LAK及TIL对靶细胞杀伤活性中效靶作用的细胞表面物质基础

在ＬＡＫ及ＴＩＬ杀伤靶细胞过程中介导效靶作用的细胞表面分子主要包括细胞粘附分子（ＣＡＭ）群类中的各种分子，其它一些尚未鉴定分属ＣＡＭ大类的糖蛋白，糖脂，层粘连层白（ｌａｍｉｎｉｎ）类似物和波形纤维蛋白（ｖｉｍｅｎｔｉｎ）类似物等。这些分子是效靶特异识别，结合的物质基础。虽然ＴＩＬ较ＬＡＫ杀伤活性呈现对靶细胞的特异性，但两者均存在不同程度的非特异性。本文以介绍ＬＡＫ及ＴＩＬ杀伤活性中效靶作用的细胞…     

淋巴因子激活的LAK／TIL细胞的表型

为研究肿瘤生物疗法中淋巴因子激活的杀伤细胞及其功能和特征与表型之间的关系，用流式细胞术（ＦＣＭ）结合抗体双荧光标记法对培养的淋巴因子激活杀伤细胞作连续的多项表型监测，发现存在着ＣＤ３－ＣＤ１６＋ＣＤ５６＋型，ＣＤ４＋型，ＣＤ８＋ＣＤ２８＋型，ＣＤ８＋ＣＤ２８－型的淋巴因子激活杀伤细胞的亚型。研究了ＣＤ３－ＣＤ１６＋ＣＤ５６＋亚型、ＣＤ８＋ＣＤ２８＋亚型的杀伤活性，显示了对不同的肿瘤靶细胞有不同的敏感性。     

LAK细胞识别,结合及杀伤肿瘤细胞分子机制的研究进展

ＬＡＫ细胞抗肿瘤机制二分复杂，可通过表面多种识别分子与肿瘤细胞表面配体分子相互识别结合，并释放多种杀伤介质直接杀伤肿瘤细胞，另外也可分泌多种细胞因子间接杀伤肿瘤细胞，本文重点介绍了ＬＡＫ细胞识别、结合、杀伤肿瘤细胞分子机制的一些最新进展，并控讨了肿瘤细胞抵抗ＬＡＫ细胞杀伤作用的分子机制。     

同种异体LAK细胞与肿瘤的LAK／IL—2疗法

近年来，同种异体ＬＡＫ细胞已被用于ＬＡＫ疗法的实践，从而有效地解决了ＬＡＫ前体的来源问题。本文就Ａｌｌｏ－ＬＡＫ过继免疫疗法依据的安全性、有效性，Ａｌｌｏ－ＬＡＫ的分布特性和Ａｌｌｏ－ＬＡＫ／ＩＬ－２在肿瘤治疗中的应用及注意的问题作一综述。     

不同取材部位的TIL得率及活力研究

为了获得更多，活力更强的ＴＩＬ细胞本实验就实体瘤肝癌，肺癌及卵巢癌取材部位进行闻取自肝癌，肺癌组织的浸润结节，交界组织，实质部位，中心及坏死区域的得率与活力，结果发现，坏死部位的细胞得率与活力最差，淋巴细胞得率也最差，肝癌，肺癌组织分别为４．０５×１０＾４／ｇ、５．５×１０＾４／ｇ；中心部位较差，分别为３．６×１０＾５／ｇ、４．１９×１０＾５／ｇ；实质部位的细胞得率最高，淋巴细胞得率，最多，肝癌，     

恶性肿瘤胸腹水中TIL生物学活性研究

从卵巢癌腹水和肺癌胸水中分离出肿瘤浸润淋巴细胞（ＴＩＬ），用ｒＩＬ－２诱导培养，观察其增殖活性。同时观察了ＰＪ－ＣＷ、ＰＯＧＰ、ＰＨＡ等对其杀伤活性的调节作用。结果表明，在培养５～１０天时ＴＩＬ增殖较快，其表面产生的ＣＤ３、ＣＤ４、ＣＤ８、Ｔａｃ等抗原，随培养时间的延长，表达量增加，尤以ＣＤ８增加较明显，ＣＤ４／ＣＤ８比值逐降。其表面产生的ＨＬＡ－ＤＲ、ＨＬＡ－ＡＢＣ抗原，在培养１５天内有增加表现，以后下降。其杀伤活性在培养５～１０天较高，其后活性下降。在培养系统中加入ＰＪ－ＣＷ、ＰＯＧＰ、ＰＨＡ等后，培养５天时，ＴＩＬ杀伤活性分别为８２．７０％，８５．５０％，８４．３５％（对照组为内保持较高的杀伤活性。     

一种高活力分离肿瘤浸润淋巴细胞方法的建立

肿瘤浸润淋巴细胞（ＴＩＬ）的增殖力、活力、杀伤力是ＴＩＬ临床治疗的基本问题。由于Ｒｏｓｅｎｂｅｒｇ方法存在着许多不足之处，本实验室对其一些重要步骤进行了改进：①用单一冷胶原酶消化肿瘤组织块；②省去影响ＴＩＬ活力的离心操作；③ＴＩＬ培养４８ｈ内除去掺杂在ＴＩＬ中的肿瘤细胞等。由于除去了影响ＴＩＬ活力的抑制因素，ＴＩＬ的增殖率、生存率、活力、杀伤力明显增加。有８０％肿瘤组织的ＴＩＬ经扩增可超过１×１０     

人LAK细胞体外抗肿瘤作用机理探讨

在对人LAK细胞体外杀伤作用研究的基础上,进一步探讨了该群细胞体外抗肿瘤的机理。实验结果证实,LAK细胞的杀伤是由多种因素参与的,杀伤时既有因子作用,又有直接接触杀伤。杀伤后靶细胞核破坏,DNA碎片释放到细胞外。在一定的培养时间内,LAK细胞的杀伤能力和细胞本身的增殖成正相关关系。     

人外周血淋巴细胞对绵羊红细胞的破坏作用

人外周血淋巴细胞,在含Ca~(2 )、Mg~(2 )的Hanks液中与绵羊红细胞(SRBC)共育,能将SRBC破坏释出血红蛋白(Hb),半数溶血值为0.1921±0.06,经胰蛋白酶抑制后淋巴细胞破坏SRBC的能力明显减弱,半数溶血值为0.0348±0.03。经尼龙棉柱法及E-玫瑰花法把淋巴细胞分离为T、B和NK等各个亚群。引起溶血作用最强的是NK细胞。对同一血源标本,同时用~(125)IUdR释放法测定NK活性和溶血分光光度法测定溶血活性,其结果呈明显正相关。本文认为人外周血淋巴细胞有破坏绵羊红细胞的能力,有这种能力的淋巴细胞是NK细胞。可望用本实验的溶血分光光度法测定NK细胞的活性。     

瘤体内注射IL－2基因后肿瘤细胞和肿瘤浸润性淋巴细胞的功能分析

将脂质体包裹的ＩＬ－２基因直接注射至Ｂ１６Ｆ１０黑色素瘤瘤体内，研究肿瘤细胞和肿瘤浸润性淋巴细胞（ＴＩＬ）的功能变化。１０ｄ后，Ｎｏｒｔｈｅｒｎｂｌｏｔ鉴定显示瘤体内注射ＩＬ－２基因后肿瘤细胞及ＴＩＬ中ＩＬ－２ｍＲＮＡ表达阳性；经Ｇ４１８筛选后的肿瘤细胞培养上清中可检测出ＩＬ－２；肿瘤细胞表面表达了较高水平的ＭＨＣⅠ类抗原（Ｈ－２Ｋ￣ｂＤ￣ｂ）；ＴＩＬ表面粘附分子ＬＡＦ－１表达也明显高于对照组，其杀伤Ｂ１６细胞的活性明显增强。结果表明瘤体内注射脂质体包裹的ＩＬ－２基因后，可在肿瘤原位将基因转移至肿瘤细胞及ＴＩＬ中，提高肿瘤细胞的ＭＨＣⅠ类抗原的表达，增强肿瘤细胞的免疫原性，并通过提高ＴＩＬ粘附分子的表达，共同促进ＴＩＬ的肿瘤特异性杀伤功能。     

THE ROLE OF ENDOTHELIAL CELLS IN THE PATHOGENESIS OF ATHEROSCLEROSIS

Using mainly rabbits and rats investigations on the endothelial permeability of the aorta, on relation between thrombus formation and endothelial damage, and on tissue culture of the endothelial cells of the rabbit aorta have been carried out.
The permeability of endothelial layer of rabbit aorta Increased by cholesterol-feeding. This seems to be derived from morphological and functional disorder of the endothelial cells. As a hemodynamic effect of blood flow, increased permeability of the endothelium of rat aorta was also found in experimental renal hypertension.
Accumulation of blood-plasma components (edema) was seen in the subendothellal area in cholesterol-fed rabbit aorta and in hypertensive rat aorta. Long-standing subendothellal edema is considered to be the cause of fibrous intlmal thickening (Grade I lesion/Nakashlma).
Endothelial defect of the aorta due to various damages is related to thrombus formation, and this seems to be accelerated by cholesterol feeding.
Morphological differences between normal endothelium and that of atherosclerotic lesion in human and in rabbit aortas can be seen.
Tissue culture of the endothelium was performed in vivo and in vitro, and some discussions were made on the characteristics of cultured endothelial cells.     

ANTIVIRAL IMMUNITY AND THE ROLE OF DENDRITIC CELLS

Using our experimental model we demonstrate the need for Th1 immune responses for recovery from influenza virus infection. Inoculation of IL-4 concurrent with infection significantly delays virus clearance and converts the immune response to a Th2 response. Immunization using live virus in the presence of IL-4 leads to generation of Th2 memory cells that fail to facilitate recovery upon subsequent virus infection. Inactivated virus expands Th2 cells, leading to responses similar to those observed following IL-4 infusion. Immunization using cultured dendritic cells incubated with live or inactivated virus mimics the results observed with direct virus injection. We demonstrate that in contrast to live virus-infected dendritic cells, inactivated virus fails to elicit Th1 immunity. This failure correlates with the inactivated viruses' inability to induce dendritic cell maturation. Thus, our data suggest that the polarity of the immune response is dictated by the nature of the antigen, and the trigger for influenza virus-induced DC maturation leading to Th1 immunity is dependent on virus replication.     

THE ROLE OF VIRUS IN THE GROWTH OF VIRAL LEUKEMIA CELLS

It is said that 'Neoplasms can proliferate autonomously without continuation of stimuli'. This is true but also true to say that some virus-induced tumors are necessary to maintain the causative virus and they can not proliferate without the continuation of virus. The authors have carried out experiments in order to elucidate the role played by viruses in the maintenance of proliferation of virus-induced neoplastic cells. The leukemia cells employed by the authors were designated as 'Friend tumors', which were originated from Friend virus-induced neoplastic lesions and have been serially transplanted in mice and rats.     

ULTRASTRUCTURAL ASPECTS OF THE ROLE OF THE MEDIA-SMOOTH MUSCLE CELLS IN ARTERIOSCLEROSIS OF MAN AND ANIMALS

In various arterial lesions including atherosclerotic lesions, the main morphological change involves smooth muscle cells. The potential sensitivity is different among the arterial smooth muscle cells, venous smooth muscle cells and smooth muscle cells of other organs. The modified smooth muscle cells characterized by the increase of rough endoplasmic reticula are considered to express their latent ability to synthesize collagen fibers, elastic fibers and other ground substances.
The foam cells noted in atherosclerosis and fatty streak consist of lipid accumulated smooth muscle cells and hematogenous macrophages. Lipid metabolism and synthesis in the latter differ from those in the former. The ratio of the two kinds of foam cells in atheroma or fatty streak varies by the stage of the lesion.
It is possible to suppose that there exists a factor which would selectively attack the media smooth muscle cells of small arteries or arterioles. This is observed electron microscopically as focal cytoplasmic necrosis (cytoplasmolysis) of smooth muscle cells and plays an important role in the histogenesis of fibrinoid necrosis.
In case of experimental periarteritis nodosa the early stage begins with cytoplasmolysis of smooth muscle cells and marked increase of rough endoplasmic reticula in adjacent smooth muscle cells.     

脾细胞在CTL克隆增殖中的作用

本文证明二次MLC上清或重组IL2均不足以维持细胞毒 T细胞(CTL)克隆的正常增殖,而同种异体、同种第三者、甚至同基因的脾细胞均可增强rIL2诱导的CTL克隆增殖应答,亦可增强 CTL由 CD3∈链抗体(145-2C11)刺激的 IL-2非依赖性增殖。脾细胞的这种辅佐效应似与可溶性因子或脾粘附细胞无关。用 rIL2 预处理的脾细胞可诱导 CTL克隆的显著增殖,这种增殖可被抗IL2 受体抗体所抑制,提示脾细胞可能将其“膜结合”rIL2递给CTL而更有效地诱导增殖反应。FD18.5(抗 LFA-1),KH1.4(抗 Ly-6)及 HK2.1(抗 Thy-1)可显著抑制脾细胞对 145-2C11 激活 CTL增殖的辅佐效应,提示脾细胞对 CTL克隆 IL2非依赖性增殖的辅佐效应与一些细胞表面分子有关。     

CD2相关蛋白在肾脏细胞中的分布及其在足细胞中的作用

目的 观察CD2相关蛋白(cD2AP)在肾脏不同细胞系中的表达分布,及其与足细胞裂孔隔膜分子nephrin和细胞骨架蛋白F-actin之间的联系.方法 以DMEM培养基培养人肾小球系膜细胞(HMC)和人肾小管上皮细胞系(HK-2),RPMI 1640培养基培养条件永生化小鼠足细胞系.以RT-PCR及Western blotting方法检测足细胞内CD2AP和nephrin的表达.间接免疫荧光结合激光共焦方法观察CD2AP在HMC、HK-2、未分化及已分化足细胞中的表达情况,及CD2AP与nephtin在足细胞中的共存.直接免疫荧光结合激光共焦方法观察F-actin在足细胞的表达及其与CD2AP的共存.结果 CD2AP均匀分布于HK-2及未分化足细胞的核周及胞浆,而不表达于HMC细胞.在足细胞分化过程中,CD2AP的分布发生了变化,出现向周边聚集的现象.CD2AP与足细胞裂孔隔膜分子nephrin及细胞骨架蛋白F-actin在足细胞中存在共定位关系.结论 CD2AP表达于上皮来源的肾脏固有细胞.CD2AP在足细胞中的分布特点,提示CD2AP可能参与足细胞的分化过程,并与裂孔隔膜分子功能及细胞骨架凋节有关.     

丝裂原激活蛋白激酶在内毒素脂多糖诱导大鼠施万细胞一氧化氮合酶表达中的作用

目的 主要研究丝裂原激活蛋白激酶(MAPK)信号途径在内毒素脂多糖(LPS)诱导大鼠施万细胞(Scs)诱导型一氧化氮合酶(iNOS)基因表达和一氧化氮(NO)产生中的作用.方法 先用3种MAPK的特异性抑制剂PD98059(ERK1/2)、SB202190(P38 MAPK)和SP600125(JNK)以不同浓度预处理细胞1h,再用LPS作用施万细胞4 h后,用RT-PCR检测细胞中iNOS mRNA、IL-6 mRNA和TNF-α mRNA的表达;Western blotting观察iNOS蛋白水平的表达变化;通过测定细胞培养液中亚硝酸盐含量来观察NO的水平.结果 LPS可显著激活施万细胞中MAPK信号通路诱导iNOS表达.MAPK的抑制剂预处理细胞后,可显著抑制细胞iNOS mRNA和NO的合成及IL-6 mRNA和TNF-α mRNA的表达.结论 MAPK信号通路参与了LPS介导的大鼠施万细胞iNOS基因表达和NO产生,通过阻断细胞内信号转导通路来减少iNOS及其他细胞因子的产生,为抑制周围神经损伤后的炎症以及免疫反应发生提供了一条新思路.     

体外培养人胎儿雪旺氏细胞的增殖及其细胞动力学研究

新鲜和液氮保存3个月的人胎儿雪旺氏细胞体外培养8天,细胞数量增加一倍,培养液中加入二丁基环磷酸腺苷和牛脑垂体浸出液,细胞增倍时间缩短近一倍.流式细胞计测定结果显示,培养早期,S期、G2+M期细胞所占比例<5%,随着培养时间的延长,其比例逐渐增加,第6天达高峰,环磷酸腺苷和垂体浸出液作用3天,S期、G2期+M期比例达15%.实验结果表明:人胎儿雪旺氏细胞在体外增殖缓慢,环磷酸腺苷和垂体浸出液能刺激雪旺氏细胞分裂;冷冻复苏后的雪旺氏细胞其生长特征以及对环磷酸腺苷和垂体浸出液的反应与新鲜细胞相同.