致敏树突状细胞活化细胞毒性T细胞抗肿瘤作用的研究

目的：研究树突状细胞（DCs）激活的细胞毒性T细胞的抗肿瘤及预防肿瘤发生的作用。 方法： 细胞因子诱生人PBMC未成熟DCs，加入肿瘤细胞抗原提取物致敏DCs产生成熟DCs；通过细胞形态、表面标记鉴定成熟DCs，MTT法测成熟DCs活化的细胞毒性T细胞(CTL)的体外杀伤活性；裸鼠体内注射活化CTL观察其抑制移植瘤生长及发生的作用。 结果： 经过7 d培养，获得大量形态典型、具有强烈刺激增殖能力、高表达CD80(63.5%)、CD83（67.6%）和CD3/ HLA-DR(83.2%)的DCs。其活化的CTL在20∶1效靶比时对抗原来源细胞株自身的杀伤率达75%以上，对同系细胞株的杀伤活性为35%-45%，对其它种系肿瘤细胞仅有微弱杀伤力（P<0.01）。CTL对裸鼠结肠癌HT-29移植瘤有特异性的生长抑制和预防生成作用（P<0.05）。CTL治疗组肿瘤组织中PCNA表达水平显著低于对照组（P<0.05）。 结论： 肿瘤细胞抗原活化的DC诱导CTL对肿瘤有特异性的杀伤作用，体内应用可特异性抑制移植结肠癌的生长或预防小鼠结肠癌移植瘤的发生。     

四种卡介苗重组疫苗对小鼠免疫原性的研究

目的研究esat6-卡介苗重组疫苗、mpt64-卡介苗重组疫苗、ag85a-卡介苗重组疫苗、ag85b-卡介苗重组疫苗的免疫原性。方法通过ELISA法检测小鼠血清中特异性抗体,用MTS法分析其脾淋巴细胞增殖指数。结果以生理盐水、卡介苗为对照,用各卡介苗重组疫苗免疫小鼠后,各组小鼠体重变化与对照组相比差异无统计学意义。应用不同抗原检测各组小鼠不同时间采集的血清,各组有其特异性抗体产生;ag85a-卡介苗重组疫苗能提高卡介苗刺激B细胞产生抗体的水平;各卡介苗重组疫苗免疫后15d,抗体水平已有升高,45d时达到最高水平。用不同抗原刺激各组小鼠脾淋巴细胞,平均刺激指数均达2.0以上,esat6-卡介苗重组疫苗组小鼠脾淋巴细胞的刺激指数稍高,其它各组之间差异无统计学意义。结论结核分枝杆菌esat6、mpt64、ag85a、ag85b基因在卡介苗中都能够表达特异的蛋白,刺激小鼠产生特异性抗体,均能刺激T淋巴细胞产生细胞免疫。     

重组卡介苗表达的MalE蛋白T细胞表位的鉴定

目的 鉴定重组卡介苗表达的MalE蛋白T细胞表位。方法 用抗原提呈试验、T细胞增殖试验、ELISPOT试验和表位作图测试重组MalE蛋白的T细胞表位。结果 重组BCG.MalE功能性表达了MalE蛋白的4种H-2^d限制性T细胞表位。结论 在4种表位中，p68-82是重组MalE蛋白的主要T细胞表位，而其余3个为次要表位。     

血吸虫重组卡介苗--Sj26GST疫苗的有关毒性研究

由于BCG免疫效果不够理想 ,以及结核分枝杆菌耐药性的产生 ,促使人们开始研制新的结核病疫苗。利用BCG独特的安全性和免疫佐剂作用和日本血吸虫中相对分子量为 2 6× 10 3 的谷光甘肽S转移酶 (Sj2 6GST)的良好免疫原性 ,已经构建出了一种重组卡介苗———Sj2 6GST(rBG Sj2 6GST)。我们就这种疫苗对小鼠的毒性进行了研究 ,从而为疫苗的安全性提供实验依据。将rBG Sj2 6GST疫苗及BCG、含质作者单位 :华中科技大学同济医学院病原生物学系 (蔡昌学 ,叶嗣颖 ,常燕子 ) ;湖北药检专科学校检验系微生物学及免疫学教研室 (胡佳杰 )通讯…     

非自身肿瘤热休克蛋白可用作抗肿瘤疫苗

科学研究业已证明。免疫系统能识别和破坏癌细胞是毋庸置疑的。事实上，大量肿瘤相关抗原或肿瘤特异性抗原都已经得到了鉴定，通常是在通过癌症患者T细胞的识别基础上得出结论的。细胞毒性T淋巴细胞(CIL)杀灭肿瘤细胞的能力被人们认为是肿瘤免疫疗法中最重要的发现之一。     

HBV S基因修饰树突状细胞疫苗诱导特异性CTL反应的探讨

目的分析腺病毒载体介导HBVS基因修饰树突状细胞（DCs）能否诱导抗HBV特异性CTL反应。方法制备携带HBVS基因的重组腺病毒，分别转染外周血诱导培养的DCs，观察腺病毒转染DCs效率和DCs中HBV抗原的表达；混合淋巴细胞反应测定HBV抗原基因修饰DCs刺激同种异体T淋巴细胞增殖能力；乳酸脱氢酶释放法检测特异性CTL细胞对HepG2 22．1．5靶细胞的杀伤能力。结果腺病毒载体能够高效介导HBVS基因在DCs中表达，且DC细胞形态完整；HBVS基因修饰DCs具有刺激同种异体T细胞增殖能力，同时能诱导抗HBV特异性CTL反应。结论HBVS基因修饰DCs疫苗具有增强抗HBV特异性CTL效应的能力，可能发展为一种新型抗病毒疫苗。     

人黑色素瘤抗原-3冲击的树突状细胞诱导抗肿瘤免疫应答

目的:观察经MAGE-3蛋白抗原冲击的树突状细胞在体外诱导抗肿瘤免疫应答的能力。方法:用粒-巨噬细胞集落刺激因子和白介素-4从人外周血分化、诱导树突状细胞,经MAGE-3蛋白冲击后,与T淋巴细胞共培养7d,收集T淋巴细胞进行免疫应答及肿瘤细胞杀伤试验。结果:与MAGE-3冲击后的树突状细胞共培养的T淋巴细胞在体外能有效地杀伤MAGE-3阳性的靶细胞。结论:经MAGE-3蛋白抗原冲击的树突状细胞,在体外能有效提呈抗原,激活抗原特异性CTL,诱导特异性抗肿瘤免疫应答。     

不同大鼠胶质瘤抗原致敏的DC体外诱导CTL活性的研究

目的: 比较大鼠骨髓来源的树突状细胞(dendritic cells, DC)体外经由大鼠C6胶质瘤细胞由不同方式制备的不同抗原致敏后,对特异性细胞毒性T淋巴细胞(cytotoxic T lymphocyte, CTL)的诱导作用。方法: 自大鼠骨髓分离DC前体细胞,经重组大鼠粒细胞巨噬细胞集落刺激因子(rrGM-CSF)+白细胞介素4(rrIL-4)诱导培养、扩增;由C6胶质瘤细胞经由反复冻融、煮沸灭活及超声破碎细胞抽提其总蛋白的方法制备各种不同抗原致敏DC,致敏的DC与T淋巴细胞进行共培养诱导CTL;以ELISA法检测CTL诱导过程中淋巴细胞趋化因子(lymphocyte chemoattractant factor)及细胞因子IFN-γ分泌水平:以 -TdR掺入法检测DC诱导T细胞增殖及其特异性CTL杀伤活性。 结果: 体外应用煮沸灭活瘤细胞制备的肿瘤抗原致敏DC,能诱导更强的刺激T细胞增殖的能力、并且可以诱导杀伤活性更强的CTL。结论: 应用煮沸灭活的瘤细胞制备瘤抗原负载DC获得瘤苗可获得更强的抗肿瘤保护作用。     

HPV 16 E6E7基因修饰树突状细胞疫苗诱导CTL致CaSki细胞凋亡体外实验研究

目的:采用人乳头状瘤-16(HPV-16)E6E7重组腺病毒(pAd-E6E7)转染树突状细胞(Dendritic cell,DC),观察基因修饰的DC疫苗诱导细胞毒性T淋巴细胞(Cytotoxic Tlymphocyte,CTL)致使CaSki细胞凋亡的效果。方法:将pAd-E6E7转染体外培养的小鼠未成熟树突状细胞制备DC疫苗,激光共聚焦显微镜观察转染的小鼠未成熟树突状细胞绿色荧光蛋白表达,流式细胞术检测转染前后小鼠树突状细胞表面标志物(CD40、CD86、MHCⅡ和CD11C)。DC疫苗诱导产生特异性细胞毒性T淋巴细胞,与CaSki细胞共培养后,采用DAPI、TUNEL及流式细胞术检测CaSki细胞凋亡情况。结果:pAd-E6E7成功转染体外培养的小鼠未成熟树突状细胞,体外转染效率约为40%～50%,成功制备了HPV16 E6E7基因修饰树突状细胞疫苗,诱导产生细胞毒性T淋巴细胞,经DAPI、TUNEL及流式细胞术检测证明CaSki出现凋亡。结论:以带有HPV16 E6E7基因的重组腺病毒载体转染DC制备基因修饰的DC疫苗,诱导CTL致使CaSki细胞出现凋亡。     

树突状细胞肿瘤疫苗的研究进展

在抗肿瘤免疫应答中,关键的一步是抗原呈递细胞将肿瘤抗原呈递给T淋巴细胞。树突状细胞是专职抗原呈递细胞,它能够有效的将肿瘤抗原呈递给T细胞,引发机体产生抗肿瘤的免疫应答。因此,利用树突状细胞制备肿瘤疫苗可望提供一种有效的肿瘤免疫治疗方法。目前,体外实验、动物实验和初期的临床实验都已经证明了树突状细胞肿瘤疫苗的抗肿瘤作用。本文主要介绍树突状细胞的生物学特征和树突状细胞肿瘤疫苗在肿瘤免疫治疗中的研究进展。     

C型CpG单链寡核苷酸对肿瘤疫苗抑瘤效果的增强作用

目的：研究C型CpG单链寡脱氧核苷酸（CpG ODN）对肿瘤疫苗抑瘤效果的增强作用.方法：利用体外淋巴细胞增生实验和病毒保护实验确定新C型CpG ODN, 采用小鼠体内抑瘤实验观察CpG ODN对肿瘤疫苗HSP65-MUC1重组蛋白抑制MUC1阳性肿瘤生长的增强作用, 并对小鼠血清中抗HSP65-MUC1抗体的类型进行了初步鉴定.结果：自行设计的CpG ODN BW005能刺激人PBMC和小鼠脾细胞增生,其刺激后的培养上清具有保护Vero E6细胞免受VSV感染的作用, 属于新型C型CpG ODN.将BW005与HSP65-MUC1联合应用于C57BL/6小鼠后, MUC1阳性肿瘤的发生明显延缓（平均44.8 d）, 其终末肿瘤发生率最低（33.33%）;荷瘤小鼠的生存期明显延长（平均49.5 d）, 其终末生存率最高（66.67%）.小鼠血清中抗HSP65和抗MUC1的IgG2a抗体水平增加.结论：C型CpG ODN可以增强肿瘤疫苗HSP65-MUC1的抑瘤效果, 考虑与小鼠体内Th1环境的形成有关.     

MAGE1纳米蛋白疫苗的研制及其抗肿瘤效应

目的制备纳米乳剂包裹基因工程MAGE1(melanoma antigen1)抗原,研究其生物学特性及抗肿瘤免疫效应。方法采用界面聚合法,制备包裹MAGE1抗原的纳米乳剂(NEMAGE1),测量粒径、包裹率、载药量。检测乳剂包裹对小鼠DC摄取抗原能力的影响。用NEMAGE1免疫小鼠,运用酶联免疫斑点法(ELISPOT)和细胞毒性杀伤实验(LDH)检测NEMAGE1激活机体细胞免疫反应的状况,并观察纳米蛋白疫苗对荷瘤小鼠的治疗作用。结果成功制备粒径为(15±5)nm的纳米乳剂,包裹率为91%,载药量0.091gPL,4℃放置6个月后性质稳定。DC摄取NEMAGE1的能力明显强于对游离蛋白抗原的摄取。ELISPOT和细胞毒性杀伤实验显示,NEMAGE1可以诱导机体产生针对MAGE1的CTL,特异性杀伤表达MAGE1的肿瘤细胞。荷瘤小鼠的治疗实验显示,NEMAGE1对表达MAGE1的肿瘤有明显疗效,其效果优于单独使用游离MAGE1蛋白。结论纳米乳剂具有较高包裹率和稳定性。纳米乳剂包裹的肿瘤特异性抗原可以有效激活机体产生抗肿瘤免疫效应,是具有临床应用前景的新型抗肿瘤疫苗。     

Plague as HIV vaccine adjuvant

Individuals homozygous for a 32-basepair deletion mutation (null mutation) in CCR5, a chemokine receptor, are nearly immune to developing HIV infection. Recently based on molecular dating evidence it has been proposed that the bacterium Yersinia pestis, cause of plague, may have been the selective pressure which selected for a high prevalence of the 32-basepair null mutation. An association of a robust chemokine response with HIV vaccine efficacy has recently been shown. I suggest that Y. pestis may be a useful adjuvant in an HIV vaccine protocol by stimulating a vigorous chemokine response.     

Mechanism of the inhibitory effect of BCG vaccine on specific antitumor immunity in Syrian hamsters

The duration of the inhibitory effect of BCG vaccine on antitumor immunity induced by SV 40 virus in Syrian hamsters and the possibility of restoring immunity after vaccination were investigated. Specific resistance in animals immunized with SV 40 virus and then inoculated with BCG remained inhibited for 1 year after vaccination. A further injection of SV 40 virus into hamsters previously subjected to combined immunization reinduced specific immunity in the animals to tumors. The results show that the phenomenon of abolition of specific antitumor resistance by BCG vaccine is probably cellular in nature.Laboratory of Immunology of Tumors, Oncologic Scientific Center, Academy of Medical Sciences of the USSR, Moscow. (Presented by Academician of the Academy of Medical Sciences of the USSR L. M. Shabad.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 86, No. 9, pp. 358–360, September, 1978.     

Immunoglobulin M as a vaccine adjuvant

Vaccines are effective in preventing disease by stimulating the immune system and sustaining an immune response towards eradication of pathogens and diseased cells. However, designing successful vaccines is not always straightforward. For a vaccine to be successful, antigen-presenting cells (APC) need to be stimulated, primarily by adjuvants, towards a sustained immune response through integration of the innate and adaptive (humoral and cellular) immune systems. Furthermore, there is an immediate need for safe and effective adjuvants. There has been significant progress in understanding the mechanisms on how vaccines work and the role of adjuvants, dendritic cells, and the toll-like receptor (TLR) pathway. Currently, different adjuvants are actively explored but the potential of the immunoglobulin M (IgM) as a vaccine adjuvant has been overlooked. This article hypothesizes how the IgM molecule could function as a vaccine adjuvant by acting as a “soluble” toll-like receptor (TLR) through the formation of an immune complex with antigen (Ag) and other components of the innate immune system. The complex should lead to sustained humoral and/or cell-mediated immune responses. Hypothetically, it is also possible that the Ag-IgM complex recruits other components of complement or other factors that can activate other members of the adaptive immune system. As it is now possible to produce commercial-scale quantities of monoclonal human IgM antibodies, understanding the role of the IgM in linking the innate and adaptive immune systems may lead to practical therapeutic applications.     

Immunostimulatory DNA as a vaccine adjuvant

Immunostimulatory DNA containing unmethylated CpG motifs is recognized by Toll-like receptor 9, resulting in the activation of innate immune responses that subsequently amplify the adaptive-immune response. Advances in the characterization of Toll-like receptor 9 signaling have identified immunostimulatory sequences (ISS) with distinct biological activities. Numerous animal models have demonstrated that synthetic ISS are effective adjuvants that enhance both humoral and cellular immune responses in diverse indications, ranging from infectious disease to cancer and allergy. An added benefit supporting the use of ISS as a vaccine adjuvant is that the specific activation of a pathway critical to the regulation of the immune response results in minimal toxicity. To date, clinical testing has largely affirmed the potency and safety of ISS-adjuvanted vaccines.     

CpG DNA as a vaccine adjuvant

Synthetic oligodeoxynucleotides (ODNs) containing unmethylated CpG motifs trigger cells that express Toll-like receptor 9 (including human plasmacytoid dendritic cells and B cells) to mount an innate immune response characterized by the production of Th1 and proinflammatory cytokines. When used as vaccine adjuvants, CpG ODNs improve the function of professional antigen-presenting cells and boost the generation of humoral and cellular vaccine-specific immune responses. These effects are optimized by maintaining ODNs and vaccine in close proximity. The adjuvant properties of CpG ODNs are observed when administered either systemically or mucosally, and persist in immunocompromised hosts. Preclinical studies indicate that CpG ODNs improve the activity of vaccines targeting infectious diseases and cancer. Clinical trials demonstrate that CpG ODNs have a good safety profile and increase the immunogenicity of coadministered vaccines.     

CpG DNA as a vaccine adjuvant

Synthetic oligodeoxynucleotides containing unmethylated CpG motifs act as immune adjuvants, accelerating and boosting antigen-specific antibody responses by up to 500-fold. CpG motifs promote the production of T-helper 1 and pro-inflammatory cytokines and induce the maturation/activation of professional antigen-presenting cells (including macrophages and dendritic cells). These effects are optimized by maintaining close physical contact between the CpG DNA and the immunogen. Coadministering CpG DNA with a variety of vaccines has improved protective immunity in animal challenge models. Ongoing clinical studies indicate that CpG oligodeoxynucleotides are safe and well-tolerated when administered as adjuvants to humans and in some cases increase vaccine-induced immune responses.