抗血吸虫感染免疫效应机理的体外研究进展

迄今已积累的研究资料表明,宿主感染血吸虫后产生的伴随免疫涉及多种免疫效应机理,其中抗体依赖的细胞介导的对童虫的细胞毒作用(ADCC),可能是主要免疫效应之一。本文综述了有关嗜酸粒细胞、中性粒细胞、单核-巨噬细胞参与的 ADCC 体外免疫效应机理的研究进展。嗜酸粒细胞依赖抗体的杀童虫作用分为两个步骤:即依赖细胞上的 Fc 受体与结合在童虫表面的抗体 Fc 段搭桥,随之细胞中颗粒的内容物作为配体促进其与童虫不可逆的结合,从而杀伤童虫。嗜酸粒细胞颗粒中的主要碱性蛋白、阳离子蛋白为主要的杀童虫物质,此外过氧化物酶-过氧化氢-卤化物系统可能也有一定的杀童虫作用。补体、肥大细胞的产物、淋巴因子等对嗜酸粒细胞介导的 ADCC 有促进作用。尽管在体外实验中,中性粒细胞有与嗜酸粒细胞相似的过氧化物酶-过氧化氢-卤化物系统杀伤童虫的机制,但其在保护性免疫表达中的作用迄今仍不明确。近年来,许多实验证明单核-巨噬细胞不仅在体外能依赖抗体特异地杀伤皮肤期童虫,而且因感染、免疫接种或无关抗原(如 BCG)的刺激激活的巨噬细胞能非特异杀伤皮肤期童虫和肺期童虫。巨噬细胞杀伤童虫的细胞毒机理尚不明确,可能与精氨酸酶和髓过氧化物酶有关。运用巨噬细胞功能缺陷的小鼠品系进行的体内诱导免疫力的研究,进一步支持巨噬细胞在抗血吸虫感染的保护性免疫的产生中可能起中心作用。     

日本血吸虫(中国大陆株)童虫杀伤机理的体外研究-抗体依赖的小鼠中性粒细胞介导的细胞毒作用

运用体外免疫效应机理分析系统,证明了抗体依赖的小鼠中性粒细胞介导对日本血吸虫（中国大陆株）童虫的细胞毒作用（ADCC）。结果表明,正常小鼠的中性粒细胞能产生对日本血吸虫童虫的依赖抗体介导的ADCC反应,这种ADCC作用不依赖补体,补体对中性粒细胞介导的这种ADCC无增强作用。     

日本血吸虫童虫杀伤机理的体外研究 抗体依赖的小鼠巨噬细胞介导的细胞毒作用

本文报告运用体外免疫效应机理分析系统,证明了对日本血吸虫(中国大陆株)童虫的抗体依赖小鼠巨噬细胞介导的细胞毒作用(ADCC)。来源于正常小鼠和感染小鼠的巨噬细胞皆能产生对日本血吸虫童虫的依赖抗体介导的 ADCC 反应;感染小鼠巨噬细胞对童虫还具有不依赖抗体的细胞毒作用,而正常小鼠巨噬细胞则无;感染小鼠巨噬细胞参与的对童虫的 ADCC 反应较其不依赖抗体的细胞毒作用强,也较正常小鼠巨噬细胞参与的 ADCC 为强。还注意到补体仅可增强感染小鼠巨噬细胞的 ADCC 反应。     

抗体和/或补体依赖嗜酸粒细胞介导的细胞毒作用对血吸虫童虫的体外杀伤试验

以日本血吸虫中国大陆株童虫为实验对象,研究比较了嗜酸粒细胞(Eos)的细胞毒效应对抗体和/或补体的依赖作用。正交设计的体外杀伤试验表明补体和抗体均可作为配体参与Eos杀伤日本血吸虫中国大陆株童虫的细胞毒效应作用;配体依赖作用的大小顺序为抗体 补体>补体>抗体;在新鲜免疫血清1∶20、培养48h、效/靶比5000∶1时童虫死亡率最高。     

白细胞三烯B_4增强中性细胞及嗜酸粒细胞——介导补体依赖的对曼氏血吸虫童虫的体外杀伤作用

以往研究表明人体嗜酸粒细胞在补体参与下能杀伤曼氏血吸虫童虫;组胺及人工合成的甲硫酰肽类f-MLP能增强这种杀伤作用;致敏肥大细胞释出的白细胞三烯B_4(LTB_4)能增强嗜酸粒细胞及中性细胞的C_3b受体。因此观察LTB_4是否影响粒细胞介导补体依赖的对童虫杀伤作用并与f-MLP及其他有关白细胞三烯比较,和观察在其单独作用时有无杀虫效应。白细胞三烯通过花生四烯酸与中性细胞一同孵育获得并经液相层析纯化,曼氏血吸虫童虫按Ramalho-Pinto(1974)机械法制     

嗜酸粒细胞在日本血吸虫(中国大陆株)感染小鼠体内的作用

以抗嗜酸粒细胞血清(AES)对嗜酸粒细胞(Eos)在日本血吸虫(中国大陆株)感染小鼠体内的作用进行了研究。小鼠经初次感染日本血吸虫尾蚴5wk后,用200条尾蚴攻击感染,并经AES处理,肺部回收童虫数明显高于用正常兔血清(NRS)处理的对照组;正常小鼠经被动转移免疫血清后感染日本血吸虫尾蚴,用AES处理,肺部回收童虫数比NRS对照组明显增多。用日本血吸虫可溶性虫卵抗原(SEA)皮下致敏小鼠,2wk后经尾静脉注射虫卵形成肺肉芽肿反应,经AES处理后,肺虫卵肉芽肿内Eos受到明显抑制,肉芽肿平均直径明显小于NRS对照组。试验结果提示,Eos是宿主体内对日本血吸虫童虫有效的杀伤细胞之一,其杀伤作用是抗体依赖的Eos介导的细胞毒作用;AES能特异性地仰制肉芽肿内的Eos,并对不同时期肉芽肿的直径有明显的抑制作用。     

血吸虫病人血清体外诱导杀童虫作用的观察

通过比较日本血吸虫感染病人与正常人血清体外诱导小鼠白细胞杀童虫的作用,了解抗体依赖的细胞介导的细胞毒作用(ADCC)在血吸虫感染宿主的获得性免疫中的作用.体外诱导小鼠白细胞杀童虫作用,日本血吸虫感染病人血清,无论是成人组还是儿童组,初次感染组还是重复感染组,均明显地强于正常人血清,表明抗血吸虫特异性抗体可以诱导细胞介导的细胞毒作用.证明日本血吸虫感染宿主体内存在着ADCC机制,该机制对于抵抗再次感染具有一定的作用,是日本血吸虫获得性免疫的重要组分.     

血吸虫病人血清体外诱导杀童虫作用的观察

通过比较日本血吸虫感染病人与正常人血清体外诱导小鼠白细胞杀童虫的作用。了解抗体依赖的细胞介导的细胞毒作用（ADCC）在血吸虫感染宿主的获得性免疫中的作用。体外诱导小鼠白细胞杀童虫作用。日本血吸虫感染病人血清,无论是成人组还是儿童组,初次感染组还是重复感染组,均明显地强于正常人血清,表明抗血吸虫特异性抗体可以诱导细胞介导的细胞毒作用。证明日本血吸虫感染宿 主体内存在着ADCC机制。该机制对于抵抗再次感染具有一定的作用。是日本血吸虫获得性免疫的重要组分。     

抗体和补体依赖嗜酸粒细胞介导的对日本血吸虫童虫作用电镜研究

本文报道了在嗜酸粒细胞(Eos)介导的细胞毒效应过程中童虫形态变化的电镜观察及受损童虫释放乳酸脱氢酶(LDH)的测定。不同配体依赖的 Eos 介导杀伤童虫作用的强度和速度不同,即补体 抗体组和补体组出现较早,且前者所致损伤较重,抗体组童虫损伤出现较晚而轻微。受损童虫的 LDH 释放多寡顺序为抗体 补体>补体>抗体。不同配体依赖的 Eos 介导细胞毒作用的速度和电镜观察相符。     

曼氏血吸虫的免疫性:抗体依赖的嗜酸粒细胞对血吸虫童虫的杀伤作用

作者用 ~(51) Cr 标记曼氏血吸虫童虫,通过测定杀伤后童虫释出的 ~(51) Cr 量来分析抗体依赖的嗜酸粒细胞对血吸虫童虫的体外杀伤作用。本文报告了两个问题,第一是用高纯度的嗜酸粒细胞制剂进行体外试验,以便比较直接地观察嗜酸粒细胞的功能;第二用各种抗代谢药物分析嗜酸粒细胞对血吸虫童虫的杀伤作用机制。     

小鼠感染后不同时间血清对日本血吸虫童虫细胞介导？..

The tests of antibody-dependent cell-mediated cytotoxicity (ADCC) were performed to observe the kinetics of in vitro killing of Schistosoma japonicum schistosomula mediated by eosinophils, macrophages or neutrophils with infected mouse sera of different duration. The results showed that when complement was not involved, the killing rates of schistosomula mediated by eosinophils or macrophages began to increase significantly at 4 wk post-infection, reached a plateau in 5-7 wk and 6-8 wk respectively, and then declined to the levels of that at 4 wk till 11 wk. In case of neutrophils, there was no significant cytotoxicity detected. When complement was involved, all the three effector cells could mediate cytotoxicity to schistosomula, with the killing rates higher than those without complement at the correspondent time intervals. These indicate that ADCC appears to be an important ingredient in the acquired resistance to S. japonicum, and demonstrate that eosinophil- and macrophage-mediated cytotoxicity to schistosomula is complement-independent, whereas neutrophil-mediated cytotoxicity to schistosomula is complement-dependent. This method may be of some value for choosing optimal time for vaccination and estimating the effectiveness of a candidate vaccine.     

Antibody-dependent cellular cytotoxicity of Trypanosoma theileri mediated by purified bovine isotypes and subisotypes

Summary Bovine neutrophils, eosinophils and macrophages mediated in vitro cytotoxicity against Trypanosoma theileri in the presence of purified IgM, IgGl and IgG2 from immune bovine serum. When the immunoglobulin fractions were assayed at similar concentrations, IgM was the most effective isotype mediating killing with all three effector cell types. Using the ELISA with monospecific antisera against the different bovine isotypes and subisotypes, IgM was shown to be contaminated by < 1%. The addition of 0·08 M 2-mercaptoethanol inhibited IgM-mediated ADCC but not that of IgGl or IgG2, and the cytotoxicity occurred in the absence of complement. The presence of isotype and subisotype specific Fc receptors on the bovine effector cells was investigated using a totally homologous erythrocyte-antibody (EA) resetting technique. FC receptors for bovine IgM, IgG1 and IgG2 were detected on bovine neutrophils. Very few FcM receptors were detected on either eosinophils or macrophages, but FcG2 receptors were detected on both cell types, and FcG1 receptors on macrophages. However, eosinophils showed very few FcG1 receptors. The failure to detect all types of Fc receptor on the three different effector cells is discussed.     

Studies on antibody-dependent cell-mediated cytotoxicity to Schistosoma japonicum schistosomula in mice

In the presence of antischistosomular serum (Ab), the adherence of macrophages (M?) eosinophils (Eos) and neutrophils(Neu) to and the killing effect on schistosomula of Schistosoma japonicum were studied with a mouse model. In vitro experiment showed that all the three kinds of effector cells could adhere to the surface of the schistosomula when opsonized with specific Ab resulting in the significant increase in the percentage of dead schistosomula. When SPA was added to the system, the percentage of schistosomula with adherent cells decreased markedly. It was revealed that the adherence of cells was at least partially through the binding of Fc fragment of IgG to Fc receptor on the cell surfaces. After having been incubated with Ab and/or cells for 18 h in vitro, the schistosomula were inoculated into the peritoneal cavity of mice. The adult recovery rate 6 weeks after inoculation in groups Eos + Ab and M? + Ab were significantly lower than that of the control group (P less than 0.01).     

Immunoglobulin G-dependent classical complement pathway activation in neutrophil-mediated cytotoxicity to infective larvae of Angiostrongylus cantonensis

The participation of antibody and complement in cell-mediated adherence and cytotoxicity to infected larvae (L3) of Angiostrongylus cantonensis was investigated in vitro. Of the different cell types involved in the reaction, neutrophils were seen to have a predominant role in immune serum--dependent adherence and cytotoxicity to L3. In the presence of immune serum, cytotoxicity to L3 by neutrophils from infected rats was twice that of neutrophils from normal rats. Although mononuclear cells and eosinophils from infected rats significantly increased the adherence to L3, they had little lethal effect on L3. A further study using gel filtration (Sephacryl S-200) and affinity chromatography (protein A) revealed that immunoglobulin G (IgG) alone was responsible for the complement activation in neutrophil-mediated killing of L3. Neither adherence nor cytotoxicity to L3 by neutrophils were affected when immune serum was heated to 50 degrees C or treated with zymosan, but they were markedly decreased when immune serum was treated with Mg2(+)-ethylene glycol-bis-(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid. The results of this study indicate that the neutrophil-mediated adherence and cytotoxicity to L3 of A. cantonensis are mediated through IgG-dependent classical complement pathway activation.     

Enhancement of eosinophil-mediated cytotoxicity to schistosomula of Schistosoma mansoni by autologous mononuclear cells from patients

Adherent mononuclear cells (monolayer), when co-cultured with autologous peripheral blood eosinophils isolated from patients treated for Schistosoma mansoni infections, enhanced the eosinophil-mediated killing of antibody coated schistosomula. The monolayer increased the activity of the eosinophils by 225%, and was observed in 80% of the patients studied. Heat labile factors other than complement, present in immune serum, further enhanced the ability of eosinophils to kill schistosomula in the presence of the monolayer. On their own the adherent cells did not mediate obvious damage to the parasite. Eosinophils that had been pre-incubated with the monolayer (100 mins) and tested separately, killed equal numbers of schistosomula as in the co-culture assay; this excludes the possibility of concurrent schistosomula cytotoxicity by the two cell populations. The ability of the monolayer to activate eosinophils was not altered by inhibitors of protein synthesis. The monolayer was largely consistent of monocytes as demonstrated by an over 96% positive staining for non-specific esterases.     

Suppression of macrophage-mediated antibody-dependent killing of Schistosoma mansoni larvae by concanavalin A stimulated spleen cell-derived factor(s)

Culture supernatants (sup) from rat spleen cells stimulated with concanavalin A (Con A) sepharose 4B suppressed schistosomulum killing by macrophages in a system of antibody-dependent cell-mediated cytotoxicity (ADCC). (a) The suppressive effect of the sup was strongest when they were added to cultures at the start of the ADCC assay, and it was decreased when sup were added to cultures in which the ADCC assay had begun more than 3 h previously. (b) From the results of various experiments in ADCC reaction, it was suggested that the suppressive factor(s) inhibited at least in part the interaction between macrophages and antibodies which had been bound to schistosomula, and that the suppressive factor(s) was bound to the macrophage side. (c) The suppressive activity was partially inactivated by treatment at 56 degrees C for 30 min, and it was lost almost completely by the treatment at 100 degrees C for 10 min. (d) The suppressive factor(s) differed from known cytokines such as interleukin 1 (IL-1), interleukin 2 (IL-2), granulocyte-macrophage-colony stimulating factor (GM-CSF), interferon-gamma (IFN-gamma), and tumour necrosis factor (TNF).     

蒿苯酯预防日本血吸虫病的实验研究

本文报道了蒿苯酯预防动物血吸虫病的实验结果。小鼠、免及犬在单次感染日本血吸虫尾蚴后第7d开始服蒿苯酯,每wkl次,连服4-6次,对宿主体内的血吸虫童虫有明显杀灭效果,减虫率达90％—100％,该药对不同感染度小鼠体内血吸虫童虫的杀灭作用相同。对连续感染14d(每d1次)的家兔减虫率亦达93.5％,蒿苯酯皮下注射给药效果比口服好,但毒性大。比较蒿苯酯、呋喃丙胺和吡喹酮3种药物杀灭血吸虫童虫的效果,以蒿苯酯为最佳。同样,蒿苯酯优于其他两种青蒿衍生物一蒿甲醚和还原青蒿素。     

东方田鼠血清蛋白组分体外杀伤日本血吸虫童虫作用的研究

目的研究东方田鼠血清蛋白组分体外杀伤日本血吸虫童虫的作用。方法采用逐级分离技术,包括硫酸铵盐析法、Sephacryl S-300分子筛层析法以及电泳洗脱法,将东方田鼠血清蛋白分离为球蛋白和白蛋白两大类,再按分子量大小依次将球蛋白分离为4个大的组分和12个小的组分,最后有重点地进行细分。然后,将分离的蛋白组分与日本血吸虫童虫共同培育48h,观察对童虫的杀伤效应。结果东方田鼠球蛋白所致的童虫死亡率为59．2％,加补体后的死亡率为68．4％,明显高于BALB／c鼠球蛋白（分别为40．8％和45．2％）;东方田鼠球蛋白中的2、3组蛋白的杀伤效果同其总球蛋白,其中的3．2、3．3、3．4组分杀伤效果分别达到45．1％、57．6％和67．2％。这3组蛋白的分子量主带为375、205kDa和100～135kDa,后者的杀伤效果接近于总球蛋白,补体有增强作用;BALB／c鼠的4组球蛋白均无明显杀伤作用。这两种鼠的白蛋白对童虫的杀伤作用无明显差异。结论东方田鼠球蛋白具有体外抗血吸虫童虫作用,其中的100-135kDa组分可能为主要效应分子。     

东方田鼠天然抗日本血吸虫感染相关蛋白质的分离和纯化

目的从东方田鼠血清中分离纯化与其天然抗日本血吸虫感染特性相关的蛋白质。方法以分子筛色谱及高效液相色谱技术对东方田鼠血清蛋白质进行分离和纯化,以体外日本血吸虫童虫杀伤实验确定具有抗日本血吸虫童虫活性蛋白质,并对活性蛋白质进行N端氨基酸序列测定,通过检索蛋白质数据库以确定活性蛋白质的性质和种类。结果利用分子筛色谱及高效液相色谱技术从东方田鼠血清中获得了三个对日本血吸虫童虫具有明显杀伤活性的蛋白质组份1-1、4-1、6-3,经对4-1的N末端氨基酸序列进行测定,其序列为DAHKSEIAHR,检索蛋白质数据库,该蛋白质为白蛋白样蛋白质,但与人或其它动物的白蛋白进行比较,存在较大程度的氨基酸残基变。结论东方田鼠血清中的白蛋白是东方田鼠天然抗日本血吸虫感染的重要效应分子。