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PubMed1974

Biochemical and morphological characterization of azurophil and specific granules of human neutrophilic polymorphonuclear leukocytes.

Postnuclear supernates from homogenates of purified neutrophil polymorphonuclear leukocytes (PMNs) from human blood were fractionated by zonal sedimentation and isopycnic equilibration in sucrose gradients. The fractions were characterized biochemically by measuring protein content and the activities of eight enzymes. Selected fractions were further analyzed by electron microscopy. In both centrifugation systems, azurophil and specific granules could be resolved almost completely. Azurophil granules sediment three to four times faster than the specifics and have an average density of 1.23. They contain all the peroxidase of the cells, large portions of four lysosomal hydrolases, and about half of the total lysozyme, and therefore appear to be, in biochemical terms, very similar to the azurophil granules of rabbit PMNs. The specific granules, which have an average density of 1.19, contain the remaining half of the lysozyme but appear to be free of the other components of the azurophil granules, and of alkaline phosphatase. Isopycnic equilibration disclosed a minor lysosomal population, which strongly overlaps the specific granules, and made possible the identification of a membrane-fraction which is characterized by the presence of the thiol-sensitive acid 4-nitrophenyl phosphatase and of alkaline phosphatase.

Acid Phosphatase↗
PubMed2003

Morphological characterization of photochemical graded spinal cord injury in the rat.

This study characterizes the histological and immunohistochemical changes in the adult rat spinal cord following photochemically induced spinal cord lesions. The spinal cord was exposed by laminectomy (T12-L1 vertebrae) and bathed with 1.5% rose bengal solution for 10 min. The excess dye was removed by saline rinse and the spinal cord was irradiated with "cold" light for 0, 1, 2.5, 5, and 10 min in different groups of rats. After 15 days a graded loss of spinal tissue was observed according to photoinduction times. Animals irradiated for 1 min showed spinal cavities involving the dorsal funiculi. The cavity became progressively larger, involving dorsal horns in animals irradiated for 2.5 min, together with the dorsolateral funiculi in animals irradiated for 5 min and the ventrolateral funiculi in those irradiated for 10 min, with loss of gray matter in these three groups. Changes in GFAP-, CGRP-, proteoglycan- and calbindin-immunoreactivity were observed in all lesioned groups when compared with control spinal cords. Hypertrophied and heavily GFAP- and proteoglycan-stained astrocytes were seen in irradiated spinal cords. Reactive microglial cells were also found. Both astroglial and microglial reactions paralleled the severity of the spinal cord lesion. A significant loss of CGRP-immunoreactive somas was seen in animals irradiated for 10 min, whereas the wider distribution of calbindin-positive neurons was found in lesioned rats. In spinal cord sections from animals illuminated for 5 min and perfused 60 min postillumination, light and electron microscopy showed cytotoxic edema with astrocytic swelling, red blood cell extravasation, and myelin degradation.

Animals↗
PubMed1988

The effect of traumatic brain injury on the visual system: a morphologic characterization of reactive axonal change.

Reactive axonal changes following fluid-percussion brain injury were studied in the highly organized afferent pathways of the cat visual system. The visual system offers several features advantageous to the study of traumatic brain injury. Specifically, as the retinal cells of origin of the optic nerve and tract are isolated from the employed fluid-percussion injury, concomitant traumatically induced somatic change is not a confounding variable. Additionally, the existence of axons of varying diameters and topographic localization within the visual pathways allows the relationship between both fiber size and distribution and their predilection for traumatically induced change to be considered. Since the visual pathway is a highly organized sensory pathway, data obtained in this system can be compared with similar studies previously conducted in motor systems. In these experiments, 20 adult cats were subjected to brain injury and killed at posttraumatic survival periods ranging from 2 to 60 days. Six cats received intravitreous injections of horseradish peroxidase (HRP) to aid in the recognition of axonal change. At the designated survival time, all animals were perfused with aldehydes. The visual system from optic chiasm to lateral geniculate was sectioned and prepared for routine light (LM) and electron microscopic (TEM) study. Animals injected with HRP were processed for the LM and TEM visualization of HRP reaction products. By the second posttraumatic day, reactive axonal swellings were observed in the optic tracts as they entered the medial intralaminar nuclei of the lateral geniculate bodies. Proximal segments of the reactive axons showed enlargement and lobulation, whereas the distal segments underwent wallerian degeneration. Over a 2 week posttraumatic course, some axonal swelling persisted unchanged, some degenerated, and others initiated regenerative sprouting. with continued survival, however, all the reactive swellings manifested only progressive degenerative change. These reactive axonal changes appeared to constitute a primary response to the traumatic episode and occurred without concomitant damage to either the related brain parenchyma or its intrinsic vasculature. Although these findings replicate many of those previously described in motor pathways, new conceptual information has been provided. These studies preclude concomitant somal damage as a confounding variable and suggest that large-caliber axons are susceptible to the shear and tensile forces of traumatic brain injury.

Animals↗
PubMed1980

Ultrastructural analysis of pancreatic carcinogenesis: morphological characterization of N-nitroso-bis(2-hydroxypropyl)amine-induced neoplasms in the Syrian hamster.

Tumours of the exocrine pancreas of the Syrian hamster, induced by lifetime weekly injections of N-nitroso-bis-(2-hydroxypropyl)amine(BHP), were studied by light and electron microscopy and their fine structure was characterized. The neoplasms were cystic and papillary adenomata and adenocarcinomata, whose development was preceded by that of cystic foci of flattened epithelial cells. The tumour epithelia consisted of flattened, cuboidal or columnar cells, the latter frequently showing mucous metaplasia. Although there was a considerable degree of ultrastructural pleomorphism within tumours, some of which was apparently related to maturation, all the neoplasms shared sufficient common cellular characteristics to indicate a common histogenesis. Although the neoplasms induced by BHP closely resembled human "ductal" pancreatic tumours, they did not appear to arise from the ducts.

Adenocarcinoma↗
PubMed1993

Functional and morphological characterization of 4F7+ spleen accessory dendritic cells.

Recently we have reported on the production of the mAb 4F7. This recognizes a molecule that is upregulated on dermal and epidermal dendritic cells after application of contact allergen. Furthermore, this antibody detects an antigen on spleen and lymph node dendritic cells. In this study, we characterize 4F7+ spleen dendritic cells and show that the mAb recognizes in situ few labeled cells in the white pulp of the spleen and approximately 1% of spleen single cell suspensions as evidenced by cell enrichment, immunoperoxidase staining and FACS analysis. Immunohistological characterization of the cells with mAbs revealed the expression of class II, class I MHC antigens, 33D1, CD11c, ICAM-1, and CD45 molecules. After enrichment and cultivation for approximately 3 days, these cells showed no adherent properties. The capacity of 4F7+ spleen dendritic cells to activate allogeneic T cells in the primary mixed lymphocyte reaction was similar to freshly isolated Ia+ Langerhans cells. With regard to the induction of a proliferative response of CD4+ naive T cells that were incubated with concanavalin A or anti-CD3 mAb, 4F7+ spleen dendritic cells were two to three times more potent than spleen microphages and B cells. Furthermore, 4F7+ cells efficiently stimulated the antigen dependent proliferation of a T helper cell line. The mAb 4F7 will be useful for the purification of dendritic cells and for functional and molecular biological studies.

Animals↗
PubMed1990

Biochemical and morphological characterization of the intra-acrosomal antigen SP-10 from human sperm.

The human sperm protein SP-10 was previously defined as a "primary vaccine candidate" by a World Health Organization Taskforce on Contraceptive Vaccines. By one- and two-dimensional immunoblots, we show that SP-10, extracted from ejaculated human sperm, demonstrated a polymorphism of immunogenic peptides from 18 to 34 kDa, a pattern that was conserved from individual to individual and was not altered by reducing agents. The majority of the antigenic peptides possessed isoelectric points of approximately 4.9. Immunocytochemistry on testis sections indicated that SP-10 was localized to round spermatids and spermatozoa within the adluminal compartment of the seminiferous epithelium. Immunofluorescence showed that SP-10 was not associated with the surface of acrosome-intact, ejaculated sperm. Light and electron microscopic immunocytochemistry localized SP-10 throughout the acrosome, and electron microscopic evidence demonstrated a bilaminar array in association with the inner aspect of the outer acrosomal membrane and the outer aspect of the inner acrosomal membrane. After induction of the acrosome reaction with the ionophore A23187, SP-10 remained displayed on the sperm head in association with the inner acrosomal membrane and equatorial segment. The results indicate that the MHS-10 monoclonal antibody may be used as a marker of acrosome development in the human and as a probe to evaluate acrosome status. The results also support the hypothesis that inhibition of sperm-egg interaction by anti-SP-10 monoclonal antibody may occur as a result of antigen exposure following the acrosome reaction.

Acrosome↗
PubMed1986

Colloid keratosis. Morphologic characterization of a nonspecific reaction pattern of squamous epithelium.

Colloid keratosis is characterized by homogeneous eosinophilic masses of variable size and number within the upper layers of squamous epithelia, including epidermis. It has been observed as the characteristic feature of many onychoses and inflammatory conditions of oral epithelium, and as an incidental finding in neoplastic and nonneoplastic lesions in the skin and respiratory tract. Its nature remains obscure, but knowledge at present suggests that it may represent a disorder of an early phase of keratinization. Current evidence supports the hypothesis that colloid keratosis represents a nonspecific cellular reaction pattern of squamous epithelium.

Adult↗
PubMed2001

Morphological characterization of Aspergillus nidulans: growth, septation and fragmentation.

The influence of the sepA gene on the growth of Aspergillus nidulans has been investigated by characterizing and comparing the parental strain A28 (pabaA6 biA1) with the sepA null mutant (sepA4DeltaBM:). The sepA gene is known to affect the septation process in A. nidulans, therefore the sepA4DeltaBM: strain does not produce any septa during the first hours of growth. During batch cultivations sepA4DeltaBM: shows an abrupt decrease in specific growth rate and more pronounced fragmentation (in response to elevated stirrer speed) than the parental strain. Higher specific fragmentation rates (q(frag)) were obtained for the sepA4DeltaBM: strain. The physiological reasons for the differences have been investigated by employing fluorescent stains. Computerized image analysis revealed that the more pronounced fragmentation in the mutant was due to the lower number and irregular spacing of septa (visualized by calcofluor white staining), which resulted in a weaker hyphal structure that is more vulnerable to shear stress and fragmentation than the parental strain. This led to a loss of active biomass (determined by Mag fura staining) from the hyphae of the mutant, which had failed to compartmentalize by formation of septa, in turn resulting in decreased specific growth rates for the culture.

Aspergillus nidulans↗
PubMed1979

Studies on the plasma membrane of normal and psoriatic keratinocytes. I. Preparation of material and morphological characterization.

A method is described for the preparation of isolated keratinocytes suitable for subsequent biochemical studies. Scanning electron microscopy showed that the maturation process is accompanied by an increase in cell size and a shortening and eventual loss of microvilli. Psoriatic keratinocytes are distinguishable by exhibiting longer microvilli at all levels of maturation.

Adolescent↗
PubMed1993

Biochemical and morphological characterizations of DU-145 cell mortality in rabbit embryo-fetal fluid.

Rabbit embryo-fetal fluid (EFF) contains regulatory factors of cell proliferation which increase the duration of the cell cycle, induce a quiescent status in some cells and lead up to cell death in others. The objective of this study was to demonstrate which of the two processes, namely necrosis or apoptosis, was responsible for the cell death. Inhibitors of protein synthesis, and nuclease and phospholipase A2 activities did not restore the viability of the cells treated with EFF. Using a combination of DNA labelling and extraction, it was possible to show that a large proportion of DNA was fragmented in the cells released in the supernatant while only a very small portion of DNA was fragmented in the monolayer cells. EFF did not induce fragmentation of DNA into nucleosome-sized subunits as analysed using polyacrylamide gel electrophoresis. Nevertheless, using cytofluorometric analysis, it was possible to demonstrate that 50% of the cells released in the supernatant contained a lower quantity of DNA per cell than in the control cells. This was also observed with EFF-treated monolayer cells but not in the control monolayer cells. The reduction of the DNA content per monolayer cell became significant at 48 h of treatment with EFF. Electron microscopic analysis did not reveal blebbing of the cells. However, depletion of glycogen, condensation of mitochondria and increasing number of lysosomes and residual bodies were observed upon treatment with EFF. From these experiments we conclude that the DU-145 cells treated with EFF do not die by apoptosis, but rather seem to die by necrosis.

Animals↗
PubMed2004

Morphological characterization and notes on the life cycle of a newly discovered variant of Loma salmonae (Putz, Hoffman & Dunbar) from a natural infection of chinook salmon, Oncorhynchus tshawytscha (Walbaum).

Two variants of Loma salmonae occur in net-pen reared chinook salmon, Oncorhynchus tshawytscha. The typical variant (OA) has a host specificity for salmonids of the genus Oncorhynchus whereas the atypical variant (SV) has a host specificity for brook trout, Salvelinus fontinalis, and in this study, the ultrastructure of the two are compared. In fish at 8 weeks post-exposure xenomas of the SV variant have a very high proportion of mature spores compared with other developmental stages, while in xenomas of the OA variant there are fewer spores and many other developmental stages. Spores of the SV variant had up to 20 turns of their polar tube whereas those of the OA variant only had 17. Furthermore, the spores of the SV variant were significantly larger than those of the OA variant. The sporophorous vesicle for both variants appears to form around a sporogonial plasmodia, which results in many spores developing within the vesicle.

Animals↗
PubMed1980

Morphological characterization of liposome suspensions by stereological analysis of freeze-fracture replicas from spray-frozen samples.

A method for size distribution analysis of liposome suspensions is described. Spray-freezing was used in order to avoid cryoprotectants and fixatives. Light scattering showed that, depending on their composition, liposomes can be modified by the spraying procedure. dipalmitoylphosphatidylcholine liposome suspensions display large modifications of light scattering upon spraying. The commonly used unilamellar phosphatidylserine/phosphatidylcholine (1:4) liposomes, with or without cholesterol, are, however, only slightly affected by the spraying procedure. For liposome suspensions stable upon spraying, size distribution analysis can be performed on freeze-fracture replicas by standard procedures and average size, area and volume of particles are obtained. The number of particles per unit volume cannot be determined by stereological procedures, since the fracture face does not correspond to a plane section through the sample. By combining stereological data with phospholipid mass and specific molecular area of phospholipids, abundance of particles, fractional volume and total membrane area can nevertheless be evaluated. Data on small unilamellar liposomes prepared by sonication are presented as an illustration of the method. In suspensions obtained by this procedure, vesicles may be considered at thermodynamic equilibrium in the aqueous phase. The experimental size distribution compared favourably with that obtained from self-assembly theory for liposomes, considering natural lecithin heterogeneity. For comparison purposes, data on large unilamellar liposomes, obtained by reverse-phase evaporation, are also presented.

Cholesterol↗
PubMed2005

Morphological characterization of receptor activator of NFkappaB ligand (RANKL) and IL-1beta expression in rodent collagen-induced arthritis.

Bone loss represents a major unsolved problem in rheumatoid arthritis (RA). The receptor activator of nuclear factor-kappaB ligand (RANKL) is essential for the development and activation of osteoclasts, which are key mediators of bone erosions. This study was performed to determine temporal and spatial expression of RANKL compared with the potentially destructive cytokine interleukin-1beta (IL-1beta), related to progression of synovitis and joint destruction in collagen-induced arthritis (CIA), a model of RA. CIA was induced in dark agouti (DA) rats, and tissue specimens were obtained for immunohistochemical analyses at various time points before and after disease onset. Arthritis was monitored visually, and joint pathology was examined histologically. No disease-preceding expression of RANKL was detected. However, a marked increase of both RANKL- and IL-1beta-expressing cells correlated with the progression of synovial inflammation and clinical disease severity. Abundant and concomitant expression of these cytokines was detected at sites of bone erosion, where a colocalization by osteoclast-like multinuclear tartrate-resistant acid phosphatase (TRAP)+ cells was noted. In contrast to the paucity of RANKL expression in cartilage, an abundant expression of IL-1beta was demonstrated, particularly in superficial cartilage layers. These data support the hypothesis that RANKL and IL-1beta are central contributors to joint destruction in CIA.

Animals↗