原位杂交检测微小病毒B19方法的建立及临床应用

目的 建立原位杂交（ＩＳＨ）方法检测人微小病毒Ｂ１９,并确定其在先天性心脏病９ＣＨＤ）心脏组织细胞的定位分布。方法 以Ｂ１９特异的衣壳蛋白ＶＰＩ ＤＮＡ内１１１２ｂｐ片段为模板,采用随机引物标记探针法,建立了ＩＳＨ检测Ｂ１９ ＤＮＡ浓度于０．１ｂｇ／μ１以上呈阳性。在６６例先天性心脏病心脏组织中,检测至Ｂ１９ ＤＮＡ阳性７例,并发现Ｂ１９ ＤＮＡ阳性信号主要定位于心肌细胞核内,３８例对照组健康心肌     

人细小病毒B19检测方法研究进展

人类微小病毒B19（HPV B19）是细小病毒属中唯一对人类有致病性的病原体。感染后可引起传染性红斑、关节病、再生障碍性贫血危象、胎儿水肿及流产，对某些免疫力低下的病人，B19病毒感染甚至是致命的。B19病毒一般通过呼吸道传播，也可经输血和血液制品传播。建立简单实用的检测方法对B19病毒相关疾病的诊断和治疗非常重要。本文拟对B19病毒检测方法作一综述。     

人细小病毒B19检测方法研究进展

人类微小病毒B19(HPVB19)是细小病毒属中唯一对人类有致病性的病原体。感染后可引起传染性红斑、关节病、再生障碍性贫血危象、胎儿水肿及流产,对某些免疫力低下的病人,B19病毒感染甚至是致命的。B19病毒一般通过呼吸道传播,也可经输血和血液制品传播。建立简单实用的检测方法对B19病毒相关疾病的诊断和治疗非常重要。本文拟对B19病毒检测方法作一综述。     

抗人细小病毒B19IgG的检测

用抗人细小病毒Ｂ１９的ＭｃＡｂ建立ＥＬＩＳＡ－抗γ捕捉法，对正常新生儿（３５例）脐血标本及１月龄～１３岁小儿（１３９例）血清标本计１７４份，进行了Ｂ１９病毒特异性ＩｇＧ检测，结果阳性率分别为２２．９％（８／３５）和５．８％（８／１３９）．总阳性率为９．０％，并且大多数阳性标本的特异性ＩｇＧ抗体水平较低，最高滴度约为１：４００。结果显示：本文建立的检测方法有高度特异性；我国小儿中有Ｂ１９病毒自然感染，并有较高数量的易感人群，应进一步深入研究Ｂ１９病毒对我国小儿的危害及其防护措施。     

类风湿关节炎患者血清IgG-IgM型类风湿因子免疫复合物检测的临床意义

目的 探讨血清IgM型类风湿因子与变性IgG抗原形成的复合物定量检测对类风湿关节炎诊断的临床意义.方法 收集类风湿关节炎(RA)患者36例、非RA患者41例和体检健康者40名作为研究对象.用包被有鼠抗人μ链抗体的ELISA孔板及HRP标记的兔抗人IgG对血清中IgM型类风湿因子(RF)与变性IgG抗原形成的免疫复合物水平进行检测,同时用胶乳凝集法检测RF,并对二者结果进行相关性分析;用ELISA试剂盒进行抗环瓜氨酸肽抗体(抗CCP抗体)的定性检测,并比较其与IgG-IgM型RF免疫复合物对于RA诊断的敏感性和特异性.结果 待检血清最佳稀释倍数为100倍.IgG-IgM型RF免疫复合物对于RA的敏感性为72.2％,特异性为95.3％.IgG-IgM型RF免疫复合物OD值与RF阳性程度相关系数为0.687(P ＜0.01).结论 血清IgG-IgM型RF免疫复合物水平对RA具有较高的敏感性和特异性,可作为RA诊断的参考指标.     

吉林省供血者人类细小病毒B19 IgG抗体的调查

目的 了解吉林省供血者人类细小病毒感染的流行病学情况,为评估我国B19病毒的感染状况提供基础数据.方法 用间接ELISA方法检测血清中的抗B19 IgG抗体.结果 在184份血清中,抗B19 IgG抗体总检出率为55.43%.女性抗体阳性率高于男性,差异有统计学意义（P＜0.05）,年龄段在35～45岁之间的献血人员抗体阳性率最高.结论 本研究数据提示吉林省地区献血人员B19病毒感染率较高,有必要进行进一步B19 DNA的调查研究,为输血安全和血液制品安全提供保障.     

类风湿性关节炎患者血清中Ⅱ型胶原抗体的检测及临床意义

<正> 自身免疫是类风湿性关节炎(RA)发病中的重要因素,患者血清中存有抗自身IgG的抗体——类风湿因子。1977年,David和Threntham首次发现了50％的患者血清中存有抗Ⅱ型胶原的抗体,此种抗体与类风湿的发病和反复出现的症状有明显联系,我们应用ELISA检测,并对实验条件做了优选,取得满意结果。     

可溶性和膜型B7-H3在类风湿关节炎患者外周血中的表达及临床意义

本实验通过检测类风湿关节炎(RA)患者外周血中可溶性B7-H3(sB7-H3)和膜型B7-H3(mB7-H3)的表达及异构体的分布,探讨该分子的异常表达在RA发病中的临床意义。通过收集RA早期患者和健康对照外周血,ELISA方法检测血清中sB7-H3的表达,统计学分析sB7-H3的表达与临床的相关性;同时分离RA患者和健康对照外周血单个核细胞(periph-eral blood mononuclear cells,PBMC),通过流式细胞术检测CD14+的单核细胞膜型B7-H3的变化;通过PCR检测外周血mRNA水平B7-H3两种异构体的表达。结果显示RA患者血清中的sB7-H3分子明显低于健康对照组,RA活动期患者sB7-H3明显低于缓解期患者,并与RA患者肿胀关节数呈负相关;RA患者外周血单核细胞上mB7-H3的表达明显高于健康对照者,RA患者PBMC在mRNA水平B7-H3明显高于健康对照者,主要表达形式为4IgB7-H3。本研究发现膜型和可溶性B7-H3在类风湿性关节炎外周血中异常表达,其可能参与了RA自身免疫性病理的调节。     

套式聚合酶链反应检测自然流产组织中人细小病毒B19

目的进一步了解我国胎儿人细小病毒Ｂ１９的感染状况及其与自然流产之间的关系。方法用所建立的套式聚合酶链反应（ＰＣＲ）技术对５０例新鲜的和６６例石腊包埋的自然流产组织及２５例同期健康孕妇人工流产组织进行人细小病毒Ｂ１９ＤＮＡ检测。结果显示病例组有３４例阳性，阳性率为２９．３％，而正常对照组有１例阳性，阳性率为４％。经统计学检验有显著性差异。结论可能人细小病毒Ｂ１９感染与自然流产有一定关联。人细小病毒Ｂ１９可能是致自然流产的重要因素之一     

Human Parvovirus B19 in Rheumatoid Arthritis

Viral arthritis occurs transiently in most cases, because the infection is self limiting. The arthropathy associated with human parvovirus B19, however, often lasts for more than 2 years and their clinical symptoms may resemble with those of rheumatoid arthritis. Data have been accumulating for the link of B19 infection with chronic polyarthropathy or rheumatoid arthritis (RA), and we discuss the possible mechanism for the role of B19 in the etiopathology of RA.     

类风湿性关节炎与自身反应性B细胞

类风湿性关节炎(Rheumatoid Arthritis,RA)是一类以人体肢端关节滑膜中炎症细胞浸润为主要表现的自身免疫性疾病,其病因至今尚未真正明确.从免疫学角度研究发现:除了免疫复合物(炎性沉淀物)在关节囊液中起着一定的作用外,其患者体内的自身抗体与RA存在着密切关联.尽管目前仍有较为明确的证据表明RA是以T细胞--特别是Th1介导为主:这是由于依赖于胶原诱导关节炎(CIA,Collagen-induced arthritis)小鼠模型研究所得的结论.然而,新近的实验研究和临床免疫干预的进展使人们重新认识到B细胞是如何受自身抗原的诱导、激活、增殖和分泌自身抗体导致了RA发病以及这些自身抗体所累及的患者临床病理变化的本质,无疑将类风湿性关节炎与自身反应性B细胞的关联性重新提到一新的重要地位.毫无疑问:患者体内自身反应性B细胞和其产生的自身抗体在RA的发病中起到关键作用.患者关节滑液中浸润着高表达IL-13受体的B细胞,它们所分泌的一系列细胞因子发挥着特殊作用.值得特别注意的是IL-13可以作用于B细胞(诱导B细胞高表达CD23,促进幼稚B细胞的增殖和分化直至这种自身反应性B细胞克隆的异常扩增),越发突显自身反应性B细胞协同自身反应性T细胞介导RA发病的重要性,同时人们将更加关注其独特的免疫病理学机制对RA的诊断、进展和疾病预后的重要意义.本文将着重综述和讨论类风湿性关节炎与自身反应性B细胞的关联性的最新进展,旨在重新评估自身反应性B细胞和其分泌的自身抗体,包括一些细胞因子特别是IL-13介导的免疫失衡在类风湿性关节炎中的重要作用.     

Human parvovirus B19 and autoimmune diseases. Review of the literature and pathophysiological hypotheses

A number of arguments support the role played by PVB19 in autoimmunity, in the broad sense of the term essentially derived from numerous clinical case reports and/or small series over the past 20–30 years in the medical literature. PVB19 can induce a very broad spectrum of autoantibody production, especially including: anti-soluble nuclear antigen antibodies, antiphospholipid antibodies anti-native DNA antibodies, antilymphocyte antibody, anticardiolipin antibodies, antinuclear antibodies and rheumatoid factor. Notably acute PVB19 infection can mimic or stimulate autoimmune systemic diseases as rheumatoid arthritis or systemic lupus erythematosus. However, at the present time, there is no formal scientific evidence demonstrating a direct role of PVB19 in autoimmunity, bearing in mind that there are also no formal arguments against it. Further large studies are needed to understand the eventual role of PVB19 in autoimmune diseases.     

Detection of IgE anti-parvovirus B19 and increased CD23+ B cells in parvovirus B19 infection: relation to Th2 cytokines

The immune profile of a parvovirus B19-infected patient (male, 8 years old) was studied on day 0 (initial presentation) and on days 14 and 210 post symptom presentation (psp). Before infection, the patient was skin test positive to various allergens, including ragweed and tree and grass pollens, and had a serum IgE level of 150 IU/mL. On day 0, the patient was diagnosed as parvovirus B19 infected, as judged by the presence of IgG anti-parvovirus Abs in serum (EIA) and presentation of "slap cheek" rash. The patient's serum IgE level increased from 150 IU/mL before infection to 256 IU/mL on day 0, was 233 IU/mL on day 14, and returned to preinfection levels on day 210. In contrast, there was little change in the levels of serum IgM, IgG, or IgA (nephelometry). IgE anti-parvovirus B19 protein (VP-N) was detected in serum (Western blot) on days 0, 14, and 210, despite the decrease in total IgE on day 210. Although there was no increase in total numbers of blood CD23+ B cells on day 0, by day 14 the numbers of these cells increased dramatically (93%), remaining high on day 210. In contrast, there were virtually no changes in total numbers of CD4+ and CD8+ T cells or CD16/56+ NK precursor cells on days 0-210. On day 0, when IgG and IgE anti-parvovirus were detected in serum, patient's peripheral blood mononuclear cells (PBMC) expressed mRNA for the Th2 cytokines IL-4 and IL-10, but not for the Th1 cytokines IFN-gamma or IL-2. However, by day 14 psp, PBMC expressed mRNA for the Th1 cytokines IFN-gamma and IL-2, as well as for IL-4 and IL-10. This is the first demonstration of the existence of IgE anti-parvovirus B19 Ab. The presence of IgE anti-parvovirus B19 Ab in serum on day 0 and its persistence in serum 7 months psp suggests that IgE anti-parvovirus may be useful in prognosis of parvovirus B19 infection. Our results reinforce the idea that IgE, in general, may play a major role in anti-viral immunity, perhaps in conjunction with CD23+ cells. The results further suggest that clearance of this infection is accompanied by a switch to Th1 cytokines.     

Detection of parvovirus B19 in the lower respiratory tract

BackgroundHuman parvovirus B19 infection generally displays a self-limiting course followed by viral clearance; although, in some cases, persistent infection may occur. Few cases of severe pulmonary disease following primary infection in both immunocompetent and immunocompromised patients were reported.ObjectivesTo investigate the prevalence and clinical impact of parvovirus B19 in the lower respiratory tract.Study designThe prevalence of parvovirus B19-DNA was evaluated by Real-Time PCR in 264 bronchoalveolar lavages (BAL) from 189 adult patients over a full-year period and related to demographic characteristics, underlying pathologies, immune status, admission to intensive care unit, mortality within 28 days, and discharge diagnosis.ResultsParvovirus B19-DNA was detected in 7/189 (3.7%) patients, without significant association to demographic characteristics, immune status, transplant versus non-transplant status, admission to intensive care unit, presence of haematological conditions. In two lung transplant recipients surveillance specimens were positive to B19. Four of the remaining five patients presented respiratory insufficiency. A significant association to mortality was found, as 3/7 (42.9%) positive patients died within 28 days. No patient presented serological evidence of recent or acute infection and viremia.ConclusionsParvovirus B19 may be detected at low frequency in BAL specimens from patients with different pathological backgrounds. This finding could be due to chronic infection with virus persistence in the lower respiratory tract, also in the absence of symptoms unequivocally attributable to B19. The high rate of mortality warrants the need for further studies to evaluate the opportunity to consider parvovirus B19 in the diagnostic work-up of lower respiratory tract infections.     

Parvovirus B19 infection in Chile: markers of infection and immunity in patients with clinical symptoms

Parvovirus B19 infection is associated with a wide variety of symptoms and signs, and given that some clinical features, such as anemia, arthropathy and rash may be attributable to other causes, laboratory diagnosis of B19 markers is necessary. The principal aims were to study the behavior of B19 infection-associated diseases in the Chilean population and to compare B19 markers for recent or active infection and for immunity status in patients with clinical symptoms suspicious of B19 infection and control individuals. Sera from a total of 267 patients with diverse clinical manifestations associated with B19 and from 69 healthy controls were tested for B19 DNA using PCR and for specific IgM and immunoglobulin G (IgG) by enzyme-linked immunosorbent assay (ELISA). Out of 267 patients examined, 89 had B19-associated disease markers: 43 had B19 DNA without IgM, 25 had IgM without B19 DNA, and 21 had both B19 DNA and IgM. Also 49 patients were positive only for IgG without B19 DNA or IgM. Out of the 69 healthy controls, only 2 had B19 DNA without IgM and 30 had IgG without B19 DNA and/or IgM. The distribution of the clinical diagnoses associated with recent B19 infection, tested by B19 DNA and/or IgM, included 38.5% with hematological illnesses, 23.4% with rheumatic diseases, 45.7% with infectious diseases, 33.3% with indications of prenatal infection, 32.3% with conditions that induce immunodeficiency, and 15.8% with other miscellaneous conditions. The use of both markers, DNA and IgM, allows a more adequate diagnosis of infection by this virus.     

Clinical significance of occult hepatitis B virus infection in chronic hepatitis C patients

Background/Aims

We investigated the frequency of occult hepatitis B virus (HBV) infection in anti-hepatitis C virus (HCV)-positive individuals and the effects of occult HBV infection on the severity of liver disease.

Methods

Seventy-one hepatitis B virus surface-antigen (HBsAg)-negative patients were divided according to their HBV serological status into groups A (anti-HBc positive, anti-HBs negative; n=18), B (anti-HBc positive, anti-HBs positive; n=34), and C (anti-HBc negative, anti-HBs positive/negative; n=19), and by anti-HCV positivity (anti-HCV positive; n=32 vs. anti-HCV negative; n=39). Liver biopsy samples were taken, and HBV DNA was quantified by real-time PCR.

Results

Intrahepatic HBV DNA was detected in 32.4% (23/71) of the entire cohort, and HBV DNA levels were invariably low in the different groups. Occult HBV infection was detected more frequently in the anti-HBc-positive patients. Intrahepatic HBV DNA was detected in 28.1% (9/32) of the anti-HCV-positive and 35.9% (14/39) of the anti-HCV-negative subjects. The HCV genotype did not affect the detection rate of intrahepatic HBV DNA. In anti-HCV-positive cases, occult HBV infection did not affect liver disease severity.

Conclusions

Low levels of intrahepatic HBV DNA were detected frequently in both HBsAg-negative and anti-HCV-positive cases. However, the frequency of occult HBV infection was not affected by the presence of hepatitis C, and occult HBV infection did not have a significant effect on the disease severity of hepatitis C.     

Tongue Diagnosis of Traditional Chinese Medicine for Rheumatoid Arthritis

Rheumatoid arthritis (RA) is a chronic, systemic autoimmune disease with unknown aetiology that causes the immune system to attack the joints (synoviums), leading to chronic inflammation. According to the traditional Chinese medicine (TCM), RA falls into the category of Impediment disease (“Bi” syndrome), that is, poor circulation of qi and blood (stasis). Tongue diagnosis is an important method of TCM to detect blood stasis. In this study, 74 RA patients, meeting the pre-set criteria, were recruited via rheumatology outpatient clinic and examined by experienced rheumatology physicians. Two images—one of the tongue and the other, sublingual vessels—of the same patient were taken by a Canon digital camera in a darkroom with uniform lighting conditions. Relevant features of the tongue were extracted by utilising image processing techniques. Every tongue was classified into corresponding patterns based on the features identified. The subjects included 62 females and 12 males with an average age of 49.86±13.81 years old, an average morbidity period of 4.56±3.92 years, an average rheumatoid factor (RF) of 225.3±373.8 IU/mL and an average erythrocyte sedimentation rate of (ESR) 40.9±31.9m/hr. According to our study, 86% of the patients with RA have tongues with sublingual vessels with a width of more than 2.7mm, a length of more than 3/5 from tongue tipto sublingual caruncle, or a count of sublingual vessels more than 2. Moreover, since RA index is highly correlated with blood stasis in TCM, a logistic regression is conducted to predict the probability of presence of RA using RF and ESR as explanatory variables. Also, the logistic regression analysis of RA with respect to the conventional tongue diagnosis criteria was performed. Based on the aforementioned studies, we concluded that tongue diagnosis is helpful in detecting blood stasis of RA.     

抗-CCP抗体、RF与CRP联合检测对类风湿性关节炎早期诊断研究

目的探讨抗环瓜氨酸多肽（抗-CCP）抗体、类风湿因子（RF）与C-反应蛋白（CRP）联合检测在类风湿性关节炎（RA）中的诊断价值。方法采用半定量酶联免疫吸附法和定量免疫散色比浊法分别检测两组人群血清中的抗一CCP抗体、RF与C一反应蛋白（CRP）浓度,并分析上述三指标与RA的相关性。结果抗-CCP抗体诊断RA的特异度为93．5％,灵敏度为72．6％,RF为77．3％和83．4％,CRP为63．6％和82．3％,三者诊断灵敏度和特异度分别高达92．5％和97．6Voo。结论抗-CCP抗体诊断RA的特异度最高,RF的灵敏度最高,而三者联合早期诊断RA效率最好,与单一诊断指标相比具有明显的统计学意义。     

Parvovirus B19 DNA detection in treatment-naïve HIV anemic patients in Lagos,Nigeria: a case control study

BackgroundParvovirus B19 (B19) has tropism for cells of the erythroid lineage, which may lead to transient inhibition of erythropoiesis. Several studies and case reports suggested that B19 infection may contribute significantly to severe chronic anemia in HIV infected persons.ObjectiveTo detect parvovirus B19 DNA in treatment-naïve HIV patients.MethodsThis was a case control retrospective study. One hundred nineteen anemic and 81 non-anemic treatment-naïve HIV infected patients participated in the study at the Lagos University Teaching Hospital, Lagos, Nigeria. Polymerase chain reaction was used to detect B19 DNA.ResultsOut of 200 patients analysed, 13(6.5%) had parvovirus B19 DNA. Eight HIV patients with anemia had B19 DNA while five non-anemic HIV patients had B19 DNA. This suggests that the presence of B19 DNA in the blood of HIV positive individuals may contribute to anemia because the majority (61.5%) who were positive for B19 DNA had anemia as compared to the non-anemic control group (38.5%).ConclusionThis study shows that the presence of B19 DNA in anemic HIV infected patients is not associated with chronic anaemia in HIV infection because no significant association exist.