胶质瘤Hsp70-肽复合物激活T淋巴细胞抗肿瘤研究

目的 研究胶质瘤Hsp70-肽复合物激活的T淋巴细胞抗肿瘤效应、探讨其用于临床治疗的可行性.方法 取胶质瘤手术标本,经组织块法培养、建株胶质瘤细胞,冻融法获取胶质瘤细胞抗原,抗原与Hsp70体外构建Hsp70-肽复合物并激活T淋巴细胞,制备特异性CTL,MTT法检测对胶质瘤细胞的体外杀伤作用.结果 Hsp70-肽复合物激活CTL对胶质瘤细胞的杀伤活性明显高于未经胶质瘤抗原激活的CTL对胶质瘤细胞的杀伤作用( P ＜ 0.01).结论 肿瘤组织中或细胞中提取的Hsp70-肽复合物作用于T淋巴细胞,能诱导CTL在体外对胶质瘤细胞能产生高效而特异杀伤作用,为临床制备肿瘤疫苗奠定理论基础.     

尤文肉瘤细胞总RNA转染的DC疫苗体外诱导尤文肉瘤患者特异性抗肿瘤免疫的实验研究

为了探讨尤文肉瘤细胞总RNA转染的DC疫苗体外诱导特异性抗肿瘤免疫的能力,采用分离尤文肉瘤患者外周血单核细胞体外诱导DC细胞,Trizol法提取患者尤文肉瘤细胞总RNA,用总RNA转染DC并诱导特异性CTL的扩增,用RT-PCR方法检测肿瘤总RNA加载的DC细胞的表达,用MTT法检测淋巴细胞的增殖和CTL的杀伤活性.结果RT-PCR测定显示尤文肉瘤细胞总RNA转染的DC细胞可以呈递肿瘤特异性抗原,并特异性地表达其特有序列EWS-FLI1.经尤文肉瘤细胞总RNA转染的DC特异性表面标志及功能相关分子表达均上调,转染后的DC可显著刺激自体T淋巴细胞增殖.诱导的特异性CTL对携带EWS-FLI1抗原的靶细胞的杀伤率显著高于LAK细胞和未经转染的DC.说明尤文肉瘤细胞总RNA转染的DC疫苗可在体外诱导出特异性抗肿瘤免疫.     

DC-CTL体外杀伤人胰腺癌细胞效应研究

研究胰腺癌患者外周血分离培养而来的树突状细胞体外对人胰腺癌细胞BXPC-3、Capan-2的抑制作用.应用各种细胞因子诱导培养从胰腺癌患者外周血分离的单核细胞,获得成熟DC及CIK.胰腺癌组织来源抗原致敏DC,激活T细胞增殖分化为CTL.MTT法检测抗原致敏的DC激活的CTL及单独的CIK对BXPC-3细胞和Capan-2 细胞的杀伤效应.结果显示:胰腺癌抗原致敏的DC,激活肿瘤抗原特异性CTL,体外对BXPC-3和Capan-2胰腺癌细胞产生杀伤作用,分别为75.85%和50.34%;CIK对BXPC-3和Capan-2胰腺癌细胞也有杀伤作用,分别达到62.06%和40.92%,但比抗原致敏的DC激活的CTL的杀伤效应低(P<0.05).胰腺癌患者外周血来源DC在体外能诱导高效的抗胰腺癌细胞免疫反应,提示肿瘤抗原激活的DC作为肿瘤疫苗在胰腺癌的免疫治疗中具有重要价值.     

VEGF局部动脉注射对断肢再植术后NF-κB、Bcl-2mRNA表达的影响

探讨血管内皮生长因子(VEGF)局部动脉注射对大鼠后肢断肢再植术后NF-κB、Bcl-2mRNA表达的影响。选用健康Wistar大鼠80只随机分为VEGF组、低分子右旋糖酐组、模型组、空白组,造后肢断肢再植模型,取腓肠肌标本,ELISA法检测的IL-6、IL-17含量变化;RT-PCR检测NF-κB、Bcl-2mRNA表达。与正常组比较,模型组IL-6、IL-17含量及NF-κB活性明显升高,Bcl-2活性明显降低,差异具有统计学意义(P0.05);与模型组比较,VEGF组、低分子右旋糖酐组IL-6、IL-17含量及NF-κB活性明显降低,Bcl-2活性明显升高,差异具有统计学意义(P0.05);与术后低分子右旋糖酐组比较,VEGF组干预后血清IL-6、IL-17含量及NF-κB、Bcl-2mRNA无差异(P0.05)。VEGF局部动脉注射能促进断肢再植成活,其机制可能与抑制NF-κB的活性,升高BCL-2活性有关。     

gD2 DNA疫苗诱导细胞免疫应答

用生殖器疱疹gD2 DNA疫苗pcDNA3-gD转染哺乳动物细胞COS-7,鉴定gD表达．再用其免疫BALB／c小鼠,测定脾细胞增殖、CTL细胞杀伤活性,以及CD4^ 和CD8^ 细胞群．结果表明：pcDNA3-gD能够在哺乳动物细胞COS-7中表达gD抗原,其免疫小鼠脾细胞增殖活性增加,CTL活性达37．69％,明显高于pcDNA3组和生理盐水组;CD4^ 占T细胞数量的33．38％,数量较pcDNA3组和生理盐水组显著增加．     

体外诱导骨肉瘤特异性抗肿瘤的免疫实验

采用分离骨肉瘤患者外周血单核细胞体外诱导DC细胞,Trizol法提取患者骨肉瘤细胞总RNA,用总RNA转染DC并诱导特异性CTL的扩增,用MTT法检测淋巴细胞的增殖和CTL的杀伤活性。探讨骨肉瘤细胞总RNA转染的DC疫苗体外诱导特异性抗肿瘤免疫的能力。经骨肉瘤细胞总RNA转染的DC特异性表面标志及功能相关分子表达均上调,转染后的DC可显著刺激自体T淋巴细胞增殖,诱导的特异性CTL对靶细胞的杀伤率显著高于单纯淋巴细胞和未经转染的DC。说明骨肉瘤细胞总RNA转染的DC疫苗可在体外诱导出特异性抗肿瘤免疫。     

树突状细胞与妇科肿瘤的免疫治疗

树突状细胞(DC)是目前所知机体内功能最强的专职抗原提呈细胞,可在体内外向T细胞提呈抗原并诱发CTL反应,在抗肿瘤免疫中发挥重要作用.近年采用DC疫苗进行抗肿瘤治疗已成为当今肿瘤生物治疗领域备受关注的焦点之一.针对DC抗肿瘤机制、妇科肿瘤的免疫逃逸及在妇科肿瘤上的应用进行了研究.     

IL-10修饰的树突状细胞的体外生物学效应研究

探讨lL-10修饰的树突状细胞（DC）对T细胞增殖和细胞毒性T细胞（CTL）胞毒活性的影响。采用乳酸脱氢酶法和ELISA法，分别测定细胞毒活性及T细胞凋亡。结果表明，IL-10对同种细胞刺激的增殖反应和特异性CTI．细胞毒活性有明显的抑制作用，修饰的DC诱导淋巴细胞增殖反应明显降低，而未修饰的DC诱导淋巴细胞增殖反应较强；IL-10和修饰的DC可诱导淋巴细胞凋亡．可见，稳定表达IL-10的DC，对T细胞增殖和对胞毒活性有明显的抑制作用，并可诱导T细胞凋亡。     

LAGE-1DNA肿瘤疫苗的抗肿瘤治疗效果研究

以LAGE-1转染EMT6构建小鼠乳腺癌肿瘤模型;将BalB/c小鼠随机分为4组,以pcDNA3.1/LAGE-1及对照pcDNA3.1分别接种实验组和对照组各三次.于末次免疫后10 d在小鼠左背部、右背部皮下分别种植EMT6肿瘤细胞和EMT6/LAGE1肿瘤细胞．荷瘤后观察成瘤时间、肿瘤大小,并对小鼠脾细胞进行CTL细胞杀伤活性实验来观察DNA疫苗引起的免疫反应.结果显示LAGE-1DNA疫苗在体内能诱导有效的特异性肿瘤免疫应答,该疫苗简便有效.     

展示肿瘤特异性抗原表位的噬菌体在小鼠体内引起的细胞免疫变化

以含有肿瘤特异性抗原表位的杂合型噬菌体(TSPA-M-A1)作为抗原,免疫C57BL/ 6J小鼠,观察免疫4周后小鼠的细胞免疫变化.结果表明:抗原TSPA-M-A1在小鼠体内能够提高T淋巴细胞对ConA刺激的反应性,使NK细胞杀伤活性明显增高,产生了特异性T淋巴细胞(CTL)反应,引起了较强的迟发型超敏反应.这些结果为肿瘤的免疫治疗提供了实验依据.     

GAGE-1 DNA肿瘤疫苗的构建及其抗肿瘤治疗效果的试验研究

目的 观察G antigen 1 (GAGE-1)核酸疫苗pcDNA3.1+/GAGE-1免疫小鼠后,对表达GAGE-1抗原的B-16/GAGE-1肿瘤细胞的保护作用.方法 将C57 BL/6小鼠随机分为4组,与0、2、4周分别接种pcDNA3.1＋/GAGE-1（实验组1）、pcDNA3.1＋/GAGE-1/白介素2（实验组2）、pcDNA3.1＋（对照组1）,pcDNA3.1＋/白介素2（对照组2）各三次.末次免疫后10d小鼠用于肿瘤细胞攻击试验：分别于左背部、右背部皮下种植B16肿瘤细胞、B16/GAGE-1肿瘤细胞.种植肿瘤细胞（荷瘤）后观察成瘤时间、肿瘤大小和荷瘤后小鼠的生存时间及生存率. 结果： pcDNA3.1+/GAGE-1/IL-2质粒免疫的小鼠在种植B16/GAGE-1、B16/pcDNA3.1+后,发现小鼠成瘤时间明显延迟,成瘤减小,生存期明显延长.结论:pcDNA3.1+/GAGE-1/IL-2 DNA 疫苗在体内能诱导出显著的GAGE-1特异性肿瘤免疫应答,且能抑制体内已经存在的少量肿瘤细胞的成瘤     

Enhancing cellular immune response to HBV M DNA vaccine in mice by codelivery of interleukin-18 recombinant

OBJECTIVE: To investigate the effect of interleukin-18 (IL-18) on immune response induced by plasmid encoding hepatitis B virus middle protein antigen and to explore new strategies for prophylactic and therapeutic HBV DNA vaccines. METHODS: BALB/c mice were immunized with pCMV-M alone or co-immunized with pcDNA3-18 and pCMV-M and then their sera were collected for analysing anti-HBsAg antibody by ELISA; splenocytes were isolated for detecting specific CTL response and cytokine assay in vitro. RESULTS: The anti-HBs antibody level of mice co-immunized with pcDNA3-18 and pCMV-M was slightly higher than that of mice immunized with pCMV-M alone, but there was not significantly different (P>0.05). Compared with mice injected with pCMV-M, the specific CTL cytotoxity activity of mice immunized with pcDNA3-18 and pCMV-M was significantly enhanced (P<0.05) and the level of IFN-Gamma in supernatant of splenocytes cul-tured with HBsAg in vitro was significantly elevated (P<0.05) while the level of IL-4 had no significant difference (P>0.05). CONCLUSION: The plasmid encoding IL-18 together with HBV M gene DNA vaccines may enhance specific TH1 cells and CTL cellular immune response induced in mice, so that IL-18 is a promising immune adjuvant.     

Immunotherapy of intracranial G422 glioblastoma with dendritic cells pulsed with tumor extract or RNA

OBJECTIVE: To investigate the anti-tumor efficacy of dendritic cell (DC)-based vaccines pulsed with tumor extracts or RNA in a mouse model of intracranial G422 glioblastoma. METHODS: Bone marrow-derived DCs were pulsed ex vivo with tumor extracts or RNA. Ninety female mice harboring 4-day-old intracranial G422 glioblastomas and 126 normal mice were treated with three spaced one week apart subcutaneous injections either with PBS, unpulsed DCs, G422 tumor extracts, RNA, DCs pulsed with G422 tumor extracts (DC/extract) or with RNA (DC/RNA). Seven days after the third immunization of normal mice, the spleens of 36 of them were harvested for cytotoxic T lyphocyte (CTL) assays and the others were challenged in the brain with G422 tumor cells. All the treated mice were followed for survival. Some mice brains were removed and examined pathologically when they died. RESULTS: Immunization using DC/extract or DC/RNA significantly induced G422-specific CTL responses compared with control groups (P<0.01). Vaccination with DC/extract or DC/RNA, either prior to G422 tumor challenge or in tumor-harboring mice, significantly prolonged survival compared with other control groups (P<0.01). CONCLUSION: DCs pulsed with tumor extracts or RNA derived from autologous tumors has potential antitumor effects via activation of cell-mediated immunity. Our results suggest a useful therapeutic strategy against gliomas.     

Influence of systemic immune and cytokine responses during the acute phase of zoster on the development of postherpetic neuralgia

Postherpetic neuralgia (PHN) is a severe sequela of herpes zoster (HZ). Until now, only age and pain severity were considered predisposing factors for the development of PHN. We evaluated 49 patients with acute phase HZ, 10 of whom developed PHN (Group A) and 39 of whom did not develop PHN (Group B). Twenty-five healthy volunteers similar in age and gender distribution to the study group were recruited as controls (Group C). Numbers of serum CD3⁺ (pan-T lymphocytes), CD4⁺ (helper/inducer), and CD8⁺ (suppressor/cytotoxic) lymphocytes were decreased significantly in Groups A and B relative to the control group, but there were no statistical differences between Groups A and B. Interleukin (IL)-1β, IL-6, tumor necrosis factor (TNF)-α, IL-8, and IL-10 were significantly elevated in Groups A and B relative to Group C. IL-6 was significantly higher in Group A than in Group B, and was significantly positively correlated with pain severity scored on a visual analog scale. Therefore, we suggest that the inflammatory response, especially that of IL-6, in the acute phase of HZ may be associated with hyperalgesia and the development of PHN.     

结肠癌患者血清中IL-8和VEGF的检测及临床意义

目的探讨白介素-8（IL-8）和血管内皮生长因子（VEGF）血清水平与结肠癌的关系．方法选择结肠癌住院患者64例,所有病例均经病理检查确诊．用ELISA法测定结肠癌患者术前、术后与健康者血清中IL-8和VEGF含量,采用方差分析进行组间两两比较．结果结肠癌患者血清IL-8和VEGF含量明显高于正常对照组（P〈0．05）,DukesC,D期结肠癌患者血清IL-8和VEGF含量明显高于A,B期（P〈0．05）;肿瘤根治术后血清IL-8和VEGF含量较术前明显降低（P〈0．05）,但仍高于正常对照组（P〈0．05）．结论检测血清IL-8和VEGF含量对于结肠癌病情估计和预后判断有重要意义．     

Immune responses of a designed HIV-1 DNA vaccine on rhesus monkeys

According to the UNAIDS/WHO report (http:// www.unaids.org/Epi2005/doc/report.html), there are about 40.3 million HIV/AIDS survivors, and the new HIV infection number amounts to about 4.9 million, while the death number is some 3.1 million in the world in…     

脑胶质瘤患者血清IL-6、IL-8、IL-10和TNF-α的表达及临床意义

[目的]探讨血清中白介素6（IL-6）、IL-8、IL-10和肿瘤坏死因子（TNF-α）对于脑胶质瘤的诊断价值。[材料与方法]分别检测健康对照组、低级别脑胶质瘤组和高级别脑胶质瘤组中IL-6、IL-8、IL-10和TNF-α的含量。[结果]与健康对照组比较,低级别脑胶质瘤组的IL-6、IL-8和TNF-α有统计学差异,高级别脑胶质瘤组的IL-6、IL-8、IL-10和TNF-α均具有统计学差异。与低级别脑胶质瘤组相比较,高级别脑胶质瘤组的IL-6、IL-10和TNF-α有统计学差异。其中区分低级别脑胶质瘤和高级别脑胶质瘤的诊断价值最好的指标为IL-10,其诊断灵敏性和特异性分别为74.90%和65.80%。IL-6、IL-10和TNF-α联合检测时其灵敏性和特异性分别为92.30%和93.10%。[结论]证实IL-6、IL-10和TNF-α联合诊断价值优于单项指标,为脑胶质瘤的临床诊断提供辅助方法。     

H-1 细小病毒感染大鼠神经胶质瘤细胞C6 机制的初步探讨

摘要: 目的探讨H-1 细小病毒感染大鼠神经胶质瘤细胞C6 后细胞因子水平的变化。方法取感染C6 细胞后不同时间点的H-1 细小病毒,同时取相同时间点的正常C6 细胞,采用ELISA 方法检测同时检测TNFα、IL-6、TGF-β1和IL-17A 的表达水平。结果正常C6 细胞中TNF-α、TGF-β1 和IL-17A 的浓度随培养时间增长而提高, IL-6 浓度无明显变化; H-1 细小病毒感染的C6 细胞中TNF-α、TGF-β1 和IL-17A 浓度无明显变化,而IL-6 浓度随感染时间推移而显著增加。结论H-1 细小病毒感染导致C6 细胞中细胞因子表达水平的变化。