The Lectin-Gold Technique: An Overview of Applications to Pathological Problems

Lectins are proteins, mainly of vegetal origin, which recognize glycosidic residues with high specificity; for this property they have been used for many studies of molecular biology. The colloidal gold represents at present the most popular electron dense marker employed in immunocytochemistry, since it offers intrinsic and unique characteristics which are superior to those displayed by the other markers. The cytochemical method which utilizes the gold-labelled lectins takes advantages from both the two systems, in order to optimize the localization of the glycoconjugates. The present paper reviews both the technical aspects of the preparation of the lectin-gold complex and its application to some selected pathological problems. In particular, the papers concerning the eye and ear tissues, the urinary, reproductive, nervous and digestive systems and the blood cells are quoted

Scanning Electron Microscopy, X-Ray Microanalysis and Immunohistochemistry on Worn Soft Contact Lenses

The deposits accumulated on the surfaces of soft contact lenses are a cause of problems for the wearer of these lenses, as the deposits are never completely removed by the available washing solutions. Therefore it appears of interest to investigate the composition of these deposits. In this paper we review the major findings in the literature and, in addition, present our personal experience. We have studied new, continuously and daily worn soft contact lenses by scanning electron microscopy (SEM), X-ray microanalysis and immunohistochemistry. We have carefully evaluated preparative methods, and we can conclude that SEM and X-ray microanalysis are best carried out on unfixed, air-dried lenses. The deposits present consist mainly of mucus, especially on the tarsal side of the lenses. Chloride and potassium, coming from the tear fluid, as well as sulfur, derived from proteins, were found. Calcium was very rarely detected. IgG, IgA, IgE and C3c complement fractions were found only on the outer surfaces and not within the lens. We believe that the best characterization of the deposits is achieved by means of correlative techniques on the same lens. In fact, this approach integrates morphology and composition

Strategies to Mitigate the Effects of Soil Physical Disturbances Caused by Forest Machinery: a Comprehensive Review

Purpose of Review: Ground-based mechanized forest operations can cause severe soil disturbances that are often long lasting and detrimental to the health of forested ecosystems. To reduce these soil disturbances, focus is being increasingly directed at identifying and using appropriate mitigation techniques. This systematic review considered 104 scientific articles and reported the main findings according to four core themes: terrain-related factors, operational planning, machine modifications, and types of amendments used to mitigate machine-induced soil impacts. Recent Findings: For terrain-related factors, most severe disturbances occur on machine operating trails exceeding 20% slope and that soil bulk density and rut depth show greater increases in fine-textured soils. When considering operational planning, trafficability maps proved to be helpful in reducing the frequency and magnitude of soil damages as well as the length of trails needed within harvest sites, especially if they are regularly updated with weather information. Machine modifications, through high flotation tires, use of extra bogie axle, lower inflation pressure, and use of steel flexibles tracks, are highly researched topics because of the considerable upside in terms of machine ground pressure distribution and increased traction. Two main types of amendments emerged to mitigate soil disturbances: brush mats and mulch cover. Brush mats created from harvesting debris can spread the load of a machine to a greater area thereby lowering peak loads transferred to the soil. Brush mats of 15–20 kg m−2 are being recommended for adequate soil protection from harvesting operations. Summary: To conclude, we outline recommendations and strategies on the use of soil mitigation techniques within cut-to-length forest operations. New research opportunities are also identified and discussed. Considering single factors causing machine-induced soil disturbances remains important but there is a pressing need for having a multi-disciplinary approach to tackle the complex problems associated with machine/soil/plant interactions

Learning to stop: a unifying principle for legged locomotion in varying environments.

Evolutionary studies have unequivocally proven the transition of living organisms from water to land. Consequently, it can be deduced that locomotion strategies must have evolved from one environment to the other. However, the mechanism by which this transition happened and its implications on bio-mechanical studies and robotics research have not been explored in detail. This paper presents a unifying control strategy for locomotion in varying environments based on the principle of 'learning to stop'. Using a common reinforcement learning framework, deep deterministic policy gradient, we show that our proposed learning strategy facilitates a fast and safe methodology for transferring learned controllers from the facile water environment to the harsh land environment. Our results not only propose a plausible mechanism for safe and quick transition of locomotion strategies from a water to land environment but also provide a novel alternative for safer and faster training of robots

Characterization of Mucus Glycoconjugates in Normal Human Conjunctiva by Lectins in Light Microscopy, Transmission and Scanning Electron Microscopy

Maintenance of tear film in normal conditions is dependent on 1) mucus layer integrity and 2) the presence and distribution of conjunctival epithelial cell microvilli. In the present work a new methodology has been developed to gain correlative information about microprojection assessment and mucus composition, from the same specimen, by Light Microscopy (LM), Transmission Electron Microscopy (TEM) and Scanning Electron Microscopy (SEM). We have characterized the glycosidic residues secreted by goblet cells in normal human conjunctiva, by means of four lectins (WGA, ConA, PNA and SBA), conjugated with FITC for LM and with colloidal gold for TEM and SEM. The cytochemical reactions were performed on histological sections of paraffin-embedded material and on semithin and ultrathin sections of both Epon embedded material directly processed for TEM and of blocks recovered from SEM and reprocessed for TEM. WGA, ConA, PNA and SBA receptors were found to be constituents of the mucus produced by goblet cells in human conjunctiva. The granules of the so-called second mucus system (SMS) cells were labelled mainly by WGA. A difference in the quality of glycoconjugates between goblet cells and SMS cells has been also demonstrated. Our results provide an improved method to evaluate alterations of tear film that occur in many conjunctival diseases

Endothelialization of a New Dacron Graft in an Experimental Model: Light Microscopy, Electron Microscopy and Immunocytochemistry

Two types of synthetic vascular grafts, Dacron Triaxial and Dacron Gelseal Triaxial, were implanted into both the common carotids of sheep. The animals were sacrificed 1, 2, 8, and 16 weeks after surgery. Multiple specimens, obtained from grafts and anastomoses, were studied by light microscopy, transmission and scanning electron microscopy. A parallel immunocytochemical analysis was performed on some specimens. Dacron Triaxial grafts failed to develop a complete neointimal coverage. Myofibroblasts and fibroblasts were the dominant cells in such synthetic graft. Moreover, focal areas of stripping, platelet deposition, and thrombosis were observed at 8 and 16 weeks. In contrast, a stable endothelial coverage developed on the Gelseal Triaxial grafts after 16 weeks

Electron Microscopy of Lipid Deposits in Human Atherosclerosis

The filipin probe associated with tannic acid stain was used to study intra-and extracellular lipids in surgically removed human atherosclerotic lesions (n = 20). In particular, intimal thickenings, fatty streaks and fibrolipidic plaques have been investigated by using mainly transmission and scanning electron microscopy. In the intimal thickenings, the lipid deposits were mainly localized in the subendothelial space as homogeneously sized particles (40-140 nm) and more heterogeneous uni-multilamellar vesicles (35-700 nm). Intermediate lipid forms were also observed. In the fatty streaks, the lipid deposits were intracellular and mainly observed in cells with a monocyte/macrophagic phenotype. Lipid inclusions, lipid lysosomal bodies and intracellular cholesterol crystals very similar to those observed in experimentally induced atherosclerosis were documented. In the fibrolipidic plaque the lipid deposits were found both in the intracellular and in the extracellular compartments. Lipids accumulated within arterial macrophages and smooth muscle cells, usually as lipid droplets. Clusters of lipoprotein-like particles (50 nm in diameter) as well as larger uni-multilamellar lipids (700 nm) with an occasional compound appearance were particularly observed bound to elastic tissue and collagen fibers. These morphological observations outline the complexity of lipid metabolism in the various histological aspects of human atherosclerosis

Correlative Scanning Electron Microscopy in the Study of Human Gastric Mucosa

We studied two aspects of the human gastric mucosa: - the surface morphology of mucous cells, as viewed by scanning electron microscopy (SEM); - the glycosidic components of intracellular mucins, characterized by means of lectins. The latter were conjugated with fluorescein isothiocyanate and with colloidal gold-silver for the visualization of the reaction products in light microscopy (LM) and in SEM (backscattered mode) respectively. The surface morphology of mucous cells appears to be correlated to the secretory state.In gastric ulcers we found a prevalence of non-secreting cells. A decrease in glycosidic receptors for fucose-binding lectin and galactose-(l-3)-N-acetyl-galactosamine-binding lectin was also observed. This suggests the presence of an impaired mucus secretion which may play a role in the pathogenesis of gastric ulcer. Spiral bacteria, supposed to be aetiologically related to peptic ulcer and gastritis, were easily detected by SEM. Intestinal metaplasia defined complete in LM showed surface morphology and glycosidic components different from those of true intestinal mucosa. This implies the necessity of taking into account also these parameters when classifying this lesion. The same applies to polyps. Our data indicate that correlative SEM may contribute further information on the pathogenesis and pathology of gastric diseases

Healing of Prosthetic Arterial Grafts

Numerous synthetic biomaterials have been developed as vascular substitutes. In vitro, ex vivo and in vivo studies have demonstrated that in animals, selected materials, i.e., Dacron and ePTFE (expanded polytetrafluoroethylene) grafts, are successfully incorporated in both the large and the small caliber host arteries through a process which is generally referred to as graft healing. Morphologically, this process consists of a series of complex events including fibrin deposition and degradation, monocyte-macrophage recruitment and flow-oriented cell-layer generation, this last event being the complete endothelialization of the arterial substitute. In contrast to experimental animals, the flow surface of synthetic vascular grafts remains unhealed in humans, particularly in the small caliber conduits. Healing in man consists of graft incorporation by the perigraft fibrous tissue response with a surface covered by more or less compacted, cross-linked fibrin. It is therefore obvious that: i) marked differences in graft healing exist between animals and man; and ii) the usual mechanisms of graft endothelialization are partially ineffective in man. In order to guarantee the patency of synthetic vascular grafts for human small artery bypass, new strategies and approaches have recently been attempted. In particular, the endothelial cell seeding approach has been successfully accomplished in animals and is being experimented in human clinical studies. The problems and results of this biological approach are outlined in this paper