Autoradiographic analysis of Schistosoma mansoni migration from skin to lungs in naive mice. Evidence that most attrition occurs after the skin phase

A study was performed to determine the extent of attrition of Schistosoma mansoni in naive mice (innate resistance) during the 1st week of infection. Each mouse was exposed to exactly 50 cercariae radiolabeled with [75Se] selenomethionine. On 1, 4, and 7 days postexposure, skin, lungs and liver were analyzed by compressed organ autoradiography for the presence of labeled larvae. Using this technique it was determined that no more than one-third of the 59% attrition that occurred between the cercarial and adult worm stages could be attributed to losses during the skin phase; most of the attrition in naive mice occurred after the migration of larvae to the lungs.     

旋毛虫机蚴抗原诱导小鼠抗日本血吸虫保护性免疫的研究

目的：探讨旋毛虫幼虫抗原免疫小鼠对日本血吸虫攻击感染的交叉保护作用。方法：用4种不同的旋毛虫幼虫抗原制剂经颈部皮下多点免疫小鼠，然后用日本血吸虫尾蚴30条或100条攻击感染，攻击后45d剖杀小鼠，收集成虫、肝组织和粪便中的虫卵，并计数。结果：4种不同制剂均可诱导不同程度的保护性免疫效应，以旋毛虫幼虫匀浆上清可溶性抗原（TsSA）效果较好，减虫率为21．3％，加用福氏佐剂时，其减虫率为29．3％，肝组织和粪便中的虫卵减少率分别达48％和58．5％，平均每鼠肝组织和粪便中的虫卵EPG减少率分别为41．7％和48．9％；当抗原剂量为10000条旋毛虫并加用福氏佐剂时，其减虫率达39．6％。结论：旋毛虫抗原免疫小鼠能诱导抗日本血吸虫攻击感染的免疫效应。     

肝片吸虫和猪蛔虫谷胱甘肽S转移酶免疫小鼠对日本血吸虫尾蚴攻击感染的保护力

[目的 ]观察天然肝片吸虫谷胱甘肽转移酶 (FhGST)和猪蛔虫谷胱甘肽转移酶 (AsGST)免疫的小鼠对日本血吸虫大陆株尾蚴攻击感染的保护力。[方法 ]从肝片吸虫和猪蛔虫中提取天然谷胱甘肽转移酶 (GST) ,免疫 3次 ,间隔时间为 14d和 7d。第 3次免疫后 5d经腹部皮肤攻击感染大陆株日本血吸虫尾蚴 30条 鼠。感染6wk后用肝门静脉灌注法收集成虫 ,分别计数肝脏、脾脏和大肠组织沉积虫卵数 ,并计算减虫率和减卵率。[结果 ]3个免疫组 (FhGST、AsGST和日本血吸虫rGST)分别与佐剂对照组和空白对照组比较的减虫率分别为2 7 8%、 37 9%、 33 1%和 31 3%、 41 0 %、 36 4% (P <0 0 1)。肝脏的EPG减卵率为 13 5 %～ 17 7% (佐剂对照组 ,P <0 0 5 )和 2 3 9%～ 2 7 6 % (空白对照 ,P <0 0 1)。脾脏的EPG减卵率为 30 4%～ 5 9 6 % (P <0 0 5 ) ,大肠的为 38 7%～ 6 2 3% (P <0 0 1)。 [结论 ]FhGST和AsGST具有明显抗血吸虫感染的保护力。     

UV致弱尾蚴免疫对小鼠血吸虫再感染的保护力观察

目的观察致弱日本血吸虫尾蚴免疫小鼠再次感染血吸虫后的减虫率、减卵率及肝脏病理损伤,为血吸虫疫苗的研制奠定基础。方法分别以400μw／cm^2×60s和422μw／cm^2×40s两种不同UV强度及时间照射的日本血吸虫尾蚴免疫C57BL／6和DBA小鼠,观察免疫小鼠对再次血吸虫感染的减虫率、肝脏减卵率及肝脏病理改变。结果400μw／cm^2×60s（A）和422μw／cm^2UV×40s（B）照射的日本血吸虫尾蚴免疫组C57BL／6小鼠再次感染血吸虫后的减虫率分别为一0．60％和0．02％,肝脏肝脏减卵率分别为2．70％和11．37％;DBA小鼠再次感染血吸虫后的减虫率分别为29．10％和25．70％,肝脏肝脏减卵率分别为59．50％和69．50％。422μw／cm^2UV×40S辐照尾蚴免疫C57BL／6小鼠,再次感染血吸虫形成的肝脏单个虫卵肉芽肿面积与对照组比较显著减小（P〈0．01）;400μw／cm^2UV×60s和422μw／cm^2UV×40S辐照尾蚴免疫DBA小鼠再次感染血吸虫造成的肝脏单个虫卵肉芽肿面积与对照组比较显著减小（P〈0．01）。结论UV致弱尾蚴免疫对C57BL／6、DBA小鼠再次感染血吸虫的保护作用较小,但能降低肝脏卵荷并减轻肝脏的病理损伤。     

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。     

紫外线致弱日本血吸虫尾蚴诱导BALB／c小鼠免疫保护作用研究

目的观察紫外线（UV）致弱日本血吸虫尾蚴免疫BALB／c小鼠免疫保护作用。方法辐照致弱日本血吸虫尾蚴,经腹部皮肤用UV致弱日本血吸虫尾蚴免疫诱导BALB／c小鼠,3周后进行攻击感染,同时设正常感染对照组。攻击感染6周后解剖小鼠,计算减虫率、减卵率,并观察虫体发育情况和肝组织细胞免疫应答情况。结果免疫组的减虫率和减卵率分别为37．90％和51．16％;免疫组小鼠体内虫体发育明显滞后于正常感染对照组内虫体;免疫组小鼠肝脏虫卵肉芽肿较正常组明显减少。结论UV致弱日本血吸虫尾蚴免疫BALB／c小鼠能诱导较好的免疫保护力。     

紫外线减活日本血吸虫尾蚴疫苗免疫小鼠肺部iNOS的表达及其与免疫保护关系的研究

目的 研究紫外线照射减活日本血吸虫尾蚴疫苗免疫小鼠后肺部诱导型一氧化氮合酶(iNOS)的动态表达及其与保护性免疫的关系。 方法 用紫外线照射减活尾蚴疫苗免疫小鼠,30 d后用尾蚴攻击感染。于感染后第4 d、9 d取免疫小鼠和未免疫小鼠肺脏,用RT-PCR半定量的方法检测各组小鼠肺部iNOS的转录水平;用免疫组化方法确定iNOS在肺部的蛋白表达定位。 结果 正常小鼠肺部无iNOS转录和蛋白表达。免疫小鼠和未免疫小鼠感染血吸虫尾蚴后第4 d肺部iNOS有较高水平的转录表达。感染后第9 d,免疫组小鼠iNOS的表达较第4 d下降,而未免疫组小鼠肺部iNOS的表达消失。免疫组化结果表明,免疫组小鼠肺部炎症病灶细胞中存在iNOS的表达。 结论 移行到肺部的血吸虫童虫能诱导肺部表达iNOS,紫外线减活尾蚴疫苗能促使小鼠肺组织较长时间表达iNOS,iNOS在肺部的持续表达可能与该疫苗使宿主产生的免疫保护力有关。     

辐照旋毛虫肌肉幼虫免疫对日本血吸虫感染的保护作用

目的　观察辐照旋毛虫肌肉幼虫 (TsML)免疫小鼠对日本血吸虫攻击感染的保护效果。　方法　用60 Co照射的TsML经腹腔免疫小鼠 15只 ,另设TsML匀浆和生理盐水免疫组作为对照。在两次加强免疫后 (1次 /周 ) ,分别用日本血吸虫尾蚴攻击感染。感染后第 40d解剖小鼠 ,计算虫荷 ,并对肝组织中的虫卵和雌虫子宫内的虫卵进行计数。同时采用数字图像处理技术对成虫形态结构进行测量分析。　结果　 1)辐照TsML免疫组与生理盐水对照组比较 ,小鼠虫荷数、肝组织和雌虫子宫内的虫卵数显著减少 ;免疫小鼠体内血吸虫虫体及睾丸、卵巢发育受到明显影响 ;2 )TsML匀浆免疫组的减虫效果较差 ,减卵效果及虫体、睾丸、卵巢发育与辐照TsML免疫组相似。　结论　辐照TsML和匀浆TsML免疫均能诱导小鼠产生对日本血吸虫攻击感染的保护作用 ,且辐照TsML较匀浆TsML具有更好的保护效果。     

日本血吸虫极低密度脂蛋白结合蛋白重组抗原的免疫保护性研究

目的?摇探讨日本血吸虫极低密度脂蛋白结合蛋白（SVLBP）重组抗原的免疫保护性及其作为候选疫苗的潜在价值。 方法 用异丙基-β-D-硫代半乳糖苷（IPTG）诱导克隆菌大量表达SVLBP重组抗原，以镍-次氮基三乙酸琼脂糖树脂（Ni-NTA）亲和层析法纯化制备SVLBP重组抗原；将纯化的重组抗原加福氏佐剂经皮下免疫C57BL/6小鼠，每隔2周免疫1次，在第3次免疫后10 d，经腹部皮肤攻击感染日本血吸虫尾蚴35±1条，感染后45 d剖杀，计数检获虫数和每克肝虫卵数（LEPG）。小鼠在免疫前和攻击感染后分别采眶窦血，用ELISA测定特异性IgG及其亚群抗体水平。 结果 相对于佐剂对照组，免疫组小鼠的减虫率为33.4%，减卵率为47.6%；免疫后小鼠产生高水平的特异性IgG（>1:6 400）；抗体亚类检测结果显示，免疫组小鼠 IgG2a、IgG2b、IgG1明显高于免疫前和佐剂对照组。 结论 日本血吸虫皮层蛋白SVLBP重组抗原能诱导小鼠产生一定的保护性免疫，是潜在的疫苗候选抗原分子。     

SIEA对感染了日本血吸虫尾蚴的小鼠诱导的免疫力研究

目的　采用尾蚴攻击感染小鼠后免疫的方式 ,研究日本血吸虫未成熟卵可溶性抗原 (SIEA)诱导抗卵胚胎发育和抗虫免疫的效果。方法　感染日本血吸虫尾蚴之后的第 2天用 SIEA免疫小鼠 ,每周 1次 ,共 5次 ,于第 48天剖杀小鼠 ,统计肝脏各期虫卵数、雌虫子宫虫卵数及每鼠成虫数。结果　与对照组比较 ,SIEA免疫诱导显著抗卵胚胎发育效果 ,肝组织成熟卵数下降 2 7.5 % ,成熟卵比例下降 11.0 % ,但 SIEA未诱导显著抗虫和抗雌虫生殖效果。结论　采取在尾蚴感染之后免疫 ,SIEA能诱导显著抗卵胚胎发育 ,但比尾蚴感染小鼠之前免疫的抗卵胚胎发育效果低     

The fate of challenge schistosomula in the murine anti-schistosome vaccine model

Mice exposed to irradiated cercariae of Schistosoma mansoni develop a partial resistance to subsequent parasite challenge. In this study we utilized histopathologic methods to investigate the fate of both the immunizing and challenge cercariae in C57BL/6J mice. After immunization by percutaneous infection, a large number of the 50 Kr irradiated organisms could be detected in tissue sections of lung. However, as early as 2 weeks after immunization, the majority of these schistosomula apparently had died, leaving residual inflammatory foci. The numbers of these foci then gradually declined during the next 4 weeks of examination. Cercarial challenge of mice vaccinated 4 weeks previously provoked an intense eosinophil-enriched inflammatory response in percutaneously exposed ear pinnae. Despite these pronounced tissue reactions, no evidence of significant parasite damage or attrition was detected in this migration site. In contrast, schistosomula arriving in the lungs of vaccinated mice produced a greater number of residual inflammatory foci than did larvae appearing in the lungs of normal mice. In addition, challenge schistosomula were cleared from the lungs of vaccinated mice at a slower rate than they were from the lungs of control mice. These observations suggest that the lung is a major site of parasite attrition for both immunizing and challenge infections in the mouse irradiated vaccine model.     

日本血吸虫pcDNA3/SjCWL01核酸疫苗的构建及其对小鼠免疫效果观察

目的构建日本血吸虫pcDNA3/SjCWL01核酸疫苗并进行免疫保护效果观察,评价其作为疫苗的潜能。方法将日本血吸虫基因SjCWL01亚克隆入真核表达载体pcDNA3,构建目的基因真核表达质粒,将pcDNA3/SjCWL01质粒转化大肠杆菌DH5α,大量制备DNA疫苗并免疫小鼠3次,间隔2w,末次免疫后2w,用ELISA法检测免疫鼠血清特异性抗体效价,日本血吸虫尾蚴进行腹部皮肤攻击感染,感染后45d剖杀冲虫,分别计算减虫率,每克肝、粪卵减少率。结果重组DNA疫苗(pcDNA3/SjCWL01)与对照组比较,虫荷、每克肝卵、每克粪卵数分别下降了27.6%,39.5%,45.9%。结论表明pcD-NA3/SjCWL01疫苗可诱导小鼠产生部分抗血吸虫感染的保护力。     

日本血吸虫云南品系SIEA对小鼠感染皖鄂品系血吸虫的免疫保护作用(英文)

目的观察日本血吸虫云南品系未成熟卵可溶性抗原SIEA免疫小鼠后攻击感染皖鄂品系血吸虫尾蚴,是否也能诱导出免疫保护效果。方法 20只ICR小鼠随机分为免疫组和对照组,经日本血吸虫云南品系SIEA免疫后攻击感染皖鄂品系血吸虫尾蚴,感染后46 d观察减虫率、粪卵减少率、雌虫子宫内虫卵数、肝表面虫卵结节数、肝脏各期虫卵数及各期虫卵构成比。结果 SIEA免疫后粪卵及雌虫子宫内虫卵减少率分别为34.45%,23.69%(P<0.05);肝表面虫卵结节密度下降29.03%(P<0.05);肝组织内虫卵减少29.07%(P<0.05),成熟虫卵减少48.41%(P<0.05)。肝组织内成熟虫卵比例下降,未成熟卵比例增加(P<0.05)。结论日本血吸虫(云南品系)未成熟卵可溶性抗原(SIEA)的抗原性较强,小鼠经云南品系SIEA免疫后攻击感染皖鄂品系血吸虫尾蚴也诱导小鼠产生了抗卵胚发育及抗雌虫生殖免疫力,结果提示将来在中国大陆使用SIEA疫苗时,似可不必考虑血吸虫品系的影响。     

Evidence that both normal and immune elimination of Schistosoma mansoni take place at the lung stage of migration prior to parasite death.

The number and distribution of autoradiographic foci observed in this and previous studies following percutaneous infection with 75Se-labeled Schistosoma mansoni cercariae indicate that the lungs are the principal site of worm elimination in both normal mice and mice immunized with irradiated cercariae. It was observed in the present study, however, that the intensities of the autoradiographic foci produced in the lungs during both the normal (early) and immune (late) phases of elimination were identical to those of foci produced in the livers of the same mice by larvae shown to be alive. In contrast, foci produced in the lungs by heat-killed, intravenously injected, lung schistosomula became smaller and fainter with time, disappearing completely between seven and 10 days after injection in normal mice and between four and six days in immunized mice. These results indicate that although the targets of both normal and immune elimination do not proceed beyond the lung stage of migration, they do not die in the lungs. A possible explanation for this paradoxical situation, for which there is some experimental evidence, is that unsuccessful migrators leave the blood stream, enter alveoli, pass up the trachea, and are eventually digested in the gastrointestinal tract or eliminated from the body intact.     

The response of mice immune to Schistosoma mansoni to a challenge infection which bypasses the skin: evidence for two mechanisms of immunity

Mice which had developed immunity to reinfection with Schistosoma mansoni following exposure to 20 cercariae and mice which had been immunized against S. mansoni by exposure to 400 highly irradiated (20 krad) cercariae, were tested for their ability to resist a percutaneous cercarial challenge and an intravenous challenge with 5-day-old lung-stage schistosomula derived from the same cercariae. Although both types of immune mice showed a marked resistance to a cercarial challenge, only the infected mice showed a comparable immunity to an intravenous challenge with lung schistosomula. These results confirm earlier studies which suggest that the major attrition of a cercarial challenge in infected mice occurs at the post-lung stage, whilst the attrition of a challenge infection in mice immunized with highly irradiated cercariae takes place in the skin. They provide further evidence for two separate mechanisms of immunity against S. mansoni in mice.     

日本血吸虫感染对小鼠肝药物代谢酶的影响

本文观察了日本血吸虫感染对小鼠肝药物代谢酶的影响。结果,感染20、40和60条血吸虫尾蚴6wk的小鼠,其成巴比妥钠的催眠时间分别为53±9、100±64和172±93min,表明随感染度的增加而明显延长。体外实验观察到各感染度小鼠的肝苯胺羟化酶(Aniline hydroxylase,AH)、氨基比林N-脱甲荃酶(Aminopyrine N-demethylase,APD)的活性和细胞色素P-450(Cy-tochrome P-450)含量均明显降低。即使相同感染度不同病程也显示不同差异,感染30条尾蚴8wk小鼠的肝AH、APD活性和P-450含量比感染4wk小鼠明显降低,分别为对照的32.7％、13.7％和7.7％。说明日本血吸虫感染能明显降低小鼠肝药物代谢酶活性,其影响程度随感染度和病程不同而呈现明显差异。     

不同免疫途径对Sj童虫细胞型免疫原诱生的保护性效果及抗体应答差异

目的比较研究小鼠经不同途径接种日本血吸虫(Schistosoma japonicum,Sj)童虫原代细胞(primary juvenile worm cells,pJCs)后所诱导的免疫保护效果,寻找其合适的免疫途径。方法将pJCs经皮下或静脉注射途径免疫昆明小鼠,间隔2w共免疫3次,末次免疫后第4w,与对照组同时经腹部皮肤感染尾蚴30±2尾/鼠,比较三组小鼠的肝脏虫卵数及虫卵肉芽肿的大小、成虫数、虫体大小。同时在第2、3次免疫前及攻击感染前1d,小鼠尾静脉采血,用ELISA法检测抗pJCs-IgG水平。结果 pJCs免疫小鼠后,与PBS对照组比,静脉免疫组的减虫率为48.53%、肝卵减少率为54.54%、虫体平均重量减少率为29.62%(P0.01)、虫卵肉芽肿面积减少率为36.8%,而皮下免疫组分别为41.28%、43.19%、23.08%、31.2%。IgG抗体水平也是静脉注射组高(PBS组,P0.01;皮下注射组,P0.05)。结论日本血吸虫童虫细胞免疫原通过皮下和静脉注射均可诱导小鼠产生对日本血吸虫的免疫保护力,其中静脉注射诱导的免疫保护效应最强,提示通过静脉注射日本血吸虫童虫细胞是可行有效的免疫途径。     

Comparative phenotypic analysis of lymph node cells in mice after infection or vaccination with normal or ultraviolet-attenuated cercariae of Schistosoma japonicum or S. mansoni

Mice were infected with 200 untreated or vaccinated with 500 ultraviolet-attenuated cercariae of either Schistosoma japonicum or S. mansoni. For three weeks, cell numbers in axillary and mediastinal lymphnodes were counted and cell populations typed by cytofluorometry. In the axillary lymphnodes, numbers of B-cells and CD3+CD4+ T-cells but not CD3+CD8+ T-cells increased. Following vaccination with either species, parasite migration was apparently delayed in the skin and interrupted at the lungs, the lymphnodes gained weight, and cell numbers of axillary lymph nodes increased more than after infection. In mediastinal lymphnodes, only immunization with S. japonicum but not S. mansoni cercariae led to an increase of CD3+CD4+ T-cells. Following infection, both schistosome species induced higher CD3+CD4+, but not CD3+CD8+ T-cells in mediastinal nodes, and the peak was earlier with S. japonicum (about seven days after infection) than with S. mansoni (about 10 days). In analogy to T-cell observations by others using a gamma-attenuated cercarial vaccine in S. mansoni, the present results suggest that CD3+CD4+ cells also play a role in the ultraviolet-attenuated vaccine against S. japonicum.     

诱变剂NTG致弱的日本血吸虫尾蚴免疫效果的进一步观察

为了进一步了解诱变剂(NTG)致弱尾蚴的免疫效果,以NTG(30μg/ml)致弱15min后,经皮感染免疫小鼠两次,间隔一周,观察免疫后不同时间攻击感染(2、4、6、8和10wk)对免疫鼠存活时间,减虫率,肝、脾肿大及肝、脾虫卵肉芽肿程度的影响。同时,还对免疫鼠体内血吸虫进行虫体表面皮层的电镜扫描观察。实验结果表明:NTG致弱尾蚴初次免疫后,8wk攻击感染时减虫率最高(77％),且肝、脾肿大及虫卵肉芽肿病变程度最轻.小鼠存活时间也长。从而提示NTG致弱尾蚴免疫小鼠后,对攻击感染确产生免疫力。但免疫后不同时间所产生的免疫力不同,初次免疫后8wk可能是最适攻击感染时间。对免疫后不同时间所产生的免疫力不同的原因进行了讨论,NTG致弱尾蚴诱导保护性免疫力产生的机制尚待进一步研究阐明。     

Recovery and distribution of Ascaris suum superimposed on a Schistosoma japonicum infection in pigs

The aim of this study was to examine the effect of a primary patent Schistosoma japonicum infection on the establishment and location of a superimposed Ascaris suum infection in pigs. The study comprised two experiments each containing two groups of pigs. In the first experiment, 7 pigs were injected intramuscular (i.m.) with 800 S. japonicum cercariae and inoculated with 1,000 A. suum eggs 11 weeks post primary infection (ppi) and 8 pigs were inoculated with 1,000 A. suum eggs at the time of challenge infection. In the second experiment, 7 pigs were injected i.m. with 1,100 S. japonicum cercariae and inoculated with 1,000 A. suum eggs 16 weeks ppi and 8 pigs were inoculated with 1,000 A. suum eggs at the time of challenge infection. All pigs were slaughtered 10 days after the A. suum challenge infection. The number of white spots caused by A. suum on the surface of the liver was significantly lower in the groups with primary infections of S. japonicum compared with the control groups. However, the present experiments did not demonstrate any effect of a primary S. japonicum infection on the total recovery and distribution of an A. suum challenge infection.