Poröse Scaffolds aus mineralisiertem Kollagen – ein biomimetisches Knochenersatzmaterial

Porous scaffolds made from mineralised collagen – a biomimetic bone graft material Using biomimetically mineralised collagen type I, a new porous bone graft material has been developed, the composition of which mimics extracellular matrix of bone tissue. The pore structure is generated by a freeze drying process, whereas the pore size can be controlled by temperature and velocity of the freezing over a wide range. The structure is stabilized by crosslinking of the collagen with the water soluble carbodiimide derivative EDC. For the seeding with bone cells, pores with diameters of about 200 μm have turned out to be optimal. These scaffolds are elastic in the wet state which allow for cell culture experiments under mechanical stimulation.     

胶原在组织工程人工皮肤中的应用

作为生物相容性良好的医用材料，胶原被广泛应用于人工皮肤。文中介绍了胶原基人工皮肤的特点、应用形式和国内外的研究情况。     

AFM analysis of the lacunar-canalicular network in demineralized compact bone

Atomic force microscopy has been successfully used to examine a wide range of cellular and biomolecular structures and interactions. The application of atomic force microscopy in the analysis of organs and tissues, however, has been limited. In this study, we present a new method for high-resolution atomic force microscopy imaging of compact bone tissue. We performed atomic force microscopy imaging on demineralized compact bone from bovine tibia to obtain structural information about the bone matrix and the lacunar-canalicular network. Knowledge of the dimensions and distributions of the network allows quantitative analysis of the microfluidics of bone tissue. Results from our study show that (1) the canalicular distribution and dimensions are homogenous in transverse, radial and longitudinal orientations; (2) the lamellae of an osteon consist of alternating high and low bands; (3) the canaliculi follow the contour of lamellar bands and (4) globular structures cover much of the bone matrix, including canalicular walls. Our work demonstrates that atomic force microscopy studies of thin-section tissue samples can provide structural details at nanometre resolution.     

The effects of refractive index heterogeneity within kidney tissue on multiphoton fluorescence excitation microscopy

Although multiphoton fluorescence excitation microscopy has improved the depth at which useful fluorescence images can be collected in biological tissues, the reach of multiphoton fluorescence excitation microscopy is nonetheless limited by tissue scattering and spherical aberration. Scattering can be reduced in fixed samples by mounting in a medium whose refractive index closely matches that of the fixed material. Using optical 'clearing', the effects of refractive index heterogeneity on signal attenuation with depth are investigated. Quantitative measurements show that by mounting kidney tissue in a high refractive index medium, less than 50% of signal attenuates in 100 μm of depth.     

Backscattered electron imaging of the undersurface of resin-embedded cells by field-emission scanning electron microscopy

In this study backscattered electron (BSE) imaging was used to display cellular structures stained with heavy metals within an unstained resin by atomic number contrast in successively deeper layers. Balb/c 3T3 fibroblasts were cultured on either 13-mm discs of plastic Thermanox, commercially pure titanium or steel. The cells were fixed, stained and embedded in resin and the disc removed. The resin block containing the cells was sputter coated and examined in a field-emission scanning electron microscope. The technique allowed for the direct visualization of the cell undersurface and immediately overlying areas of cytoplasm through the surrounding embedding resin, with good resolution and contrast to a significant depth of about 2 μm, without the requirement for cutting sections. The fixation protocol was optimized in order to increase heavy metal staining for maximal backscattered electron production. The operation of the microscope was optimized to maximize the number of backscattered electrons produced and to minimize the spot size. BSE images were collected over a wide range of accelerating voltages (keV), from low values to high values to give ‘sections' of information from increasing depths within the sample. At 3–4 keV only structures a very short distance into the material were observed, essentially the areas of cell attachment to the removed substrate. At higher accelerating voltages information on cell morphology, including in particular stress fibres and cell nuclei, where heavy metals were intensely bound became more evident. The technique allowed stepwise ‘sectional’ information to be acquired. The technique should be useful for studies on cell morphology, cycle and adhesion with greater resolution than can be obtained with any light-microscope-based system.     

Lymphatic networks in the periodontal tissue and dental pulp as revealed by histochemical study.

The structural organization and fine distribution of the lymphatic networks in the periodontal tissues (gingiva, periodontal membrane, and alveolar process) and dental pulp of animals and humans were reviewed with special reference to histochemical examination by light and electron microscopy. The distinction between lymphatics and blood vessels was made on cryostat sections of undecalcified and calcified teeth treated with EDTA solution and whole mount preparations of periodontal membranes using 5'-nucleotidase (5'-Nase)-alkaline phosphatase (ALPase) double staining. This staining procedure allowed lymphatic vessels in the periodontal tissue and dental pulp to be differentiated from blood vessels. The specificity and localization of the enzyme reactions were confirmed by comparative histochemical studies of the same specimen with light microscopy and scanning or transmission electron microscopy. Well-developed 5'-Nase-positive lymphatic networks were observed on the tissue sections and whole mount preparations of the gingiva, periodontium, and dental pulp. More lymphatic vessels were seen in the root area of the periodontium than in the cervical area. In the dental pulp, lymphatic vessels were more numerous in the central part than in the peripheral odontoblastic layer. These distributions of the lymphatic capillary networks are discussed in relation to their ability to supply lymph to the teeth.     

Low-temperature X-ray microanalysis of the differentiating vascular tissue in root tips of Lemna minor L

The fracture faces of bulk-frozen tissue offer a number of advantages for the analysis of diffusible elements. They are easy to prepare, remain uncontaminated, and, unlike most frozen-hydrated sections, can be shown to exist in a fully hydrated state throughout examination and analysis. Root tips of Lemna minor briefly treated with a polymeric cryoprotectant are quench frozen in melting nitrogen. Fractures are prepared using the AMRAY Biochamber, lightly etched if necessary to reveal surface detail and carbon coated while maintaining the specimen at 110 K. The frozen-hydrated fracture faces are analysed at 110 K using the P/B ratio method which is less sensitive to changes in surface geometry and variations in beam current. The method has been used to investigate the distribution of seven elements (Na+, Mg⁺⁺, P, S, Cl^?, K+ and Ca⁺⁺) in the developing vascular tissue of the root tip. The microprobe can measure relative elemental ratios at the cellular level and the results from this present study reveal important variations in different parts of the root. The younger, more actively dividing cells, appear to have a slightly higher concentration of diffusible ions in comparison to the somewhat older tissues which have begun to differentiate into what are presumed to be functional vascular elements.     

A preparation method for observing intracellular structures by scanning electron microscopy

In order to observe intracellular structures by scanning electron microscopy, excess cytoplasmic matrix must be removed from the fractured surface of cells. Previously we reported an Osmium-DMSO-Osmium method devised for this purpose. This method is very effective in revealing intracellular structures, but requires osmium tetroxide for initial fixation with some consequent disadvantages. In the present study, a revised Osmium-DMSO-Osmium method is reported, in which an aldehyde mixture is used as the initial fixative instead of osmium tetroxide. As fixation is carried out by perfusion in this revised method, better preservation of fine structures is achieved than by the original method, especially in the central nervous tissue which tends to suffer from post-mortem degeneration. Moreover this method can be applied to cytochemical studies of intracellular structures with a scanning electron microscope (SEM). In this study, acid phosphatase of lysosomes is demonstrated in a coloured SEM micrograph.     

基于小波系数Hu矩的生物组织损伤监测方法

提出了一种基于B超图像的小波系数Hu矩特征值并结合支持向量机监测生物组织损伤的方法。利用高强度聚焦超声(HIFU)对新鲜离体猪肉组织进行辐照,实时获取辐照前后的B超图像,并对其进行预处理获取减影图像。提取减影图像的Hu矩、小波系数均值和基于小波系数的Hu矩3个特征值,分别利用支持向量机对生物组织样本进行学习、分类处理。结果表明:小波系数Hu矩特征值比Hu矩和小波系数均值的总辨识率分别高出2.70%和2.05%,从而可以更有效地监测HIFU治疗中生物组织损伤情况。该方法可以帮助临床医生客观地监控HIFU治疗过程,对提高HIFU疗效有实际意义。     

Structural heterogeneity and its effects on the properties of an in situ TiB2–TiC/Cu composite synthesised by hot pressing

ABSTRACT

In situ TiB₂ and TiC reinforced copper matrix composites with tailored heterogeneous structure were fabricated via high-energy ball milling of Cu, TiH₂ and B₄C powders followed by hot pressing. The microstructures of both ball-milled powders and hot-pressed composites were compared. Although the dislocation density of Cu matrix was changed after hot pressing, the mode of distribution of ceramic phases in the Cu matrix was noted to transmit from the ball-milled powders to hot-pressed composites in case of the TiH₂ particles synthesised by the in situ reactions. The structural inheritance between the ball-milled powders and hot-pressed composites could be used to control microstructural features and thus to tune properties. The hot-pressed TiB₂–TiC/Cu composites with tailored heterogeneous structure exhibited better performance than those of homogeneous counterparts.