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。     

益肾蠲痹丸及雷公藤对日本血吸虫虫卵肉芽肿的影响

对感染15条日本血吸虫尾蚴的雌性小鼠.分别于感染后25d及45d起喂服益肾蠲痹丸和雷公藤.结果表明益肾蠲痹丸对虫卵肉芽肿发展具明显抑制作用,肉芽肿平均直径和肉芽肿内嗜酸粒细胞均明显减少.雷公藤对急性期虫卵肉芽肿可有促进发展作用,而对慢性期虫卵肉芽肿则有抑制作用.     

小鼠感染后不同时间血清对日本血吸虫童虫细胞介导的细胞毒作用

用抗体依赖、细胞介导的细胞毒试验(ADCC)观察了小鼠感染后不同时间血清在体外杀伤系统中对日本血吸虫童虫的作用规律。未加补体时,嗜酸粒细胞或巨噬细胞介导的童虫杀伤作用于感染后4wk出现明显作用,分别于5—7wk和6—8wk达高峰,然后下降,至11wk时降至4wk时的水平;而未加补体时无明显的中性粒细胞介导的杀伤作用。在补体参与下,这三种细胞均能介导对童虫的杀伤作用,且作用增强。证实ADCC是日本血吸虫获得性抵抗力的一个重要组分,嗜酸粒细胞、巨噬细胞介导的对童虫的细胞毒作用是不依赖补体的,而中性粒细胞介导的作用是依赖补体的。结果对选择免疫预防适宜时间和评价候选疫苗效果有参考意义。     

紫外线减毒日本血吸虫尾蚴免疫小鼠的皮肤组织细胞反应

目的 :探讨紫外线减毒血吸虫尾蚴免疫宿主的皮肤组织在抗攻击感染中的作用。方法 :88只小鼠分为 4组 ,1、2组分别以 50± 3条紫外线减毒日本血吸虫尾蚴免疫或日本血吸虫尾蚴感染 ,5wk后以 10 0 0± 10 0条尾蚴进行攻击感染 ;3、4组分别以减毒尾蚴或尾蚴 10 0 0± 10 0条免疫或初次感染。各组均于感染或接种后 6- 12 0 h定时取局部皮肤组织作病理观察。结果 :( 1)免疫攻击组皮肤组织内炎症细胞杀伤童虫现象最明显、炎症反应最强且持续时间最长 ,嗜酸性粒细胞( EOS)浸润百分数最高 ;( 2 )感染攻击组呈现相似炎症反应但程度略轻 ;( 3)免疫对照组减毒童虫在皮肤内滞留时间延长 ,至 12 0 h皮下仍可见较多童虫 ;( 4 )感染对照组皮肤内的童虫数于 72 h后明显减少 ,童虫周围轻微炎症反应。电镜观察可见免疫攻击组童虫内部结构破坏 ,虫体周围 EOS、淋巴细胞增多 ,肥大细胞颗粒溶解。结论 :提示皮肤组织的细胞免疫反应在血吸虫减毒尾蚴免疫的宿主抗攻击感染保护性免疫中起重要作用。     

在体外人的嗜酸粒细胞和日本血吸虫童虫的相互作用:IgG依赖细胞的粘附和杀虫效果

作者旨在通过体外实验证明,血清中嗜酸粒细胞能粘附于日本血吸虫童虫,有杀灭童虫的作用。嗜酸粒细胞、中性粒细胞的制备:从5例嗜酸粒细胞增多症的患者,于治疗前抽静脉血加入肝素,然后按Gartner方法(1980)纯化。血吸虫童虫的收集:以钉螺逸出的日本血吸虫山梨株的尾蚴经肤感染小鼠,用Clegg     

在体外人嗜酸粒细胞对埃及血吸虫童虫的抗体依赖的杀伤

本文报道在体外人嗜酸粒细胞对埃及血吸虫童虫具有抗体依赖的杀伤作用。童虫的制备:从感染区儿童尿中获得埃及血吸虫虫卵,孵出毛蚴,感染螺蛳,然后使尾蚴自螺体逸出,制备皮肤期童虫。童虫活力通过甲苯胺蓝在玻片上检测。嗜酸粒细胞的制备:按Vadas等(1979)法纯化自愿者外周血中的嗜酸粒细胞,纯度达95％以上。血清的制备:静脉穿刺或手指取血,     

小鼠感染日本血吸虫后iNOS的表达及NO介导的杀虫作用研究

目的研究小鼠在感染日本血吸虫后肺部诱导型一氧化氮合酶(iNOS)的动态表达及一氧化氮(NO)对童虫的杀伤作用。方法在感染日本血吸虫后不同时间点剖杀小鼠并取其肺组织,用半定量RTPCR的方法检测肺部iNOS的转录水平;用NOS的抑制剂AG(Aminoguanidine,氨基胍)处理感染小鼠,比较处理组与对照组虫荷的高低以确定NO对童虫的杀伤作用。结果未感染小鼠肺部无iNOS转录表达,感染后第4d肺部iNOS有较高水平的转录表达,第9d表达消失;AG处理小鼠的虫荷较未处理组高(P<0.05)。结论移行到肺部的童虫能诱导肺组织表达iNOS,由iNOS产生的NO能对童虫发挥杀灭作用。     

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。     

小鼠初次和攻击感染日本血吸虫尾蚴后皮肤内的细胞反应

对正常和慢性感染鼠,经耳廓皮肤分别初次和攻击感染日本血吸虫尾蚴。攻击感染15b后,慢性感染鼠皮肤内的嗜酸粒细胞(EOS)较初次感染鼠明显增加.两组鼠炎症细胞在童虫附近或包围完整及解体的童虫,形成粒细胞性微脓肿。超微结构显示,扁平的EOS紧贴童虫表面,并脱颗粒。粘附处局部皮层可见破坏现象。中性粒细胞(NEU)的粘附不如EOS紧密,未见脱颗粒现象,童虫皮层完整。结果提示,EOS是对日本血吸虫皮肤期童虫有效的杀伤细胞.     

青蒿琥酯治疗小鼠日本血吸虫病的血清学研究

目的观察青蒿琥酯对日本血吸虫感染小鼠的治疗作用及血清IgG的动态变化，探讨血清抗体与体外杀童虫效果的相关性。方法用日本血吸虫感染C57／BL6小鼠，在感染后8、15、22和29d分别一次性灌服300mg／kg青蒿琥酯，对照组小鼠灌服1％羧甲基纤维素钠。最后一次给药后7d剖杀小鼠，计算成虫数和虫卵数，以ELISA法检测小鼠血清中日本血吸虫特异性IgG抗体的水平，观察接受青蒿琥酯治疗的小鼠血清与巨噬细胞体外协同杀伤日本血吸虫童虫的效应。结果青蒿琥酯治疗组小鼠未检获成虫和虫卵；血清中日本血吸虫尾蚴抗原特异的IgG抗体水平在感染后21d达高峰，此后下降；成虫抗原特异的IgG抗体水平呈上升态势，二者与对照组比较差异均无显著性；青蒿琥酯治疗小鼠血清中虫卵抗原特异的IgG抗体呈低水平状态，显著低于对照组。青蒿琥酯治疗组小鼠血清和对照组小鼠血清均具有较强的体外杀童虫效果。结论青蒿琥酯对日本血吸虫感染的小鼠具有显著的治疗效果，其血清与巨噬细胞在体外具有协同杀伤童虫的作用。     

日本血吸虫感染小鼠血清中虫体核酸来源的实验研究

目的探讨日本血吸虫感染小鼠血清中核酸的来源,为血吸虫病核酸诊断方法的早期诊断及疗效考核提供理论依据。方法用PCR法检测单、双性感染小鼠1w~12w血清中的特异的日本血吸虫DNA片断。结果在单性感染小鼠1w~4w的血清可通过PCR扩增到特异性DNA片段,感染小鼠5w后的血清中则未能测出阳性结果;在双性感染小鼠组中,应用PCR从1w~12w的血清中均可扩增到特异的阳性条带。结论在日本血吸虫感染早期,小鼠血清中的核酸可能主要来自在体内移行的日本血吸虫童虫及虫体在发育过程中的代谢产物;在感染的急性期和慢性期,血清中的核酸则可能主要来自小鼠体内崩解死亡的虫卵组织。     

Cellular responses elicited by challenged schistosomula of Schistosoma japonicum in the skin of naive and chronically infected mice

Naive and chronically infected C57BL/6 mice were challenged percutaneously over the ear pinna with Schistosoma japonicum cercariae. After 15 hours, the number of EOS increased significantly in the skin of chronically infected mice. Inflammatory cells aggregated in the vicinity of schistosomula or entrapped intact and disintegrated schistosomula, forming granulocytic micro-abscesses in both groups. Ultrastructure studies revealed that flattened EOS tightly attached to the schistosomulum surface and degranulated. Local tegument damage occurred in the area of attachment. NEU adherence did not seem to be as intimate as EOS, and degranulation was not seen. The tegument of the attached schistosomulum remained normal. The result suggested that EOS appeared to be the efficient killer cell against skin phase schistosomula of S. japonicum.     

On the interaction between macrophages and developmental stages of Schistosoma mansoni: effect of muramyl tripeptide phosphatidyl ethanolamine (MTP-PE) treatment on mice survival and the generation of schistosomulicidal macrophages

Schistosomiasis is a chronic disease afflicting hundreds of millions of people throughout the world against which there is as yet no effective vaccine. In the present study we tested the effect of the immunomodulator muramyl tripeptide phosphatidyl ethanolamine (MTP-PE) on the survival of Schistosoma mansoni-infected mice and on the induction in them of schistosomulicidal macrophages. Mice exposed to 80 cercariae each and then treated with MTP-PE showed prolonged survival following either single or repeat infection. The treatment with MTP-PE, when initiated 70 days post the schistosome infection, diminished significantly the mortality of infected mice over an observed period of 110 days. In terms of treatment efficacy there was no evident difference between the intravenous and intraperitoneal mode of administration of the drug. MTP-PE treatment significantly reduced granuloma size and markedly diminished liver damaged as judged by the lower levels of alkaline phosphatase in the serum. Such treatment exerted no significant effect on the spleen or liver weight in infected mice nor on the worm burden resulting from either a single or double infection. In infected and non-treated mice, schistosomulicidal macrophages appeared after 8-10 weeks of infection. In infected mice treated with MTP-PE there was an accelerated appearance of such macrophages and these exhibited a greater cidal effect on the schistosomula. These immunostimulatory and life-prolonging effects of MTP-PE on S. mansoni-infected mice might indicate an effect of this reagent on cells involved in the granulomatous process.     

共刺激分子B7-1介导Th1/Th2免疫应答对日本血吸虫虫卵肉芽肿病变的影响

目的观察B7-1激发型mAb的体内注射对日本血吸虫感染小鼠Th1/Th2免疫偏移的影响及对虫卵肉芽肿病变的调节作用,探讨干预共刺激信号调控Th1/Th2免疫偏移控制血吸虫虫卵肉芽肿病变的新途径。方法小鼠感染日本血吸虫尾蚴后4w给予腹腔内注射B7-1激发型mAb,于6、8wk摘眼球取血收集血清,用ELISA双抗体夹心法测定血清中抗体IgGI、gG1I、gG2a的水平;同时取脾淋巴细胞培养72h,同法测定培养上清中细胞因子IFN-γ和IL-4的表达水平;并且检测感染小鼠虫卵肉芽肿体积。结果B7-1激发型mAb能显著下调感染小鼠血清IgG水平,并能显著上调IFN-γ的表达水平,下调IL-4的表达水平,同时能使虫卵肉芽肿体积显著减小。反映Th1/Th2免疫偏移的IgG1/IgG2a有下降趋势。结论B7-1激发型mAb能调节Th1/Th2免疫应答,诱导免疫反应向Th1方向偏移,从而为控制日本血吸虫虫卵肉芽肿病变的免疫治疗新途径提供了实验依据。     

Detection of high molecular weight eosinophil chemotactic factor in murine schistosomiasis sera

Eosinophil chemotactic activity in sera from mice undergoing an acute stage of schistosomiasis japonica and mansoni was examined. Eosinophilotactic activity in the serum was dependent on the dose and time of infection. Eosinophilotactic activity in sera from S. japonicum-infected mice was higher than that from S. mansoni-infected mice when they were compared at the comparable dose and time of infection. After gel chromatography on Sephadex G-200, eosinophilotactic activity in sera from mice infected with 30 cercariae of S. japonicum for 5 weeks was detected in the high molecular weight component. On the other hand, when sera from mice infected with 30 cercariae of S. japonicum for 8 weeks was chromatographed through Sephadex G-200 columns, eosinophilotactic activity was segregated into high (greater than 455,000) and low (less than 13,000) molecular weight components. High molecular weight ECF in sera from mice infected with 30 cercariae of S. japonicum for 8 weeks had high affinity to Con A, and was stable to heating or pronase digestion, but was sensitive to periodate oxidation, indicating its polysaccharide or glycoprotein nature. This high molecular weight ECF could be adsorbed by, and eluted from immunoaffinity beads coated with rabbit IgG anti-S. japonicum adult worm antibody. Thus, at least some part of circulating high molecular weight ECF would be derived from adult parasites.     

STAT4和STAT6对日本血吸虫发育生殖及虫卵肉芽肿形成的影响

目的观察STAT4或STAT6缺陷对日本血吸虫发育生殖及宿主虫卵肉芽肿形成的影响。方法设正常BALB/cJ小鼠及遗传背景相同的STAT4-/-、STAT6-/-小鼠3组,每组10只,每只感染日本血吸虫尾蚴25条。于感染后第42天剖杀小鼠,收集虫体,计数总虫荷及雌、雄虫荷;另取小鼠肝组织,计数肝内虫卵;制作肝组织切片,观察虫卵肉芽肿病理学变化,测量单个虫卵肉芽肿直径。结果 STAT4-/-和STAT6-/-小鼠体内总虫荷、雌雄虫配对数及肝组织中每对虫体平均产卵数与正常BALB/cJ小鼠无明显差异（P均〉0.05）。STAT6-/-小鼠肝脏中单个虫卵肉芽肿直径为（213.3±68.6）μm,与感染组BALB/cJ小鼠（319.5±71.9）μm比较,差异有统计学意义（P〈0.05）,且虫卵肉芽肿较不典型、纤维化较不明显。STAT4-/-小鼠肝脏单个虫卵肉芽肿直径及其病理学改变与正常BALB/cJ小鼠相比无明显差异。结论 STAT4或STAT6缺陷对日本血吸虫发育生殖无明显影响。STAT6缺陷不利于虫卵肉芽肿的形成及纤维化,STAT4缺陷对虫卵肉芽肿的形成无明显影响。     

氯化钆对血吸虫肉芽肿期肝脏调节性T细胞的抑制作用

目的探讨氯化钆封闭Kupffer细胞功能对日本血吸虫病肉芽肿期CD4+CD25+T细胞的影响。方法6～8w龄雌性C57BL/6小鼠分为3组,即对照组、感染组与感染/氯化钆组,其中氯化钆为15 mg/kg,尾静脉注射,每w2次;感染第8w流式细胞仪检测CD4+CD25+T细胞的数量;免疫组织化学染色检测Foxp3的分布;酶联免疫吸附试验检测血细胞因子IL-4、IL-5、IL-10、TGF-β1与IFN-γ的表达水平。结果感染小鼠CD4+CD25+T细胞数量为13.8%,感染/氯化钆组为9.3%(p<0.05),对照组为6.4%;IL-10在感染组为41.4 pg/mL,感染/氯化钆组22.6 pg/mL;此外,氯化钆还能下调Foxp3的分布以及减轻血吸虫肉芽肿周围的炎症反应。结论氯化钆封闭Kupffer细胞的功能后可减少日本血吸虫感染肉芽肿期CD4+CD25+T细胞的产生以及减轻肉芽肿周围的炎症反应。     

日本血吸虫肺期童虫收集方法的实验研究

目的建立一种收集纯净肺期童虫的方法。方法用日本血吸虫尾蚴感染昆明小鼠,分别于感染后第1、2、3、4、5、6、7、10、12、14、21天解剖,从皮肤、肺脏、肠系膜静脉及肝门静脉处收集童虫。结果肺期童虫出现的高峰期为感染后第3天,获得最佳收获肺期童虫的时间,建立了回收大量纯净童虫的方法。     

microRNA在小鼠血吸虫病及吡喹酮治疗中的表达特征

目的 目的 探究炎症调节基因miR?155和miR?146a及炎症相关基因在小鼠血吸虫病及吡喹酮治疗中的表达特征, 为 进一步阐明吡喹酮治疗血吸虫病的作用机制奠定基础。方法 方法 以BALB/c小鼠为研究对象, 建立日本血吸虫感染的动物模 型; 小鼠随机分为4组： 正常组、 感染6周组、 感染12周组和吡喹酮 （PZQ, 300 mg/kg 1次灌胃杀虫治疗） 治疗组。以HE染色 观察肝脏病理变化; 以Real?time PCR检测肝脏miR?155和miR?146a及炎症相关基因TNF?α、 IL?1β和IL?6的mRNA水平。 结果 结果 感染6周组小鼠的肝脏miR?155、miR?146a及TNF?α、 IL?1β和IL?6的mRNA水平均显著高于正常组和感染12周组 （P＜0.05）; 与感染12周组小鼠相比, PZQ治疗组小鼠肝脏虫卵肉芽肿反应减轻, 且肝脏miR?155、 miR?146a及TNF?α、 IL?1β 和IL?6的mRNA水平有所升高（P＜0.05）。结论 结论 本研究发现miR?155和miR?146a可能与血吸虫病肝脏炎症的发生发展 有关, 并且参与了吡喹酮对炎症的调节。     

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).