淡水湖泊微囊藻毒素的污染和毒理学研究

微囊藻毒素是淡水湖泊蓝藻产生的一类肽类毒素 ,它的产生受到藻类的遗传和环境因素共同影响。微囊藻毒以肝脏为靶器官 ,通过多种作用引起肝细胞坏死 ,抑制蛋白磷酸酶 1(PP1)和蛋白磷酸酶 2A(PP2A)的活性 ,具有致癌性 ,是肿瘤促进剂。微囊藻及其毒素的污染已成为一个受人关注的公共卫生问题。由于微囊藻毒素污染的广泛性、持续性以及它所具有的热稳定性和水溶性 ,可能存在潜在的食品安全问题。本文综述了微囊藻及其毒素的污染、微囊藻毒素的产生原因和毒理学研究现状 ,预防的基本原则 ,并就其在食品安全方面将要开展的工作提出了一些建议     

水中微囊藻毒素的定量酶联免疫法研究

目的建立高灵敏度的水中微囊藻毒素的酶联免疫吸附试验（ELISA）方法。方法使用微囊藻毒素检测试剂盒,利用抗原与抗体间免疫化学反应,进行定量。结果微囊藻毒素在0.1～2.0μg/L范围内呈线性,标准曲线相关系数-0.995～-0.999。向样品中分别添加0.3、0.7、1.5μg/L 3个水平的微囊藻毒素,平均回收率分别为77.8%、85.7%和90.0%,相对标准偏差6.5%。结论该方法灵敏度高,适合水中痕量微囊藻毒素检测,并可实现大批量样品的筛选。     

014 微囊藻毒素污染状况、检测及其毒效应

随着水体污染逐渐加重,出现富营养化而导致的蓝藻水华日趋普遍,微囊藻毒素污染已成为一个全球关注的环境问题.微囊藻毒素是淡水湖泊、水库蓝藻产生的一类具有生物活性的环状七肽物质,它的产生受到藻类的遗传和环境因素共同影响,它能抑制蛋白磷酸脂酶1(PP1)和2A(PP2A)的活性,是一种强烈的肝脏肿瘤促进剂,并具有心、肾及胃肠毒性.本文概述了近年来在微囊藻毒素的结构特性、污染状况、检测及其毒效应等方面的研究进展.     

微囊藻毒素污染状况、检测及其毒效应

随着水体污染逐渐加重，出现富营养化而导致的蓝藻水华日趋普遍，微囊藻毒素污染已成为一个全球关注的环境问题。微囊藻毒素是淡水湖泊、水库蓝藻产生的一类具有生物活性的环状七肽物质，它的产生受到藻类的遗传和环境因素共同影响，它能抑制蛋白磷酸脂酶1(PP1)和2A(PP2A)的活性，是一种强烈的肝脏肿瘤促进剂，并具有心、肾及胃肠毒性。本文概述了近年来在微囊藻毒素的结构特性、污染状况、检测及其毒效应等方面的研究进展。     

酶联免疫吸附法在肉制品快速检测中的应用

简单介绍了酶联免疫吸附法(ELISA),并且就其在肉制品分析中的应用进行了主要包括用于肉制品微生物、毒素、农药残留、肉类鉴别和重金属的检测。     

晨尿ELISA法检测男性淋病的结果分析

目的用酶联免疫吸附法(ELISA)检测晨尿诊断男性淋病的效果作出评价。方法应用酶联免疫吸附法(ELISA)法对疑似淋病的男性患者的晨尿进行检测,同时取尿道拭子标本进行酶联免疫吸附法(ELISA)检测及淋球菌培养,三者进行比较,判断晨尿酶联免疫吸附法(ELISA)检测诊断男性淋病的效果。结果110例确诊病例,培养法96例阳性(87.3%),拭子酶联免疫吸附法(ELISA)法108例(98.2%),晨尿酶联免疫吸附法(ELISA)106例(96.4%)阳性。结论酶联免疫吸附法(ELISA)检测晨尿与拭子酶联免疫吸附法(ELISA)具有较好的一致性,检出率高于培养法,而晨尿酶联免疫吸附法(ELISA)取材方便,可代替拭子标本诊断男性淋病。     

微囊藻毒素检测方法与福州市售水产品污染调查

目的报道微囊藻毒素(MCs)检测方法,调查福州市售水产品MCs污染情况。方法于2015年夏季采集福州市售8种水产品44份,用固相萃取-高效液相色谱法,检测水产品中微囊藻毒素MC-LR、MC-RR、MC-YR含量。结果检测的44份水产品样品中,MCs检出率为27.3%(12/44);在花蛤、贻贝、田螺、鲫鱼、鲢鱼、草鱼样品中均有检出,以贝类污染率较高;MCs污染检出率9月(41.7%)高于7月(5.0%)。结论福州市售水产品存在微囊藻毒素污染,应加强对养殖水源污染的防治。     

水体中蓝绿藻分子标志物--mcyD基因的检测

目的建立一种筛选自然水体中产微囊藻毒素藻株的方法.方法应用编码微囊藻毒素合成酶mcyD基因的特异引物结合藻类保守序列16 S rRNA对所培养的纯种蓝绿藻(铜绿微囊藻FACHB 469、铜绿微囊藻PCC7806、微囊藻属FACHB 569、浮游蓝丝藻属小颤藻FACHB247、泡沫节球藻FACHB377)和采集的水样进行聚合酶链反应,观察扩增结果,并以酶联免疫吸附方法检测其微囊藻毒素浓度.结果在产微囊藻毒素(铜绿微囊藻PCC7806、微囊藻属FACHB569)的藻株和水样均可以扩增出1条约870 bp的单一条带,而不产微囊藻毒素的藻株(铜绿微囊藻FACHB469、浮游蓝丝藻属小颤藻FACHB247、泡沫节球藻FACHB377)和水样未扩增出此条带.结论以mcyD基因为微囊藻毒素分子标志物的分子生物学方法鉴定自然水体中的产微囊藻毒素和非产微囊藻毒素蓝绿藻是可行和实用的.     

乙型脑炎血清学早期诊断方法的探讨

本文用IgM抗体捕捉酶联免疫吸附试验(MAC ELISA)、酶联免疫吸附试验检测IgG(ELISA-IgG)、反向被动血凝抑制试验(RPHI)和血凝抑制试验(HI)四种方法检测1986～1988年从我省各地收集的78例临床疑似乙脑患者双相或单相血清,其确诊率分别是:MAC ELISA和RPHI均为69.7％,ELISA-IgG为48.5％,HI为33.3％;几种试验检测血清抗体滴度的相关性分析表明,它们之间存在正相关。根据试验结果作者认为MAC ELISA和RPHI试验均具有敏感、特异、简便和快速之优点,是目前乙脑临床早期实验诊断较理想的方法。     

微囊藻毒素对鱼类免疫毒性的研究进展

微囊藻毒素(microcystins,MCs)是富营养化水体中最常见、毒性最强、研究最广泛的一类藻毒素,已经引起了广泛的关注。MCs除具有典型的肝毒性外,还具有免疫毒性。笔者主要从免疫器官的病理变化、免疫细胞、免疫相关酶以及基因等方面综述了MCs对鱼类免疫毒性的研究现状,进而系统梳理了MCs对鱼类的免疫毒性作用机制,可为有效预警环境中MCs的潜在风险提供支持。     

Investigation of Microcystin Congener–Dependent Uptake into Primary Murine Neurons

Background

Contamination of natural waters by toxic cyanobacteria is a growing problem worldwide, resulting in serious water pollution and human health hazards. Microcystins (MCs) represent a group of > 80 cyclic heptapeptides, mediating cytotoxicity via specific protein phosphatase (PP) inhibition at equimolar concentrations (comparable toxicodynamics). Because of the structure and size of MCs, active uptake into cells occurs via organic anion-transporting polypeptides (OATP/Oatp), as confirmed for liver-specific human OATP1B1 and OATP1B3, mouse Oatp1b2 (mOatp1b2), skate Oatp1d1, and the more widely distributed OATP1A2 expressed, for example, at the blood–brain barrier. Tissue-specific and cell-type–specific expression of OATP/Oatp transporters and specific transport of MC congeners (toxicokinetics) therefore appear prerequisite for the reported toxic effects in humans and other species upon MC exposure. Beyond hepatotoxicity induced by the MC-LR congener, the effects of other MC congeners, especially neuronal uptake and toxicity, are unknown.

Objectives

In this study we examined the expression of mOatps and the uptake of congeners MC-LR, MC-LW, and MC-LF in primary murine neurons.

Methods

Intracellular MC accumulation was indicated indirectly via uptake inhibition experiments and directly confirmed by Western blot analysis and a PP inhibition assay. Neuronal mOatp expression was verified at the mRNA and protein level.

Results

MCs can cross neuronal cell membranes, with a subsequent decrease of PP activity. Of 15 mOatps, 12 were expressed at the mRNA level, but we found detectable protein levels for only two: mOatp1a5 (Slco1a5) and the known MC-LR transporter mOatp1b2 (Slco1b2).

Conclusions

These data suggest mOatp-mediated uptake of MC congeners into neurons, thus corroborating earlier assumptions of the neurotoxic potential of MCs.     

Necessity of Screening Water Chestnuts for Microcystins After Cyanobacterial Blooms Break Out

Water chestnut is one of the most popular vegetables in Asian countries that grows in shallow water. Eighteen water chestnut samples were collected from Lake Tai and six samples were bought at markets in Wuxi, China, in October 2007. Extraction solution of water chestnut was cleaned up with a solid phase extraction column and immunoaffinity chromatography cartridges, then the microcystin (MC) level was detected by indirect competitive enzyme-linked immunosorbent assay (ELISA) and liquid chromatography-mass spectrometry (LC-MS). The results of ELISA showed that there were six samples collected from Lake Tai which contained MCs; the highest level of total MCs was 7.02 ng/g. The results of LC-MS confirmed that MC-LR and MC-RR were present in five samples. The highest level of MC-LR was 1.02 ng/g and that of MC-RR was 4.44 ng/g. Heavy cyanobacterial blooms had occurred, and MCs were detected in water at the points in Lake Tai where MCs occurred in water chestnuts collected in 2007. MCs were not detected in the six samples bought at Wuxi markets. The results suggest that MCs can accumulate in water chestnuts, which is a potential hazard for human health.     

Short-term effects of air temperature on blood pressure and pulse pressure in potentially susceptible individuals

Background

Only few epidemiological studies have investigated the association between air temperature and blood pressure (BP) or pulse pressure (PP), with inconsistent findings. We examined whether short-term changes in air temperature were associated with changes in BP or PP in three different populations.

Methods

Between March 2007 and December 2008, 371 systolic and diastolic BP measurements were collected in 30 individuals with type-2 diabetes mellitus (T2D), 30 persons with impaired glucose tolerance and 42 healthy individuals without a metabolic disorder from Augsburg, Germany. Hourly means of ambient meteorological data were obtained from a fixed measurement station. Personal temperature measurements were conducted using data loggers. Temperature effects were evaluated using additive mixed models adjusting for time trend and relative humidity.

Results

Decreases in air temperature were associated with an increase in systolic BP, diastolic BP and PP in individuals with T2D. For example, a 1 °C decrease in ambient temperature was associated with an immediate increase in systolic BP of 1.0 mmHg (95%-confidence interval: [0.5;1.4] mmHg). Effects of personally measured air temperature were similar. Temperature effects were modified by age, body mass index, gender, antihypertensive medication and whereabouts, such as being indoors.

Conclusions

We observed associations between decreases in air temperature and increases in BP as well as PP in persons with T2D indicating that these people might be potentially more susceptible to changes in air temperature. Our findings may provide a hypothesis for a mechanism between air temperature decreases and short-term increases of cardiovascular events.     

Influences of long-term antibiotic administration on Peyer's patch lymphocytes and mucosal immunoglobulin A levels in a mouse model

BACKGROUND: Long-term antibiotic administration is sometimes necessary to control bacterial infections during the perioperative period. However, antibiotic administration may alter gut bacterial flora, possibly impairing gut mucosal immunity. We hypothesized that 1 week of subcutaneous (SC) antibiotic injections would affect Peyer's patch (PP) lymphocyte numbers and phenotypes, as well as mucosal immunoglobulin A (IgA) levels. METHODS: Sixty-one male Institute of Cancer Research mice were randomized to CMZ (cefmetazole 100 mg/kg, administered SC twice a day), IPM (imipenem/cilastatin 50 mg/kg x 2), and control (saline 0.1 mL x 2) groups. After 7 days of treatment, the mice were killed and their small intestines removed. Bacterial numbers in the small intestine were determined using sheep blood agar plates under aerobic conditions (n = 21). PP lymphocytes were isolated to determine cell numbers and phenotypes (CD4, CD8, alphabetaTCR, gammadeltaTCR, B220; n = 40). IgA levels in the small intestinal and bronchoalveolar washings were also measured with ELISA. RESULTS: Antibiotic administration decreased both bacterial number and the PP cell yield compared with the control group. There were no significant differences in either phenotype percentages or IgA levels at any mucosal sites among the 3 groups. CONCLUSIONS: Long-term antibiotic treatment reduces PP cell numbers while decreasing bacterial numbers in the small intestine. It may be important to recognize changes in gut mucosal immunity during long-term antibiotic administration.     

Extracellular signal-regulated kinase 1/2 and protein phosphatase 2A are involved in the antiproliferative activity of conjugated linoleic acid in MCF-7 cells

Conjugated linoleic acid (CLA) has protective properties in breast cancer. Here, we studied the mechanisms underlying the effects of CLA on MCF-7 breast cancer cell proliferation, especially in correlation with the involvement of the extracellular signal-regulated kinase 1/2 (ERK1/2) pathway and protein phosphatase 2A (PP2A). CLA inhibits MCF-7 cell growth in a concentration- and time-dependent manner, without triggering apoptosis. In assessing expression levels of proteins that play obligatory roles in the ERK cascade, we evidenced that CLA down-regulated Raf-1 and decreased levels of phospho-ERK1/2, as well as c-myc expression. Increase in PP2A expression rates were additionally observed after CLA treatment of MCF-7 cells. The above effects, as well as CLA-induced inhibition of cell growth, were reversed by okadaic acid, a specific inhibitor of PP2A. Thus, PP2A likely participates in deactivation of ERK1/2, and its up-regulation may represent a novel mechanism for CLA-induced inhibition of cell proliferation.     

Validation of an indirect immunoenzyme assay for the detection of antibodies against Trypanosoma evansi in horses in Argentina

An indirect enzyme-linked immunosorbent assay (ELISA) to detect antibodies against Trypanosoma evansi was evaluated using 90 different sera, obtained from naturally-infected horses. As negative controls, 218 sera from the T. evansi-free zone of Argentina, and 90 uninfected sera from the enzootic zone were used. The results of the ELISA were expressed in terms of percent positivity (PP) when compared with a positive primary reference serum, obtained from a horse experimentally-infected with T. evansi. The inter-assay coefficient of variation (CV), expressed as PP, was 44.7% for the negative control serum, 8.8% for the mildly positive reference serum, and 9.2% for the secondary positive control serum, while the intra-assay CV for each of the above sera was 6%, 2.8% and 5%, respectively. Positive and negative serological results were differentiated using a histogram of the distribution of the results obtained using sera from infected and uninfected animals from the enzootic zone (expressed in PP). A PP of 50 indicated a sensitivity of 95.5% for a confidence interval (CI) of 91.3% to 99.7%, and a specificity of 98% for a CI between 95% and 100%. Positive and negative predictive values were established for each rate of prevalence between 0.01% and 25%. The use of reference control sera in each assay enabled reproducible results to be obtained. The author recommends that this methodology be used whenever certification of the T. evansi status of horses is required, and particularly when animals are to be moved from an infected to a disease-free area.     

An observer blinded,randomized, placebo-controlled,phase I dose escalation trial to evaluate the safety and immunogenicity of an inactivated West Nile virus Vaccine,HydroVax-001, in healthy adults

BackgroundWest Nile virus (WNV) is the most common mosquito-borne infection in the United States. HydroVax-001 WNV is a hydrogen peroxide inactivated, whole virion (WNV-Kunjin strain) vaccine adjuvanted with aluminum hydroxide.MethodsWe performed a phase 1, randomized, placebo-controlled, double-blind (within dosing group), dose escalation clinical trial of the HydroVax-001 WNV vaccine administered via intramuscular injection. This trial evaluated 1 mcg and 4 mcg dosages of HydroVax-001 WNV vaccine given intramuscularly on day 1 and day 29 in healthy adults. The two dosing groups of HydroVax-001 were enrolled sequentially and each group consisted of 20 individuals who received HydroVax-001 and 5 who received placebo. Safety was assessed at all study days (days 1, 2, 4 and 15 post dose 1, and days 1, 2, 4, 15, 29, 57, 180 and 365 post dose 2), and reactogenicity was assessed for 14 days after administration of each dose. Immunogenicity was measured by WNV-specific plaque reduction neutralization tests (PRNT₅₀) in the presence or absence of added complement or by WNV-specific enzyme-linked immunosorbent assays (ELISA).ResultsHydroVax-001 was safe and well-tolerated as there were no serious adverse events or concerning safety signals. At the 1 mcg dose, HydroVax-001 was not immunogenic by PRNT₅₀ but elicited up to 41% seroconversion by WNV-specific ELISA in the per-protocol population (PP) after the second dose. At the 4 mcg dose, HydroVax-001 elicited neutralizing antibody responses in 31% of the PP following the second dose. In the presence of added complement, PRNT₅₀ seroconversion rates increased to 50%, and 75% seroconversion was observed by WNV-specific ELISA.ConclusionsThe HydroVax-001 WNV vaccine was found to be modestly immunogenic and well-tolerated at all dose levels.     

Diabetes mellitus and Helicobacter pylori infection

Alterations of glucose metabolism in diabetes have been suggested as promoting Helicobacter pylori colonization. We performed a cross-sectional sero-prevalence study of diabetic patients (insulin-dependent, or type 1, and non-insulin-dependent, or type 2, diabetes mellitus) with H. pylori and compared them with a control group. Consecutive diabetic outpatients aged 12 to 75 y and with disease duration of greater than 1 y were enrolled. Helicobacter pylori status was evaluated by using an enzyme-linked immunosorbent assay for anti-H. pylori immunoglobulin G. Demographic data were obtained from each individual, and socioeconomic class was assessed by occupation and education level. A total of 891 individuals participated (240 with type-2 diabetes, 145 with type-1 diabetes, and 506 control subjects). After controlling for age, there was no significant difference in the prevalence of H. pylori infection in any age group. In fact, the prevalence of H. pylori was numerically higher among children in the control group than among children with type-1 diabetes (25% versus 9%, respectively; P = 0.1). Previous associations of H. pylori and diabetes may have arisen from failure to consider socioeconomic status or age. Because childhood is the most common period for acquisition of H. pylori infection, the higher prevalence of infection among the normal children as opposed to those with type-1 diabetes confirms the lack of an association.     

Viral antigen production in cell cultures on microcarriers Bovine parainfluenza 3 virus and MDBK cells

Viral antigens can be obtained from infected mammalian cells cultivated on microcarriers. We have worked out parameters for the production of bovine parainfluenza 3 (PI-3) virus by Mandin-Darby Bovine Kidney (MDBK) cells cultivated on Cytodex 1 microcarriers (MCs) in spinners flasks and bioreactor using fetal bovine serum (FBS) supplemented Eagle minimal essential medium (Eagle-MEM). Medium renewal during the cell culture was shown to be crucial for optimal MCs loading (>90% MCs with confluent cell monolayers) and cell growth (2.5 x 10(6)cells/mL and a micro(x) (h(-1)) 0.05). Since cell cultures performed with lower amount of MCs (1g/L), showed good performances in terms of cell loading, we designed batch experiments with a lower concentration of MCs in view of optimizing the cell growth and virus production. Studies of cell growth with lower concentrations of MCs (0.85 g/L) showed that an increase in the initial cell seeding (from 7 to 40 cells/MC) led to a different kinetic of initial cell growth but to comparable final cell concentrations ((8-10)x10(5)cells/mL at 120 h) and cell loading (210-270 cells/MC). Upon infection with PI-3 virus, cultures showed a decrease in cell growth and MC loading directly related to the multiplicity of infection (moi) used for virus infection. Infected cultures showed also a higher consumption of glucose and production of lactate. The PI-3 virus and PI-3 antigen production among the cultures was not significantly different and attained values ranging from, respectively, 7-9 log(10) TCID(50)/mL and 1.5-2.2 OD. The kinetics of PI-3 virus production showed a sharp increase during the first 24h and those of PI-3 antigen increased after 24h. The differential kinetics of PI-3 virus and PI-3 antigen can be explained by the virus sensitivity to temperature. In view of establishing a protocol of virus production and based on the previous experiments, MDBK cell cultures performed under medium perfusion in a bioreactor of 1.2L were infected and the PI-3 virus production in 12L attained 12 log(10) TCID(50). Other than establishing a protocol for PI-3 production in MDBK cell cultures on Cytodex 1, the experiments are proposed as a basis for approaching the development of a virus production protocol in mammalian cells cultivated on microcarriers in bioreactors.     

GDF-15、PLA2R及CysC在2型糖尿病肾病早期诊断中的价值

目的探讨生长分化因子-15(GDF-15)、M型磷脂酶A2受体(PLA2R)及血清膀胱抑素C(Cys C)在2型糖尿病肾病早期诊断中的价值。方法选取于我院2013年12月—2016年3月就诊的2型糖尿病患者120例,根据其24 h尿微量白蛋白(m Alb)水平分为正常白蛋白尿组(A组)、微量白蛋白尿组(B组)及大量白蛋白尿组(C组),每组患者均为40例。分别对三组患者GDF-15、PLA2R及Cys C水平进行检测,探讨临床2型糖尿病肾病早期诊断中的应用价值。结果 A组患者GDF-15、PLA2R及Cys C指标水平依次低于B组、C组,存在显著差异(P0.05);三组患者GDF-15、PLA2R和Cys C指标水平组间对比差异显著(P0.05);经ROC曲线分析,临床早期诊断2型糖尿病肾病中GDF-15、PLA2R和Cys C指标均存在应用价值。结论 GDF-15、PLA2R及Cysc指标可作为临床早期诊断2型糖尿病肾病的辅助指标,具有一定临床推广价值。