Dense chromatin plates in metaphase chromosomes

In a previous work we observed multilayered plate-like structures surrounding partially denatured HeLa chromosomes at metaphase ionic conditions. This unexpected finding has led us to carry out an extensive investigation of these structures. Our results show that plates can also be found in metaphase chromosomes from chicken lymphocytes. We have used atomic force microscopy (AFM) to image and investigate the mechanical properties of plates in aqueous solution. Plates are thin (~6.5 nm each layer) but compact and resistant to penetration by the AFM tip: their Young’s modulus is ~0.2 GPa and the stress required for surface penetration is ~0.03 GPa in the presence of Mg²⁺ (5–20 mM). Low-ionic strength conditions produce emanation of chromatin fibers from the edges of uncrosslinked plates. These observations and AFM results obtained applying high forces indicate that the chromatin filament is tightly tethered inside the plates. Images of metal-shadowed plates and cryo-electron microscopy images of frozen-hydrated plates suggest that nucleosomes are tilted with respect to the plate surface to allow an interdigitation between the successive layers and a thickness reduction compatible with the observed plate height. The similarities between denatured plates from chicken chromosomes and aggregates of purified chromatin from chicken erythrocytes suggest that chromatin has intrinsic structural properties leading to plate formation. Scanning electron micrographs and images obtained with the 200-kV transmission microscope show that plates are the dominant component of compact chromatids. We propose that metaphase chromosomes are formed by many stacked plates perpendicular to the chromatid axis.     

Unique digestive system,trophic specialization,and diversification in the deep‐sea gastropod genus Scaphander

Dietary specialization is known to be important for the evolution of Cephalaspidea gastropods, but still little is known about the overall trophic interactions of the group and the putative role of trophic ecology on diversification. The genus Scaphander is a group of predominantly deep‐sea, infaunal cephalaspids with about 40% of its species (eight) occurring on the Atlantic Ocean. They are carnivorous and have a unique digestive system with a large, strongly muscularized gizzard containing three sizable and heavily calcified plates. This work aims to describe the diet of Scaphander, to evaluate if there is a functional relation between the anatomy of the digestive tract and exploitation of novel food resources, and to assess if dietary specialization may have played a role in the diversification of the Atlantic species of Scaphander. Gut contents were studied from 31 specimens representing seven of the Atlantic species using scanning electron microscopy and light microscopy. The chemical composition of the gizzard plates was analysed by X‐ray microanalysis and X‐ray powder diffraction. Foraminiferans, including agglutinating forms, were shown to be the most important food item for Scaphander; bivalves, gastropods, scaphopods, and polychaetes with calcareous tubes were also found to form part of the diet. The gizzard plates were shown to consist of a phosphate‐rich amorphous component and a crystalline component identified as fluorite (hardness 4; Moh's scale). It is suggested that the ability of Scaphander gastropods to prey upon organisms with hard tests, particularly agglutinating foraminiferans, is not only due to the hardness of the gizzard plates but to the cumulative effect of shape and hardness of the gizzard plates and relative size of the gizzard and associated musculature. No interspecific differences were found in the diet and morphology of the digestive tract, indicating that dietary specialization probably has not played a significant role in the diversification of Atlantic species of Scaphander. © 2013 The Linnean Society of London, Biological Journal of the Linnean Society, 2013, 109 , 512–525.     

Systematic and ecological wood anatomy of Neotropical Schefflera (Araliaceae), with an emphasis on the Didymopanax group

Wood anatomy has been investigated from 35 species belonging to the Neotropical clade of the polyphyletic genus Schefflera (Araliaceae), representing three of the five subgroups (Didymopanax, Crepinella and Sciodaphyllum). The species examined are rather uniform in their wood structure, sharing the presence of scalariform and simple perforation plates, septate fibres and scanty paratracheal axial parenchyma. The observed variation in many wood characters showed statistically significant differences relative to latitude, climate and, especially, vegetation types. In particular, the intervessel pits are larger in species from higher latitudes and in seasonally dry habitats than those from lower latitudes and rainforests. Latitudinal and ecological trends in the variation of vessel element lengths, bar numbers on perforation plates, intervessel pit sizes and ray widths may be at least partially explained as effects of adaptation to drier environments in the course of dispersal outside the Amazonian region and diversification in the Atlantic Forest subclade and the Savannic subclade within the Didymopanax group. The occurrence of a granular annulus on the intervessel pit membranes in S. chimantensis and S. sprucei (both of the Crepinella group) is the first record of this feature in Araliaceae. In comparisons of Neotropical Schefflera with the other major clades of Schefflera sensu lato, wood anatomical diversity is consistent with the polyphyly of this genus based on molecular phylogenetic analyses. © 2013 The Linnean Society of London, Botanical Journal of the Linnean Society, 2013, 173 , 452–475.     

Fit optimisation of a distal medial tibia plate

An iterative method for the fit optimisation of a pre-contoured fracture fixation plate for a given bone data set is presented. Both plate shape optimisation and plate fit quantification are conducted in a virtual environment utilising computer graphical methods and 3D bone and plate models. Two optimised shapes of the undersurface of an existing distal medial tibia plate were generated based on a dataset of 45 3D bone models reconstructed from computed tomography image data of Japanese tibiae. The existing plate shape achieved an anatomical fit on 13% of tibiae from the dataset. Modified plate 1 achieved an anatomical fit for 42% and modified plate 2 a fit for 67% of the bones. If either modified plate 1 or plate 2 is used, then the anatomical fit can be increased to 82% for the same dataset. Issues pertaining to any further improvement in plate fit/shape are discussed.     

虻科成虫雌性尾器亚生殖板的研究

应用亚生殖板细微结构的鉴别方法,研究了虻科20种雌性成虫雌性尾器的亚生殖板。结果表明:虻科雌性成虫不同种其尾器亚生殖板整体大小,颜色,有无骨化现象、骨化部位及骨化程度,端缘、基缘凹陷深浅及宽窄,侧突位置、大小,基角大小及形状,毛和鬃的有无、着生位置及多少,皱褶的有无及位置等特征均有变化。虻科成虫雌性尾器亚生殖板的细微结构有种间鉴别的特异性。     

Inappropriate analysis does not reveal the ecological causes of evolution of stickleback armour: a critique of Spence et al. 2013

In a recent paper in this journal, Spence et al. (2013) sought to identify the ecological causes of morphological evolution in three‐spined sticklebacks Gasterosteus aculeatus, by examining phenotypic and environmental variation between populations on the island of North Uist, Scotland. However, by using simple qualitative assessments of phenotype and inappropriate measures of environmental variation, Spence et al. have come to a conclusion that is diametrically opposite to that which we have arrived at in studying the same populations. Our criticisms of their paper are threefold: (1) using a binomial qualitative measure of the variation in stickleback armour (“low” versus “minimal” (i.e., “normal” low‐plated freshwater sticklebacks versus spineless and/or plateless fish)) does not represent the full range of phenotypes that can be described by quantitative measures of the individual elements of armour. (2) Their use of unspecified test kits, with a probable accuracy of 4 ppm, may not be accurate in the range of water chemistry on North Uist (1 to 30 ppm calcium). (3) Their qualitative assessment of the abundance of brown trout Salmo trutta as the major predator of sticklebacks does not accurately describe the variation in brown trout abundance that is revealed by catch‐per‐unit‐effort statistics. Repeating Spence et al.'s analysis using our own measurements, we find, in direct contradiction to them, that variation in stickleback bony armour is strongly correlated with variation in trout abundance, and unrelated to variation in the concentration of calcium in the lochs in which they live. Field studies in ecology and evolution seldom address the same question in the same system at the same time, and it is salutary that in this rare instance two such studies arrived at diametrically opposite answers.     

桡骨远端骨折治疗的选择

目的：探讨不同类型的桡骨远端骨折的有效治疗方法。方法：分析106例桡骨远端骨折,分别采用闭合手法复位,切开复位或有限切开复位内固定方法,分别对骨折复位比较及功能评分。结果：完整随访106例桡骨远端骨折病例,随访时间3～21个月。对保守治疗组与手术治疗组进行骨折复位测定及改良Garland和Werley评分,A,B型骨折无显著性差异;C型骨折中,手术组明显优于保守治疗组。结论：对于C型骨折,建议行切开复位内固定治疗;对于A,B型需根据实际情况选择治疗方式。     

前后联合入路微创治疗合并神经损伤的不稳定骶骨骨折

目的:探讨前后联合入路锁定加压钢板(LCP)微创治疗合并神经损伤的骶骨不稳定骨折的效果。方法:前后联合入路按照杜明奎等方法[1]采用LCP固定治疗合并神经损伤的不稳定骶骨骨折患者5例:L5神经根损伤2例,骶丛神经损伤3例。前方入路暴露骨盆前环重建钢板固定,后方入路经皮下锁定加压钢板骨折复位固定术,椎管探查减压以MRI显示有无神经压迫为据。结果:5例均获随访,时间3～20(14.8±7.2)个月。2例L5神经根损伤和3例骶丛神经损伤者Frankel分级由C级恢复至E级,术后功能根据Matta评分标准评定:优3例,良2例。结论:前后联合入路LCP微创治疗合并神经损伤的骶骨不稳定骨折是一种简单微创有效的方法,值得临床推广。     

Harvesting and Cryo-cooling Crystals of Membrane Proteins Grown in Lipidic Mesophases for Structure Determination by Macromolecular Crystallography

An important route to understanding how proteins function at a mechanistic level is to have the structure of the target protein available, ideally at atomic resolution. Presently, there is only one way to capture such information as applied to integral membrane proteins (Figure 1), and the complexes they form, and that method is macromolecular X-ray crystallography (MX). To do MX diffraction quality crystals are needed which, in the case of membrane proteins, do not form readily. A method for crystallizing membrane proteins that involves the use of lipidic mesophases, specifically the cubic and sponge phases^1-5, has gained considerable attention of late due to the successes it has had in the G protein-coupled receptor field^6-21 (www.mpdb.tcd.ie). However, the method, henceforth referred to as the in meso or lipidic cubic phase method, comes with its own technical challenges. These arise, in part, due to the generally viscous and sticky nature of the lipidic mesophase in which the crystals, which are often micro-crystals, grow. Manipulating crystals becomes difficult as a result and particularly so during harvesting^22,23. Problems arise too at the step that precedes harvesting which requires that the glass sandwich plates in which the crystals grow (Figure 2)^24,25 are opened to expose the mesophase bolus, and the crystals therein, for harvesting, cryo-cooling and eventual X-ray diffraction data collection.The cubic and sponge mesophase variants (Figure 3) from which crystals must be harvested have profoundly different rheologies^4,26. The cubic phase is viscous and sticky akin to a thick toothpaste. By contrast, the sponge phase is more fluid with a distinct tendency to flow. Accordingly, different approaches for opening crystallization wells containing crystals growing in the cubic and the sponge phase are called for as indeed different methods are required for harvesting crystals from the two mesophase types. Protocols for doing just that have been refined and implemented in the Membrane Structural and Functional Biology (MS&FB) Group, and are described in detail in this JoVE article (Figure 4). Examples are given of situations where crystals are successfully harvested and cryo-cooled. We also provide examples of cases where problems arise that lead to the irretrievable loss of crystals and describe how these problems can be avoided. In this article the Viewer is provided with step-by-step instructions for opening glass sandwich crystallization wells, for harvesting and for cryo-cooling crystals of membrane proteins growing in cubic and in sponge phases.     

Scale-up of Escherichia coli growth and recombinant protein expression conditions from microwell to laboratory and pilot scale based on matched k(L)a

Fermentation optimization experiments are ideally performed at small scale to reduce time, cost and resource requirements. Currently microwell plates (MWPs) are under investigation for this purpose as the format is ideally suited to automated high-throughput experimentation. In order to translate an optimized small-scale fermentation process to laboratory and pilot scale stirred-tank reactors (STRs) it is necessary to characterize key engineering parameters at both scales given the differences in geometry and the mechanisms of aeration and agitation. In this study oxygen mass transfer coefficients are determined in three MWP formats and in 7.5 L and 75 L STRs. k(L)a values were determined in cell-free media using the dynamic gassing-out technique over a range of agitation conditions. Previously optimized culture conditions at the MWP scale were then scaled up to the larger STR scales on the basis of matched k(L)a values. The accurate reproduction of MWP (3 mL) E. coli BL21 (DE3) culture kinetics at the two larger scales was shown in terms of cell growth, protein expression, and substrate utilization for k(L)a values that provided effective mixing and gas-liquid distribution at each scale. This work suggests that k(L)a provides a useful initial scale-up criterion for MWP culture conditions which enabled a 15,000-fold scale translation in this particular case. This work complements our earlier studies on the application of DoE techniques to MWP fermentation optimization and in so doing provides a generic framework for the generation of large quantities of soluble protein in a rapid and cost-effective manner.