicity. The addition of interleukin-12 to the culture medium did not augment the cytotoxicity of CTLs. Overall, our results suggest that DC-tumor hybrids effectively induce human tumor-specific CTLs and may thus be applicable for clinical trials of adoptive immunotherapy.     

紫杉醇诱导胃癌细胞凋亡的实验研究

 目的　探讨紫杉醇对胃癌细胞的诱导凋亡作用及其诱导的胃癌细胞凋亡的周期时相性。方法　用Sub-G1法检测紫杉醇诱导胃癌细胞MKN-28的凋亡,API法检测紫杉醇诱导胃癌细胞凋亡的周期时相性,并分选后激光共聚焦显微镜技术（PSC）观察形态学,MTT法检测紫杉醇对临床胃癌组织细胞的敏感性。结果　Sub-G1法结果显示紫杉醇能诱导胃癌细胞的凋亡,紫杉醇（浓度10 mg／L）诱导胃癌细胞凋亡在10 h后达到高峰;API法检测结果显示紫杉醇诱导胃癌细胞发生凋亡的时相在G2／M期,PSC形态学观察到G2／M期凋亡特征;MTT法结果显示紫杉醇诱导的20例临床胃癌组织标本中,有16例抑制率大于50 %。结论　紫杉醇能诱导胃癌细胞发生凋亡,相对较敏感,其诱导的胃癌细胞凋亡具有周期时相性。     

维生素E琥珀酸酯诱导人胃癌细胞发生未折叠蛋白反应的研究

目的：探讨维生素E琥珀酸酯（VES）对人胃癌细胞未折叠蛋白反应的影响。方法：采用Western blot法分析VES不同剂量（0、5、10和20 μg/mL）处理人胃癌SGC-7901细胞24 h和20 μg/mL VES处理SGC-7901细胞不同时间（0、6、12、18和24 h）对内质网膜上的跨膜蛋白PKR样内质网蛋白激酶（PERK）磷酸化和活化转录因子6（ATF6）活化的影响;采用RT-PCR法检测VES不同剂量（0、5、10和20 μg/mL）处理SGC-7901细胞24 h和20 μg/mL VES处理SGC-7901细胞不同时间（0、3、6、9、12、15、18和24 h）对活化转录因子4（ATF4）mRNA表达和X盒结合蛋白1（XBP1）mRNA剪切的影响。结果：在翻译水平上VES对磷酸化PERK的诱导作用具有时间依赖关系,与阴性对照组比较,10 μg/mL VES的诱导作用最强（P < 0.05）;在翻译水平上VES对ATF6活化片段的诱导作用具有时间依赖关系和剂量依赖关系;在转录水平上,随着作用时间的延长,VES对ATF4 mRNA的诱导作用也不断增强,18 h达到高峰,而XBP1 mRNA从VES作用6 h即开始出现剪切。结论：VES诱导人胃癌SGC-7901细胞发生内质网应激过程中,未折叠蛋白反应相关信号分子被活化。     

树突状细胞诱导的LAK细胞对肺癌的生长抑制作用

目的　研究肺癌细胞裂解物负载的树突状细胞 (DC)诱导的淋巴因子激活杀伤细胞 (LAK )对肺腺癌裸鼠移植瘤的生长抑制作用。方法　肺腺癌患者手术切除标本接种裸鼠皮下建立肺腺癌移植瘤模型 ,同时将手术切除标本用反复冻融的方法制备肿瘤细胞裂解物 ;同一患者外周血分离得到的单个核细胞中 ,贴壁的细胞加入DC生长因子 (DCGF)培养得DC ,未贴壁的细胞加入rhIL 2培养得LAK细胞。用肿瘤细胞裂解物负载自体DC ,诱导LAK细胞生成DC LAK细胞 ,注射荷瘤裸鼠腋下以观察DC LAK细胞对肺癌移植瘤的生长抑制作用。结果　用肺癌手术标本能成功建立裸鼠移植瘤模型。DC LAK组、LAK组、DC组和生理盐水对照组肿瘤瘤重平均值分别为 0 .47、1.0 5、1.3 0和 1.5 8g ,DC LAK组、LAK组和DC组肿瘤生长抑制率分别为 70 .3 %、3 3 .5 %和 17.9% ,DC LAK细胞具有抑制裸鼠肺癌移植瘤生长的作用而且强于LAK细胞 (P <0 .0 5 )。结论　通过荷瘤裸鼠体内抗瘤实验证实了DC LAK细胞的抗肺癌作用明显高于LAK细胞 ,提示DC LAK细胞治疗是更为有效的抗肺癌生物治疗方法 ,为DC疫苗用于肺癌的临床治疗提供依据     

肺癌可溶性抗原联合超抗原修饰致敏ＤＣ诱导抗瘤免疫的研究

目的　观察经肺癌肿瘤可溶性抗原（ＴＳＡ）和超抗原金黄色葡萄球菌肠毒素Ａ（ＳＥＡ）联合修饰致敏树突状细胞（ＤＣ）体外诱导抗肺癌的免疫效应。方法　３ｍｏｌ／Ｌ氯化钾提取法获得人肺癌细胞ＧＬＣ ８２的可溶性抗原;从人外周血单个核细胞（ＰＢＭＣ）中诱导扩增ＤＣ,并用流式细胞仪（ＦＣＭ）检测表型;以肺癌ＴＳＡ和ＳＥＡ联合修饰致敏的ＤＣ、单纯肺癌抗原致敏的ＤＣ和未经抗原修饰的ＤＣ分别与同种异体外周血Ｔ淋巴细胞共同孵育,刺激Ｔ淋巴细胞活化增殖（作为效应细胞分别称为ＴＳＡ-ＳＥＡ-ＤＣＬ、ＴＳＡ-ＤＣＬ、ＤＣＬ）,直接活细胞计数法观察增殖倍数;ＭＴＴ法检测不同ＤＣ∶Ｔ淋巴细胞比例的效应细胞对靶细胞ＧＬＣ-８２的体外杀伤效应。结果　诱导出高表达ＣＤ１ａ、ＣＤ８０、ＨＬＡ-ＤＲ的ＤＣ,光镜下具有典型的ＤＣ特性;联合抗原修饰后的ＤＣ具有较强的免疫刺激活性,少量致敏ＤＣ即可强烈激发Ｔ细胞的增殖;ＴＳＡ-ＳＥＡ-ＤＣＬ对靶细胞ＧＬＣ-８２的杀伤率明显高于ＴＳＡ-ＤＣＬ及ＤＣＬ;联合抗原修饰的ＤＣ以１∶１００与Ｔ淋巴细胞共孵后的杀伤肿瘤细胞效应最强。结论　经肺癌ＴＳＡ和ＳＥＡ联合修饰致敏的ＤＣ可强烈激发同种异体Ｔ淋巴细胞活化增殖;肺癌ＴＳＡ联合超抗原ＳＥＡ诱导的ＤＣ疫苗对肺癌细胞有高效杀伤作用,经肺癌ＴＳＡ与超抗原ＳＥＡ联合修饰ＤＣ的活性明显强于单用肺癌ＴＳＡ。     

Induction of higher-avidity human CTLs by vector-mediated enhanced costimulation of antigen-presenting cells.

The efficacy of antigen-specific CD8(+) CTLs depends not only on the quantity of CTLs generated but also perhaps, more importantly, on the avidity of the CTLs. To date, however, no strategy has been shown to preferentially induce higher-avidity human CTLs. In the present study, antigen-presenting cells (APC) generated from human peripheral blood mononuclear cells were infected with a recombinant avipox vector (rF-) containing the transgenes for a triad of costimulatory molecules (human B7.1, intercellular adhesion molecule-1, and LFA-3, designated as rF-TRICOM) and then used to elicit peptide-specific CTLs from autologous T cells. Compared with peptide-pulsed noninfected APCs or peptide-pulsed APCs infected with wild-type vector, peptide-pulsed APCs infected with rF-TRICOM induced not only more CTLs but also higher-avidity CTLs; this was shown by tetramer staining, tetramer dissociation, IFN-gamma production, and cytolytic assays. Peptide-pulsed rF-TRICOM-infected dendritic cells were also shown to induce CTLs with a >10-fold higher avidity than CTLs induced using CD40L-matured dendritic cells; the use of peptide-pulsed CD40L-matured dendritic cells infected with rF-TRICOM as APCs induced CTLs of even greater avidity. To our knowledge, these studies are the first to show a methodology to induce higher-avidity human CTLs and have implications for the development of more efficient vaccines for a range of human cancers.     

细胞因子诱导杀伤细胞对裸鼠胃癌移植瘤的靶向抑制作用

目的: 利用蛋白质组学基质辅助激光解析离子化飞行时间质谱(matrixassisted laser desorption/ionization timeofflight mass spectrometry, MALDITOF MS)技术检测食管癌患者血清蛋白指纹图谱,建立食管癌蛋白指纹图谱诊断模型,探讨其临床应用价值。方法: 收集河北医科大学第四医院2008年5月至9月间胸外科食管癌患者血清32例和健康志愿者血清28例,采用弱阳离子交换蛋白质芯片（WCX磁珠）提纯血清蛋白,MALDITOF MS技术进行血清蛋白质质谱检测,所得结果用ZUCI蛋白芯片数据分析系统进行分析。运用遗传算法结合支持向量机运算建立食管癌蛋白指纹图谱诊断模型;将60例标本随机分成训练组和盲法测试组,训练组为21名食管癌患者和19名健康人,盲法测试组为11名食管癌患者和9名健康人,验证诊断模型的特异性和敏感性。结果:采集食管癌患者和健康对照者的血清蛋白指纹图谱,经数据对比分析找到44个有显著性差异的质荷比峰（P<0.05）;从中筛选出差异最显著的6个蛋白质荷比峰（m/z分别为2 210、2 864、6 634、4 068、2 083和8 131）,并以此建立食管癌蛋白指纹图谱诊断预测模型;验证该模型诊断食管癌的特异性为88.9%、敏感度为100%。结论: 应用MALDITOF MS技术检测食管癌患者血清蛋白指纹图谱建立的食管癌蛋白指纹图谱诊断模型在食管癌的诊断中具有较高的敏感度和特异性。