Alteration of cartilage matrix morphology with histological processing

An interlacunar network in the extracellular matrix of femoral head articular cartilage of neontal rats was seen by light microscopy to: (1) consist of elements, 0·5 μm thick, which occurred as individual elements, as bundles of elements, and as fused elements, (2) stain intensely with toluidine blue, methylene blue, and safranin O, and (3) connect chondrocytes by inserting on the chondrocyte capsules which were composed of morphologically and cytochemically similar material. By electron microscopy, the single elements were seen to be composed of thicker, denser staining areas of the honeycomb appearing matrix and the fused elements appeared as non-membrane bound channels containing granular material. Articular cartilage was processed using combinations of fixatives, dehydrating agents, and embedding media. Regardless of fixation, demineralization, or embedding, the network was not seen after dehydration of the cartilage with methanol, ethanol, acetone or tert-butanol but was seen after dehydration with aqueous solutions of glycol methacrylate, propylene oxide, 2-propanol or 2,2-dimethoxypropane. Network visualization following a variety of methods demonstrated that no single fixative, dehydrating agent, or embedding medium caused its formation. The presence of the network in different cartilage zones, its consistent morphology by light and electron microscopy, the uniformity of the elements in their connection with the chondrocytes, and presence in fresh-frozen sections suggest the network may be real, but rigorous evidence for its existence in vivo is still required. Since cartilage morphology was altered by histological methods, especially dehydration, common methods used in studying connective tissue matrix should be evaluated to determine their effect on matrix morphology.     

The effects of glycol methacrylate as a dehydrating agent on the dimensional changes of liver tissue

The dimensional changes of liver sections during the course of processing with glycol methacrylate (GMA) or with ethanol are described. Tissue processing with ethanol served as a control. During prolonged processing steps (24 h each), linear shrinkage of tissue specimens dehydrated with GMA at room temperature was 13.2%. Subsequent infiltration with GMA resulted in trivial swelling, and polymerization in slight shrinkage (2.3%). In comparison, processing with cold GMA resulted in shrinkage during dehydration (about 10.8%), a slight swelling in pure GMA, followed by shrinkage during polymerization (2.2%). Short routine processing schedules resulted in similar shrinkage/swelling patterns, although precise values differed slightly. In all experiments, ethanolic dehydration resulted in smaller dimensional tissue changes than did GMA dehydration. The dimensional changes of tissue sections during stretching on water, mounting and drying compensated for the major part of the shrinkage manifested during processing.     

The biological reality of the interlacunar network in the embryonic,cartilaginous, skeleton: A thiazine dye/absolute ethanol/LR White resin protocol for visualizing the network with minimal tissue shrinkage

Third toe phalanges of chicks aged 8–13 days in ovo and 7-day post-natal rat femoral growth plate were examined to determine whether the interlacunar network (IN), a structure with no lipoprotein membrane component or cytoplasmic organelles, is a genuine component of young growth cartilage. In chick phalanges dehydrated by 70% (v/v) ethanol and LR White resin, variable metachromatic staining of the interlacunar network by toluidine blue and red staining by picro-Sirius red indicate the presence of glycosaminoglycans and collagen. The network in phalanges dehydrated by 80% (v/v) ethanol appears little different; however, the network is much less widely detectable in phalanges dehydrated by 90% (v/v) ethanol and, after dehydration by absolute ethanol, is almost completely undetectable. In contrast, when the young cartilage is permeated by a thiazine dye such as toluidine blue, using a solution of dye in the aldehyde fixative, the network is widely detectable, following dehydration by absolute ethanol, both in chick phalanges and in rat growth plate. Comparison of projected areas shows that the extent to which whole chick feet are found to have shrunk, by the time that they are photographed under LR White resin, is determined principally by the extent of dehydration, by 70% (v/v) or absolute ethanol; post-shrinkage areas are 33% or 35% of areas measured in buffer for 70% (v/v) ethanol/LR White resin and 71% or 75% for absolute ethanol/LR White resin (the higher value in each is for the toluidine blue treatment). The network is thus present in radically shrunk tissue, but, significantly, is also fully represented in tissue shrunk by only a conventional margin and is therefore not produced as an artefact by exceptional tissue shrinkage as has been suggested.     

Effect of the polymeric cryoprotectant dextran on fluid secretion in the isolated rabbit pancreas

Physiological effects of the polymeric cryoprotectant dextran on an ion-transporting epithelium were investigated. In the isolated rabbit pancreas, dextran caused inhibition of fluid secretion and an increase of the concentrations of Na⁺, K⁺ and Cl^? in the secreted fluid. Dextran did not affect the basal or pancreozymin-stimulated enzyme secretion. These effects of dextran can partially be explained by the fact that it is osmotically active and does not permeate through the epithelium. The effect of dextran on water transport can be compensated by lowering the ion concentrations in the solvent of the cryoprotectant. It is concluded that in cryoprotected ion-transporting epithelia the absolute ion concentration values obtained by X-ray microanalysis of frozen-hydrated specimens may not be completely correct, but that valid conclusions about intracellular ion distribution may still be drawn.     

Practical considerations in the use of polarized light microscopy in the analysis of the collagen network in articular cartilage

Polarized light microscopy is a traditional method for visualizing the collagen network architecture of articular cartilage. Articular cartilage repair and tissue engineering studies have raised new demands for techniques capable of quantitative characterization of the scar and repair tissues, including properties of the collagen network. Modern polarized light microscopy can be used to measure collagen fibril orientation, parallelism, and birefringence. New commercial instruments are computer controlled and the measurements are easy to perform. However, often the interpretation of results causes difficulties, even errors, because the theoretical aspects of the technique are demanding. The aim of this study was to describe the instrumentation and properties of a modern polarized light microscope, to point out some sources of error in the interpretation of the results, and to recall the theoretical background of the polarized light microscopy.     

选煤厂PLC应用系统设计选型浅析

以近几年实践经验为基础,对选煤厂基于PLC的控制系统进行了分析和研究,探讨了选煤行业PLC应用系统的设计选型原则,期望能推动行业自动化系统设计整体水平再上新台阶.     

The use of freeze-substitution in the preparation of plant tissue for ion localization studies

A method, utilizing freeze substitution, is described for the preparation of plant tissue for analytical electron microscopy. The fine structure of the cytoplasm was adequately preserved after freezing leaf tissue in 2-methylbutane at ?170°C. Furthermore, following substitution in ether, losses of sodium and potassium from the tissue were less than 4% of the original ion content and the loss of chloride was less than 1%. The merits of the procedure as a means of tissue preparation for ion localization studies are discussed.     

Treatment of banana fiber for use in the reinforcement of polymeric matrices

Recent studies on the use of vegetable fibers in polymers have led to the conclusion that the fibers must be treated to improve their adherence to plastic matrices. The present study investigated the effect of physical and chemical treatments on the suitability of fiber from the Canary banana tree for use as reinforcement for polymers in injection molding processes. This fiber has the advantage of being derived from the vegetable waste that is produced by farms involved in banana cultivation in the Canary Islands. Sodium hydroxide and maleic anhydride were used to treat the fiber under different conditions of pressure and temperature, and then the fiber was examined by thermogravimetric analysis and Fourier transform infrared spectroscopy. The best treatment for improving the thermal properties of banana fiber, with no significant decrease in mechanical behavior, for use in a composite was a combination of 1 N NaOH and saturation pressure.     

Preservation of ultrastructure and immunoreactivity in cryosections of brain tissue stored in a sucrose-gelatin solution at freezing temperatures

We evaluated the preservation of ultra-structure and immunoreactivity in cryosections of central nervous system tissue mounted with and stored in a sucrose–gelatin solution for one month at −20°C or −80°C. The ultra-structure of synaptic structure in these sections was well preserved and comparable to that of freshly cut cryosections. Quantitative analysis of mitochondrial ultra-structure demonstrated gradually lower degrees of preservation in sections stored at −20°C and −80°C compared with that in freshly cut sections. We observed distinct metabotropic glutamate receptor 1 (mGluR1)-immunogold labelling at peri-synaptic sites in freshly cut sections and also in those stored at −20°C and −80°C. Quantitative analysis of mGluR1 immunoreactivity revealed that the total number of immunogold particles per synapse and the number of non-specifically bound particles were similar under all three conditions. However, the percentage of gold particles bound to a specific synaptic region was greatest in freshly cut sections (79.0%) and progressively lower in sections stored at −20°C (76.1%), in which sections were not frozen, and in sections stored at −80°C (68.0%). These data indicate that ultra-thin cryosections may be conveniently stored in a sucrose–gelatin solution at −20°C for cryoultramicrotomy-immunolabelling.     

Staining methods for the detection of laticifers in plant tissue

Simple, easy and inexpensive methods have been evolved to detect laticifers in plants with the help of bright-field and fluorescence microscopy using Oil Red O, dansyl chloride, neutral red, Rhodamine B and Ponceau S stains.     

Countercurrent plunge cooling: A new approach to increase reproducibility in the quick freezing of biological tissue

Conditions fulfilling all the accepted criteria for optimum cryoquenching are realized for the first time in a plunge freezing device. Samples are plunged down through a smoothly rising cryogen stream that is maintained within ± 0.25 K of a preselected temperature. Dry nitrogen gas at ambient temperature removes evaporated cryogen molecules from the plunging path immediately above the liquid coolant to prevent pre-cooling of the sample. Symmetrical instrument design and conditions of flow ensure symmetrical freezing under optimum conditions in a totally repeatable fashion. The device has been used with halocarbons, propane and ethane. It provides consistent cooling rates comparable with the best so far obtained by any other plunging method.     

The use of rhodamine 6G and fluorescence microscopy in the evaluation of phospholipid-based polymeric biomaterials

A technique is described that allows the staining and subsequent visualization of polymers that contain the phosphorylcholine (PC) group. These materials are useful as bulk materials or coatings for the fabrication of medical devices. The staining method employs rhodamine 6G, which can be simply and rapidly applied to the polymer coating and imaged using fluorescence microscopy. The specificity of the staining for the PC polymers makes this technique suitable for the evaluation of a wide range of substrates and provides qualitative information on coating uniformity, coverage and morphology. It can be used to examine the durability of, and defects in, the coating. Statistical analysis of the fluorescent intensity by measuring the pixel value during imaging can allow for the method to be used as a quality control tool.     

Advantages of the use of intact plant tissues in freeze-fracture electron microscopy

The use of whole, intact plant tissue for freeze-fracture electron microscopy provides important information that cannot be obtained from the use of isolated biological membranes or of artifical (phospholipid) membrane preparations. This is not to imply that these exminations of such preparations are not useful, since it would be difficult to interpret our observations of intact cells and tissues without the analysis of these model systems. Analysis of intact tissue and cells reveals the relative densities of membrane proteins of the different membranes within a cell; the three-dimensional organization of various organelles, especially the endoplasmic reticulum; changes in intramembranous particle (IMP) distribution due to stress or injury; and, in conjunction with the use of filipin, membrane sterol content and relative distribution. It is our intention that this survey of freeze-fracture images of intact plant tissues will illustrate the uniqueness of the information gained from an analysis of whole plant tissues compared to isolated membrane fractions.     

Histochemical evidence of the initial chondrogenesis and osteogenesis in the periosteum of a rib fractured model: implications of osteocyte involvement in periosteal chondrogenesis

We have examined cellular events at the early stages of periosteal chondrogenesis and osteogenesis induced by bone fracture, using a well-standardized rib fracture model of the mouse. The initial cellular event was recognized as considerable proliferation in the deeper layer referred to as the "cambium layer" of the periosteum, as evidenced by numerous proliferating cell nuclear antigen-positive cells. The periosteal cartilage and bone were then regenerated directly from the region of the most-differentiated cell, i.e., mature osteoblasts of the cambium layer both close to and distant from the fracture site. Therefore, periosteal osteoblasts appeared to have the potential to differentiate into chondrogenic and osteoblastic lineages. CD31-positive blood vessels were uniformly localized along the periosteum that was regenerating cartilage and bone, being therefore indicative of less influence on the initiation of osteochondrogenesis. In contrast, however, the regenerated periosteal cartilage or bone extended from the cortical bones included dead or living osteocytes, respectively. Empty lacunae and lacunae embedded with amorphous materials were found close to the regenerated cartilage, while intact osteocytes persisted adjacent to the regenerated bone. The embedded lacunae with amorphous materials would render the tissue fluid, nutrients, oxygen, and several secretory factors such as dentin matrix protein-1 impossible to be delivered to the periosteal osteoblasts that interconnect osteocytes via gap junctions. Our study thus provides two major clues on initial cellular events in response to bone fracture: the potentiality of periosteal osteoblastic differentiation into a chondrogenic lineage, and a putative involvement of osteocytes in periosteal cartilage and bone regeneration.     

Some limitations of the use of tannic acid as a marker of damaged skeletal muscle fibres

Mouse biceps brachii muscles containing single transverse incisions were treated with low molecular weight tannic acid. In the absence of fibre degeneration, intracellular spread of tannic acid was confined to a short distance along the fibres from the damaged portions of the cell membranes. Beyond this, damaged and undamaged fibres could not be distinguished from one another. Tannic acid is not a reliable marker of localized trauma to skeletal muscle fibres.     

现场总线技术在火电厂的应用前景探讨

本文以现场总线基金会组织开发的FF总线为例,从网络结构、网络设备特性、通信技术、系统组态等多个方面对现场总线技术作了介绍,并通过与当前被广泛应用的DCS系统及PLC构成的控制系统的比较,从网络安全及经济性两个方面对现场总线技术在火电厂的应用前景作了探讨,说明了现场总线的优越性及在火电厂应用的光明前景。     

净水厂最佳投药量的神经网络控制系统

混凝剂的投加是净水厂水处理工艺的重要环节，本文通过对目前水厂实际运行过程中混凝剂投加量的影响因素的分析，利用神经网络预测理论，建立了净水厂混凝剂投加量的预测模型，并利用该模型对某水厂的实际运行情况进行了预测，对网络预测模型的性能进行了验证。结果表明，该模型具有很强的自学习性，自适应性和容错性，依靠网络的在线自学习，可使预测结果的准确度明显提高。     

Control of chemical processes using neural networks: implementation in a plant for xylose production

This work demonstrates the use of artificial intelligence for control of xylose reactor performance in a paper factory. Two types of neural networks are used, a perceptron for the temperature controller and an adaptive formulation for the noise filter. The results show an improvement in the temperature stabilization time with respect to a classic PID control.     

New elements in the interreceptor matrix: a comparative study of Megavilli and Landolt's club

Megavilli and Landolt's clubs were reported in the interreceptor matrix of the retina of the goldfish and neonatal rat for the first time. Megavilli encroach onto the inner or outer segments of photoreceptors and were different from microvilli. Landolt's clubs were not present in the adult rat but only in the adult goldfish.     

Ultrastructural localization of nucleic acids in plant tissues following the use of malachite green or neutral red in the fixative solution

Malachite green and neutral red, when added to glutaraldehyde for fixation of various tissues, yielded high-contrast images of cell ultrastructure. Malachite green, in acid conditions, appeared to increase contrast of heterochromatin material in the nucleus whereas neutral red gave greater clarity to the nucleolus and to cytoplasmic ribosomes. Control tissue fixed under acid conditions showed little damage but there were ‘crystalline’ areas at the periphery of the nucleolus. RNase did not digest cytoplasmic ribosomes from tissue after neutral red glutaraldehyde fixation. These results suggested that neutral red became bound to RNA in the tissues. Fixation with malachite green, at a pH below 6, did not affect the digestion of RNA by RNase but did protect chromatin against the bleaching action of the chelating agent EDTA. The addition of malachite green (pH < 6) or neutral red to glutaraldehyde are useful techniques for the investigation of the ultrastructure of nuclear material and cytoplasmic ribosomes.