Bile crystallography and viscosimetry in cholelithiasis

High prevalence of cholelithiasis necessitates development of an easy screening diagnostic method, and therefore structural analysis of a dried drop of biological fluid attracts special attention. A new method, clinical crystallography, has been used for analysis of bile from patients with cholelithiasis. A relationship between the crystal morphology and biological fluid viscosity has been revealed.     

A new,fast, and simple DNA extraction method for HLA and VNTR genotyping by PCR amplification

In the present study a new DNA extraction method is described. The new protocol, which uses caprylic acid for isolating DNA, is technically simple and very fast, as it enables us to obtain DNA from peripheral blood in only 10 minutes. Moreover, DNA preparations obtained with this procedure can be effectively used for HLA class II and variable number tandem repeat genotyping by polymerase chain reaction, so the new method is well suited for routine clinical use in any type of analysis requiring DNA typing for individual characterization. © 1996 Wiley-Liss, Inc.     

用液相芯片方法检测禽流感病毒H5N1亚型的研究

目的基于液相芯片（MASA）技术建立一种对H5N1亚型禽流感病毒进行快速检测的新方法。方法对GenBank上已有H5和N1核酸片段进行比对分析,设计简并引物和探针,将合成的探针与荧光编码微球进行偶联,建立检测方法。利用该方法检测H5N1亚型禽流感病毒和其他常见亚型（H1N1、H2N2、H3N2、H9N2）的标本。结果利用该方法能够特异性地检测出H5N1标本,检测信号具有较高荧光强度,对其他亚型标本的检测则为阴性结果。该方法对H5NI亚型RNA的最低检出量为10pg。结论利用液相芯片技术研制的H5N1亚型禽流感病毒检测方法能够用于禽流感病毒H5N1亚型的快速、灵敏、特异性检测及鉴定。     

Interference of bamifylline with the Zimmerman reaction for 17-ketosteroids.

γ-Carboxyglutamic acid, the amino acid responsible for the vitamin K dependent, Ca²⁺-binding structures of some of the blood coagulation proteins, has been identified in human urine. The amino acid was isolated and its identity was proved by comparing it with synthetic γ-carboxyglutamic acid by electrophoretic and Chromatographic methods and by mass spectrometry. The isolated compound was also converted to glutamic acid by heat decarboxylation, a reaction consistent with its anticipated structure.A method for the quantitation of free γ-carboxyglutamic acid in human urine was developed. The method consisted of an anion-exchange concentration step followed by automatic amino acid analysis using a pH 2.0 lithium citrate buffer. In three non-fasting adult males the urinary excretion of γ-carboxyglutamic acid ranged between 27 and 42 μmol/24 h and in six non-fasting adult females it ranged between 19 and 32 μmol/24 h. One fasting adult male excreted 36 μmol/24 h.     

Determination of quinidine in human serum in the presence of diuretics.

A new method for the determination of quinidine in human serum by fluorometry is described. The interference of triamterene encountered with other methods is greatly reduced, whereas a number of other diuretics do not interfere at all. The specificity is based on quenching of the fluorescence of cinchona alkaloids by chloride ions. The method has good reproducibility and linearity, and gives a recovery of 98%. A serum sample of 0.1 ml is sufficient for the assay. Comparison with two other methods showed that this procedure yields the total amount of cinchona alkaloids present in serum, which makes it extremely useful for routine clinical analysis.     

A rapid, sensitive method for accurate analysis of individual bile acids in biological fluids by high performance thin-layer chromatography and densitometry

1. A rapid new micromethod for quantitative analysis of individual bile acids in duodenal juice by high performance thin-layer chromatography (HPTLC) and densitometry is described and evaluated by comparison with standard TLC and spectrophotometry. 2. Advantages of HPTLC over TLC include more rapid separation, better resolution and more sensitive detection (5 - 10 fold), without the need for prior extraction. Densitometry provides simple, direct and rapid quantitation. 3. The method is accurate and reliable over a range of bile acid concentrations. In the 0.5 mM range, recovery was greater than 89%, and coefficients of variation for within-day analysis were 2 - 12% and for between-day analysis were 6 - 18% for the individual bile acids. Twenty analyses can be performed by one worker in a single day. 4. We conclude that the method offers several advantages over most currently described techniques, is suitable for routine use and is deserving of wider application.     

野生型与突变型PS1真核表达载体的构建及其在SH-SY5Y细胞中的表达

目的为了对我们发现的中国人家族性阿尔茨海默病早老素-1(presenilin1,PS1)基因新的点突变进行蛋白功能研究,分别构建野生型和突变型PS1(G289T)与绿色荧光蛋白(EGFP)共表达载体,并检测其在SHSY5Y细胞内的表达。方法利用含人全长PS1cDNA的pcDNA3·1(zeo ),采用定点突变技术,构建PS1(G289T)-pcDNA3·1(zeo )载体。采用基因重组技术构建野生型和突变型PS1与绿色荧光蛋白共表达载体。应用脂质体将携带野生型和突变型PS1的质粒转染至SH-SY5Y细胞,检测报告基因表达,RT-PCR检测PS1mRNA表达。结果经限制性内切酶酶切图谱分析及DNA测序证实融合蛋白表达载体构建成功;RT-PCR产物经测序显示突变型PS1mRNA在SH-SY5Y中有表达。结论成功构建了人野生型和突变型PS1与EGFP共表达载体并成功转染至SH-SY5Y细胞,为进一步的研究工作奠定了基础。     

贲门癌组织中annexin 1的表达及意义

目的：研究贲门癌组织中annexin1的表达情况，探讨其表达与贲门癌临床病理特征的联系。方法：应用免疫组织化学方法检测贲门癌度癌旁组织中annexin1的表达。结果：贲门癌和癌旁组织annexin1表达的阳性率分别为90％（45／50），15％（3／20），annexin1的表达与肿瘤分化程度、浸润深度、淋巴结转移程度厦有无远处转移有关。结论：annexin1在贲门癌组织中的表达率较正常贲门组织显著增高，可能是反映贲门癌的生物学特性，评估预后的较好指标。     

重组水蛭素克隆的表达及其产物的分离纯化

目的：在原核细胞中表达具有生物活性的重组水蛭素变异物１（ｒＨＶ１）并进行重组产物的分离纯化研究。方法：以质粒ｐＢＶ２２０为载体，在大肠杆菌ＤＨ５α中表达ｒＨＶ１，发色底物法测定其抗凝血酶生物活性；利用超滤、ＤＥＡＥ－ＳｅｐｈａｄｅｘＡ－５０以及凝血酶亲合层析进行重组产物的分离纯化。结果：在大肠杆菌中获得了ｒＨＶ１的活性表达，表达量约占总蛋白的１６．９％，活力达到２０～３０ＡＴＵ／ｍｌ培养液；初步的纯化研究得到了具抗凝生物活性的ｒＨＶ１纯品，ＳＤＳ聚丙烯酰胺凝胶电泳呈单一条带。结论：重组水蛭素变异物１的活性表达及其分离纯化研究填补了国内空白，为研制国产高效重组抗凝药物带来希望。     

Genetic polymorphisms in MDR1 and CYP3A5 and MDR1 haplotype in mainland Chinese Han, Uygur and Kazakh ethnic groups

BACKGROUND AND OBJECTIVE: The drug transporter MDR1 and the drug metabolizing enzyme CYP3A are the two major biological factors determining the pharmacokinetics of many drugs. The functional MDR1 single nucleotide polymorphisms (SNPs) and a prevalent CYP3A5 SNP show marked interethnic variation among Orientals, Caucasians and Africans. In this study, we investigated the distribution of MDR1 and CYP3A5 SNPs among mainland Chinese Han, Uygur and Kazakh ethnic groups. METHODS: Genotypes of the MDR1 C1236T, G2677T/A and C3435T, and CYP3A5*3, CYP3AP1*3 SNPs were determined in 434 unrelated healthy subjects (165 Chinese Han, 161 Chinese Uygur and 108 Chinese Kazakh) using polymerase chain reaction followed by restriction fragment length polymorphism analysis. RESULTS AND DISCUSSION: A significantly higher MDR1 3435T variant frequency was observed in Uygur (52.8%), than in Kazakh (39.8%) and Han (37.9%) Chinese (P < 0.01, Fisher's exact test). There was no significant difference in MDR1 1236T and 2677T/A variant frequencies between Han, Uygur and Kazakh. CYP3A5*3 (G) allele was observed at intermediate frequencies in Uygur (84.8%) and Kazakh (86.6%), relative to Han (72.7%) and values previously reported in Caucasians (91.7%). The CYP3AP1*3 (A) allele was strongly linked to CYP3A5*3 in Chinese Han, Uygur and Kazakh. CONCLUSION: Significant interethnic differences in MDR1 haplotype and CYP3A5 variant frequencies exist between mainland Chinese Han and Caucasians, and the intermediate frequencies observed in Chinese Uygur and Kazakh might be due to the genetic admixture of Eurasians and Orientals.     

Relative exchangeable copper: a new highly sensitive and highly specific biomarker for Wilson's disease diagnosis

Wilson disease (WD) is an autosomal recessive inherited disorder of copper metabolism. Failure to diagnose WD can be dramatic leading to irreversible damages. The molecular genetic analysis of ATP7B gene is the reference test for diagnosis but the number of reported mutations of the ATP7B gene is on the rise. The analysis is cumbersome and requires tedious work. Other clinical and biological tests are proposed but it is often difficult to interpret some patients' results. A rapid and reliable biological test for WD diagnosis is still needed. Analytical reliability of Exchangeable copper (CuEXC) determination procedure is examined by studying the repeatability, the short term stability and stability in frozen serum. Relative exchangeable copper (REC=CuEXC/total copper%) is proposed and evaluated as a new diagnostic test and compared to classic tests used for WD diagnosis. Sixteen new Wilson disease patients were diagnosed in our institution between January 2009 and May 2011. The different biological tests used for WD diagnosis yielded lower sensitivity and specificity compared to our new biomarker, the REC. We show that REC is an excellent discriminatory tool for the diagnosis of WD offering 100% sensitivity and 100% specificity.     

巨噬细胞抑制细胞因子-1蛋白在胃癌中的表达和临床意义

目的研究胃癌组织中巨噬细胞抑制细胞因子-1（MIC-1）蛋白的表达特点和意义。方法常规石蜡包埋行MIC-1免疫组织化学染色。结果MIC-1在Ⅲ／Ⅳ期、癌细胞侵润浆膜层和淋巴结转移的胃癌组织中的阳性表达明显高于Ⅰ／Ⅱ期、癌细胞未侵润至浆膜层和淋巴结未转移的胃癌组织,组间比较差异均有显著性（P〈0．01）;复发组与非复发组胃癌组织MIC-1的表达之间存在差异（P〈0．01）。结论MIC-1表达与胃癌的发生发展、生物学行为和预后可能有关,可作为重要的生物学标记物。     

Imaging analysis for multiple paramagnetic agents using OMRI and electrophoresis

Nitroxides have been widely used as a molecular probe for analysis of various diseases models. This article describes an analytical method for separation and semi-quantification of multiple paramagnetic contrast agents with simple procedure combining electrophoresis and Overhauser enhancement magnetic resonance imaging (OMRI) imaging. We used three nitroxides, 3-carbamoyl PROXYL, 3-carboxy PROXYL, and CAT-1, which have different ionic charges in the molecule. In addition, we showed that this method could apply for in vitro measurement using biological sample. The results showed the nitroxides were successfully separated with electrophoresis depending on their charge, and their separation was visualized with OMRI after electrophoresis. Vehicle media such as whole blood did not affect the electrophoresis results and OMRI enhancement factor. Thus, the method can be used to analyze the redox status of biological samples without preprocessing. This analytical method enables in vitro measurement of biological samples to determine the redox status of specific tissue layers using paramagnetic agents, which is helpful for detailed analysis of redox-related diseases.     

Anti-cytokines: promising tools for diagnosis and immunotherapy

Anti-Cytokines belong to a new family of biological response modifiers which interfere with the biological functions of Cytokines. An imbalance between Cytokines and natural anti-Cytokines may represent an important factor to explain the pathogenesis of some human diseases. Indeed, dysregulation of anti-cytokine serum levels was observed in cancers and auto-immune and infectious diseases. It was sometimes associated with disease activity and in some circumstances they may help to predict clinical response to treatment. Control of disease activity in septic shock and auto-immune diseases was demonstrated after administration of anti-Cytokines such as soluble TNF receptors and ILlra inhibitors. Therefore these agents are promising tools for diagnosis and immunotherapy.     

Rac 1在胃癌组织中的表达及其临床意义

目的 观察胃癌和正常胃粘膜组织中Ras相关的C3肉毒素底物1（ras-related C3 botulinum toxin substrate 1,Rac 1）蛋白的表达情况并探讨其临床意义。方法 应用链酶亲和素-生物素过氧化物酶复合物（StreptAvidin-Biotin Com-plex,SABC）免疫组化法检测60例福尔马林固定、石蜡包埋之胃癌及正常胃粘膜组织标本中Rac 1蛋白的表达情况并分析其与胃癌临床病理参数间的关系。结果 Rac 1蛋白在正常胃粘膜组织中呈弱阳性染色（11/60,18.33%）,但在胃癌组织中则呈强阳性染色（43/60,71.67%）,两者间统计学差异非常显著（P〈0.01）;且随胃癌组织分化程度的降低、浸润深度的增加、TNM分期的升高及伴有淋巴结转移的发生,Rac 1蛋白染色阳性率逐渐升高,统计学意义显著（P〈0.05或P〈0.01）。但Rac 1蛋白表达水平与胃癌患者的性别、年龄、癌变原发部位及癌肿大小无关,计统学意义不显著（P〉0.05）。结论 Rac 1蛋白的表达与胃癌浸润、转移密切相关并有可能作为反映其生物学行为的一种新型标志物。     

Analyzing biological rhythms in clinical trials

BackgroundThe human body exhibits a variety of biological rhythms. There are patterns that correspond, among others, to the daily wake / sleep cycle, a yearly seasonal cycle and, in women, the menstrual cycle. Sine/cosine functions are often used to model biological patterns for continuous data, but this model is not appropriate for analysis of biological rhythms in failure time data.MethodsWe consider a method appropriate for analysis of biological rhythms in clinical trials. We present a method to provide an estimate and confidence interval of the time when the minimum hazard is achieved. A motivating example from a clinical trial of adjuvant of pre-menopausal breast cancer patients provides an important illustration of the methodology in practice.ResultsAdapting the Cosinor method to the Weibull proportional hazards model is proposed as useful way of modeling the biological rhythm data. It presents a method to estimate the time that achieves the minimum hazard along with its associated confidence interval. The application of this technique to the breast cancer data revealed that the optimal day for pre-resection incisional or excisional biopsy of 28-day cycle (i.e. the day associated with the lowest recurrence rate) is day 8 with 95% CI 5–10. We found that older age, fewer positive nodes, smaller tumor size, and experimental treatment are important prognostic factors of longer relapse-free survival.ConclusionsThe analysis of biological/circadian rhythms is usually handled by Cosinor rhythmometry method. However, in FTD this is simply not possible. In this case, we propose to adapt the Cosinor method to the Weibull proportional hazard model. The advantage of the proposed method is its ability to model survival data. This method is not limited to breast cancer data, and may be applied to any biological rhythms linked to right censored data.     

SDF-1/CXCR4与血液系统恶性肿瘤的关系研究进展

基质细胞衍生因子（SDF—1）及其受体CXCR4相互作用转导特定信号，在许多生理和病理过程中都发挥了重要的效应，CXCR4在多种血液系统肿瘤中高表达，与疾病的预后、耐药、复发密切相关。用SDF-1抗体或CXCR4抗体能有效的抑制肿瘤细胞的生长，为治疗血液系统肿瘤开辟了新途径。本文就SDF—I／CXCR4在血液系统肿瘤中的表达，及其与预后、耐药、复发和治疗关系的研究进展作一综述，     

Influence of the paraoxonase‐1 Q192R genetic variant on clopidogrel responsiveness and recurrent cardiovascular events: a systematic review and meta‐analysis

Summary. Background: A poor biological response to clopidogrel is associated with an increased risk of major cardiovascular ischemic events (MACE). Paraoxonase 1 (PON1) enzyme activity is modulated by the PON1‐Q192R variant (rs662) and was recently suggested to be strongly involved in clopidogrel bioactivation, but the influence of the PON1‐Q192R variant on the risk of MACE in clopidogrel‐treated patients is controversial. Objectives: To determine whether the PON1‐Q192R variant influences clopidogrel biological responsiveness and the risk of MACE in patients treated with clopidogrel. Methods: Systematic review and meta‐analysis of studies of the association between the PON1‐Q192R polymorphism and the biological response to clopidogrel and/or the risk of MACE during clopidogrel administration. Results: Seventeen studies were included. In the 12 studies of the biological response to clopidogrel (n = 5302 patients), there was no significant difference between 192QQ and 192QR + 192RR subjects, whatever the laboratory method used (global mean standardized difference = 0.10 [?0.06; 0.25], P = 0.22). Eleven studies assessed the risk of MACE, four using a case–control design (n = 2739 patients) and seven a prospective design (n = 5353 patients). Overall, MACE occurred in 19% of patients in case–control studies and in 6% of patients in prospective cohort studies, with no significant difference between 192QQ and 192QR + 192RR patients (OR = 1.28 [0.97; 1.68], P = 0.08). Similar results were obtained when study design was taken into account. Heterogeneity was mainly driven by one publication. Conclusions: This meta‐analysis suggests that the PON1‐Q192R polymorphism has no major impact on the risk of MACE and does not alter the biological response to clopidogrel in clopidogrel‐treated patients.     

Modification of the chemiluminescent analysis method for evaluation of the whole blood phagocyte activity in sensibilized animals

A modified method of chemiluminescence analysis was developed in experiments with animals. Whole blood of guinea pigs was made use of as the biological sample. BaSO4 and another new substance, i.e. aerosol, were used in the phagocytes' stimulation of whole blood. Optimal concentrations of reaction components, the incubation period of whole blood samples and of specific allergens and the duration of chemiluminescence registration were defined. The discussed modified method of chemiluminescence analysis can be used to monitor the specific treatment schemes by immunotherapy and to choose adequate schemes for the mentioned therapy.