Use of silicone fluids in studies of cellular amino acids

In numerous cellular studies, cells labeled with radioisotopes have been separated from the labeling medium by an aqueous solution in order to determine the quantity of internalized labels; however, the aqueous wash tends to remove significant labeling from the cells. Therefore, in order to preserve all of the internalized labels, non-aqueous medium such as silicone fluids may be used. The termination of the labeling is achieved in the silicone method when, upon centrifugation, the cells separate from the medium and enter the silicone fluid to sediment to the tube bottom. This sedimentation of cells placed above a layer of silicone fluid exhibits a critical dependence on the centrifugal force, and gives rise to an uncertainty of only 2 s in determining the time of separation of cells from the medium using General Electric F-50 silicone fluid and a modified Beckman J2-21 centrifuge. It is therefore possible to determine the kinetics of incorporation of labeled amino acids into intracellular pools and proteins. In particular, since this silicone wash method determines the size of the total pool and the aqueous wash method determines the size of the acid-extractable pool, the simultaneous measurements of the size of both pools leads to the determination of the kinetics of labeling of the free amino acid pool. Among many possible applications and extensions of these methods, the studies of formation of intracellular pools and relations among different pools of transported molecules, such as water and amino acids, appear promising.     

一个可能引起歧义的概念——“条件致病菌”辨析

"条件致病菌"是国内医学微生物学教材中普遍应用的一个基本概念,但此概念在建立与应用中都存在明显缺陷,其使用可能对医学学生理解"人体共生微生物群落"与"机会性感染"等重要的医学基础理念造成误解与歧义。谨就此提出一些粗浅的认识与观点,希望通过同行间切磋,就此建立更完善的概念与更合理的表述。     

Determination of cefazedone in human plasma by high performance liquid chromatography–tandem mass spectrometry: Application to a pharmacokinetic study on Chinese volunteers

A rapid, sensitive and simple high performance liquid chromatography–tandem mass spectrometry (HPLC–MS/MS) method was developed for determination of cefazedone in human plasma using metronidazole as internal standard (IS). The chromatographic separation was achieved on an Ultimate XB-CN column (2.1 mm × 150 mm, 5 μm) with an isocratic mobile phase of acetonitrile and 20 mM ammonium acetate in 0.1% formic acid in water (15:85, v/v). Detection was performed using electrospray ionization in positive ion multiple reaction-monitoring mode (SRM), monitoring the transitions m/z 548.2 → 344.1 for cefazedone and m/z 172.2 → 128.1 for IS. Calibration curves were linear over a wide range of 0.20–401.12 μg/mL for cefazedone in plasma. The lower limit of quantification (LLOQ) was 0.20 μg/mL. The intra- and inter-day precisions were less than 7.2%. The average recovery of cefazedone was 90.8–91.0%. The validated method was successfully applied to the pharmacokinetic study of cefazedone in Chinese healthy volunteers following intravenous (IV) administration of 500, 1000 and 2000 mg cefazedone injection.     

鼎突多刺蚁群体结构和生活史的研究

本文报告了鼎突多刺蚁(Polyrhachis vicina Roger)的群体结构和生活史。通过研究表明,鼎突多刺蚁一年发生一代,以蚁后、雄蚁、工蚁、幼虫和卵越冬。卵和幼虫在冬天发育停滞(或极慢)。到了春天,随着气温的上升,卵和幼虫又恢复正常发育。据室内人工饲养观察,卵的发育历期为23.8±2.5天(平均温度26℃)幼虫为20.4±4.4天(26℃),工蚁蛹为19.8±5.5天(27℃)。成长工蚁在5—11月出现;8—11月,雄蚁从蛹中羽化;10月,雌蚁从蛹中羽化。雌蚁分飞交尾后,进入邻近蚁巢或回到原巢,脱翅成为蚁后。蚁巢内存在着多后现象。     

Compositional changes in cell wall polysaccharides from chilled and non-chilled cucumber fruit

Cell wall carbohydrate composition and 1-aminocyclopropane-1-carboxylic acid (ACC) content have been determined in chilled (2.5°) and non-chilled (12.5°) cucumber fruit. The major compositional change that accompanied the increased capability for ACC synthesis during chilling was a diminished loss of galactose residues, relative to the loss which occurred at 12.5°. However, the loss of galactose residues increased markedly when fruit were transferred from 2.5° to 20°, and wall galactose levels eventually declined to similar levels in both chilled and non-chilled fruit. Rhamnose, arabinose, xylose, mannose and cellulose content of walls was similar in chilled and non-chilled fruit and did not change substantially upon transfer of fruit to 20°. Upon transfer of chilled fruit from 2.5° to 20°, an increase in the relative amount of galacturonic acid in cell walls occurred; this change did not occur in non-chilled fruit. Thus, chilling stress results in a rapid change in the neutral sugar and galacturonic acid composition of cell wall pectic polysaccharides upon warming.     

Draft Genome Sequence of Fusobacterium nucleatum subsp. fusiforme ATCC 51190T

Fusobacterium nucleatum, one of the major causative bacteria of periodontitis, is classified into five subspecies (nucleatum, polymorphum, vincentii, animalis, and fusiforme) on the basis of the several phenotypic characteristics and DNA homology. This is the first report of the draft genome sequence of F. nucleatum subsp. fusiforme ATCC 51190(T).     

盐度和CO2倍增环境下碱蓬幼苗呼吸酶活性的变化

研究了生长在正常大气ＣＯ２和ＣＯ２倍增环境中的盐生植物碱蓬（Ｓｕａｅｄａｓａｌｓａ）幼苗呼吸酶活性对ＫＣｌ和ＮａＣｌ的反应．结果表明,在ＣＯ２倍增（７００μｌ·Ｌ－１）和正常大气ＣＯ２（３５０μｌ·Ｌ－１）下,３００ｍｍｏｌ·Ｌ－１ＫＣｌ和ＮａＣｌ均能抑制琥珀酸脱氢酶（ＳＤＨ）和苹果酸脱氢酶（ＭＤＨ）活性,而异柠檬酸脱氢酶（ＩＤＨ）活性为ＮａＣｌ抑制、ＫＣｌ促进;ＮａＣｌ和ＫＣｌ明显抑制细胞色素氧化酶（ＣＯ）和光呼吸中乙醇酸氧化酶（ＧＯ）、羟基丙酮酸还原酶（ＨＰＲ）活性;并指出在ＫＣｌ胁迫下,ＣＯ２使三羧酸循环（ＴＣＡＣ）的运行变慢,ＮａＣｌ胁迫下使其加快,ＴＣＡＣ运行限速步骤与ＭＤＨ无关,ＣＯ为盐对呼吸代谢影响的重要位点．另外,Ｋ＋、Ｎａ＋对蛋白表达的影响有差异,ＣＯ２可使盐胁迫下的碱蓬幼苗蛋白表达降低．     

Macrophage migration inhibitory factor (MIF) interacts with Bim and inhibits Bim-mediated apoptosis

The pro-apoptotic Bcl-2 family member Bim acts as a sensor for apoptotic stimuli and initiates apoptosis through the mitochondrial pathway. To identify novel regulators of Bim, we employed the yeast two-hybrid system and isolated the human gene encoding macrophage migration inhibitory factor (MIF), a ubiquitously expressed proinflammatory mediator that has also been implicated in cell proliferation, the cell cycle and carcinogenesis. The interaction between MIF and Bim was confirmed by both in vitro and in vivo protein interaction assays. Intriguingly, protein complexes between MIF and the three major Bim isoforms (BimEL/BimL/BimS) could be detected in HEK293 and K562 cells, especially in cells undergoing apoptosis. Moreover, exogenous expression of MIF partially inhibited Bim-induced apoptosis in HEK293 cells. SiRNA-mediated knockdown of MIF increased apoptosis in K562 cells exposed to the chemical oxidant diamide. Endogenous MIF may regulate the pro-apoptotic activity of Bim and inhibit the release of cytochrome c from mitochondria.     

Buckwheat trypsin inhibitor with helical hairpin structure belongs to a new family of plant defence peptides

A new peptide trypsin inhibitor named BWI-2c was obtained from buckwheat (Fagopyrum esculentum) seeds by sequential affinity, ion exchange and reversed-phase chromatography. The peptide was sequenced and found to contain 41 amino acid residues, with four cysteine residues involved in two intramolecular disulfide bonds. Recombinant BWI-2c identical to the natural peptide was produced in Escherichia coli in a form of a cleavable fusion with thioredoxin. The 3D (three-dimensional) structure of the peptide in solution was determined by NMR spectroscopy, revealing two antiparallel α-helices stapled by disulfide bonds. Together with VhTI, a trypsin inhibitor from veronica (Veronica hederifolia), BWI-2c represents a new family of protease inhibitors with an unusual α-helical hairpin fold. The linker sequence between the helices represents the so-called trypsin inhibitory loop responsible for direct binding to the active site of the enzyme that cleaves BWI-2c at the functionally important residue Arg(19). The inhibition constant was determined for BWI-2c against trypsin (1.7×10(-1)0 M), and the peptide was tested on other enzymes, including those from various insect digestive systems, revealing high selectivity to trypsin-like proteases. Structural similarity shared by BWI-2c, VhTI and several other plant defence peptides leads to the acknowledgement of a new widespread family of plant peptides termed α-hairpinins.