A sensitive automated procedure for the determination of glucuronic acid.

A newly automated method for the detection of uronic acids, particularly glucuronic acid is described. Data showing the limitations of this method are also presented. The advantages of the method are: (1) greatly increased sensitivity compared to the carbazole methods and increased range; (2) lowered interference from other sugars, proteins and salts; (3) variability to allow small sample volumes and/or continuous flow; (4) a more stable color reagent solution.     

Differences in vancomycin MIC among MRSA isolates by agar dilution and E test method

In this study, the correlation between vancomycin minimum inhibitory concentration (MIC) obtained by the E test technique and the Clinical And Laboratory Standards Institute (CLSI) agar dilution method was evaluated. A total of 53 Methicillin Resistant Staphylococcus aureus (MRSA) strains were tested by both the methods in the present study. MICs of vancomycin obtained by the E test method were consistently higher (+0.5 to 2 log2 dilutions) than those obtained by the agar dilution method. Out of 53 MRSA isolates, 49 isolates showed higher MIC results by E test than by agar dilution method. Three isolates showed same MIC result by both methods. Since many studies have demonstrated increased clinical failure with MRSA isolates for which vancomycin MICs are increased (>1 µg/ml) but still within the susceptibility range (≤ 2 µg/ml), our findings suggest the requirement to re-look into the breakpoints for vancomycin for determining sensitivity of MRSA isolates. Guidelines should also specify the method to be used for determining the MIC.     

Bronchoprovocation methods: direct challenges

Inhalation challenges with direct-acting stimuli histamine and methacholine are widely used to measure airway responsiveness. Three widely used methods (2-min tidal breathing method, breath-activated dosimeter method, hand-held manual nebulizer) are described. Careful standardization is important so as to best differentiate normal from increased airway responsiveness and to permit comparison between methods. With current methods standardized as suggested by the ATS, a methacholine (or histamine) PC(20) > 16 mg/mL is considered normal. A PC(20) < 16 mg/mL is highly sensitive for current symptoms of asthma. Interpretation of methacholine or histamine inhalation test requires that symptoms be current (within a few days) and that FEV(1) be normal.     

FLOW LOADED CANINE CAROTID ARTERY: I. A Morphometric Study of Microfilament Bundles in Endothelial Cells

To estimate microfilament bundles in the endothelial cells of flow loaded arteries quantitatively, blood flow change was produced in the common carotid arteries of eleven female beagle dogs (1–2 years of age, 8–10 kg) employing the arteriovenous shunt method between the common carotid artery and external jugular vein. After 1 week, the amount of microfilament bundles was measured with a point counting method on transmission electron microscopical photographs. In the arteries loaded by highly elevated blood flow, the average thickness of microfilament bundles increased over 0.1 μm (non-operative control: 0.01–0.02 μ) and the average longitudinal cut area of a microfilament bundle grew over 0.7 μm² (non-operative control: 0.2 μm²). The number of bundles increased over 1.0 piece (non-operative control: 0.2 pieces) per unit length (μm) in transverse section and increased over 0.5 pieces (non-operative control: 0.2 pieces) in longitudinal section. The average thickness of the bundles significantly correlated to the blood flow increase ratio (the ratio of the blood flow rate of the final measurement to that before anastomosis). We consider that the microfilament bundles are the structure developed to combat the wall shear stress induced by the elevated blood flow.     

The methodological aspects of nasal and exhaled nitric oxide levels in adult Japanese asthmatics

Background: Because both allergic rhinitis and asthma are caused by eosinophilic airway inflammation, using the same method to measure the eosinophilic inflammation of both the upper and lower airway would be advantageous. The levels of nitric oxide in exhaled air (FeNO) and nasal air (nNO) are useful as noninvasive markers of eosinophilic airway inflammation. Although the off-line method of measuring these parameters is easier and more useful than the on-line method, studies using the off-line method are rare in Japan. Methods: In Study 1, we measured the levels of nNO and FeNO in 9 healthy controls and 9 subjects with allergic rhinitis, to validate the methodology for using the off-line method to measure nNO. In Study 2, we measured the nNO and FeNO levels of and performed spirometry on 69 stable asthmatics treated with inhaled corticosteroid. Results: In Study 1, nNO levels were significantly increased in patients with allergic rhinitis compared with healthy subjects (31.0 [20.8 to 41.2] versus 7.4 [0.0 to 14.8] ppb {median [95% confidence interval]}, p=0.018). The 69 patients with asthma that comprised the study population in Study 2 were classified as asthmatics with rhinitis (treatment-na?ve, n=14; treated with antiallergic drugs, n=11; treated with intranasal corticosteroid, n=19) and asthmatics without rhinitis (n=15). Although FeNO did not differ among groups, nNO was significantly increased in treatment-na?ve asthmatics with rhinitis compared with patients with asthma only (26.5 [17.1 to 35.9] versus 8.0 [-1.1 to 17.1] ppb, p=0.033). Conclusion: nNO levels measured by the off-line method are useful markers of allergic rhinitis.     

A titrographic method for determining the glycolysis flow rate with Plasmodium berghei-infected red blood cells

A procedure is described for the elucidation of the glycolytic flux rate of red blood cells infected with the malarial parasite Plasmodium berghei. It is based on the titration of the protons originating from the glycolytic lactate accumulation. Compared with traditional methods of biochemical measurements of glucose consumption or accumulation of lactate the proposed procedure shows the following advantages: continuously measurement is possible; constancy of the pH-value during the measurement; lower amounts of biological material are necessary (increased sensitivity) increased accuracy; The method can also be applied to studies on other cells and species with normal or increased glycolytic flux rate.     

A simple technique for increased sensitivity in immunoelectrophoresis by the use of antiserum against IgG

Precipitates in two-dimensional immunoelectrophoresis (2D-IE) plates can be amplified by the application of an antiserum against IgG after the second dimension run. Thereby, the sensitivity of the IE method is increased; faint precipitates and antigen-antibody reactions close to but below the level of forming recognizable precipitates are made visible. The advantage of the method was demonstrated for a multilinear Mycobacterium phlei system.     

A sensitive staining method for NORs

A sensitive staining method for nucleolar organizer regions (NORs) is described, using blue toning of AgNORs. NORs are loops of DNA which are transcribed into ribosomal RNA. NORs can be demonstrated by staining with silver nitrate, since NOR-associated proteins are argyrophilic, producing structures termed AgNORs. Normal blood lymphocytes were stained with both methods. The number and resolution of NORs increased 2–3 times by blue toning (30 mmol/l FeCl₃, 11 mmol/l potassium hexacyanoferrate(III), and 33 mmol/l oxalic acid) compared with silver staining. A significant difference in the number of NORs was noticed between silver-stained and blue-toned cells (P<0.001). The blue toning technique thus appears to be more sensitive in detecting NORs than the AgNOR method and may prove a useful alternative for applications in histopathology.     

Role of individual serum pentitol concentrations in the diagnosis of disseminated visceral candidiasis

A technique was developed for separating total serum pentitols into individual arabinitol, adonitol and xylitol and determining their relevance for the diagnosis of disseminated candidiasis. Pentitols were separated as trimethylsilyl derivatives on two 25 m long, bonded methyl silicone columns with a 0.32 mm i. d., and quantified by selected ion monitoring of their protonated molecular ions obtained in chemical ionization. The 173 samples studied were divided into culture-positive, culture-negative, and no-culture groups. Twelve percent of all samples were false positives by the total pentitols method due to increased adonitol and/or xylitol. The continued use of the total pentitols method is, nevertheless, recommended because of its convenience; however, samples with increased total pentitols (and normal creatinine) should be reanalyzed for individual pentitols. Increased arabinitol and normal creatinine are indicative of candidiasis even when blood cultures are negative.     

运动条件下健康人循环内皮祖细胞的数量和功能*****☆

背景：运动条件对内皮祖细胞数量和功能是否会产生的影响目前尚无公识。 目的：观察急性运动对健康成人循环内皮祖细胞数量和功能的影响。 方法：健康男性成人志愿者12例参加急性平板运动锻炼(9.3±2.1) min。用流式细胞仪测定运动前后CD34和KDR双标阳性循环内皮祖细胞水平,ac-LDL及lectin荧光标记方法评估体外培养内皮祖细数量,检测内皮祖细胞的黏附、迁移和增殖能力,并测定运动前后血浆和内皮祖细胞分泌一氧化氮、血管内皮生长因子和粒-巨噬细胞集落刺激因子水平变化。 结果与结论：流式细胞仪检测显示健康志愿者运动后循环内皮祖细胞水平较运动前增加(P < 0.05)。荧光标记法证实运动后ac-LDL及lectin抗体双阳性内皮祖细胞数量较运动前增加(P < 0.05),内皮祖细胞迁移和增殖能力明显增强(P < 0.05),但黏附能力无明显变化。急性运动能明显增加健康志愿者的血浆一氧化氮水平(P < 0.05),健康志愿者运动前后血浆一氧化氮水平和循环内皮祖细胞数量及功能的增加倍数呈明显的直线相关回归关系(P < 0.05)。结果证实,运动可明显增加健康成人循环内皮祖细胞的数量和功能,其机制与其诱导一氧化氮释放增多有关。     

Improved sensitivity of nucleic acid amplification for rapid diagnosis of tuberculous meningitis

Early diagnosis of tuberculous meningitis (TBM) is essential for a positive outcome; but present microbiological diagnostic techniques are insensitive, slow, or laborious. We evaluated the standard BDProbeTec ET strand displacement amplification method (the standard ProbeTec method) for the detection of Mycobacterium tuberculosis complex organisms in parallel with the ProbeTec method with a modified pretreatment procedure with 101 prospectively collected cerebrospinal fluid specimens from 94 patients with suspected TBM. By the modified method, the sample-washing step was omitted. A definitive diagnosis was attained by culture. Thirteen specimens from 12 patients were culture positive for M. tuberculosis complex organisms; three specimens (23%) were microscopy positive for acid-fast bacilli. Among the culture-positive specimens, the standard ProbeTec method was positive for 8 (61.5%) and the modified assay was positive for 10 (76.9%). The overall specificity by both procedures was 98.8% compared to the results of culture. After discrepancy analysis, conducted by reviewing the patients' previous laboratory data, the specificity increased to 100%. If the cutoff value for respiratory specimens was adjusted from the recommended value of 3,400 to 1,000, the sensitivity of the modified procedure increased to 84.7%, with unchanged specificity. Results were obtained in 3 to 4 h. The new pretreatment procedure with the ProbeTec assay described here provides a rapid, simple, and sensitive tool for the diagnosis of TBM.     

A single step immunoelectrophoresis method for the quantitation of complement C3c in biological fluids

An intermediate gel rocket immunoelectrophoresis (IRI) suitable for direct quantitation of complement split products with C3c specificity is described. The technique represents a simple, quick, and reproducible method for the assessment of complement activity in biological fluids in diseases. The advantages of the quantitative approach of the IRI method as compared to the conventional crossed immunoelectrophoresis (CIE) are the increased sensitivity, specificity, and precision of the method; also, the assay capacity is superior to CIE. The IRI-C3c assay is therefore highly suitable as a scientific tool, and may also be useful for routine laboratory investigations.     

Effect of Light on Anthocyanin Levels in Submerged, Harvested Cranberry Fruit

Anthocyanins are a group of plant antioxidantsknown for their therapeutic use. The effects of natural light,red light, and far-red light on individual as well as totalanthocyanin content in cranberry fruit (Vacciniummacrocarpon Ait) were examined in an experimental settingdesigned to mimic water-harvesting conditions. The reversed-phasehigh-performance liquid chromatography (HPLC) method was used toseparate and analyze the anthocyanins. In contrast to thecase of the control sample that was kept in the dark,natural light increased the total anthocyanin level by 75.3%and 87.2% after 24 and 48 hours of water immersion,respectively. Red light and far-red light increased the totalanthocyanin level by 41.5% and 34.7%, respectively. Theamount of each individual anthocyanin increased differently undernatural light, red light, and far-red light, suggesting thatexpressions of enzymes that catalyze the anthocyanin biosynthesisare regulated differently by environments.     

Sodium bicarbonate secretion indicated by ultrastructural cytochemical localization of HCO3 −, Cl−, and Na+ ions on rat bile duct brush cells

Brush cells are widely distributed in the digestive and respiratory apparatus, but their function is still unknown. Because brush cells (BC) are found in organs secreting NaHCO₃, it was hypothesized that these cells may secrete NaHCO₃. To test this possibility, rat common bile duct epithelia were examined by ultrastructural cytochemical methods for localizing HCO₃⁻, Cl⁻, and Na⁺ ions. All three ion precipitates were few in or on BCs of rats without stimulation. Lead carbonate precipitates, which localized HCO₃⁻ ions by the lead nitrate-osmium method, increased markedly on the surface of the microvilli (MV) of BCs after secretin or meal stimulation, but similar precipitates were few on the luminal surface of principal cells (PCs). Silver chloride precipitates, which indicate the presence of Cl⁻ ions by the silver-osmium method, increased in the apical cytoplasm and in MV of BCs after secretin or meal stimulation, but they were few in PCs. Sodium pyroantimonate precipitates, which localize Na⁺ ions by the potassium pyroantimonate-osmium method, increased on the surface of the MV, along the basolateral membrane, and in the apical cytoplasm of BCs after secretin or meal stimulation, but they were few in PCs. These results strongly suggest that BCs may be a significant source of NaHCO₃ secretion.     

脑外伤小鼠海马水通道蛋白-9表达的变化

目的探讨脑外伤小鼠海马水通道蛋白-9(aquaporin 9,AQP9)表达的变化。方法40只Balb/c小鼠随机分为假手术组(10只)和脑外伤组(30只)。脑外伤组依据脑外伤的不同时间点再分6 h、1 d、3 d三个小组,每组10只。免疫组化和Western blot检测各组小鼠海马AQP9蛋白的表达。RT-PCR方法检测各组小鼠海马AQP9 mRNA的表达变化。结果免疫组化和Western blot结果发现,脑外伤组小鼠海马AQP9蛋白表达量明显高于假手术组(P<0.05);RT-PCR结果也发现,脑外伤组小鼠海马AQP9 mRNA表达量明显高于假手术组(P<0.05)。结论脑外伤小鼠海马AQP9表达明显升高。     

Multiparameter Analysis of EEG in Old Wistar Rats after Bilateral Carotid Artery Ligation

We showed that 302 of 840 EEG parameters in old Wistar rats significantly change 3 h after bilateral carotid artery ligation. All animals (n=7) died 250±47 min after ligation. EEG power spectra had signs of suppression (increased δ-activity, decreased τ-activity, and increased low-frequency α-activity) or activation of CNS (decreased high-frequency α-activity and paroxysmal increase in β-activity). The coherence, synchrony, and entropy of potentials were reduced in the majority of brain regions. This method and scheme of experiments are suitable for the search and study of anti-stroke drugs.     

黄芪总皂苷对心衰大鼠心肌细胞凋亡和线粒体膜电位的影响

目的研究黄芪总皂苷(ASTs)对心衰大鼠心肌细胞凋亡和线粒体膜电位的影响和机制。方法将大鼠分成对照组(normal)、心衰组(model)、AST低、中、高剂量干预组(10、20、40 mg/kg灌胃),每组12只,检测大鼠心功能指标。取大鼠心肌组织,TUNEL法检测细胞凋亡;黄嘌呤氧化酶法检测SOD活性;二硫代二硝基甲基苯法检测GSH-PX活性;硫代巴比妥酸法检测MDA含量;JC-1法检测线粒体膜电位;Western blot检测c-caspase-3、c-caspase-9、p-AKT、p-p38MAPK蛋白和线粒体、胞质中cyt-c蛋白表达水平。结果与对照组比较,model组大鼠LVSP、+dp/dt_max和-dp/dt_max降低(P<0.05);细胞凋亡率和c-caspase-3、c-caspase-9蛋白水平升高(P<0.05);SOD、GSH-PX活性降低(P<0.05);MDA含量升高(P<0.05);线粒体膜电位降低(P<0.05);线粒体中cyt-c蛋白水平降低(P<0.05);胞质中cy...     

The pathology curriculum in US medical schools. 1986 survey by the Association of Pathology Chairmen

Responses to questionnaires on teaching pathology to medical students from 71.6% of US medical schools indicate the following: (1) average class size continues to decrease; (2) in very few schools is instruction entirely interdisciplinary; (3) use of small-group teaching has increased slightly since 1982; (4) average curriculum time remains 250 hours per year; (5) clinical pathology instruction is frequently integrated with systemic pathology (mean time allocation, 40 hours per year); (6) lecture and study of gross and microscopic specimens remain mainstays of instruction; (7) honors/pass/fail grading systems are used by 54.7% of respondents (none use class rank); (8) multiple-choice examinations are a major method of evaluation; (9) graduates seeking post-medical degree training in pathology range from 0% to 6% of each class (mean, 2.13%); and (10) problems most often encountered are that there are too many students per class, and that students are unprepared for pathology.     

脑室注射组胺对下丘脑室旁核促皮质素释放激素神经元的作用

目的:观察第三脑室注射组胺对下丘脑室旁核促皮质素释放激素(CRH)神经元活动的影响。方法:Fos癌蛋白免疫组化LSAB法结合双抗原标记法;半定量逆转录聚合酶链反应(RT-PCR)方法。结果:第三脑室注射组胺后,(1)下丘脑室旁核Fos阳性神经元数目明显增加(P＜0.05);(2)室旁核内的Fos阳性神经元中约有31.78%同时呈CRH阳性反应;(3)室旁核CRHmRNA含量明显升高,且有量效关系。结论:中枢组胺可以激活下丘脑室旁核的CRH神经元,并使CRH基因表达增加。     

影像增强剂对食管癌边缘靶区剂量的影响

目的:研究调强放射治疗中使用影像增强剂后靠近心脏及胸主动脉等血流丰富组织的食管癌边缘靶区的剂量是否会降低,是否会出现欠量照射的情况。方法:选取20例食管癌患者病例,在相同体位下进行平扫及增强CT扫描,在增强CT上完成靶区勾画及调强治疗计划设计。使用两种方法进行对比,方法（1）:在增强及平扫CT上确定PTV & 5 mm Aorta和PTV & 5 mm Heart两个边缘靶区,然后进行图像配准并将治疗计划移植到平扫CT上计算剂量,对比上述两个边缘靶区在增强及平扫CT中剂量的变化。方法（2）:因为增强和平扫CT是在不同时间点扫描的,配准时两幅图像之间会存在一定差异,为了避免体位偏差的影响,使用合成CT与增强CT对比。所谓合成CT,即在复制的增强CT中将心脏、胸主动脉等血流丰富组织的CT值人工修改为平扫CT的平均CT值,操作过程与方法（1）相同,仍然观察上述两个边缘靶区的剂量变化。实验中也对比了其它组织器官在平扫与增强CT中剂量的变化。结果:方法（1）和方法（2）均显示PTV & 5 mm Aorta和PTV & 5 mm Heart两个边缘靶区剂量有所增加,其中方法（1）中两个边缘靶区的剂量分别增加1.28%（P<0.01）和1.64%（P<0.01）,方法（2）中分别增加0.41%（P<0.01）和0.83%（P<0.01）;正常组织器官（心脏、肺、脊髓等）剂量也有所增加,但均在临床可接受范围以内。结论:使用影像增强剂后靠近心脏及胸主动脉等血流丰富组织的食管癌边缘靶区剂量有所增加,不会出现欠量照射情况。