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A case of an unusual foreign body (a tip fragment from a plastic chopstick) embedded in one of two unerupted supernumerary teeth in the premaxillary region of a 12-year-old Japanese boy is presented.
Voluntary muscle activity in chick embryos was paralysed by administration in ovo of tubocurarine hydrochloride, administered in single or multiple doses, from day 9 to day 13 after fertilization. Control eggs were given saline instead of tubocurarine or were simply incubated without operative interference. Embryos were killed at 9, 13, 14, 16 and 19 days after fertilization. Flexor digitorum tendons were removed, fixed in glutaraldehyde, embedded in plastic, sectioned, and stained with phosphotungstic acid for electron microscopy. The diameters of the tendon collagen fibrils were measured, on electron micrographs, using a Magiscan Mk II programme. Tendon collagen fibril expansion was not inhibited by tubocurarine treatment. It is concluded that the rapid increase of collagen fibril diameters, which coincides in the normal embryo with the first onset of use of the associated muscle, is not dependent on muscle activity. There remains a possibility that other ways of producing tension in the tendon could provide sufficient stimulus to fibril expansion.
A spermatocele refers to the cystic accumulation of semen in the male reproductive tract. Although it is thought to be caused by narrowing of the lumen of the excurrent duct with resultant cystic dilatation of the duct, the pathogenesis of the narrowing remains unknown. In the present study, we histologically examined spontaneous spermatoceles in C3H/He mice to elucidate the pathogenesis of the lesions. Testes, efferent ducts, epididymides and vas deferens obtained from young and aged C3H/He mice were embedded in plastic for histological observation at the light microscopic level. It was found that spontaneous spermatoceles were localized in the rete testis and efferent ducts of aged mice, as seen in man. The dilated rete testis and efferent ducts contained many degenerated and aggregated germ cells derived from the exfoliated seminiferous epithelium in the aged testis. In particular, it was noted that the agglutinated germ cells obstructed the narrow lumen of the efferent ducts, resulting in the failure of transport of germ cells to the caput epididymis, and spermatoceles were consistently found in the region between the rete testis and the obstructed site in the efferent ducts. However, no inflammatory cell infiltration, traumatic injury or spermatic granulomas were found in the occluded region. These results suggest that agglutinated germ cells may occupy the narrow lumen of the efferent ducts, resulting in the formation of a spermatocele. It may be that a senile change to the seminiferous epithelium, which releases immature germ cells into the lumen of the seminiferous tubules, is the cause of this type of spermatocele.
The marginal integrity and microleakage of pressed glass ceramic inlays were evaluated using autoradiography. IPS/Empress ceramic inlays were fabricated for 10 human molar mandibular teeth. After adjusting the inlays, they were etched with 37% phosphoric acid gel for 30 s and silanized with Monobond S for 30 s. Before cementation with dual cure resin cement the inlays and cavity walls were gently covered with a thin layer of bonding agent. When the cementation process was completed the samples were cycled 300 times between a 55 degrees C hot bath and a 5 degrees C cold bath. The samples were placed in each bath for 60 s, with 5 s intervals between immersions, then the specimens were immersed in an aqueous solution of Ca-45. After 24 h the inlay and tooth assemblies were removed, rinsed with water and placed in cold-cured acrylic resin, then sectioned through the long axis for autoradiographic analysis. According to the penetration of Ca-45, the microleakage level was scored for each section. The results indicated slight penetration of Ca-45 on autoradiographic films.
Seventeen multiparous cross-bred sows (Swedish Land-race x Swedish Yorkshire) were inseminated in their second oestrus after weaning and divided into two groups. One group (ACTH, n = 9) was given an intravenous injection of adrenocorticotropin hormone (ACTH) every 6 h commencing 4-8 h after ovulation, whereas another group (control, n = 8) was given saline solution at the same times. The sows were slaughtered 35-53 h after ovulation. Uterine samples, taken from the mesometrial side of the uterine horns immediately after slaughter, were fixed, embedded in plastic resin and stained with toluidine blue. The endometrium was then examined by light microscopy. There was no significant effect of the ACTH treatment on the distribution of lymphocytes and macrophages, but there was a tendency of an effect on the distribution of neutrophils (P = 0.1) in the sow endometrium.
In previous studies, the localization of a pertussis toxin-sensitive G protein was demonstrated in ependymal cilia, but the identification of the subtype of G protein was inconsistent. To clarify this issue, we studied the localization of Goalpha, Gi1alpha, Gi3alpha and Gi2alpha in the ciliated ependymal cells and in the cilia of some other tissues of rats using specific antibodies. The cilia of the ependymal cells that line the ventricular cavity of the brain were intensely immunoreactive for Gi2alpha, but not for Goalpha, Gi1alpha or Gi3alpha. Immunoblot analysis demonstrated higher levels of Gi2alpha in the ependymal cilia-rich pellet than in the motor area of the parietal cortex. At the ultrastructural level, the immunoreactivity specific for Gi2alpha was found predominantly in the cilia, but rarely in the microvilli or the basal bodies of ependymal cells. In cross-sections, the immunoreactivity specific for Gi2alpha was observed only in cell membranes, in particular, in the inner electron-dense leaflet of the trilaminar structure. In addition to that in the ependymal cilia, such specific localization of Gi2alpha was observed in the motile cilia in other tissues, including the oviduct and trachea. By contrast, the stereocilia in the ductus deferens were not immunopositive for Gi2alpha. These findings suggest that Gi2 might play an important role in the signal transduction in ciliary membrane-associated function(s) of the ependymal cells, oviduct and trachea.
BACKGROUND: We have previously shown that, as in human adenine phosphoribosyltransferase (APRT) deficiency, Aprt knockout mice form 2,8-dihydroxyadenine (DHA) renal stones. The disease develops earlier and is more severe in male than in female mice. To examine the biological bases for these differences, the area occupied by DHA crystals was quantified in kidney sections from male and female mice (strain 129) aged one day to eight months and this parameter was correlated with changes in renal histopathology. Aprt heterozygous and wild-type mice were used as controls. METHODS: Following anesthesia, the left kidney was removed and immediately frozen in dry ice. Unstained cryosections were examined by polarized light to determine total area of birefringent particles. The right kidney was perfused and embedded in plastic, and stained sections were viewed by light microscopy to examine the histopathology and to determine the location of the birefringent particles. A pathological score was assigned to the histological findings. The scores from the right kidney were compared with crystal/particle area in the left kidney, and the data were analyzed using two-way analysis of variance. The chemical composition of the particles was determined by x-ray diffraction analysis. Several stone fragments from the bladder were also examined by scanning electron microscopy (SEM). RESULTS: Crystals were detected in kidney sections from one- to two-day-old Aprt knockout mice. The crystal burden remained low in both sexes throughout the study except in males at the 120- to 240-day period. Furthermore, there was a substantial degree of renal pathology, primarily seen as interstitial fibrosis, in those males with a very high level of stone formation. The crystalline material was identified as 6-amino-2,8(3,9)-purine dione, a tautomeric form of DHA. SEM indicated that the crystals were spherical, with a diameter of 10 to 20 microm. Tissue staining and fixation procedures dramatically reduced the amount of birefringent material in kidney sections. Aprt heterozygotes of both sexes had low levels of crystalline material in the kidneys and no pathology. Birefringent material or pathological changes were not seen in kidneys from wild-type mice. CONCLUSIONS: Both male and female Aprt knockout mice accumulate DHA. However, the area occupied by DHA crystals was significantly greater in 120- to 240-day-old males compared with the females of similar age. Also, substantial renal pathology was detected in kidneys of male mice that had very high levels of stone material.
BACKGROUND: migration and kinking of stent-grafts can occur late after endovascular aneurysm repair. It is unknown if endovascular grafts incorporate enough to be permanently anchored. In this report, healing of aortic stent-grafts was assessed in humans. PATIENTS AND METHODS: we retrieved 23 Dacron stent-grafts from patients treated for an aortic aneurysm since 1993. Twelve stent-grafts were explanted at late conversion to open repair and 11 at autopsy. The deaths were unrelated to graft fixation. The median age of the patients was 74 years (IQR 55-84 years) and the grafts were explanted 9 months (1-31 months) after insertion. Microscopic slides were prepared by conventional techniques or by cutting and grinding arterial specimens embedded in plastic with the stent-grafts in situ. RESULTS: the stent-grafts detached readily from the native arteries at surgery or autopsy, except when the stents had hooks or barbs which engaged the vessel wall. A space filled with poorly organised blood components persisted between the graft and the aortic wall 2.5 years after implantation. No firm incorporation of the grafts was observed proximally in the aneurysm neck or distally in the iliac segment. A friable neo-intimal layer covered parts of the luminal aspect of the grafts. CONCLUSIONS: endovascular healing provides poor fixation of Dacron stent-grafts in humans. At present, fixation relies on the mechanical properties of the stent-grafts.
The difficulties in obtaining human brain material for teaching neuroanatomy have increased the demand for more durable brain specimens. In this paper, we describe results obtained by preparing large, plastinated, dissected human brain specimens and Mulligan-stained sections of the human brain. The brains were fixed in formalin, washed and dissected in order to visualize the fibre tracts and larger nuclei in the central nervous system. This was followed by dehydration at -20 degrees C in acetone. The specimens were then impregnated with silicone, Biodur S10, in vacuo and hardened in Biodur S6 vapour. The grey and white substance in the central nervous system as well as the larger fibre tracts and nuclei were clearly visible in the dissected, plastinated specimens. Coronal and sagittal sections of the human brain were stained according to Tompsett's modification of the Mulligan method. The sections were then dehydrated in cold acetone followed by forced impregnation with Biodur S10 and hardening. The plastinated sections stained distinctly and strongly and the nuclei in the forebrain, cerebellum and brain stem could be identified easily. The sections did not fade when exposed to light and could be easily handled in the classroom without damage. Therefore, the distinct visualization of neuroanatomical structures, the improved durability of the specimens, as well as the lack of odour make plastinated specimens and stained sections of the central nervous system a valuable tool for teaching neuroanatomy that compliments the use of wet preparations.
Kidney specimens, obtained from 11 dogs of diverse species that had contracted spontaneous leishmaniasis, were submitted to a histopathological investigation using a classical preparation of sections embedded in paraffin, semi-thin and ultra-thin sections embedded in plastic, and sections frozen under immunofluorescence. Eight animals presented moderate to severe renal insufficiency. Two major categories of glomerular lesions were identified in the kidneys examined: acute glomerulonephritis and extra-membranous glomerulonephritis. These two categories were never associated and no transition from from one to the other was found. Immune complex deposits prevailed sharply over proliferative phenomena, which remained infrequent. Study of these glomerulonephrities could represent a model for better understanding of the immunological aspects of leishmaniasis.
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Alterations in the numerical density and structure of asymmetrical synapses were examined in thin sections through barrel D4 in six CD/1 mice, including three controls and three sensory deprived animals. Sensory deprivation was effected by once daily trimming of all large mystacial vibrissae on the contralateral side of the snout from P0. The mice were perfuse-fixed at P20, several days following the termination of rapid synaptic growth during barrel development (White et al., Somatosens Mot Res 14: 34-55, 1997). Cerebral hemispheres contralateral to the deprived side were osmicated, sectioned at 40 microm and embedded in plastic for thin sectioning. Sterio's (J Microsc 134: 127-136, 1984) procedure combined with serial thin section analysis (Braendgaard and Gundersen, J Neurosci Meth 18: 39-78, 1986), was applied blindly to systematic random samples of neuropil in barrel hollows and septa. No significant difference in the numerical density, estimated total number, or in the proportion of perforated postsynaptic densities was observed. However, a significant decrease in the diameters of asymmetrical postsynaptic densities was observed in hollow (P < 0.05) and septal (P < 0.05) neuropil of deprived animals. These results demonstrate a significant morphological alteration in asymmetrical synapses of a type consistent with a reduction in synaptic activity consequent to sensory deprivation.
While long-term fixation and storage of specimens is common and useful for many research projects, it is particularly important for space flight investigations where samples may not be returned to Earth for several months (International Space Station) or years (manned mission to Mars). We examined two critical challenges of space flight experimentation: the effect of long-term fixation on the quality of mouse bone preservation and the preservation of antigens and enzymes for both histochemical and immunohistochemical analyses, and how the animal/sample processing affects the preservation. We show that long-term fixation minimally affects standard histological staining, but that enzyme histochemistry and immunolabeling are greatly compromised. Further, we demonstrate that whole animal preservation is not as suitable as whole leg or stripped leg preservation for long-term fixation and all histological analyses. Overall, we recommend whole leg processing for long-term storage of bone specimens in fixative prior to embedding in plastic for histological examination.
The leading tips of elongating nerve fibers are enlarged into "growth cones" which are seen in tissue culture to continually undergo changes in conformation and to foster numerous transitory slender extensions (filopodia) and/or a veillike ruffling sheet. After explantation of 1-day-old rat superior cervical ganglia (as pieces or as individual neurons), nerve fibers and tips were photographed during growth and through the initial stages of aldehyde fixation and then relocated after embedding in plastic. Electron microscopy of serially sectioned tips revealed the following. The moving parts of the cone, the peripheral flange and filopodia, contained a distinctive apparently filamentous feltwork from which all organelles except membranous structures were excluded; microtubules were notably absent from these areas. The cone interior contained varied forms of agranular endoplasmic reticulum, vacuoles, vesicles, coated vesicles, mitochondria, microtubules, and occasional neurofilaments and polysomes. Dense-cored vesicles and lysosomal structures were also present and appeared to be formed locally, at least in part from reticulum. The possible roles of the various forms of agranular membranous components are discussed and it is suggested that structures involved in both the assembly and degradation of membrane are present in the cone. The content of these growing tips resembles that in sensory neuron growth cones studied by others.
The localization of cytokinins in corn root tips was investigated using antibodies or antibody fragments directed against dihydrozeatin riboside and labeled with rhodamine or colloidal gold. Roots were sectioned at -30 degrees to -40 degrees for immunofluorescence or freeze-substituted in ethanol or acetone and embedded in plastic for electron microscopy. Meristematic cells surrounding the quiescent center as well as root cap cells were specifically labeled using direct immunofluorescence techniques, whereas cells of the quiescent center did not bind label. Tissue sections treated with colloidal gold-labeled antibody fragments had gold particles widely distributed in the cytoplasm. The results show that the quiescent center is not the major site of cytokinin localization in root tips.
Layer IVA of rhesus monkey striate cortex contains pyramidal cells arranged in distinct groups. Their cell bodies are in a configuration of flat cones, each with an average diameter of 60 microns, and their apical dendrites aggregate into bundles that ascend toward the pial surface. Nissl-stained sections suggest that these pyramidal cell cones have their bases in layer IVB, with their tops extending into layer IVA. The neurons in the cones are readily apparent in MAP2 antibody-stained material, and in cytochrome oxidase-reacted tissue it is evident that the pyramidal cell cones occupy the pale spaces that are surrounded by the darkly reactive honeycomb lattice. This lattice of neuropil around the cones contains some axons and boutons that are immunoreactive for parvalbumin, and it is within the lattice that other investigators have shown afferents from the parvocellular (P)-layers of the dLGN to terminate. Because of this input, it is likely that the pyramidal cell cones of layer IVA are involved with color and form perception. The relationship between the layer IVA cones of neurons and the underlying system of previously described pyramidal cell modules (Peters and Sethares, 1991) is discussed, as well as the possibility that the pyramidal cell cones might represent aggregations of neurons, which receive input from basic sets of P-like afferents originating from color-responsive ganglion cells of the retina, as described by Schein and de Monasterio (1987).
An examination of cortex of area 46 in the floor of the principal sulcus in the frontal lobe of the rhesus monkey has been carried out using three young (4-6 years of age), one middle-aged (12 years of age), and five old (25-32 years of age) rhesus monkeys. Light microscopic examination revealed no age-related change in the thickness of the cortex, and no changes in the frequency of profiles of neurons displaying nuclei and contained in 250-micron-wide strips of 1-micron-thick sections. Since the diameters of the nuclei of the neurons were found to be the same in the young and old monkeys, it was concluded that there was no change in the numbers of neurons beneath similar areas of cortical surface of area 46 with age. This conclusion was reinforced by an electron microscopic examination, since there was no suggestion of degeneration of the cell bodies of the neurons, which accumulated but little lipofuscin in the old monkeys. However, there were signs of degeneration in some of the dendrites in the upper layers of the cortex in the old monkeys, especially in layer 1, in which many of the dendrites had lost organelles from their cytoplasm. The other notable change was a degeneration of myelinated axons in the deep layers and white matter in some of the old monkeys. In contrast to the neurons, the effects of aging on the neuroglial cells and pericytes were very obvious, since in the old monkeys each type of neuroglial cell accumulated large inclusions within its cytoplasm. Prior to fixation, these monkeys had been behaviorally tested using a series of spatial and visual recognition tasks, which revealed that relative to the young monkeys, the old monkeys as a group displayed memory impairment. On one task, the extent of the impairment for each old monkey correlated well with the extent of degeneration of myelinated fibers in the cortex and white matter. Consequently, it is suggested that age-related cognitive changes are unlikely to be a result of a loss of neurons, but might be due to an alteration in connections between the cortex and other brain structures.
Marmosets are New World small primates phylogenetically close to humans and are commonly used in biomedical research. Although the reproductive biology of the common marmoset Callithrix jacchus is fairly well investigated, there are few data available for testis function for its close relative, Callithrix penicillata. In this regard, the present study was performed to investigate testis structure, spermatogenic cycle length, and spermatogenic and Sertoli cell efficiencies in eight captive C. penicillata. These animals received (3)H-thymidine injections and had their testes perfused-fixed with glutaraldehyde and embedded in plastic at different time periods after (3)H-thymidine injections, for histomorphometric and autoradiographic evaluation. The analysis of the different germ cell associations showed that two or more stages were observed in about 30% of the seminiferous tubule cross sections investigated. The values found for spermatogenic cycle length and for total duration of spermatogenesis in the marmoset C. penicillata, 15.4 and 69.3 days respectively, were very close to those cited in the literature for humans. However, the results observed for Sertoli cell efficiency (number of round spermatids per Sertoli cell; 8:1) and spermatogenic efficiency (daily sperm production per gram of testis; 18.4 million) were substantially higher than those observed for humans. The results found in the present investigation suggest that the black tufted-ear marmoset C. penicillata might represent an alternative and useful experimental model to perform comparative studies regarding the spermatogenic process, particularly in investigations related to the expansion of spermatogonial stem cells and the establishment of spermatogenic waves.