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PubMed1976

Platelets, endothelial cells and macrophages in the spleen. An ultrastructural study on perfusion-fixed organs.

Rabbit spleens have been examined after perfusion fixation with and without prior washing with various fluids. The platelets were stored in the splenic sinuses and in the cord spaces as single platelets, or in loosely packed aggregates which appeared to be anchored to the endothelium by one or a few platelets. After washing prior to fixation most of the platelets disaggregated and regained their normal shape. Some platelets adhered to morphologically normal endothelium even after prolonged perfusion. Occasionally, platelets were observed inside splenic endothelial cells. Others were closely associated with macrophages, many of which also contained engulfed platelets. There was no morphological evidence of a particular platelet population being retained in the spleen after washing. In the sinuses special granule-rich cytoplasmic structures were observed. They were interposed between ordinary endothelial cells and contained a large number of small lysosome-like granules. Nuclei were never observed in these structures, probably because they consisted of pseudopod-like protrusions. Their origin and function are discussed. They may represent actively phagocytizing elements.

Animals↗
PubMed1987

Ultrastructural demonstration of exocytosis in intact and saponin-permeabilized cultured bovine chromaffin cells.

Exocytosis is the release of intracellular vesicular contents directly to the cell exterior after fusion of the vesicular and plasma membranes. It is generally accepted as the process by which transmitters and hormones are released from neurons and neurosecretory cells. There is overwhelming biochemical evidence that exocytosis is the mechanism by which catecholamines are released from adrenal chromaffin cells. With the exception of the hamster, however, there is little ultrastructural evidence to support such a mechanism. We have used a modified in vitro tannic-acid method to visualize exocytosis by transmission electron microscopy in intact and saponin-permeabilized bovine chromaffin cells. When cells are exposed to tannic-acid-containing medium, the content of vesicles involved in exocytosis is coagulated in situ as the vesicle opens to the exterior. Numerous exocytotic profiles were observed. The exposed vesicle contents appeared more granular than those of vesicles in the cell interior. Tannic acid also made the plasma membrane more distinct. Small holes were apparent in the plasma membrane of saponin-treated cells, with little disruption of underlying cytoplasmic structure. Furthermore, when these cells were stimulated with calcium, exocytosis was evident only at regions of intact plasma membrane, not at the holes. Parallel measurements of secretion showed no secretion in the presence of tannic acid. Pretreatment with tannic acid prevented subsequent secretion by intact cells and markedly reduced that of permeabilized cells, indicating a probable change in the nature of the plasma membrane.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1976

A scanning electron microscope study of the interstitial tissue of the canine testis.

Scanning electron microscopy (SEM) is a potent tool that is especially valuable in interpreting the three-dimensional relationships of cells within tissues. This type of information is obtainable from thin sections in transmission electron microscopy (TEM) only by reconstructions of serial sections. The arrangement of the interstitial cells of the testis in relation to the capillaries and lymphatic channels, in particular, is easier to visualize in SEM than in TEM. Cytoplasmic constituents, as well as cell surface modifications, are demonstrable by this technique. The presence of droplets, presumably lipid droplets, both within and on the Leydig cells and the lymphatic endothelial cells, is quite evident. Other cytoplasmic structures are also apparent. For example, the possible functional significance of "openings" that are seen by SEM on the septa that surround lipid droplets is discussed relative to the appearance of the same area as seen in thin sections or in freeze-fracture replicas. SEM should become a very useful method for studying cytological and morphological alterations that occur in testicular tissue that is subjected to physical or chemical manipulation.

Animals↗
PubMed1989

Teratogenic antibodies are directed against a coated-pit glycoprotein.

It has been well established that heterologous antibodies against certain tissue components may cause congenital abnormalities when injected into pregnant rats during the critical period of organogenesis. A glycoprotein antigen (gp340) of rat renal proximal tubules was isolated (C.C.K. Leung: (J. Exp. Med., 156:372-384, 1982); antibodies against gp340 were teratogenic. Indirect colloidal gold immunocytochemical method was utilized to study the ultrastructural localization of gp340. For comparative studies, both preembedding and postembedding immunostaining procedures were used. The results indicate that gp340 is a resident of coated pits and possibly also of coated vesicles of the rat renal proximal tubules and visceral yolk-sac (VYS) endodermal cells. It appears that gp340 may also be associated with the microvilli and some as-yet-unidentified cytoplasmic structures of the same tissues. However, gp340 is absent on the epithelium of the small intestine. It is hypothesized that the teratogenic antibodies may interact with gp340 on the coated pits and interfere with receptor-mediated endocytosis, causing yolk-sac placenta dysfunction which in turn causes abnormal embryonic development.

Animals↗
PubMed1992

Supra-neuroectodermal cells and fibers on the primary nasal cavity and in the fourth ventricle of mouse and human embryos: scanning and transmission electron microscopic studies.

Neuroectoderm-derived epithelia of the primary nasal cavity and the fourth ventricular floor and roof were observed by scanning (SEM) and transmission electron microscopy (TEM) and SEM-TEM correlative views in mouse embryos of 9th to 13th days of gestation, and in 38 externally normal human embryos ranging at Carnegie stages from 13 to 18 (about 5 to 7 weeks of gestation). Smooth-surfaced spindle-shaped cells with one or more cytoplasmic processes and cord-like cytoplasmic structures were observed by SEM on the wall of the primary nasal cavity of both species. They had morphological features similar to those of neuronal type 1 supraependymal (SE) cells and SE fibers on the floor and roof of the fourth ventricle in both species. Type 1 SE cells, SE fibers, and corresponding structures in the primary nasal cavity were localized in relation to the underlying developing nerve and vascular systems. Furthermore, their processes and fibers ran roughly parallel to these underlying structures and they penetrated the epithelial layer at the ends, suggesting a connection with underlying structures. From TEM and SEM-TEM correlative observations, SE fibers in the fourth ventricle and cord-like structures in the primary nasal cavity, both with a larger diameter, were deduced as single axon-like processes or bundles of processes. Those fibers and cord-like structures of smaller diameters were interpreted as elongated telophase bridges; both contained parallel packed microtubules and connected distant cells. Since these processes and fibers were generally longer and became fewer at later developmental stages, they appeared to be transient neuronal structures. They may play a development-related role in such morphogenetic cell movements as in the developing nerve and vascular systems in the epithelial and/or subepithelial layers, but not as direct rudiments of adult nerve tissues.

Animals↗
PubMed1994

Extractions reveal specific argentophilic proteins in rat and bull sperm heads.

BACKGROUND: Silver-stainability (argentophilia) of cytoplasmic structures occurring in spermatids have been localized into the organizing perinuclear theca, but the biochemical nature and structural associations of these proteins with the cytoskeletal and membranous elements are unresolved and, therefore, were the aim of the present study. METHODS: Light and electron microscopic analysis of the silver-stainability in the rat spermatids and spermatozoa was carried out in the intact testis tissue and epididymal spermatozoa and after their chemical and mechanical extraction. Correlation of argentophilia with specific proteins of rat and bovine spermatids and spermatozoa was investigated using a recently developed technique for silver nitrate staining of proteins on nitrocellulose. RESULTS: Sequential formation of the silver-stainable domains seemed to proceed from the argentophilic acrosomal ring. Various extractions indicated that argentophilia in the spermatids and spermatozoa was mainly associated with the perinuclear theca and to some extent to the plasma membrane. Hyamine-soluble extract from spermatozoa of rat and bull revealed only a single argentophilic protein of 130 kDa. Hyamine and SDS-soluble extracts of rat testis tissue contained an additional group of argentophilic polypeptides of lower molecular weight (115, 94, 36, 23, and 21 kDa). CONCLUSIONS: Reduction in the number of argentophilic proteins appears to be involved in a series of changes in the cyto-architecture of developing spermatids. Tentative cytoskeletal nature of argentophilic proteins remains to be identified. Nevertheless, they may have important physical relations with the higher-order organization of the sperm head cytoskeleton and overlying membranes.

Animals↗
PubMed1988

Arthritis-associated changes in flow cytometric characteristics of cultured synovial fibroblasts.

Synovial fibroblasts cultured from patients with rheumatoid or reactive arthritis and from controls were studied by flow cytometry, spectroscopy, and electron microscopy. Analysis of 29 cell lines revealed consistent differences between arthritic and normal fibroblasts. Cells cultured from inflamed synovial tissue exhibited higher autofluorescence than did control fibroblasts, and displayed exceptional light scatter properties in flow cytometry, indicating changes in cytoplasmic structures. Electron microscopic examination of the fibroblasts from arthritic synovial tissue revealed large numbers of round, swollen, laminated, mitochondrion-like bodies, which were not observed in the control fibroblasts. The changes observed by flow cytometry (light scatter and autofluorescence) coincided with the presence of the mitochondrion-like organelles. The strong autofluorescence observed in the arthritic fibroblasts resembled the fluorescence spectrum of mitochondrial flavoproteins. These data suggest that persistent metabolic and structural changes have occurred in the mitochondria of synovial fibroblasts and inflammatory synovial tissue. The usefulness of flow cytometry in identifying such cells is described.

Arthritis↗
PubMed1988

The molecular mobility of alpha-actinin and actin in a reconstituted model of gelation.

Dictyostelium discoideum alpha-actinin (D.d. alpha-actinin) is a calcium and pH-regulated actin-binding protein that can cross-link F-actin into a gel at a submicromolar free calcium concentration and a pH less than 7 [Fechheimer, et al., 1982]. We examined mixtures of actin and D.d. alpha-actinin at four pH and calcium concentrations that exhibited various degrees of gelation or solation. The macroscopic viscosities of these mixtures were measured by falling ball viscometry (FBV) and compared to the translational diffusion coefficients measured by gaussian spot and periodic-pattern fluorescence photobleaching recovery (FPR) of both the actin filaments and D.d. alpha-actinin. A homogeneous, macroscopic gel was not composed of a static actin network. Instead, the filament diffusion coefficient decreased to approximately 65% of the control value. If the D.d. alpha-actinin concentration was increased, the solution became inhomogeneous, consisting of domains of higher actin concentration. These domains were often composed of a static actin network. The mobility of D.d. alpha-actinin consisted of a major fraction that freely diffused and a minor fraction that appeared immobile under the conditions employed. This suggested that D.d. alpha-actinin binding to the actin filaments was static over the time course of measurement (approximately 5 sec). Under solation conditions, there was no apparent interaction of actin with D.d. alpha-actinin. These results demonstrate that 1) actin filaments need not be cross-linked into an immobile, static array in order to have macroscopic properties of a gel; 2) interpretation of the rheological properties of actin:alpha-actinin gels are complicated by spatial heterogeneity of the filament concentration and mobility; and 3) a fraction of D.d. alpha-actinin binds statically to actin in undisturbed gels. The implications of these results are discussed in relation to cytoplasmic structure and contractility.

Actinin↗
PubMed1993

Preferential dendritic localization of pericentriolar material in hippocampal pyramidal neurons in culture.

Centrosomes are unique cytoplasmic structures which serve as microtubule organizing centers (MTOC). In most animal cells centrosomes consist of one or more pair of centrioles surrounded by electron dense amorphous pericentriolar material (PCM) responsible for nucleation of microtubules. In the present study we analyzed the pattern of induction and localization of proteins of the PCM at different stages of neuronal development in cell cultures prepared from the embryonic hippocampus. For this purpose we used a human polyclonal antibody that recognizes two proteins of the PCM (100 kd and 60 kd, respectively). The results indicate that in mature neurons, pericentriolar immunoreactive material is preferentially localized in dendritic processes, and that throughout the course of neurite development and differentiation it is systematically excluded from the neuron's axon. Western blot analysis showed that during neuronal development in situ, there is an increase in the immunoreactivity for both proteins recognized by this antibody. In contrast, in hippocampal pyramidal neurons that develop in culture, there is an increase in the 60 kd polypeptide, while the 100 kd one is not detected after 7 days in vitro.

Animals↗
PubMed1979

Endocytic activity of subependymal microglial cells in the toad brain: a cytochemical study of peroxidase uptake.

A population of microglial cells that rapidly incorporate extracellular material introduced into the ventricular system has been identified just beneath the ependyma of all four cerebral ventricles in the toad (Bufo marinus). In untreated tissue these cells appear to be scattered, possess few processes and have an elongate shape with their long axes lying parallel to the ventricular surface. Their most distinctive ultrastructural features are nuclei containing clumps of chromatin, cytoplasmic dense bodies and single strands of granular endoplasmic reticulum. When horseradish peroxidase (HRP) is perfused through the ventricular system and the tissue processed using the DAB cytochemical method, the cells change shape and incorporate HRP into cytoplasmic structures. Even after very short perfusion periods (2-5 minutes) cells become rounded, the surface is ruffled and pseudopodia develop that contain characteristic flocculent material. Reaction product for HRP is contained in plain and coated vesicles, tubules, vacuoles and long structures composed of two closely apposed membranes. At these early times, relatively few multivesicular bodies and dense bodies contain reaction product, but when the cells are viewed at longer time periods after the ventricular perfusion of HRP an increasing proportion of the multivesicular bodies and dense bodies contain reaction product. By 320 minutes reaction product is found almost exclusively in these two organelles. In addition, many pseudopodia containing dense bodies with peroxidase activity are found in the neurophile; some, but not all, can be traced from the subependymal microglial cells. The cell bodies have resumed their flattened shape. When compared to the subependymal microglial cells, other brain cells--oligodendrocytes, astrocytes, ependymal cells and neurons--contain relatively little reaction product at short time intervals; only by 320 minutes are moderate amounts of HRP present. Because of the position of the microglial cells and their ingestive capacity, it is suggested that they function to protect the brain from foreign substances entering from the CSF.

Animals↗
PubMed2003

Cytopathological grading, as a predictor of histopathological grade, in ductal carcinoma (NOS) of breast, on air-dried Diff-Quik smears.

Fine-needle aspiration cytology (FNAC) is a widely practiced technique in the diagnosis of breast carcinoma, and it is the only diagnostic procedure performed before definitive treatment, at most institutions. While the histological grading of breast carcinoma has become routine in many centers worldwide, the cytopathological grading of breast carcinoma is not commonly used. Grading of breast carcinoma, while the tumor is still in vivo, would be the most ideal and desirable situation, as it would be helpful in the selection of patients for appropriate therapy. The objective of this study, therefore, was to devise a simple system for grading breast carcinoma, based on the cytological features alone. We reviewed 125 cases of breast carcinoma retrospectively, which were initially diagnosed by FNAC, with subsequent histopathological confirmation. These included 105 ductal, 6 lobular, 2 tubular, 1 papillary, and 1 medullary carcinoma. There was 1 ductal carcinoma in situ. Nine cases were rendered insufficient for grading. Thus 105 cases of ductal carcinoma (NOS) were evaluated for final cytological grading. Air-dried Diff-Quik-stained smears were reviewed at least twice independently by four histopathologists and were then compared with the original histological grades. Six cytological features used for grading were found to be statistically significant: cellular pleomorphism, nuclear size, nuclear margin, nucleoli, naked tumor nuclei, and mitoses. A scoring system based on these six essential parameters was used, to classify ductal carcinoma into three cytological grades, which showed close correlation with the established histological grades. In addition, two less consistent, but still important, features were the presence or absence of necrosis and stromal invasion. Another six parameters, including smear cellularity, degree of cell dispersion or clustering, lymphoplasmacytic infiltrate, presence of tubular structures, cytoplasmic appearance of the tumor cells, and smear background, were not statistically significant. However, these additional parameters were found helpful in assigning the correct grade, in cases with borderline scores. The concordance rate with histology was 100% for grade 1, 98% for grade 2, and 93% for grade 3.

Biopsy, Fine-Needle↗
PubMed1994

Developmental arrest of fertilized eggs from the B6.YDOM sex-reversed female mouse.

When the Y chromosome of a Mus musculus domesticus mouse strain is placed onto the C57BL/6J (B6) inbred background, the XY progeny develop ovaries or ovotestes but never normal testes during fetal life. While some of the hermaphroditic males become fertile, none of the XY females produces litters. Here, we examined the fertility and development of oocytes derived from the XY female mouse. With or without preceding injection of gonadotropins, female mice were mated with normal B6 males, and their embryos were recovered at various developmental stages. In vitro fertilization was performed with the eggs recovered from the oviduct after treatment with gonadotropins. Development of embryos was examined by both light and electron microscopy. The results indicate that the oocytes released from the B6.YDOM ovary were efficiently fertilized and often initiated the first cell cleavage, but all embryos died during early preimplantation periods. Even when oocytes were fertilized in vitro, minimizing their exposure to the XY oviduct/uterus environment, most embryos died at the 1- or 2-cell stage. A few exceptional embryos reached the 4- or 8-cell stage, but abnormalities were evident in both nuclear and cytoplasmic structures of all embryos. After cleavage, neighbouring blastomeres were only loosely associated, and microvilli were abundant at the intercellular interfaces. We postulate that oocytes of the B6.YDOM female mouse become defective during XY ovarian differentiation, and, hence, fail to proceed through normal embryonic development.

Animals↗
PubMed2001

The arrest gene is required for germline cyst formation during Drosophila oogenesis.

In Drosophila, oogenesis is initiated when a germline stem cell produces a differentiating daughter cell called the cystoblast. The cystoblast undergoes four rounds of synchronous divisions with incomplete cytokinesis to generate a syncytial cyst of 16 interconnected cystocytes, in which one cystocyte differentiates into an oocyte. Strong mutations of the arrest (aret) gene disrupt cyst formation and cause the production of clusters of ill-differentiated germline cells that retain cellular and molecular characteristics of cystoblasts. These mutant germ cells express high levels of BAM-C and SXL proteins in the cytoplasm but do not accumulate markers for advanced cystocytes or differentiating oocytes, such as the nuclear localization of SXL or the accumulation of osk mRNA, orb mRNA, and cytoplasmic dynein. However, the mutant germ cells do not contain spectrosomes, the cytoplasmic structure that objectifies the divisional asymmetry of the cystoblast. The aret mutant germ cells undergo active mitosis with complete cytokinesis. Their mitosis is accompanied by massive necrosis, so that the number of germ cells in a stem cell-derived cluster ranges from one to greater than 70. These defects of aret mutants reveal a novel function of aret as the first gene with a defined function in the cystoblast to cyst transition during early oogenesis.

Animals↗
PubMed2004

Dystrophic microglia in the aging human brain.

We have studied microglial morphology in the human cerebral cortex of two nondemented subjects using high-resolution LN-3 immunohistochemistry. Several abnormalities in microglial cytoplasmic structure, including deramification, spheroid formation, gnarling, and fragmentation of processes, were identified. These changes were determined to be different from the morphological changes that occur during microglial activation and they were designated collectively as microglial dystrophy. Quantitative evaluation of dystrophic changes in microglia revealed that these were much more prevalent in the older subject (68-year-old) than in the younger one (38-year-old). Thus, we conclude that microglial dystrophy is a sign of microglial cell senescence. We hypothesize that microglial senescence could be important for understanding age-related declines in cognitive function.

Adult↗
PubMed1985

A dual expression of cytokeratin and neurofilaments in bronchial carcinoid cells.

Intermediate filaments (IF) are ubiquitous cytoplasmic structures which, by virtue of their cell- and tissue-type-specific characteristics, are widely used as markers of tissue derivation and as differential diagnostic aids in surgical pathology. In contradistinction to other IFs, vimentin filaments, characteristic of mesenchymal cells, may be co-expressed with other cell-type specific IFs--cytokeratin filaments, desmin filaments, glial filaments and neurofilaments--in some tumor cells, embryonic cells, and cells in vitro. In this study we describe a novel type of IF co-expression which does not involve vimentin-filaments, viz. the presence of both cytokeratin filaments and neurofilaments in human bronchial carcinoid tumor cells.

Bronchial Neoplasms↗
PubMedDate not supplied

Changes in testicular morphology in boars actively immunized against gonadotropin hormone-releasing hormone.

Alterations in testicular morphology were studied in boars actively immunized against gonadotropin hormone releasing hormone (GnRH). Ten boars were divided equally into two experimental groups (five GnRH-immunized, and five controls). Antibody production was achieved by conjugating GnRH to human serum globulin (hSG). The GnRH-hSG conjugate was emulsified in complete Freund's adjuvant, and administered to boars at 12 weeks of age. Boars were given a booster in incomplete Freund's adjuvant on week 18 and 20. The presence of high antibody titers to GnRH caused luteinizing hormone and testosterone to decline to nondetectable levels. Morphometric examination showed a reduction in percentage volume in Leydig cells/unit testis, seminiferous tubule diameter and seminiferous epithelial height, and an increase in non-Leydig cell interstitial tissue in GnRH-immunized boars compared with controls. Histologic evaluation displayed severe damage of the seminiferous epithelium, absence of spermatids, incomplete cell associations, disruption of Sertoli cells, formation of multinucleated giant cells, and a striking reduction in size and cytoplasmic structures of Leydig cells in GnRH-immunized animals. These results demonstrate the potent inhibitory effects of GnRH immunoneutralization on the boar reproductive system.

Animals↗
PubMedDate not supplied

Morphological and functional alterations in human proximal tubular cell line induced by low level inorganic arsenic: evidence for targeting of mitochondria and initiated apoptosis.

The kidney is a known target organ for arsenic and is critical for both arsenic biotransformation and elimination. Previous studies have demonstrated that at high doses (ppm) inorganic arsenic is toxic to mitochondria primarily by affecting cellular respiration. However, the effect of inorganic arsenic on mitochondria after low level exposures is not known, particularly in the kidney. Thus the functional and morphological effects of low level inorganic arsenic were investigated in a human proximal tubular cell line, HK-2. Mitochondrial function was assessed at subcytotoxic concentrations of arsenite (< or = 10 microm) by examining the alteration of the mitochondrial membrane potential using MitoTracker Red, a mitochondrion selective dye. In a subset of cells, subcytotoxic arsenite led to mitochondrial membrane depolarization, which could subsequently lead to permeability transition and apoptosis. Subcytotoxic arsenite also induced translocation of phosphatidylserine, indicative of early-stage apoptosis. To confirm whether subcytotoxic arsenite induces cellular and/or mitochondrial morphological alterations consistent with initiated apoptosis, HK-2 cells were evaluated with transmission electron microscopy. Classic morphology of apoptosis was not observed with subcytotoxic arsenite exposures; however, evidence of necrotic changes in the cytoplasmic structure and mitochondrial morphology were apparent. Therefore, based on depolarization of mitochondria and the externalization of phosphatidylserine, HK-2 cells appear to initiate apoptosis following subcytotoxic arsenite insult, but morphological changes indicate that HK-2 cells fail to complete apoptosis and ultimately undergo necrosis. Therefore, subcytotoxic arsenite can be sufficiently toxic to mitochondria that they lose their ability to keep the cell on course for apoptotic cell death.

Apoptosis↗
PubMed1984

A monoclonal antibody recognizes an epitope common to an avian-specific nuclear antigen and to cytokeratins.

X3, a monoclonal antibody of unusual specificity, is described. This antibody reacts with one or more cytokeratin polypeptides and also reacts with an avian (chicken, quail) nuclear antigen that appears to be present in all cell types (chicken) tested, although with variable staining pattern and intensity. This antigen is distinct from the cytokeratins but does have an epitope in common with this class of proteins. It disappears from the nucleus during the early stages of cell division and reappears during anaphase as a granular cytoplasmic structure. In late telophase the antigen is relocated in the nucleus. This antigen, which we have designated as avian-specific nuclear antigen ( AVNA ), is not associated with chromatin or ribonucleoproteins. From immunoblotting experiments on chicken fibroblast nuclei, AVNA is probably a complex composed of one or several polypeptides, one of which has a molecular weight of approximately 60 kD. The proteins were identified as nuclear matrix proteins rather than pore complex-lamina proteins by immunoblotting experiments on the purified nuclear matrix of chicken erythrocytes. The major polypeptide had a molecular weight of 60 kD and the minor polypeptide a molecular weight of 69 kD.

Animals↗