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PubMed1979

Immunocytochemistry with osmium-fixed tissue. I. Light microscopic localization of growth hormone and prolactin with the unlabeled antibody-enzyme method.

Growth hormone and prolactin were localized on thin plastic sections of rat anterior pituitary gland and mammosomatotropic tumor MtTW15 that were fixed with osmium tetroxide (alone,mixed with aldehydes, or after aldehydes). Intense immunocytochemical staining for both antigens was obtained after plastic was removed from sections with an alcoholic solution of sodium hydroxide. The results indicated that antigenic determinants of rat prolactin and growth hormone were not completely destroyed or inactivated by fixation with osmium and embedment in epoxy resin, and that removal of the polymerized epoxy resin was necessary to obtain light microscopic postembedding immunocytochemical staining of these antigens. The results also demonstrated that tissues which have been conventionally processed for morphological evaluation by electron microscopy may be suitable for postembedding immunocytochemical staining of some antigens for light microscopy.

Animals↗
PubMed1993

Morphological plasticity of synaptic mitochondria during aging.

A morphometric investigation has been carried out on the synaptic mitochondria of cerebellar glomeruli in young, adult and old rats by means of a computer-assisted image analysis technique. Mitochondrial volume density (Vv), numerical density (Nv), average volume (V) and average length (Skeleton = Sk) were investigated in tissue samples fixed, embedded and sectioned according to conventional electron microscopic methods. Vv was unchanged in the three groups of age taken into account. Nv was significantly increased in adult vs. young animals, whereas it was decreased in the old group as compared to both the other two groups investigated. V and Sk showed the same age-dependent changes: they significantly decreased in the adult vs. the young and the old groups of rats while increased significantly in the old rats vs. both the adult and young animals. A percentage distribution of Sk demonstrated that in the old group 20.6% of the population of synaptic mitochondria accounts for elongated organelles (> 5 microns) as compared to 8.6% and 5.3% in young and adult animals, respectively. The present findings match the changes previously reported by us on the ultrastructure of synaptic contact zones both in rats and human beings, and support the idea of an age-dependent dynamic adaptation in the morphology of synaptic mitochondria to cope with the metabolic needs of the pattern of synaptic connectivity they subserve.

Aging↗
PubMed2004

A comparative study of crack development in stainless-steel hedstrom files used with step-back or crown-down techniques.

The purpose of this study was to evaluate the effect of step-back (SB) and crown-down (CD) techniques on crack development of stainless-steel Hedstrom files (H-files). Ninety-seven mesial root canals of freshly extracted human mandibular molars were selected. Six sets of H-files (#15, #20, and #25 ISO size each) were used with SB and CD techniques for the chemomechanical preparation of 8, 16, and 24 root canals using 2.5% NaOCl as an irrigant and EDTA gel as a chelating agent. Files were sterilized in an autoclave between successive uses. A seventh unused set served as control. After macroscopic examination all files were embedded in epoxy resin, ground, polished, and studied under a metallographic microscope. The maximum crack size of each file was measured by means of standard image-analysis procedures. Macroscopically the files presented no signs of plastic deformation, apart from a file #15, used with SB technique, which fractured after 16 root-canal preparations. The microscopic examination showed no cracks: on control files, on files used with the CD technique for 8 and 16 cycles, on file #20 used with the SB technique for 8 and 16 cycles, as well as on file 25 used with the SB technique for 8 cycles. Cracks were found on H-file #15 used with the SB technique for 8 and 16 cycles, on file 25 used with the SB technique for 16 cycles, and on all files used for 24 cycles with both SB and CD techniques. Under the conditions of this study it was concluded that the instrumentation technique is deeply implicated in the crack development. Crack propagation was accelerated when SB technique was used.

Chelating Agents↗
PubMed1988

Polarity and differentiation of human rectal adenocarcinoma cells in suspension and collagen gel cultures.

HRA-19a1.1. cells, derived from a primary human rectal adenocarcinoma, form polarized monolayers when grown on tissue-culture plastic. The apical membrane of the cells is in contact with the culture medium while the basolateral surface is attached to the plastic substratum. Cells cultured on non-tissue-culture plastic form floating colonies. Cells within these colonies are orientated so that their apical membrane is in contact with the culture medium while the basolateral membrane faces the centre of the colony. When these colonies are embedded in collagen gel the cells organize to form glandular structures similar to those observed in xenografts of this cell line. In addition a reversal in the orientation of cell polarity is observed with the basolateral membrane now in contact with the collagen gel, while the apical membrane faces lumina within the colony. This interaction is specific to collagen gel, as in a control experiment where colonies are embedded in agarose gel, neither a reversal in polarity nor the formation of glandular structures is observed. These results demonstrate an induction of glandular organization in human rectal adenocarcinoma cells by collagen gel. In addition an increased absorptive cell differentiation and reversal in cell polarity is observed in response to collagen gel.

Adenocarcinoma↗
PubMed2005

Efficacy of a Bacillus thuringiensis israelensis tablet formulation, vectobac DT, for control of dengue mosquito vectors in potable water containers.

VectoBac DT, a tablet formulation of Bacillus thuringiensis israelensis (Bti) was evaluated for the potential control of dengue vectors in various types of potable water containers. On introduction to containers, the tablet sinks to the bottom and the Bti toxins are found concentrated at the sides and the base, while the treated water column is free of Bti toxins within 24 hours after tablet introduction. In a simulated study, earthen, HDPE and plastic containers were kept covered and laboratory-bred larvae were introduced to determine the control by the tablet. The efficacy and persistence of the tablet, with a control of > 90%, was significantly longer in earthen containers in comparison to the HDPE and plastic containers. Efficacy and persistence were observed in earthen containers for a minimum period of 5.5 months (166 days) both without water replenishment and with weekly, 50% water volume, replenishment, and for a maximum period of 2.2 months (66 days) with daily, 50% water volume, replenishment. In plastic and HDPE containers, the tablet activity had a persistence of 2.1 months (63 days) without water replenishment and 1.8 months (54 days) with weekly water replenishment. The efficacy and persistence of the VectoBac DT was significantly longer in the earthen containers, with or without regularly treated water exchange, due to the Bti toxins being embedded in the porous earthen container surfaces, which protects them from rapid degradation. Lesser toxin amounts are removed from the water column during water exchange. The efficacy of VectoBac DT was also evaluated for the control of natural infestation of Aedes larvae which were resistant to temephos at the WHO diagnostic dosage of 0.012 mg/l. The tablet significantly reduced the pupal density by 8 fold in earthen containers for 67 days and 5 fold in HDPE containers for 55 days in comparison to untreated containers (p < 0.05). However, the tablet was effective for a shorter period of 25 days post-tablet-introduction due to fungal infestation in the treated plastic containers. There is a need to determine the capacity of the VectoBac DT to reduce the dengue vector population to a threshold which will prevent dengue outbreaks in dengue endemic areas.

Aedes↗
PubMed1984

Scanning and transmission electron microscopy as tools for the study of phagocytosis of bacteria adherent to hard surfaces.

A technique for studying the phagocytosis of bacteria colonizing hard surfaces is described. Rabbit peritoneal macrophages were allowed to settle on the surface of high-molecular-weight polyethylene which had been previously colonized by Escherichia coli. To ascertain the presence of bacteria on the surface of the polyethylene and the degree of spreading of the attached macrophages, the preparations were observed by scanning electron microscopy. The ingestion of E. coli by the macrophages was studied by transmission electron microscopy on ultrathin sections of resin-embedded monolayers after their separation from the polyethylene surface. Numerous intracellular bacteria were located near the area of attachment of the macrophages to the substrata, suggesting that the phagocytosis of bacteria adherent to the surface of the plastic had taken place.

Animals↗
PubMed1982

Unconventional application of standard light and electron immunocytochemical analysis to aldehyde-fixed, araldite-embedded tissues.

A simple procedure for the immunocytochemical analysis of glutaraldehyde/formaldehyde-fixed, Araldite- or Epon-embedded tissues by either light or electron microscopy is presented. Retention of immunoreactive antigen in deplasticized sections was achieved by use of a low concentration of glutaraldehyde in the fixative in combination with a seldom-used plastic solvent. This protocol produced good ultrastructural preservation in tissues and large, high-quality, 2-micrometers thick, plastic-free sections. These semithin sections provided a level of structural and antigenic preservation, image resolution, and labeling intensity that surpassed all other conventional sectioning methods used for immunocytochemistry. The capacity to use a single tissue sample in studies designed for light and electron immunocytochemistry, in conjunction with existing autoradiographic and cytochemical techniques, makes this a very desirable method for routine tissue preparation in research and clinical applications.

Animals↗
PubMedDate not supplied

A simple method for the preparation of paraffin-embedded cell blocks from fine needle aspirates, effusions and brushings.

A simple method for the preparation of paraffin-embedded cell blocks from cytologic specimens obtained by fine needle aspiration, by brushing or from effusions is described. The cells are fixed in suspension in 50% ethanol for one hour and pelleted by centrifugation in a 50-mL plastic tube. The fixative is removed, and the pellet is suspended in 3 mL of acetone for dehydration for ten minutes and thereafter repelleted. The acetone is then removed, and the cell pellet is dried at 60 degrees C for one hour. Melted paraffin is added onto the dry warmed cell mass and allowed to solidify at room temperature. A conical paraffin block with the cells in the top is obtained and can be handled as a routine tissue block.

Biopsy, Needle↗
PubMed1997

Synaptic plasticity of 5-hydroxytryptamine-immunoreactive terminals in the phrenic nucleus following spinal cord injury: a quantitative electron microscopic analysis.

The present study was conducted to examine the plasticity of 5-hydroxytryptamine (5-HT)-immunoreactive terminals in the rat phrenic nucleus following an ipsilateral C2 spinal cord hemisection and 30-day survival period. A retrograde horseradish peroxidase (HRP) labeling technique was used to identify the phrenic motoneurons at the electron microscopic (EM) level. After employing a pre-embedding immunocytochemical technique, the ultrastructural characteristics of 5-HT-immunoreactive terminals were qualitatively and then quantitatively analyzed with a computerized morphometric system before and after injury in separate groups of rats. The results indicated that the majority of the 5-HT-labeled terminals formed axodendritic contacts, but some 5-HT-labeled terminals made axosomatic contacts. 5-HT terminals were associated with either asymmetrical or symmetrical synapses, and some displayed postsynaptic dense bodies. Approximately 2% of the 5-HT terminals had dense-core vesicles. Although the total number of labeled and unlabeled terminals in the phrenic nucleus was reduced after hemisection, the number of 5-HT terminals in the hemisected group was greater than that of the control group. Moreover, the total number and length of asymmetrical and symmetrical synaptic active zones per 5-HT terminal were significantly greater after injury. Finally, the total number of 5-HT terminals with multiple synapses was significantly greater in the hemisected group as compared to controls. It is possible that 5-HT synaptic plasticity may be part of the morphological substrate for the unmasking of the latent crossed phrenic pathway which mediates recovery of the ipsilateral hemidiaphragm paralyzed by C2 spinal cord hemisection.

Animals↗
PubMed1984

The dystrophic murine skeletal muscle cell plasma membrane is structurally intact but "leaky" to creatine phosphokinase. A freeze-fracture analysis.

Skeletal muscle cells of genetically dystrophic mice (dy/dy) of the REJ-129 Bar Harbor strain exhibit reduced cytoplasmic levels of the enzyme creatine phosphokinase (CPK) when compared with normal (+/+) mice following SDS-gel electrophoresis of sarcoplasmic proteins. This observation has been thought to reflect "leakage" of CPK from dystrophic muscle cells through lesions in the sarcolemma. The present study has employed the freeze-fracture method to examine vast expanses of sarcolemma fracture face for determination of whether lesions do exist in the membrane or an alternate route is present for extravasation of CPK from dystrophic muscle cells. Most of the dystrophic cells examined in this study appeared intact and were therefore presumed viable. The intramembrane lipoprotein particles characteristic of PF-fracture face membrane were reduced in dystrophic as compared with normal murine skeletal muscle, and the plasmalemma possessed a greatly amplified population of caveolae as compared with nondiseased sarcolemma. No abnormal structural feature of these dystrophic muscle plasma membranes could be interpreted as a perforating focal "delta" lesion, such as the structures seen in thin plastic sections by other investigators. However, a second group of cells, generally few in number, that exhibited features indicative of necrosis (and loss of viability), were seen in both thin sections and platinum replicas. These moribund cells were usually embedded in dense sheaves of connective tissue along with other dystrophic cells that lacked signs of necrosis. The cytoplasm of the necrotic muscle cells was disorganized, as was the contractile machinery. The sarcolemma showed numerous perforations, through which CPK could escape into the tissue extracellular compartment. We conclude on the basis of our observations that the "focal lesions" reported by other investigators are not a structural feature of viable dystrophic muscle cell plasma membranes and are found only in necrotic or dying cells, and that the elevated serum levels of CPK associated with muscular dystrophy may result either from escape of the enzyme through lesions present in necrotic or dying cells or by extravasation along avenues provided by the hyperplastic mass of membrane caveolae present in dystrophic sarcolemma.

Animals↗
PubMed1997

Long-term results of hydroxyapatite-fibrin glue implantation in plastic and reconstructive craniofacial surgery.

The bone tissue formed in orthotopic or heterotopic implants of granular, porous hydroxyapatite and fibrin glue was examined several (2 1/2-8) years after implantation. The results showed distinct ossification in all cases and the functional situation and external appearance were also satisfactory. The reconstituted spongy and compact bone underwent a remodelling process similar to that of normal bone. Most of the hydroxyapatite granules were embedded in the bone tissue; a few were very close to the fibrillar connective tissue of the intertrabecular spaces and were either covered by osteoid-like collagenous borders or were in contact with osteoclast-like giant cells. Even if the results refer only to a few patients, and were obtained from a cross-sectional study, they allow the conclusion to be reached that the implantation of porous hydroxyapatite and fibrin glue leads to the formation of long-lasting bone whose hardness is equal to, or greater than, that of normal bone.

Adult↗
PubMed1980

Proliferation of epithelial cells in the adult primate choroid plexus.

An adult rhesus monkey was injected intraperitoneally with [H3] thymidine (2.3 microCi/gram body weight) and perfused 90 minutes later with a mixture of aldehydes. One and a half micrometer plastic sections were then cut and dipped into liquid emulsion for radioautography. Labeled cells were observed in the choroid plexus of the anterior lateral ventricle; cell identification was evaluated using electron micrographs taken from serial thin sections of re-embedded. radioautographic 1.5-micron sections. The ultrastructure and location of both mitotic figures and labeled cells confirmed the presence of undifferentiated basal choroid plexus epithelial cells in the adult primate central nervous system.

Animals↗
PubMed2002

Comparative genomic analysis of the MHC: the evolution of class I duplication blocks, diversity and complexity from shark to man.

The major histocompatibility complex (MHC) genomic region is composed of a group of linked genes involved functionally with the adaptive and innate immune systems. The class I and class II genes are intrinsic features of the MHC and have been found in all the jawed vertebrates studied so far. The MHC genomic regions of the human and the chicken (B locus) have been fully sequenced and mapped, and the mouse MHC sequence is almost finished. Information on the MHC genomic structures (size, complexity, genic and intergenic composition and organization, gene order and number) of other vertebrates is largely limited or nonexistent. Therefore, we are mapping, sequencing and analyzing the MHC genomic regions of different human haplotypes and at least eight nonhuman species. Here, we review our progress with these sequences and compare the human MHC structure with that of the nonhuman primates (chimpanzee and rhesus macaque), other mammals (pigs, mice and rats) and nonmammalian vertebrates such as birds (chicken and quail), bony fish (medaka, pufferfish and zebrafish) and cartilaginous fish (nurse shark). This comparison reveals a complex MHC structure for mammals and a relatively simpler design for nonmammalian animals with a hypothetical prototypic structure for the shark. In the mammalian MHC, there are two to five different class I duplication blocks embedded within a framework of conserved nonclass I and/or nonclass II genes. With a few exceptions, the class I framework genes are absent from the MHC of birds, bony fish and sharks. Comparative genomics of the MHC reveal a highly plastic region with major structural differences between the mammalian and nonmammalian vertebrates. Additional genomic data are needed on animals of the reptilia, crocodilia and marsupial classes to find the origins of the class I framework genes and examples of structures that may be intermediate between the simple and complex MHC organizations of birds and mammals, respectively.

Animals↗
PubMed1975

[Substitution of the tendon sheath by autologous vein in the "no-man's land" of the hand].

Reconstructive surgical interventions on the level of the proximal phalanges and performed after the lesions of the flexor tendons on the distal palm are discussed. In order to reduce the adhesions "bridging of the no man's land" by means of an autotransplanted vein is recommended by the authors. The vein-graft cannulated on a silicone bar is embedded within a few weeks, and after the removal of the silicone the "no man's land" is "over-bridged" by a pseudotendon sheath, stuffed with endothelium of good sliding surface. Thus, in the second sitting each of the plastic "bridge" methods (Bunnell, Paneva--Holevitch) may be performed. The operative technique and the first 4 cases are shortly discussed by the authors.

Hand Injuries↗
PubMedDate not supplied

Olfactory bulb lesions in Alzheimer's disease.

The olfactory bulb (OB), with its comparatively simple and well-delineated connectivity, presents an interesting system for examining cell-specific pathology in neurologic degenerative disorders such as Alzheimer's disease (AD). We have found that in AD the large, efferently projecting neurons (mitral cells) of the OB degenerate, typically without classical Alzheimer neurofibrillary changes. In some cases, with less severe neocortical pathology, the terminal arborizations of olfactory nerve appear hyperplastic and are associated with focal accumulations of A-4 (beta-amyloid) immunoreactivity that are not detectable by standard amyloid stains. These abnormalities may represent a pathologic manifestation of normally occurring plasticity in the olfactory system.

Aged↗