IL-12对小鼠肥大细胞瘤基因疫苗的免疫学作用

目的 研究小鼠肥大细胞瘤P815基因疫苗和鼠IL-12对该疫苗的免疫学作用。方法 将小鼠肥大细胞瘤P815特异抗原基因P1A克隆到真核表达质粒pCI-neo中；用P815细胞对DBA/2小鼠右腹侧皮下注射，构建P815小鼠肿瘤模型；以重组基因疫苗单独或与鼠IL-12真核表达质粒一起肌肉注射，观察肿瘤的消长，特异细胞毒T淋巴细胞激活和抗全的生成情况。结果 重组基因疫苗在体外有很好的表达，注射后CTL的杀伤效率为40%，IL-12共注射的CTI，杀伤效率达到60%，免疫后，30%小鼠的肿瘤出现消退；同IL-12共注射则有50%的小鼠的肿瘤出现消退，2种情况下都不能检测到任何特异抗体的产生。结论 重组P1A肿瘤疫苗能有效激活机体的肿瘤特异免疫应答；基因疫苗对小鼠P815肿瘤的治疗作用主要归因于细胞免疫；IL-12有增强这种免疫应答的作用。     

095 肿瘤特异性抗原基因克隆技术的新进展

肿瘤特异性抗原的获得、鉴别一直是肿瘤免疫研究的热点之一,分子生物学技术的飞速发展,给这一领域带来了勃勃生机.本文综述了应用分子生物学技术分离肿瘤特异性基因,并用免疫学方法鉴定其抗原性,为获取肿瘤特异性抗原开避新途径.     

肿瘤标志物检测技术研究进展

肿瘤标志物的联合检测为肿瘤的筛查和早期诊断带来了新的发展机遇.寻找具有较高灵敏度和特异性的肿瘤标志物一直是肿瘤研究领域的热点之一.随着分子生物学、免疫学诊断技术的飞速发展,通过多种检测技术的联合应用,互相补充,以实现对肿瘤的早期诊断和治疗.     

肿瘤疫苗的研究进展

肿瘤疫苗是用肿瘤细胞、肿瘤细胞裂解物或肿瘤抗原激活机体免疫系统产生特异性抗肿瘤细胞免疫效应.它是一种治疗性的、新型的肿瘤治疗方法,也是一种主动性免疫疗法.随着肿瘤免疫学和分子生物学的发展,肿瘤与机体之间的相互作用、肿瘤免疫耐受以及肿瘤抗原鉴定都取得了很大的进展,这也促进了肿瘤疫苗的发展.     

基于α-病毒复制酶的高效小鼠P815肿瘤疫苗的构建和表达

目的：克隆小鼠肥大细胞瘤P815细胞株的肿瘤特异PIA基因于含α－病毒复制酶的真核表达载体PSMART中，以制备P815肿瘤DNA疫苗。方法：RT－PCR方法扩增P1A基因，以含α－病毒复制酶的哺乳细胞高效表质质粒PSMART为载体，构建重组肿瘤DNA疫苗，重组体经酶切和测序后，再用脂质体转化293人胚肾细胞，ECL western-blot法和免疫组化法鉴定转化细胞中PIA基因的表达。结果：正确构建了P1A／PSMART重组质粒，并且在转化此质粒的293细胞中检测出了P1A的表达。结论：成功构建了重组P1A／ pSMART肿瘤DNA疫苗，可以进行下一步的肿瘤动物模型的疫苗接种及疗效观察。     

肿瘤特异性抗原基因克隆技术的新进展

肿瘤特异性抗原的获得、鉴别一直是肿瘤免疫研究的热点之一 ,分子生物学技术的飞速发展 ,给这一领域带来了勃勃生机。本文综述了应用分子生物学技术分离肿瘤特异性基因 ,并用免疫学方法鉴定其抗原性 ,为获取肿瘤特异性抗原开避新途径     

颗粒性肽-DNA复合疫苗抗肿瘤免疫的研究

诱导有效的抗肿瘤免疫应答是肿瘤免疫治疗的主要目标 ,由于肽疫苗和DNA疫苗具备较高的安全性和实用性 ,因而成为肿瘤疫苗学领域中发展最为迅速的肿瘤免疫治疗方案之一。但是这两种疫苗形式仍然具有各自的缺点 ,其抗原呈递动力学存在显著差异 ,提示这两种疫苗可能存在互补性。本研究对多肽疫苗和DNA疫苗进行综合 ,设计出新型肽 DNA复合疫苗 ,并以P815A为模式抗原 ,证实了该种新型疫苗激发抗肿瘤免疫的有效性     

HBV与HCV融合基因免疫的抗肿瘤作用研究

目的 :观察基因疫苗SpcDNA3.1、CpcDNA3.1及融合基因疫苗CSpcDNA3.1诱导BALB c小鼠 (H 2 d)的特异性细胞免疫应答及其对稳定表达HBsAg和HCcAg的小鼠肥大细胞瘤P815细胞 (H 2 d)接种动物后成瘤性的影响。方法 :3种重组质粒SpcDNA3.1、CpcDNA3 1和CSpcDNA3 1分别肌注免疫小鼠 ,3w后背部皮下接种质粒CSpcDNA3 1转染的P815细胞 ,观察小鼠成瘤和存活时间。LDH法检测免疫小鼠的脾淋巴细胞CTL活性。结果 :融合质粒CSpcDNA3 1可显著抑制肿瘤出现和生长 ,小鼠生存时间明显延长 ,生存率提高 ,CSpcDNA3 1免疫的小鼠脾淋巴细胞体外对转染的P815肿瘤细胞有明显的杀伤作用。结论 :融合基因疫苗CSpcDNA3 1免疫的小鼠能诱发特异性抗肿瘤细胞免疫。     

Th17细胞在肿瘤免疫中的作用研究进展

自从2005年发现产生IL-17的Th17细胞以来,作为辅助性T细胞的新亚群,Th17细胞受到了广泛的关注,并成为免疫学研究的热点之一.Th17细胞在自身免疫病、感染性疾病以及移植排斥反应中发挥非常重要的作用,但是Th17细胞在肿瘤免疫中的作用却知之甚少.目前认为Th17细胞在肿瘤免疫中具有双重作用.现就Th17细胞及其在肿瘤免疫中的作用做一综述.     

2017 年国内外免疫学研究重要进展

2017 年,国内外免疫学研究在基础研究和临床应用多个分支领域取得令人激动的新成果、新进展,如发现了表观遗传修饰、RNA 修饰、蛋白质翻译后修饰及能量代谢等多种胞内调控机制在免疫系统发育和功能的新作用,对于诸多免疫学科学问题有了更加深入的本质性认识。在转化医学及免疫治疗方面也取得重大进展,特别是在肿瘤发生发展机制探索和肿瘤免疫治疗领域取得了一系列重要进展,对于寻找肿瘤诊断、治疗及疗效预判新型靶标、建立新型联合肿瘤免疫治疗方法具有重要的意义。国内免疫学研究者近年来研究水平和创新能力不断提高,涌现出一批受到国际同行认可的创新性科研成果,成为国际免疫学研究的重要力量。此文中,我们梳理总结了2017 年国内外免疫学研究领域较为代表性的理论研究及转化应用新成果,与各位同行共同探讨免疫学研究的最新前沿和重大挑战。     

小鼠肥大细胞瘤P815模型的建立与初步应用

目的 建立小鼠肥大细胞瘤Ｐ８１５种植瘤模型，并对其体内诱发特异性ＣＴＬ应答机理进行初步的研究。方法 经有限稀释法筛选Ｐ８１５单克隆瘤系，用ＲＴ－ＰＣＲ及产物ＤＮＡ测序鉴定肿瘤抗原Ｐ１Ａ基因的表达；在此基础上，用活瘤苗免疫同系小鼠，经标准＾５１Ｃｒ释放试验检测在体特异性ＣＴＬ的活性。结果 筛选出了一个Ｐ８１５单克隆成瘤系Ｐ８１－Ｆ３，并鉴定该瘤存在Ｐ１Ａ基因的表达，用活瘤疫苗免疫６只步鼠，在３只体内     

Experimental gene therapy of cancer using tumor cells engineered to secrete interleukin-13

Cytokines locally delivered to the site of a tumor boost both specific and nonspecific host anti-tumor defenses. Interleukin (IL)-13 is a recently described cytokine produced by mouse type 2 helper T lymphocytes. The aim of this study was to evaluate the inhibition of tumor growth induced by IL-13 delivered locally within or around transplanted tumor cells in mice. We observed that local administration of IL-13 at the site of transplanted tumor cells in vivo had potent inhibitory effects on growth of both immunogenic (P815 mastocytoma, H-2^d) or nonimmunogenic (3LL lung carcinoma, H-2^b) tumor cells. Mice injected with transfected P815 cells secreting large amounts of IL-13 rejected the P815 tumor and developed systemic specific anti-tumor immunity leading to long-lasting specific anti-tumor protection. Less efficient anti-tumoral effects were obtained with the nonimmunogenic 3LL tumor model when local administration of IL-13 was achieved by co-inoculating xenogeneic chinese hamster ovary (CHO) IL-13 cells. Several local injections of CHO IL-13 cells were needed to obtain rejection of 3LL tumors and no induction of long-lasting anti-3LL memory was obtained. Several studies were performed to elucidate the IL-13 anti-tumoral effects. Experiments with nude mice indicated that IL-13 can also stimulate nonspecific anti-tumor defenses. The histological examination of P815 IL-13 cells undergoing rejection showed monocytic cells and neutrophils infiltrating the tumor. Studies indicated that IL-13 administered in vitro did not directly stimulate the cytotoxicity of peritoneal macrophages and natural killer cells. However, experiments with Boyden chemotaxis chambers indicated that IL-13 was chemotactic for macrophages. Finally, preliminary experiments in vitro suggest that IL-13 improved antigenic presentation of P815 membranes. Thus, anti-tumor effects of IL-13 in vivo most probably result from pleiotropic effects including recruitment of nonspecific cells and improved stimulation of immune-specific anti-tumor effectors.     

Quantitation of tumor growth in the ears of mice

Transplantation of ascitic P-815 mastocytoma cells intradermally in the ears of syngeneic DBA/2J mice allows a simple and precise quantitation of tumor growth by ear swelling. Tumor growth was retarded and then arrested in allogeneic DBA/1J mice, whereas syngeneic hosts died as a result of tumor dissemination in draining lymph nodes, liver and spleen.     

Protein tumor antigens.

Immunogenic tumor antigens have been sought through a variety of paradigms for several past decades. Recent developments in antigen presentation have radically changed the prism through which we view these enigmatic antigens. This article discusses the small but growing treasure chest of these antigens--stress-induced proteins, the P1AB antigen of the P815 mastocytoma, the p53 tumor suppressor protein, the gp95/p97 antigen of human melanoma, mucins and others.     

BALB/c小鼠肥大细胞瘤/白血病模型的建立

目的研究建立BALB/c小鼠肥大细胞瘤/白血病模型。方法 BALB/c小鼠尾静脉注入小鼠肥大细胞瘤P815细胞,实验按每只小鼠接种细胞数量分为1×10^7/只组、5×10^6/只组、2.5×10^6/只组、1×10^6/只组、5×10^5/只组、1×105/只组和溶剂（RPMI1640培养液）对照组。观察小鼠的生存状态、体重及肝肺脾（重量、形态、病理）、染色体、血涂片、骨髓涂片、白细胞及血小板计数。结果注入P815细胞≥1×106/只的所有组小鼠均死亡,〈1×10^6/只的所有组生存良好,且死亡小鼠的生存时间与注入细胞数量呈负向依赖关系（P〈0.05）。死亡组小鼠体重较注射前相当或减轻、肝肺脾重较对照组及未死亡组显著增加（P〈0.05）;肝质地变硬,表面可见大量大小不一的白色结节突起,部分脾肺可见出血梗死灶,病理示肝脾有大量异形肥大细胞瘤细胞浸润;脾细胞染色体分析可见肥大细胞瘤细胞的非正常染色体核型;白细胞和血小板计数较对照组降低（P〈0.01）,血涂片可见少量肥大细胞瘤细胞,骨髓涂片示骨髓原始细胞比例增高。结论 P815细胞通过静脉注射能使BALB/c小鼠形成肥大细胞瘤/白血病,可作为肿瘤实验研究的一种模型构建方法。     

A radio (51Cr) micro-tube leukocyte adherence inhibition assay: specific tumor-associated immunity in 3 murine tumor systems.

A radio (51Cr) micro-tube leukocyte adherence inhibition assay is described. In this study, murine mononuclear cells were labeled with 51Cr, plated into tissue culture plates with different tumor extracts and counts/min (cpm) of the non-adherent cells were used as a parameter of adherence inhibition. This assay was used to measure anti-tumor immunity, in vitro, in 3 murine tumor systems: MCA-38 colon adenocarcinoma, L1210 lymphoma and P815 mastocytoma. Tumor immunity was detected using 3 doses (0.01-0.001 mg/ml) of tumor extract in the MCA-38 tumor model, and using 2 doses (0.1-0.05 mg/ml) of tumor extract in both the L1210 and P815 tumor models. It was observed that specific tumor-associated adherence inhibition could be measured in the MCA-38 tumor model between days 7 and 22 of tumor growth and in the L1210 and P815 tumor models between days 7 and 17 of tumor growth. The radio-LAI assay described is an easy, specific and reproducible way to measure tumor-associated adherence inhibition, in vitro.     

In vitro and in vivo production of chemotactic inhibitors by tumor cells.

The ascites fluid or peritoneal washings of DBA/2 mice bearing the P815 mastocytoma have been found to contain a chemotactic factor inactivator (CFI) which inactivates the bacterial chemotactic factor as well as the chemotactic activity associated with the C3 and C5 fragments when assayed on rabbit neutrophils. The amount of CFI is proportional to the number of tumor cells in the peritoneal exudate. The inactivator is also found in tumor cell hemogenates as well as in culture fluid from tumor cells growing in vitro. The activity is heat-labile but is not affected by protease inhibitors. Its molecular weight is greater than 50,000 daltons, based on Sephadex chromatography and sucrose density gradient ultracentrifugation studies. In C57BL/6 mice, which reject the mastocytoma, CFI levels decrease in proportion to the decreasing numbers of tumor cells.