Some observations on the subfibrillar structure of collagen fibrils as noted during treatment with NKISK and cathepsin G with mechanical agitation

We observed the structure of collagen fibrils in rat tail tendons after treatment with NKISK and cathepsin G. NKISK is a pentapeptide that has been previously shown to bind fibronectin, while cathepsin G is a serine protease that cleaves fibronectin but not type I collagen. In tendons treated with NKISK, fibrils were seen to extensively dissociate into smaller-diameter subfibrils. These subfibrils were homogeneous in diameter with an average diameter of 26.3?±?5.8?nm. Similar, although less extensive, dissociation into subfibrils was found in tendons treated with cathepsin G. The average diameter of these subfibrils was 24.8?±?4.9?nm. The ability of NKISK and cathepsin G to release subfibrils at physiological pH without harsh denaturants may enhance the study of the subfibrillar structure of collagen fibrils.     

Phase‐Separation‐Induced Micropatterned Polymer Surfaces and Their Applications

We report a route to fabricate micropatterned polymer films with micro‐ or nanometer‐scale surface concavities by spreading polymer solutions on a non‐solvent surface. The route is simple, versatile, highly efficient, low‐cost, and easily accessible. The concavity density of the patterned films is tuned from 10⁶ to 10⁹ features cm^–2, and the concavity size is controlled in the range from several micrometers to less than 100 nm, by changing the film‐forming parameters including the polymer concentration, the temperature of the non‐solvent and the interactions between polymer, solvent, and non‐solvent. We further demonstrate that these concavity‐patterned films have significantly enhanced hydrophobicity, owing to the existence of the surface concavities, and their hydrophobicity could be controlled by the concavity density. These films have been used as templates to successfully fabricate convex‐patterned polymer films, inorganic TiO₂ microparticles, and NaCl nanocrystals. Their other potential applications are also discussed.     

High-resolution ultrastructure of amyloid fibrils in familial amyloid polyneuropathy

The ultrastructure of amyloid fibrils in familial amyloid polyneuropathy (FAP) was clearly demonstrated. Amyloid of three patients with FAP caused by the point mutation of the 30th amino acid of transthyretin (ATTR Val30Met) and one patient with FAP caused by two point mutations of the 30th and 104th amino acid of transthyretin (ATTR Val30Met/Arg104Cys) were partially isolated, stained negatively and examined with an electron microscope. Amyloid fibrils of both types were composed of two protofilaments and twisted at 180 degrees to the right and left alternately with a periodicity of 125-135 nm. This is the first report demonstrating such unique alternating twist structure of amyloid fibrils. There were no ultrastructural differences between the fibrils caused by the ATTR Val30Met and ATTR Val30Met/ Arg104His; therefore, it is suggested that the point mutation of the 30th amino acid of transthyretin might play an important role in the formation of amyloid fibrils. Further biochemical study on the mechanism of this alternating twist formation should be undertaken.     

A sulphur matrix complex,elastic fibril composed of a fine core of amorphous elastin and microfibrils was largely accumulated in the aortic intima of aged rats

We examined elastic fibrils in the aortic intima of aged rats with elastin staining, elastase digestion and X-ray microanalysis. The innermost elastic lamina, heavily stained by a brief treatment with azure II-toluidine blue without heating, was thin and fragmented, and an amorphous substance that stained weakly to moderately was accumulated in the thick subendothelial space of 26-28-month-old rat aortas. The substance was always present in the intimas; to a large extent in the aged rats but rarely in 5-month-old rats, and disappeared after digestion with elastase. The amorphous substance was identified as elastic fibrils approximately 0.1-0.2 microm in diameter under an electron microscope with tannic acid-uranyl acetate/lead citrate staining. Elastase digestion revealed that elastic fibrils were composed of a number of microfibrils, which were 10-12 nm in diameter, in and around a fine core of amorphous elastin. X-ray microanalysis revealed a clear peak of sulphur in the elastic fibrils.     

Development of the Fibrillar and Microfibrillar Structure During Biomimetic Mineralization of Wood

Wood is a hierarchical composite, consisting at its lowest hierarchy level of crystalline cellulose elementary fibrils with diameters of 2–4 nm embedded in a matrix of hemicelluloses and lignin. At the micrometer scale, it has a cellular architecture resembling a honeycomb structure. The transformation of the hierarchical wood structure into a silica replica has been reported recently. Its formation process and structural details are studied in this contribution. First, a silica/biopolymer composite is prepared by wood delignification and cell‐wall modification, followed by silica precursor infiltration and condensation. The calcination process is monitored to gain insight into the structure development upon decomposition of the biopolymers. The material changes its architecture gradually from fibrillar structures of 10–20 nm in diameter with homogeneous electron density, into fibrils of 8–10 nm in diameter with inhomogeneous electron density, exhibiting internal sub‐fibrillar structures of about 2 nm in diameter. The steps of the successful replication of the cellulose elementary fibrils into nanopores of similar diameter and orientation in a fibrillar silica matrix are demonstrated. These nanopore replicas of the original cellulose are wound in a steep helix within the macropore walls. These advanced materials may have lightweight structural applications and the nanopores may be advantageous for molecular separation.     

Perichromatin granule-like granular structures in the nuclei of antral-follicular oocytes of the Chinese hamster

In the nuclei of antral-follicular oocytes of the Chinese hamster (Cricetulus griseus), numerous granular structures ranging from 60 nm to 250 nm in diameter were present in the nucleolar area, and a few were observed on the surface of heterochromatin materials and within the interchromatin space. When treated with bismuth staining en bloc after glutaraldehyde fixation (GA-Bi staining), these granular structures were shown to be composed of fine fibrils intensely contrasted with bismuth, indicating that these may be regarded as one type of perichromatin granule.     

Thermo‐Mechanical Responses of Liquid‐Crystal Networks with a Splayed Molecular Organization

Films of liquid‐crystal networks with a splayed molecular alignment over their cross‐section display a well‐controlled deformation as a function of temperature. The deformation can be explained in terms of differences in thermal expansion depending on the average molecular orientation of the mesogenic centers of the monomeric units. The thermal expansion of the anisotropic polymers has been characterized as a function of their molecular structure and the polymerization conditions. As a reference, films with an in‐plane 90° twist have also been studied and compared with the splayed, out‐of‐plane molecular rotation. The twisted films show a complex macroscopic deformation owing to the formation of saddle‐like geometries, whereas the deformation of the splayed structured is smooth and well controlled. The deformation behavior is anticipated to be of relevance for polymer‐based microelectromechanical system (MEMS) technology.     

Abnormal formation of the glucan network from regenerating protoplasts in Schizosaccharomyces pombe cps8 actin point mutant

To study the close relationship between the actin cytoskeleton and cell wall formation, the process of cell wall formation in reverting protoplasts of the fission yeast, Schizosaccharomyces pombe, cps8 actin point mutant was investigated by ultra-high-resolution low-voltage scanning electron microscopy (UHR-LVSEM) and transmission electron microscopy (TEM). The protoplast of the cps8 mutant began to form a glucan network in a unipolar manner and to secrete alpha-galactomannan. The site of cell wall formation grew in a cylindrical shape in the wild-type protoplast. The alpha-galactomannan did not fill in the intrafibrillar spaces completely, however, and the fibrils were exposed on the cell surface. UHR-LVSEM images indicated that the glucan fibrils were thin and rope-shaped, forming a looser network than the wild-type. TEM images indicated the finest fibrils were approximately 1.5 nm in diameter, the same diameter as the wild-type. These results suggest that the cps8 mutant was insufficient in developing cross-linkage with the glucan fibrils up to the wide ribbon shape as found in the wild-type [Osumi M et al. (1989) J. Electron Microsc. 38: 457-468; Osumi M (1998) Micron 29: 207-233]. These findings appear to indicate that the actin cytoskeleton controls formation of the glucan network and secretion of beta-1,6-glucan, and confirm the close relationship of the actin cytoskeleton and glucan formation.     

Snakes, shapes, and gradient vector flow

Snakes, or active contours, are used extensively in computer vision and image processing applications, particularly to locate object boundaries. Problems associated with initialization and poor convergence to boundary concavities, however, have limited their utility. This paper presents a new external force for active contours, largely solving both problems. This external force, which we call gradient vector flow (GVF), is computed as a diffusion of the gradient vectors of a gray-level or binary edge map derived from the image. It differs fundamentally from traditional snake external forces in that it cannot be written as the negative gradient of a potential function, and the corresponding snake is formulated directly from a force balance condition rather than a variational formulation. Using several two-dimensional (2-D) examples and one three-dimensional (3-D) example, we show that GVF has a large capture range and is able to move snakes into boundary concavities.     

A New Active Contour Model Based on Distance-Weighted Potential Field

Snakes, or active contours, have been widely used in various image processing applications. Typical problems of snakes, including limited capture range, poor convergence to concavities, noise sensitivity, and initialization sensitivity, have limited their applications. For solving these problems, we propose a new potential for the active contour model. In this proposed potential field, each location’s potential is computed by integrating the feature information from all the pixels in the image with the distances as the weights. The external forces are computed as the gradients of this proposed potential field, and the computed external forces are static and have global capture range. Experiments and also the comparisons with the snake using gradient vector flow (GVF) as external forces are conducted to examine the performances of this proposed snake. The results show that the proposed snake has a large capture range and an excellent convergence to boundary concavities, and also the proposed snake is more robust to noise, more time efficient, and less sensitive to the initialization, compared with the GVF snake.     

基于3D区域增长法和改进的凸包算法相结合的全肺分割方法

肺实质分割结果的准确性在实际临床应用中具有非常重要的意义。但由于肺结节的位置、大小、形状的不规则性,肺部病变的多样性,以及人体胸部解剖结构的明显差异等,使得各类分割方法不能统一地适用于所有的胸部CT图像,所以对于肺实质分割方法的研究仍具有很大的挑战。该文在国内外研究分析的基础上提出基于3D区域增长法与改进的凸包修补算法相结合的全肺分割方法。在3D区域增长法的粗分割基础上,对分割的结果进行细化工作,通过连通域标记法与形态学方法相结合去除气管和主支气管,得到初步的肺实质掩膜,最后应用改进的凸包算法对肺部轮廓进行修补平滑,最终得到肺部分割结果。通过与凸包算法及滚球法相对比,证明该文所提改进的凸包算法能够有效地修补肺部轮廓凹陷,修补后的结果分割精度较高。     

Ultra-Small ATP-Decorated Gold Nanoparticles for Targeting Amyloid Fibrils in Neurodegenerative Diseases

Ultrafine Gold nanoparticles (Au NPs) functionalized with various biomolecules constitute an alternative to antibodies as anti-amyloidogenic agents. However, generating stable ultrafine Au NPs with high surface activity is challenging. Here, the capacity of phosphate groups in biomolecules is used to stabilize Au NPs. The characteristics of Au NPs decorated with adenosine mono-, di-, and tri-phosphate are compared as well as adenosine and peptide nucleic acid-containing adenosine as controls. Among them, ATP-Au NPs are found to be superior having small size (2–4 nm) and stability (for several months) when analysed by spectroscopy and electron microscopy. Spectroscopy analysis also revealed that each ATP-stabilized Au NP is decorated with 7–8 molecules of ATP. ThT binding analysis and TEM imaging showed that the ATP-Au NPs efficiently prevented amyloid fibril formation in vitro by Aβ-42, α-Synuclein as well as by the Glucosylceramide metabolite, and disaggregated their pre-formed fibrils. NMR analysis revealed the interaction of the ATP-Au NPs with the amyloid fibrils. The ATP-Au NPs are safe toward cultured SH-SY5Y cells and when co-incubated with α-Synuclein amyloids inhibited their cytotoxicity and readily enter the cells to inhibit formation of amyloid fibrils within them. The results indicates the pharmacological potentials of ATP decorated Au NPs.     

系统性淀粉样变性的超微病理诊断

目的:应用光镜(LM)及透射电镜(TEM)观察不同部位淀粉样物质的沉积及超微结构特点,为系统性淀粉样变性提供可靠的病理诊断依据.方法:对21例临床拟诊为系统性淀粉样变性病例的肾脏,腹壁脂肪和直肠粘膜活检组织按常规进行LM及TEM标本制备及病理检查.结果:LM观察显示淀粉样物质在肾脏,腹壁脂肪和直肠粘膜沉积阳性率分别为7...     

Appearance of electron-dense segments: indication of possible conformational changes of pre-mineralizing collagen fibrils in the osteoid of rat bones

To elucidate precise mechanisms of appositional mineralization of bone, structural features of mineralizing collagen fibrils of the osteoid in normal and hypocalcaemic rats were examined in detail by transmission electron microscopy. Ultrathin sections of the osteoid of various types of bones of the rats fed with regular or normal calcium diet often displayed electrondense segments in the specific regions of the collagen fibrils located immediately adjacent to the mineralization front or to the mineralization nodules. Such dense segments appeared only after Ur-Pb staining and were more distinct in undecalcified specimens. Dense segments were undetectable in ultrathin sections picked up on ethylene glycol instead of water in the trough, even after Ur-Pb staining. Collagen fibrils in the widened osteoid of hypocalcaemic rats fed with calcium-free diet failed to show electron-dense segments. A careful comparison between the hydrously or anhydrously processed adjacent sections of a normal rat bone indicated a drastic dissolution of electron-dense material from the bone matrix near the mineralization front in hydrously processed sections and, thus, implicated the presence of labile mineral-matrix complexes in the recently mineralized bone matrix. Such labile sediments were readily dissociated within the ultrathin sections while the sections were floating on water and immediately adsorbed onto the pre-mineralizing collagen fibrils, where some conformational changes might have occurred. These data indicate that highly electron-dense segments appearing in the osteoidal collagen fibrils are a type of process-induced product, which indirectly represent possible structural alterations in the segmental portions of pre-mineralizing collagen fibrils in the osteoid of rat bones.     

R-Snake:一种基于边缘与区域信息的图像主动轮廓提取模型

 传统Snake模型存在着对轮廓的初始化敏感,对高噪声图像易陷入局部极小值,以及对具有狭长深度凹陷区域的图像无法获得正确轮廓等问题.本文提出了一种基于边缘与区域信息的主动轮廓模型R-Snake(Region Snake).该模型通过文中设计的图像变换算子,并结合区域积分与曲线积分间转化的Green公式,导出了包含目标图像区域信息的区域力.然后由力平衡方程将该区域信息自然直接地引入到主动轮廓提取模型中,从而实现图像的轮廓提取.由于该模型同时利用了图像的区域信息和梯度信息来引导轮廓曲线的演化,使得本文方法不仅扩大了轮廓初始化的范围,降低了对图像噪声的敏感性,而且还增加了轮廓曲线收敛到真实边界的能力.实验结果表明,本文方法具有很强的适应性和鲁棒性,尤其是对高噪声图像和具有狭长深度凹陷的图像获得了优于传统Snake模型的结果.     

Ultrastructure and behavior of actin cytoskeleton during cell wall formation in the fission yeast Schizosaccharomyces pombe

Fluorescence microscopy has shown that F-actin of the fission yeast Schizosaccharomyces pombe forms patch, cable and ring structures. To study the relationship between cell wall formation and the actin cytoskeleton, the process of cell wall regeneration from the protoplast was investigated by transmission electron microscopy (TEM), immunoelectron microscopy (IEM) and three-dimensional reconstruction analysis. During cell wall regeneration from the protoplast, localization of F-actin patches was similar to that of the newly synthesized cell wall materials, as shown by confocal laser scanning microscopy (CLSM). In serial sectioned TEM images, filasomes were spherical, 100-300 nm in diameter and consisted of a single microvesicle (35-70 nm diameter) surrounded by fine filaments. Filasomes were adjacent to the newly formed glucan fibrils in single, cluster or rosary forms. By IEM analysis, we found that colloidal gold particles indicating actin molecules were present in the filamentous area of filasomes. Three-dimensional reconstruction images of serial sections clarified that the distribution of filasomes corresponded to the distribution of F-actin patches revealed by CLSM. Thus, a filasome is one of the F-actin patch structures appearing in the cytoplasm at the site of the initial formation of the cell wall and it may play an important role in this action.     

基于负染－电镜的Aβ体外聚集表征测试要点分析

本文应用电镜负染色技术,对阿尔茨海默病（ Alzheimer′s disease, AD）中所涉及的β淀粉样肽（β?amyloid peptide,Aβ）的聚集规律进行了分析。通过对样品的浓度、pH值、温度和时间等设置系列梯度,以及调整负染试剂的种类、浓度和染色时间,优化了Aβ寡聚体样品制备以及电镜观察的最佳条件,获得了清晰的Aβ纤维图像并用以估算纤维长度。实验结果表明：经过六氟异丙醇给予单体化处理,并溶于无水二甲基亚砜（ DMSO）的Aβ（100μmol·L－1）,在pH7?2的PBS中4℃孵育24 h,Aβ寡聚体形态清晰,且Aβ42寡聚体比Aβ40寡聚体聚集趋势更明显。在pH 8?0的硼酸缓冲液中,Aβ42（40μmol·L－1）在37℃孵育72 h后,纤维形成明显;而Aβ纤维样品经过煮沸后再制备负染样品,所得电镜图像更为清晰,便于对纤维长度和结构进一步分析。因此电镜负染色技术,可作为一种快捷,直观的Aβ体外聚集形貌表征的质控方法。     

A Nonconservative Flow Field for Robust Variational Image Segmentation

We introduce a robust image segmentation method based on a variational formulation using edge flow vectors. We demonstrate the nonconservative nature of this flow field, a feature that helps in a better segmentation of objects with concavities. A multiscale version of this method is developed and is shown to improve the localization of the object boundaries. We compare and contrast the proposed method with well known state-of-the-art methods. Detailed experimental results are provided on both synthetic and natural images that demonstrate that the proposed approach is quite competitive.      

Fluid Vector Flow and Applications in Brain Tumor Segmentation

In this paper, we propose a new approach that we call the ldquofluid vector flowrdquo (FVF) active contour model to address problems of insufficient capture range and poor convergence for concavities. With the ability to capture a large range and extract concave shapes, FVF demonstrates improvements over techniques like gradient vector flow, boundary vector flow, and magnetostatic active contour on three sets of experiments: synthetic images, pediatric head MRI images, and brain tumor MRI images from the Internet brain segmentation repository.