大鼠心肌缺血再灌注损伤模型的实验研究

目的 制备一种新型的心肌急性缺血再灌注损伤模型,以探讨一种更符合临床实际需求的实验方法.方法 将20只雌性SD(Sprague-Dawley)大鼠随机分成2组(对照组、实验组),采用结扎主动脉根部引起心肌缺血5min再灌注30 min建立心肌急性缺血再灌注模型;通过应用透射电镜观察心肌细胞超微结构的改变,同时检测心肌组织匀浆丙二醛(Maleic Dialdehyde,MDA)含量、超氧化物歧化酶(Superoxide Dismutase,SOD)活力.结果 透射电镜下超微结构显示实验组较对照组明显加重了心肌组织结构和线粒体的损害;实验组心肌组织MDA明显高于对照组(P<0.01),而SOD明显低于对照组(P<0.01).结论 本实验成功建立了方法简便、易于操作、取材范围广泛的心肌缺血再灌注损伤模型,为心肌缺血再灌注损伤研究提供了一种更为可行的模型.     

Beneficial effect of fluorocarbon emulsion media on the function of neuromuscular preparations in vitro

The effects of liquid fluorocarbons as bathing media were determined by use of in vitro neuromuscular preparations. Rat hemidiaphragms were bathed in either oxygenated fluorocarbon (FC) emulsion or standard oxygenated Krebs solution. Contractile force in response to simple supramaximal nerve stimuli as well as to high frequency stimulation was greater, while twitch:tetanus ratio was smaller in FC emulsion. With such medium, post-tetanic potentiation of contraction was also more consistently observed. Indirectly stimulated diaphragms survived longer in FC emulsion. After cessation of oxygenation, oxygen tension (ρO(2)) of the medium declined more rapidly with Krebs than with FC emulsion; ρO(2) directly correlated with force of contraction. Similarly, in the chick biventer cervicis preparation, FC emulsion enhanced nerve-stimulated force of contraction; returning the preparation to standard Krebs solution reversed this phenomenon. Dose-resonse curves of muscle contraction in response to acetycholine and KCl administration were shifted upward during FC emulsion superfusion. Frequency of miniature endplate potentials was lower in FC emulsion than that observed in Krebs solution, measured from the same cell of the rat diaphragm. Resting membrane potentials were also greater in muscle cells sampled from FC emulsion-bathed preparations. These data suggest that FC emulsion is superior to standard Krebs solution as a bathing medium for in vitro neuromuscular preparations by virtue of the high solubility of oxygen in it.     

Metabolic responses of microbiota to diesel fuel addition in vegetated soil

The effects of trees and contamination on microbial metabolic activity, especially that of hydrocarbon degrading bacteria, were compared during phytoremediation to find which conditions increase diesel fuel removal. Diesel fuel utilisation, microbial extracellular enzyme activities and utilisation of Biolog ECO plate carbon sources by soil bacteria were determined during phytoremediation experiments consisting of two separate diesel applications. Diesel fuel removal after 28 days of second diesel application was 20–30% more than after the first application 1 year earlier. Soil microbiota utilised 26–31 of the 31 Biolog ECO plate carbon sources. Carbon source utilisation profiles indicated minor differences in microbiota in soil vegetated with pine compared to microbiota in soil vegetated with poplar. The potential maximum rates of aminopeptidase activity were 10–10² M AMC/h/g dry soil prior to and after second diesel application, except 14days after the second diesel addition, where the rates were at the scale of 10³M AMC/h/g dry soil. The potential maximum rates of esterase activity were 10³–10⁴M MUF/h/g dry soil. The presence of plants did not influence the activity of esterases. The utilisation of diesel by soil bacteria in Biolog MT2 plate assay was higher in contaminated soil, especially when vegetated, than in uncontaminated soil, measured both as lag times and maximum specific utilisation rates. MT2 plate assay detected the biological response after diesel fuel addition better than general activity methods.     

Software for quantification of labeled bacteria from digital microscope images by automated image analysis

Automated image analysis software, CellC, was developed and validated for quantification of bacterial cells from digital microscope images. CellC enables automated enumeration of bacterial cells, comparison of total count and specific count images [e.g., 4',6-diamino-2-phenylindole (DAPI) and fluorescence in situ hybridization (FISH) images], and provides quantitative estimates of cell morphology. The software includes an intuitive graphical user interface that enables easy usage as well as sequential analysis of multiple images without user intervention. Validation of enumeration reveals correlation to be better than 0.98 when total bacterial counts by CellC are compared with manual enumeration, with all validated image types. The software is freely available and modifiable: the executable files and MATLAB source codes can be obtained at www. cs. tut.fi/sgn/csb/cellc.     

Thesaurus-based disambiguation of gene symbols

Background  

Massive text mining of the biological literature holds great promise of relating disparate information and discovering new knowledge. However, disambiguation of gene symbols is a major bottleneck.     

Functional studies of signaling pathways in peri-implantation development of the mouse embryo by RNAi

Background

Studies of gene function in the mouse have relied mainly on gene targeting via homologous recombination. However, this approach is difficult to apply in specific windows of time, and to simultaneously knock-down multiple genes. Here we report an efficient method for dsRNA-mediated gene silencing in late cleavage-stage mouse embryos that permits examination of phenotypes at post-implantation stages.

Results

We show that introduction of Bmp4 dsRNA into intact blastocysts by electroporation recapitulates the genetic Bmp4 null phenotype at gastrulation. It also reveals a novel role for Bmp4 in the regulation the anterior visceral endoderm specific gene expression and its positioning. We also show that RNAi can be used to simultaneously target several genes. When applied to the three murine isoforms of Dishevelled, it leads to earlier defects than previously observed in double knock-outs. These include severe delays in post-implantation development and defects in the anterior midline and neural folds at headfold stages.

Conclusion

Our results indicate that the BMP4 signalling pathway contributes to the development of the anterior visceral endoderm, and reveal an early functional redundancy between the products of the murine Dishevelled genes. The proposed approach constitutes a powerful tool to screen the functions of genes that govern the development of the mouse embryo.     

Characterization of iron- and sulphide mineral-oxidizing moderately thermophilic acidophilic bacteria from an Indonesian auto-heating copper mine waste heap and a deep South African gold mine

Iron- and chalcopyrite-oxidizing enrichment cultures were obtained at 50°C from acidic, high-temperature, copper/gold mine environments in Indonesia and South Africa. Over 90% copper yield was obtained from chalcopyrite concentrate with the Indonesian enrichment in 3 months with 2% solids concentration, when pH was maintained at around 2. Neither addition of silver cations nor an enhanced nutrient concentration influenced chalcopyrite leaching. Excision and sequencing of bands from denaturing gradient gel electrophoresis of the amplified partial 16S rRNA gene showed that the enrichment cultures from different environments in South Africa and Indonesia were very simple, and similar. Chalcopyrite concentrate supported a simpler and different community than Fe²⁺. The members of the enrichment cultures were closely related to Sulfobacillus yellowstonensis and Sulfobacillus acidophilus.     

Behavioural and physiological consequences of acute social defeat in growing gilts: effects of the social environment

Endocrine, behavioural and immunologic processes, together with body growth, were evaluated in gilts that were defeated at 10 weeks of age in resident-intruder tests. Immediately after defeat, gilts were either separated from or reunited with a familiar conspecific (litter-mate; always a barrow). Gilts were assigned to one of four treatments: (a) DI: defeat, followed by isolation (separation from original litter-mate; n=8); (b) I: no defeat, isolation (control group; n=9); (c) DP; defeat, followed by pair-housing (reunion with original litter-mate; n=8); and (d) P: no defeat, pair-housing (control group; n=8). The following general conclusions were derived: (1) social defeat caused pronounced short-term elevations in hypothalamic-pituitary-adrenal (HPA) and sympathetic-adrenal medullary activities, and of prolactin levels. Moreover, as soon as 1h after defeat, percentages of blood lymphocytes and neutrophilic granulocytes were, respectively, decreased and increased; (2) social defeat had some long-lasting influence on behaviour and physiology, but isolation predominantly determined responses in the longer term. Defeat, as well as isolation, resulted in increased cardiovascular activities compared to P controls, as observed in a novel object test (NOT: +7 days) and an aversion test (AVT: +14 days). Moreover, defeated as well as isolated gilts did not habituate to a repeated novel environment test (NET: -7, +2 and +7 days) in terms of frequencies of vocalising, whereas P controls did. Isolation, through the separation from any other pig, was responsible for the other observed long-term characteristics, which developed progressively. Isolated gilts showed high mobilities and high cortisol responses in the repeated NET (+7 days), not being habituated. This contrasted the reactions of pair-housed gilts, which were much reduced. In addition to their high cardiovascular activities in the NOT and the AVT, isolated gilts also displayed higher heart rates in the repeated NET and during human presence following the NOT, compared to pair-housed gilts. Finally, isolated gilts were more inhibited to approach a novel object (in the NOT) than pair-housed pigs; and (3) stress responses of defeated gilts were modulated by the subsequent social environment. Stimulation of the HPA-axis (plasma- and salivary cortisol) was prolonged in those defeated gilts which were isolated (observed in the first hour). Changes in leucocyte subsets were still observed after 3 days in DI, but were 'normalised' within 1 day in DP gilts. Two days after defeat, habituation to the repeated NET in terms of mobility and salivary cortisol responses occurred in control and DP gilts, but not in DI gilts. We argue that these effects of the social environment shortly after defeat were related to a stress-reducing effect of a stable social relationship, i.e. social support.