肝泡状棘球蚴组织中HIF-1α、VEGFA的表达及对血管生成的作用

目的 探讨HIF-1α、VEGFA在沙鼠肝泡状棘球蚴组织的表达及血管生成过程中的作用。方法 126只沙鼠随机分成空白组(6只)、假手术组(60只)、模型组(60只),采用开腹直视肝脏穿刺法建立泡状棘球蚴动物模型,假手术组接种同体积的PBS,术后第3 d、7 d、14 d、28 d、42 d、56 d、70 d、84 d、98 d、112 d随机取6只沙鼠,取临近病变的边缘区组织及正常肝组织。采用HE观察病理改变;qRT-PCR、原位杂交、免疫组化检测HIF-1α、VEGFA表达,CD34标记微血管进行MVD计数。结果 HE染色根据病理特点将病程分为早期(14 d内)、中期(14 d~56 d)、晚期(56 d后)。模型组泡状棘球蚴组织随感染时间不同,其HIF-1αmRNA随感染时间呈动态改变,术后第14 d其表达量高于空白组、低于假手术组(F=82.732,P< 0.001);术后第42 d、112 d其相对表达量均高于空白组与假手术组(χ²=11.536,χ²=15.189,P< 0.01);模型组术后第14 d VEGFA mRNA相对表达量低于空白组及假手术组(χ²=15.174,P< 0.01);术后第42 d、112 d,VEGFA mRNA表达量均高于空白组与假手术组(χ²=15.158,χ²=15.158,P< 0.01)。模型组术后第14 d、42 d、112 d HIF-1α表达均高于空白组、假手术组(χ²=8.627,χ²=9.000,F=15.690,P< 0.01);模型组术后第14 d、42 d VEGFA表达高于空白组、假手术组(F=11.250, F=70.059,P< 0.001);模型组术后第14 d、42 d、112 d,其MVD-CD34均高于空白组、假手术组(χ²=12.517,P< 0.01,χ²=13.157,P< 0.01;χ²=13.220,P< 0.01)。结论 沙鼠感染肝泡状棘球蚴中期,病变边缘区HIF-1α、VEGFA表达均升高,同时伴有大量微血管生成,可能存在HIF-1α转录因子激活并上调VEGFA的表达,促进肝泡状棘球蚴组织边缘区的血管新生。;     

复杂性肝棘球蚴病外科治疗策略研究进展

肝棘球蚴病是一种由棘球属绦虫幼虫所致的人畜共患性疾病。我国主要致病绦虫类型为细粒棘球绦虫和多房棘球绦虫,分别引起细粒棘球蚴病和多房棘球蚴病。目前,棘球蚴病治疗已经取得重大进展,但对于部分就诊时已出现一种或多种并发症以及病灶侵及肝门、重要血管及胆管的复杂性棘球蚴病病例的治疗仍存在一定困难。本文根据近年来文献报道,结合临床经验,对复杂性肝棘球蚴病的外科治疗策略进行综述。［关键词］     

79例肝棘球蚴病患者影像学分析

目的 回顾性分析79例肝棘球蚴病患者的影像学表现,为该病诊断与鉴别诊断提供参考。方法 收集2014-2017年在青海省人民医院行影像学检查并经病理检查证实的79例肝棘球蚴病患者病历资料,对其影像学表现进行回顾性分析。结果 79例肝棘球蚴病患者中,细粒棘球蚴病57例,多房棘球蚴病22例;细粒棘球蚴病患者中,单囊型21例,多子囊型16例,内囊塌陷型9例,实变型4例,钙化型7例。79例患者中,62例为常见影像学征象。单囊型细粒棘球蚴病表现为肝内囊性水样病灶,囊壁薄厚均匀、无强化;多子囊型表现为“囊中囊”、“玫瑰花瓣”、“轮辐征”等;当内囊塌陷分离时表现为“飘带征”、“双环征”等征象;囊壁钙化时呈弧线状、蛋壳状,囊内容物呈现絮状或者整个病灶钙化。多房棘球蚴病表现为肝内实性肿块,密度及信号不均匀,边缘不规则;病灶强化不明显,病灶内散在或者群簇状分布的“小囊泡”,常伴有钙化,整个病灶呈“地图样”外观。另外17例患者表现为复杂少见的影像征象;其中6例细粒棘球蚴病囊内含脂肪,影像表现为囊内单发或多发脂肪密度结节灶,CT值为-28～-84 HU;4例病灶破入胆管,邻近胆管密度增高,胆管壁增厚,周围胆管扩张;4例合并原发性肝癌,影像表现为肝内细粒或多房棘球蚴病合并实性强化肿块,增强呈“快进快出”表现;3例合并感染,影像学表现为囊壁明显增厚且强化明显,其中2例囊内见气体影,1例囊肿合并感染并侵及腹壁。结论 肝棘球蚴病影像学表现复杂多样,在临床工作中需认真分析,做好鉴别诊断。     

人源细粒棘球蚴喙钩形态学参数分析

目的 测定本地区人源细粒棘球蚴喙钩的形态学参数,为棘球蚴的分类及流行病学研究提供依据。方法 以外科手术的方法从患者肝脏分离肝包虫,取囊液等内容物离心沉淀制成涂片标本,光学显微镜下观察,根据形态确定棘球蚴原头蚴的喙钩,应用计算机图像分析系统采集喙钩总长、总宽、柄长、刀长4个形态学参数数据,用统计学软件进行统计分析。结果 棘球蚴的原头节上具有上下两层喙钩,上层为大钩,下层为小钩。分别采集到游离状态的大钩56个,小钩49个,测量并获得了两种喙钩的4个形态学参数:大钩的总长、总宽、柄长、刀长的平均值分别为12.38 μm、4.58 μm、5.51 μm和6.51 μm;小钩的总长、总宽、柄长、刀长的平均值分别为10.41 μm、3.76 μm、5.04 μm和4.72 μm。大钩和小钩的各项形态学参数比较结果具有统计学意义(P<0.05)。结论 本研究应用计算机图像分析系统获得本地区人源细粒棘球蚴喙钩的形态学参数,为棘球蚴的分类鉴定及流行病学研究提供了可靠的形态学资料。     

肝细粒和多房棘球蚴混合感染3例

目的探讨肝细粒和多房棘球蚴混合感染患者诊断及手术治疗,为该类患者临床诊治提供经验。方法回顾性分析2017-2018年青海省人民医院诊断为肝细粒和多房棘球蚴混合感染患者的临床资料。结果共确诊3例肝细粒和多房棘球蚴混合感染患者。其中1例经术前CT检查确诊为细粒和多房棘球蚴混合感染,并在术中得到证实;另2例经术前彩超及影像学检查诊断为细粒棘球蚴病,但根据术中病灶形态和术后病理学确诊为细粒和多房棘球蚴混合感染。2例患者行根治性手术治疗,1例探查后仅行肝细粒棘球蚴内囊摘除、外囊次全切除术。结论肝细粒和多房棘球蚴混合感染患者术前易漏诊、误诊,影像学检查联合术后组织病理学检查结果才能最终确诊。肝细粒和多房棘球蚴混合感染患者手术相对复杂、困难,针对不同感染类型患者应采取个体化手术治疗方案。     

细粒棘球蚴和多房泡球蚴在人体内蛋白质表达谱初步分析

目的 初步掌握流行于青海省细粒棘球绦虫幼虫棘球蚴(原头节、囊壁、囊液)和多房棘球绦虫幼虫泡球蚴在中间宿主人体内蛋白质表达情况。方法 利用SDS-PAGE和 Western-blot分析棘球蚴原头节、囊壁、囊液蛋白质和泡球蚴总蛋白表达谱。结果 细粒棘球蚴原头节的蛋白质浓集在分子量72 kDa处,囊壁的蛋白质浓集在72 kDa、26 kDa和17 kDa 处,囊液的蛋白浓集在72 kDa、43~55 kDa、26 kDa和17 kDa;泡球蚴总蛋白质浓集在分子量72 kDa、55~72 kDa和26 kDa处。结论 不同地区细粒棘球蚴在人体内蛋白的表达存在差异;不同亚种泡球蚴原头节蛋白的表达存在差异。     

微囊法棘球蚴继发感染小鼠动物模型的建立

目的 通过体外培养原头蚴发育成微囊,经腹腔注射建立稳定的细粒棘球蚴和多房棘球蚴继发感染小鼠动物模型。方法 无菌条件下采集羊源细粒棘球绦虫原头蚴和鼠源多房棘球绦虫原头蚴,经胃蛋白酶消化后检测虫体活力并计数,于37℃、5%CO₂条件下进行体外培养至发育成微囊,以每鼠50个微囊的剂量经腹腔注射的途径分别接种BALB/c小鼠。接种6个月后,通过腹部解剖大体观察和病理检测分析各组小鼠的感染情况及包虫囊的生长情况。结果 原头蚴在体外培养60d时发育成微囊,显微镜下观察Eg具有明显的透明角质层结构,而Em微囊角质层较薄。小鼠细粒棘球蚴和多房棘球蚴的感染率均为100%,Eg包虫囊为游离单囊,成囊率达70%,囊内无原头蚴;Em包虫囊为类似肿瘤的团块状组织,病灶内有生发囊及原头蚴。结论 采用微囊法可建立稳定的棘球蚴继发感染小鼠动物模型,为疫苗研制、药物筛选和疗效判定提供研究动物模型。     

甘氨鹅脱氧胆酸对体外细粒棘球蚴原头节活力及ROS影响

目的 研究甘氨鹅脱氧胆酸(glycochenodeoxycholic acid, GCDCA)对体外细粒棘球蚴原头节活力及活性氧(Reactive oxygen species,ROS)水平的影响。方法 将细粒棘球蚴原头节分为对照组及GCDCA处理组(500、1 500、2 500 μ mol/ L),应用体外培养技术,通过伊红染色法每天定时在光学显微镜下观察原头节的活力并记录存活和死亡数目。使用试剂盒检测不同浓度GCDCA处理 24 h后各组原头节体内caspase-3酶活性变化;DCFH- DA染色法荧光显微镜观察 GCDCA作用24 h后原头节内 ROS水平变化。结果 500、1 500、2 500 μ mol/ L GCDCA处理后均可导致原头节活力减弱,且最高浓度组在第7 d时原头节全部死亡。500、1 500、2 500 μ mol/ L GCDCA组作用原头节24 h后 caspase-3酶活性与对照组相比明显升高(F=555.162, P<0.05)。500、1 500、2 500 μ mol/ L GCDCA作用原头节24 h后原头节内 ROS水平与对照组相比明显升高(F=216.901, P<0.05)。结论 GCDCA 能抑制体外原头节生长,促使原头节死亡,可能与活化caspase-3及提高原头节内ROS 水平相关,具体机制有待进一步研究。     

阿苯达唑联合IFN-α抗小鼠细粒棘球蚴病的实验研究

目的 在BALB/c小鼠体内评价阿苯达唑(ABZ)联合干扰素(interferon, IFN)-α对于CE的治疗效果。方法 Balb/c小鼠原头蚴腹腔继发感染5个月后,小鼠被随机分配到4组:ABZ组、IFN-α组、ABZ+IFN-α组和未经处理对照组。不同处理组分别给药2个月。在治疗的第0、7、14、28、36、48、60 d从小鼠尾静脉采血检测血清中抗体水平变化。治疗结束后处死小鼠,检测相关指标评价治疗效果。结果 与对照组(P<0.01)或ABZ组(P<0.05)相比,ABZ+IFN-α组包囊的数量、大小及重量都显著减少。对不同处理组的包囊进行透射电镜观察(TEM)发现,ABZ+IFN-α组的包囊的超微结构发生了明显的改变。ELISA实验结果表明,ABZ+IFN-α组的血清与脾细胞分泌的白细胞介素(interleukin, IL)-10显著下降(P<0.01);血清中IgE、IgG抗体及其亚型浓度与对照组相比显著降低(P<0.01)。结论 本研究证实ABZ联合IFN-α可能成是一种有效的CE治疗方案。     

细粒棘球蚴特异性诊断抗原Eg-07279的制备及免疫原性研究

目的 筛选出包虫病特异性诊断抗原分子并验证其免疫原性。方法 对国家人类基因组南方研究中心公布的细粒棘球绦虫测序数据进行分析,利用生物信息学的方法筛选出六钩蚴中不表达且原头蚴中高表达的抗原分子Eg-07279,经重组克隆、表达后用亲和层析法纯化获得重组蛋白rEg-07279;重组蛋白免疫小鼠检测其特异性 IgG 水平并使用Western blot 验证其免疫原性。结果 筛选出的抗原分子Eg-07279经克隆、表达、纯化获得重组蛋白。利用该重组蛋白免疫小鼠,ELISA检测结果显示Eg-07279产生的特异性IgG水平(2.559 ± 0.125)明显高于空白组(0.319 0±0.01),差异有统计学意义(P<0.01);Western blot检测结果显示原头蚴继发感染组和重组蛋白免疫组血清均可识别该重组蛋白而空白对照组鼠血清不能识别。结论 获得细粒棘球蚴差异表达抗原Eg-07279并证实该重组蛋白具有较好的免疫原性,是较好的诊断抗原候选分子。     

Hepatic echinococcosis: clinical and therapeutic aspects

Echinococcosis or hydatid disease (HD) is a zoonosis caused by the larval stages of taeniid cestodes belonging to the genus Echinococcus. Hepatic echinococcosis is a life-threatening disease, mainly differentiated into alveolar and cystic forms, associated with Echinoccus multilocularis (E. multilocularis) and Echinococcus granulosus (E. granulosus) infection, respectively. Cystic echinococcosis (CE) has a worldwide distribution, while hepatic alveolar echinococcosis (AE) is endemic in the Northern hemisphere, including North America and several Asian and European countries, like France, Germany and Austria. E. granulosus young cysts are spherical, unilocular vesicles, consisting of an internal germinal layer and an outer acellular layer. Cyst expansion is associated with a host immune reaction and the subsequent development of a fibrous layer, called the pericyst; old cysts typically present internal septations and daughter cysts. E. multilocularis has a tumor-like, infiltrative behavior, which is responsible for tissue destruction and finally for liver failure. The liver is the main site of HD involvement, for both alveolar and cystic hydatidosis. HD is usually asymptomatic for a long period of time, because cyst growth is commonly slow; the most frequent symptoms are fatigue and abdominal pain. Patients may also present jaundice, hepatomegaly or anaphylaxis, due to cyst leakage or rupture. HD diagnosis is usually accomplished with the combined use of ultrasonography and immunodiagnosis; furthermore, the improvement of surgical techniques, the introduction of minimally invasive treatments [such as puncture, aspiration, injection, re-aspiration (PAIR)] and more effective drugs (such as benzoimidazoles) have deeply changed life expectancy and quality of life of patients with HD. The aim of this article is to provide an up-to-date review of biological, diagnostic, clinical and therapeutic aspects of hepatic echinococcosis.     

Past to present: echinococcosis in Turkey

Echinococcosis is a zoonotic infection caused by Echinococcus spp. and is one of the most important helminthic diseases worldwide. Two forms of echinococcosis occur in Turkey, i.e. Echinococcus granulosus and E. multilocularis. The life cycle of E. granulosus is predominantly in dogs and sheep, and most sheep farmers in Turkey keep a dog or two. Stray dogs are numerous and prevention or treatment of infection in these dogs is very difficult. Cystic echinococcosis (CE) occurs throughout Turkey whereas alveolar echinococcosis (AE) predominantly occurs in the eastern Anatolian region of the country. Both CE and AE are known to be endemic but few surveys have been performed. Most data on human CE and AE have been collected from hospital records. The first reference of echinococcosis, "Kyste hydatique multiloculaire", in Turkey dates as far back to 1872 by an Ottoman Physician, C.R. Katibian. The results of the first Turkish study on E. granulosus were published in 1928. According to Ministry of Health records, 21303 patients had operations to treat or confirm CE in the period 1987-1994 which corresponds to approximately 2663 patients per year. The estimated surgical case rate of CE is 0.87-6.6 per 100000 in Turkey. The prevalence of E. granulosus infection in dogs in Turkey is between 0.32 and 40% and varies widely with geographical location. The reported prevalence of CE in domestic animals in Turkey has ranged from 11.2 to 50.7% and has varied widely with geographical location. Although no detailed information has been published on AE in domestic and wild animals in Turkey, the main definitive and intermediate hosts of E. multilocularis are assumed to be rodents and red foxes, respectively. However, there has been only a single published report of E. multilocularis in a wild animal (fox) in the northwest in 1965 by Merdivenci. The first human case of AE in Turkey was reported by Mutlu in 1939, and total cumulative reported case number is 202 between 1980 and 1998.     

Molecular confirmation of human alveolar echinococcosis in Poland

Infections of humans with Echinococcus multilocularis, the causative agent of alveolar echinococcosis (AE), a zoonosis, have been described with increasing frequency in Poland since 1994. In the attempt to verify these reports, we analyzed specimens obtained from a representative group of Polish patients. Liver lesions in patients with AE that was diagnosed on the basis of results of histological and serological tests contained E. multilocularis DNA, as shown by the presence of specific microsatellite sequences and mitochondrial 12S rDNA. The same tests clearly distinguished between AE and cystic echinococcosis, which is caused by Echinococcus granulosus. These data are unequivocal proof that human infections with E. multilocularis occur in Poland.     

人肝细粒棘球蚴病致病的研究进展

人肝细粒棘球蚴病是棘球蚴绦虫引起的一种人兽共患寄生虫病,也称肝包虫病,细粒棘球蚴的致病机制主要是对器官挤压,其次是细粒棘球蚴分泌的毒素对人体的毒性反应.目前,肝包虫病主要通过手术进行根治治疗.随着微创手术及精准手术标准的引用,腹腔镜手术治疗肝包虫病得到了长足的发展,本文将从腹腔镜肝包虫病适应症的选择、腹腔镜手术治疗肝包...     

Serological prevalence of echinococcosis and risk factors for infection among children in rural communities of southern Ningxia, China

Objective  To assess the usefulness of serology as an indicator of Echinococcus transmission for developing preventive measures against echinococcosis in rural communities.
Methods  Cross-sectional survey in 2002 among 861 children aged 7 to 18 years in Xiji County, Ningxia, China. Before ultrasound abdominal examination, a questionnaire was used to identify socioeconomic, sanitary and hygiene risk factors for echinococcal infection; filter paper blood samples were collected from each child for specific antibody detection using EmP ( Echinococcus multilocularis ) and EgB ( Echinococcus granulosus ) antigens.
Results  Transmission of both E. multilocularis and E. granulosus occur in this area. Serological prevalence was far higher than disease prevalence in this age range. We found no gender-related differences of seroprevalence among the children, suggesting an equal chance of exposure to echinococcal egg-contaminated environments. The seroprevalence distribution was associated with changes in the ecology of wild hosts for E. multilocularis contamination, and with changes in socio-geographic features of the communities for E. granulosus contamination.
Conclusions  Serological data obtained for children in mass surveys of echinococcosis appear to be a comprehensive and useful tool to monitor changes of transmission dynamics in humans and provide 'warning signals' to decision makers for the instigation of specific control measures against the disease.     

泡球蚴持续感染对小鼠肝脏纤维化的影响

目的 评价泡球蚴持续感染对小鼠肝脏纤维化的影响,为研究泡型棘球蚴病肝纤维化进展及其治疗方法提供参考.方法 以泡球蚴感染长爪沙鼠血清(25、50、100 μL)和泡球蚴及其生发层细胞、原头节分别对肝星状HSC-T6和LX-2细胞进行体外刺激48 h,采用CCK-8法检测细胞增殖,应用酶联免疫吸附试验(enzyme-lin...     

Development of Em18-immunoblot and Em18-ELISA for specific diagnosis of alveolar echinococcosis

Extensive experience has documented that Em2(plus)-ELISA, Em10-ELISA and Em18-immunoblot and Em18-ELISA are reliable serologic methods for detection of alveolar echinococcosis (AE) caused by the metacestodes of Echinococcus multilocularis. Among these, tests based on detection of antibodies to the specific Em18 antigen, either immunoblot or ELISA, appears to be the most specific for AE. Between 90 and 97% of AE cases with characteristic hepatic lesions detectable by image analysis have been positive in Em18-serology. In contrast Antigen B (8 kDa)-immunoblot is the most sensitive for all forms of echinococcosis, although it can not differentiate AE from cystic echinococcosis (CE). Primary serologic screening for echinococcosis, especially for CE using hydatid cyst fluid of Echinococcus granulosus appears to be highly sensitive in endemic areas. Glycoproteins (GPs) purified from cyst fluid of Taenia solium are highly specific for diagnosis of T. solium neuorcysticercosis (NCC). Using currently available antigens it is not difficult to differentiate these three larval cestodiases serologically. We recommend that (1) primary screening of CE in endemic areas should be carried out using hydatid cyst fluid of E. granulosus prepared from cysts in either sheep, human or mouse for immunoblot and from sheep or mouse for ELISA, (2) both primary screening and confirmation of AE in endemic areas should be carried out using Em18-ELISA, Em18-immunoblot or Em2(plus)-ELISA. Serodiagnosis in areas where both AE and CE are endemic, such as in China, should be carried out as a combination of (1) and (2), and (3) serology of NCC should be carried out using GP-ELISA or GP-immunoblot. All samples showing antibody to Em18 are exclusively from echinococcosis cases. There have been no false positive test reactions with sera from other diseases. Strongest Em18 responders are all from patients with AE but some weaker responses may be found in sera of persons with advanced complex lesions of CE. These highly reliable serodiagnostic methods using native, recombinant and synthetic antigens are briefly summarized and experiences with these methods in Japan is reviewed. We believe that use of these specific antigens in screening and confirmation programs for AE in Japan will improve specificity and reduce the confusion, anxiety and expense in persons whose sera give false positive reactions with crude echinococcal antigens.     

肝包虫病与肝囊肿CT表现比较*

目的 探讨使用256排CT检查在鉴别肝包虫病与肝囊肿方面的临床价值。方法 2017年4月～2020年11月我院诊治的肝囊肿患者77例和肝包虫病患者34例接受256排CT增强扫描检查,采用ELISA法检测血清抗囊液抗原抗体(EgCF)、抗头节抗原抗体(EgP)、抗囊液半纯化抗原抗体(EgB)和抗泡球蚴抗体(Em2)。结果 本组肝包虫病患者术前血液嗜酸性粒细胞计数为(0.3±0.1)×10⁹/L,显著高于肝囊肿患者【(0.1±0.1)×10⁹/L,P＜0.05】;肝包虫病患者血清抗EgCF抗体和抗EgB抗体阳性率分别为85.3%和61.8%,均显著高于肝囊肿患者【分别为31.2%和0.0%,P＜0.05】;增强CT扫描,肝包虫病患者肝内存在类圆形、圆形或分叶状囊性灶,囊壁钙化;在入组的111例患者中,CT检查将3例(2.7%)肝包虫病误诊为肝囊肿,1例(1.0%)肝包虫病被误诊为肝转移癌,2例(1.8%)肝囊肿被误诊为肝包虫病。结论 CT检查能很好地显示肝包虫病囊性病灶的影像学特征,对鉴别肝包虫病与肝囊肿有帮助,为临床治疗提供可靠的影像学依据。     

1995-2005年青海省棘球蚴病流行病学调查分析

目的分析1995—2005年青海省人与动物棘球蚴病的流行病学调查结果。方法人群棘球蚴病感染情况以Bu-ELISA、EM18-ELISA和B超、X线进行检查和评价。动物棘球蚴病／棘球绦虫感染调查采用解剖学方法和寄生虫学方法。结果①女性人群的血清阳性率和患病率显著高于男性人群（14．41％和4．81％、10．22％和3．25％）；随年龄增长，人群血清阳性率和患病率升高（见表5）；以牧民、喇嘛的血清阳性率和患病率最高（17．73％和9．33％、18．56％和10．0％）。②青海省人与动物棘球蚴病／棘球绦虫的血清阳性率和感染／患病率以青南高原的果洛、玉树、黄南三州最高（见表1、7、8），祁连山地和河湟谷地的海南、海北两州次之，海东地区、西宁及柴达木盆地的海西州较低。③青海省是以囊型棘球蚴病／细粒棘球绦虫为主的囊型和泡型棘球蚴病的混合流行区，并在其南部发现存在石渠棘球绦虫的动物感染。结论青海省棘球绦虫的生活史循环链十分复杂，家养动物相互之间、野生动物相互之间、家养动物和野生动物之间均参与其中。