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PubMed2006

Ultrastructural dynamics of human reproduction, from ovulation to fertilization and early embryo development.

This study describes the updated, fine structure of human gametes, the human fertilization process, and human embryos, mainly derived from assisted reproductive technology (ART). As clearly shown, the ultrastructure of human reproduction is a peculiar multistep process, which differs in part from that of other mammalian models, having some unique features. Particular attention has been devoted to the (1) sperm ultrastructure, likely "Tygerberg (Kruger) strict morphology criteria"; (2) mature oocyte, in which the MII spindle is barrel shaped, anastral, and lacking centrioles; (3) three-dimensional microarchitecture of the zona pellucida with its unique supramolecular filamentous organization; (4) sperm-egg interactions with the peculiarity of the sperm centrosome that activates the egg and organizes the sperm aster and mitotic spindles of the embryo; and (5) presence of viable cumulus cells whose metabolic activity is closely related to egg and embryo behavior in in vitro as well as in vivo conditions, in a sort of extraovarian "microfollicular unit." Even if the ultrastructural morphodynamic features of human fertilization are well understood, our knowledge about in vivo fertilization is still very limited and the complex sequence of in vivo biological steps involved in human reproduction is only partially reproduced in current ART procedures.

Embryonic Development↗
PubMed1998

Testicular effects of vasectomy in rats: an ultrastructural and immunohistochemical study.

OBJECTIVES: The correlation between infertility and morphofunctional alterations following vasal occlusion is not clearly understood. Although a correlation has been found between the expression of a high titer of antisperm antibodies and the status of infertility, the relationship between the immunoglobulin (Ig) depositions in the testis and ultrastructural alterations of the peritubular structures has not been clearly established. The objective of this study is to explain the etiopathogenesis of diffuse and focal testicular lesions caused by vasal obstruction. METHODS: Unilateral vasectomy was performed on adult male rats. Ultrastructural changes of testicular tissues and immunostaining affinity of peritubular structures with anti-actin, anti-vimentin, anti-laminin, and anti-immunoglobulin G (anti-IgG) antibodies were analyzed. RESULTS: In an ultrastructural study, vasectomized animals presented abundant collagen fibril accumulation in the peritubular area. In testis with intense IgG antibodies, staining revealed weak immunostaining of peritubular myoid cells with anti-actin antibodies, but intense immunostaining with anti-vimentin antibodies. The tubules of rats with no IgG deposition on peritubular structures revealed intense immunostaining with anti-actin antibodies but scant immunostaining with anti-vimentin antibodies. Anti-laminin deposits decreased severely in most of the tubules demonstrating intense IgG deposition. CONCLUSIONS: Our findings suggest that spermatogenesis deteriorates more severely in testes with dense IgG deposition. The sclerosis of the lamina propria in cases with vasal occlusion is secondary to alterations in the peritubular myoid cells. With the progressive changes that occur in the peritubular myoid cells or differentiation of the peritubular myoid cells that acquire active cell characteristics, collagen accumulation adjacent to these cells increases markedly. The alterations of the peritubular myoid cells reported here may be caused by alterations in basement membrane structures.

Animals↗
PubMed2003

Ciliary beat pattern is associated with specific ultrastructural defects in primary ciliary dyskinesia.

BACKGROUND: The main symptoms of primary ciliary dyskinesia (PCD) are nasal rhinorrhea or blockage and moist-sounding cough. Diagnosis can be difficult and is based on an abnormal ciliary beat frequency, accompanied by specific abnormalities of the ciliary axoneme. It is unknown whether determining ciliary beat pattern related to specific ultrastructural ciliary defects might help in the diagnosis of PCD. OBJECTIVE: We sought to determine ciliary beat pattern and beat frequency (CBF) associated with the 5 common ultrastructural defects responsible for PCD. METHODS: Nasal brushings were performed on 56 children with PCD. Ciliary movement was recorded using digital high-speed video imaging to assess beat frequency and pattern. Electron microscopy was performed. RESULTS: In patients with an isolated outer dynein arm or with an outer and inner dynein arm defect, 55% and 80% of cilia were immotile, respectively. Cilia that moved were only flickering. Mean CBF (+/- 95% CI) was 2.3 Hz (+/- 1.2) and 0.8 Hz(+/- 0.8), respectively. Cilia with an isolated inner dynein arm or a radial spoke defect had similar beat patterns. Cilia appeared stiff, had a reduced amplitude, and failed to bend along their length. Immotile cilia were present in 10% of cilia with an inner dynein arm defect and in 30% of radial spoke defects. Mean CBF was 9.3 Hz (+/- 2.6) and 6.0 Hz (+/- 3.1), respectively. The ciliary transposition defect produced a large circular beat pattern (mean CBF, 10.7 Hz [+/- 1.1]). No cilia were immotile. CONCLUSIONS: Different ultrastructural defects responsible for PCD result in predictable beat patterns. Recognition of these might help in the diagnostic evaluation of patients suspected of having PCD.

Adolescent↗
PubMed1999

The influence of enucleation on the ultrastructure of in vitro matured and enucleated cattle oocytes.

The enucleation of recipient oocytes in nuclear transfer experiments is generally carried out by aspirating one third of the ooplasm adjacent to the first polar body. It was supposed that this enucleation step affects the ultrastructure of the remaining cytoplast, resulting in a decline or destruction of its cellular compartments. Even if the transferred nucleus had the potential to support the development of a single-cell nucleus transfer embryo to the blastocyst stage, meiotic division could be stopped at any stage if the destruction of the ultrastructure of host cytoplasm resulted in a limited metabolism. The present study was conducted to investigate the influence of the enucleation procedure on the ultrastructure of the remaining ooplast. In vitro matured oocytes; in vitro matured and enucleated oocytes; and in vitro matured and enucleated oocytes that were subsequently cultivated in vitro for additional 4 h were prepared for transmission electron microscopy (TEM). An examination of ultra-thin sections showed that the arrangement of organelles in all matured oocytes was in accordance with that already described for normal oocyte development. Immediately after enucleation no major differences in the arrangement of cortical granules, mitochondria, smooth endoplasmic reticulum (SER), lipid droplets and vacuoles were found compared with nonmanipulated oocytes. After enucleation and 4 h of culture, 24- and 36-h matured oocytes differed from each other in the arrangement of large aggregates of SER surrounded by a wall of mitochondria and lipid droplets. These complexes were still found in the 24-h but not in 36-h matured, enucleated and cultivated oocytes. Clusters of SER, mitochondria and lipid droplets were described by different authors as having metabolic activity. The results of this study in connection with results from nuclear transfer experiments suggest that these aggregates and their metabolic activity can be transferred with cytoplasm from 24- but not 36-h matured oocytes. Only cytoplasm from the 24-h matured oocytes showed a development-supporting effect when fused to enucleated recipient cells before nuclear transfer.

Animals↗
PubMed2000

Clinical, histologic, and ultrastructural features of the corneal dystrophy caused by the R124L mutation of the BIGH3 gene.

OBJECTIVE: This study was designed to describe the clinical, histologic, and ultrastructural features of the corneal dystrophy associated with the R124L mutation of the BIGH3 gene. DESIGN: Retrospective clinical and histologic review of a new genetic mutation. PARTICIPANTS: Thirty-four patients from five unrelated French families with corneal dystrophy caused by the R124L mutation of the BIGH3 gene were studied at the clinical, histologic, and ultrastructural levels. Records of patients carrying this mutation were compared with those from three unrelated patients with corneal dystrophy of Bowman's layer (CDB) type 2 (R555Q mutation) and from three unrelated patients with classic corneal granular dystrophy (R555W mutation). INTERVENTION: The mutational genetic status of the BIGH3 gene was determined for each patient, and the histologic and ultrastructural data available after corneal graft were analyzed. MAIN OUTCOMES MEASURES: Genomic DNA was extracted from peripheral blood leukocytes. Exons 4 and 12 of the BIGH3 gene were amplified by the polymerase chain reaction (PCR), and the PCR products were directly sequenced. RESULTS: All 34 patients with the R124L mutation displayed the clinical, histologic, and electron microscopic features of the dystrophy previously described as a superficial variant of corneal granular dystrophy. Combining molecular genetics with clinical and histologic findings established a clear distinction between the R555Q and R555W dystrophies. CONCLUSIONS: The R124L mutation of the BIGH3 gene is associated with specific clinical and morphologic criteria. This indicates that molecular studies are needed for an adequate classification of corneal dystrophies. All criteria are presently available to segregate the dystrophy caused by the R124L mutation (known as CDB1) from the dystrophy caused by the R555Q mutation (known as CDB2).

Adult↗
PubMed1992

Retinal hemangioblastoma. A histologic, immunohistochemical, and ultrastructural evaluation.

The authors evaluated the histologic, immunohistochemical, and ultrastructural characteristics of two eyes with retinal hemangioblastoma from patients with von Hippel-Lindau and von Hippel disease. Results of histologic evaluation showed the eyes to have degenerative changes and residual retinal hemangioblastoma. Immunohistochemical stains performed for MAC-387, factor XIIIa, lysozyme, alpha 1 anti-chymotrypsin (histiocyte markers), factor VIII-associated antigen, ulex europeaus (endothelial markers), neuron-specific enolase, chromogranin, neurofilament (neuroectodermal/neural/neuroendocrine markers) and glial fibrillary acid protein (glial marker) showed normal retinal vascular endothelium, neurons, and glial cells to stain where expected. Vascular endothelium in the retinal hemangioblastomas stained for factor VIII and ulex europeaus. Interstitial cells in the stroma of the tumors failed to stain for the histiocyte markers, chromogranin, and neurofilament. The stromal cells stained for glial fibrillary acid protein and neuron specific enolase. Ultrastructural findings in both eyes included endothelial/pericyte-lined vascular channels, elongated stromal cells, and plump, vacuolated stromal cells with ultrastructural features consistent with glial cells. This study supports the concept that retinal hemangioblastoma is composed of a proliferation of capillaries and glial cells.

Adolescent↗
PubMed1999

Pleomorphic adenocarcinomas of the ciliary epithelium. Immunohistochemical and ultrastructural features of 12 cases.

OBJECTIVE: To examine the immunohistochemical and ultrastructural features of the rare pleomorphic adenocarcinomas of the ciliary epithelium (CE). DESIGN: Retrospective case series. PARTICIPANTS: The study materials included 12 cases of adenocarcinoma of the ciliary epithelium: 9 cases of CE hyperplasia and 3 cases of CE adenomas. INTERVENTION: Histologic sections were stained with hematoxylin-eosin, alcian blue, periodic acid-Schiff, and occasionally with Masson trichrome. Additionally, the following immunohistochemical markers were used: Kermix (ae1/ae3 + ck1), cytokeratin 7 (CK7), cytokeratin 20 (CK20), epithelial membrane antigen, CAM 5.2, S-100 protein, neuron-specific enolase, glial fibrillary acid protein, smooth muscle actin, and vimentin. Five lesions were studied ultrastructurally. Clinical data were available in all cases, and follow-up was obtained in 9 of the 12 patients. RESULTS: Nine tumors occurred in phthisical eyes in adults. The tumor cells were arranged in tubular and solid patterns and surrounded by thick basement membrane (BM) material and fibrous stroma. Immunohistochemistry (IM) of adenocarcinomas showed positivity with kermix (8 of 12 lesions), CAM 5.2 (7 of 12), and CK7 (5 of 12). Ultrastructurally, the tumor cells were surrounded by a thick, homogeneous, and/or multilaminar BM and attached to each other by junctional complexes. CONCLUSIONS: Clinically, this intraocular neoplasm should be considered in adults with a longstanding blind eye with an epibulbar mass and/or proptosis of recent duration. Fatal cases only occurred in tumors with extraocular extension. Adenocarcinomas of CE should be differentiated from amelanotic melanoma and metastatic lesions by the presence of a thick BM material around the tumor cells and intraocular fibrosis. Immunohistochemistry is helpful in differentiating from melanomas but not helpful in cases of metastatic carcinomas.

Adenocarcinoma↗
PubMed1998

Ultrastructural changes in liver damage induced by carbon tetrachloride in spontaneously hypertensive rats and Wistar-Kyoto rats.

The effects of carbon tetrachloride (CCl4) on the liver in spontaneously hypertensive rat (SHR) and Wistar-Kyoto rat (WKY) were investigated ultrastructurally in this study. Repetition of CCl4 treatment twice a week for four weeks, revealed liver cirrhosis in SHR, whereas, only bridging fibrosis was observed in the WKY, histologically, by reticulin silver impregnation. Scanning electron microscopy revealed that the liver surface of SHR was irregularly nodulated, while it was relatively smooth in case of the WKY. Transmission electron microscopy revealed that the mitochondria cristae were peripherally located in both strains. Vesicular change of the rough endoplasmic reticulum was observed to be more severe in SHR than in WKY. Further, local hypertrophy of the smooth endoplasmic reticulum and large lipid droplets were more easily observed in WKY than in SHR. The present results hence indicate that chronic chemical CCl4 intoxication induces severe ultrastructural organelle damage in hypertensive rats, leading to liver cirrhosis. On the other hand, CCl4 induced fatty liver and a relatively mild fibrosis were observed in normotensive rats. These findings suggest that the liver ultrastructural organelles are more susceptible to CCl4 in SHR than in WKY.

Animals↗
PubMed1988

Ultrastructural and functional studies of pancreatic B cells in Wistar rats treated with immunotherapeutic doses of cyclosporin.

Cyclosporin (CYA) was administered to Wistar rats at immunotherapeutic doses (20 mg/kg) and the functional and morphological effects on pancreatic B cells were observed. Serum CYA levels were kept in the range that is often encountered in clinical immunotherapy in humans. The treated rats had slightly elevated plasma glucose levels; on treatment days 6 and 13, their serum immunoreactive insulin and pancreatic insulin contents were low. Plasma immunoreactive glucagon levels were slightly elevated during treatment. Glucose tolerance was impaired on day 13. These functional abnormalities disappeared 2 weeks after drug withdrawal. B cells showed ultrastructural evidence of CYA toxicity, including cytoplasmic degranulation, nuclear inclusions, and cisternal dilatation of both the rough endoplasmic reticulum and the Golgi apparatus. These changes were observed on the third day of treatment and became more pronounced on the 6th and 13th days of treatment. By the second week after drug withdrawal, most of the B cells presented a normal ultrastructure. We confirmed that a dose of CYA similar to that used in clinical immunotherapy in humans produces ultrastructural disturbances in the pancreatic B cells of Wistar rats after relatively short-term administration.

Animals↗
PubMed1986

Fulminant hepatitis. An ultrastructural study.

Ultrastructural changes were observed in 23 consecutive patients who died from fulminant hepatic failure due to hepatitis B virus (4 cases), sporadic non-A, non-B (7), or paracetamol (acetaminophen) overdose (12) and in 3 patients with subacute hepatic necrosis of unknown cause. The findings are described in detail in 12 of these patients. Fatal fulminant hepatitis was characterised by massive confluent necrosis accompanied by collapse of reticulin framework and sudden drop-out of liver cells. No aetiological distinction could be made between different viral causes of fulminant hepatitis on the basis of ultrastructural pathology. Parenchymal changes in viral cases varied from reversible non-specific necrosis to irreversible changes where fragmentation of endoplasmic reticulum, mitochondria and nuclei had occurred. Differences in ultrastructural pathology between non-viral (paracetamol overdose-induced) and viral fulminant hepatitis were apparent. Modification of endoplasmic reticulum with enlarged attached polyribosomes, breakdown of plasma membrane, accumulation of cytoplasmic amorphous material and karyorrhexis and karyolysis of nuclei were the most prominent features in non-viral cases.

Acetaminophen↗
PubMed2000

Structural and ultrastructural analysis of root primordia in vitro cultures (RPC) of Solanum lycopersicoides Dun.

The structure and ultrastructure of cell aggregates and derived root primordia were analysed in established suspension cultures of Solanum lycopersicoides Dun. A total of four modified Murashige and Skoog (1962) (MS) media were used, two containing alpha-naphthaleneacetic acid (NAA) and two containing 2,4-dichlorophenoxyacetic acid (2,4-D). Considerable differences were observed in the size and structure of the aggregates regardless of the medium. The largest aggregates had a four-zone structure (cover, starch, dividing and differentiation zones) with distinct ultrastructural organization. The degree of histological and ultrastructural differentiation in the aggregates indicated rhizogenesis initiation. It begins with the protrusion of mounds of root primordia, which are the result of radial growth of cells in defined zones of the aggregate. Further growth from the primordium forms a root meristem with three tiers initial centre and wholly distinguished histogens (dermatogen, periblem, plerome). Rhizogenesis was more regular on media with NAA than on media with 2,4-D. Primordia and fully organized roots were liberated due to fragmentation of the aggregate and underwent changes in the permanently active developmental cycle of the culture which gave rise to successive generations of aggregates. Growth regulators had different influences on the organellar composition and phenolic compounds presence in the different zones of aggregates.

Journal Article↗
PubMedDate not supplied

Ultrastructural and immunohistochemical aspects of carcinoma in mixed tumors.

Four cases of malignant mixed tumor (carcinoma in pleomorphic adenomas) were studied for ultrastructural appearance and for the presence of cytokeratins and vimentin. Ultrastructurally, both squamous and glandular epithelial differentiation were found not only in the same tumor but also within the same cell. One tumor showed mainly mesenchymal differentiation with fibroblast-like cells. The intermediate filament expression of benign mixed tumors (i.e., both cytokeratin and vimentin content) were found in two of the three malignant tumors investigated. In the third tumor, only cytokeratins were found. Thus, the filament content of mixed tumors may change when the tumor becomes malignant. This change does not always parallel a change in morphology. Although one tumor was clearly epithelial ultrastructurally and another mostly mesenchymal, both did contain both cytokeratins and vimentin.

Adenoma, Pleomorphic↗
PubMedDate not supplied

Ultrastructural and permeability features of microvessels in the hippocampus, cerebellum and pons of senescence-accelerated mice (SAM).

We previously reported that the accumulation of blood-borne radiolabelled serum albumin in brain parenchyma increased with aging, especially in senescence-accelerated mice (SAMP8), which showed age-related deficits in learning and memory. In this study, in order to examine morphological events related to the age-related increase of the brain accumulation of serum albumin, the transvascular passage of blood-borne horseradish peroxidase (HRP) and ultrastructural features of microvessels were examined in the hippocampus, cerebellum and pons of SAMP8 and SAMR1 (control) mice. Ultrastructural examination of the hippocampus showed that the staining for HRP was occasionally spreading throughout the parajunctional cytoplasm of the endothelial cell of aged SAMP8 mice, but not in young SAMP8 mice nor in SAMR1 mice. The number of vessels showing the staining reaction for HRP in the parajunctional cytoplasm of the endothelial cells in aged SAMP8 mice increased significantly compared with that in the others. Electron microscopic morphometry showed that there were no significant differences among the number of HRP-positive vesicles per unit area of the endothelial cell cytoplasm in young and old mice of both strains. The staining reaction for HRP was not seen in the basal lamina of microvessels and the perivascular neuropil in all mice examined. Perivascular lipofuscin-like granules and collagen deposits, swelling of astroglial perivascular endfeet and perivascular cells containing foamy, lipid-like droplets were frequently found in several brain regions of aged SAMP8 mice. The perivascular cells with a few lipid-like droplets and more electron-homogeneous lysosomes were occasionally seen in SAMR1 and young SAMP8, while the other findings were scarcely observed in SAMR1 and young SAMP8 mice. These findings suggest that the blood-brain barrier to HRP was preserved in microvessels in three brain regions of SAM mice but the blood microvessels showed some age-related ultrastructural alterations in SAMP8 brains. Uncontrolled passage of HRP through the parajunctional cytoplasm of the endothelial cells may partly contribute to the age-related increase of accumulation of serum albumin in SAMP8 brains.

Aging↗
PubMed1991

Effect of acute lead intoxication on the ultrastructure of neutrophils in the peripheral blood of the rat.

The influence of lead acetate on the ultrastructure of peripheral neutrophils was investigated in adult male rats. It was found that a single intraperitoneal administration of lead at dose 150 mg Pb/kg b.w. led to ultrastructural changes in the neutrophils at 3 and 6 h post injection. At the time of testing the exposed population had a mean (+/- SD) blood lead concentration from 206.2 +/- 24.8 micrograms/100 ml to 75.2 +/- 9.9 micrograms/100 ml compared with a mean value of 4.0 +/- 0.7 micrograms/100 ml for the control groups. The ultrastructural alterations such as irregular nuclei with deep invaginations, plasma membrane pockets, the presence of vacuoles with a heterogeneous material and an increasing amount of RER cisternae, were most clearly expressed 6 h after lead administration. In addition there appeared sometimes nuclear pockets, and a prominent crystalline inclusion placed in the cytoplasmic matrix of some neutrophils. No differences in the mitochondrial morphology and cytoplasmic granule pattern between exposed and control rats were observed.

Animals↗
PubMed1990

Correlation study of serum prolactin levels, pituitary weights, number and ultrastructure of pituitary lactotrophs in female Long-Evans rats.

Twenty-five female, 23-month-old Long-Evans rats were sacrificed for investigation of correlations between serum prolactin levels, pituitary weights, numbers and ultrastructural features of pituitary lactotrophs. Serum prolactin levels measured by radioimmunoassay varied from 7.1 ng/ml to 1,534.0 ng/ml. Pituitary weights ranged from 16.0 mg to 255.0 mg. Serum prolactin levels were increased when pituitary weights were high; statistical analysis revealed a correlation between the two. No significant correlation was evident between serum prolactin levels and lactotroph percentage and ultrastructure. Light microscopy revealed a large number of lactotrophs in almost every pituitary. Values for nuclear, cell and cytoplasmic areas, nuclear/cytoplasmic ratios and nucleolar diameters remained similar in the lactotrophs of all animals. Electron microscopy disclosed no lactotroph damage or vascular injury. Ultrastructural morphometry of lactotrophs in the pituitaries studied showed no major differences in cytoplasmic volume densities of endoplasmic reticulum, Golgi complex, mitochondria, secretory granules and lysosomes as well as secretory granule diameters. It is reasonable to conclude that elevations of serum prolactin levels are due to an increase in pituitary mass, resulting from accumulation of lactotrophs. Thus hyperprolactinemia appears to be secondary to an increase in the number of lactotrophs and not hyperactivity of individual cells.

Animals↗
PubMed1986

Idiopathic microscopic polyarteritis nodosa: ultrastructural observations on the renal vascular and glomerular lesions.

Although the glomerulonephritis (GN) and renal vasculitis in polyarteritis nodosa (PAN) are generally considered to be immune-mediated, the pathogenesis of the renal injury and the role of immune complex (IC) deposition are unclear. To better define the nature of the glomerular and vascular injury in PAN, we performed a detailed ultrastructural study of 27 renal biopsies from 20 patients with histologically confirmed PAN of the microscopic or overlap (microscopic/macroscopic) type. A total of 48 arteries and arterioles were studied ultrastructurally, including 20 vessels with recognizable vasculitis in 1 micron-thick survey sections. By immunofluorescence, glomerular and vascular immunoglobulin deposits were generally scanty, primarily located in areas of necrosis or sclerosis. Fibrinogen, C3 and C1 were more commonly detected, often in the absence of demonstrable immunoglobulin. By electron microscopy, discrete electron-dense deposits of probable immune-type were found in the glomeruli of five initial biopsies. No electron-dense deposits were identified in any of the arteries or arterioles studied. In both glomeruli and vessels, endothelial injury and subendothelial fibrin deposition were the earliest detectable ultrastructural changes. The pathogenetic implications of these findings are discussed.

Adult↗
PubMed2001

Of mice and men: comparison of the ultrastructure of megakaryocytes and platelets.

OBJECTIVE: Mice provide an excellent model for studying platelet and megakaryocyte (Mk) biology in vivo. Given the increasing use of transgenic and knockout mice, it is important that any similarities and differences between murine and human platelet/Mk biology be well defined. Therefore the objective of this study was to compare and contrast in detail any significant morphological differences between Mks, platelets, and mechanisms of thrombopoiesis in humans and mice. METHODS: The distinctive structural and ultrastructural features of murine and human platelets and Mks are reviewed. Several platelet and Mk glycoproteins were also localized in murine cells by immunoelectron microscopy using polyclonal antibodies directed against human platelet proteins and compared to existing human data. Finally, the ultrastructure of maturing murine and human Mks in culture and bone marrow were examined in detail to facilitate a comparison of either in vivo or in vitro platelet production. RESULTS: Human and murine platelets exhibit significant but well-established morphological differences. Murine platelets are smaller and more numerous and display much greater granule heterogeneity than their human counterparts. Immunoelectron microscopy also demonstrated that murine platelet alpha-granules are highly compartmentalized. In fact, they are remarkably similar to human alpha-granules, with asymmetrical distribution of von Willebrand factor (vWF), and labeling of alpha(IIb)beta(3) and P-selectin (CD62P) in the granule limiting membrane. In vivo, murine but not human Mks are also consistently localized within the spleen. Subcellular events accompanying platelet formation and release by murine Mks are presented for the first time, and compared to human. Consistent differences were found in the pathway of redistribution of demarcation membranes preceding platelet formation, which may be important for the clarification of the mechanism of platelet release. CONCLUSION: Human and murine platelets and Mks display several characteristic ultrastructural differences (size, number, histological distribution, platelet shedding) which have been emphasized and analyzed in this report. Nevertheless, since there are also many close similarities (organelle and glycoprotein subcellular distribution) mice offer an excellent in vivo model to study various aspects of human Mk and platelet biology.

Animals↗
PubMed2000

Benzodiazepines protect hippocampal neurons from degeneration after transient cerebral ischemia: an ultrastructural study.

The ability of full and partial benzodiazepine receptor agonists to prevent DNA fragmentation and neuronal death after transient cerebral ischemia was investigated in the Mongolian gerbil. Diazepam (10mg/kg, i.p.) or the partial agonist imidazenil (3mg/kg, i.p.) was administered 30 and 90min after transient forebrain ischemia produced by occlusion of the carotid arteries for 5min. Treatment with diazepam completely protected CA1b hippocampal pyramidal neurons in 94% of the animals and partially protected pyramidal neurons in 6% of the animals, as assessed with a standard Nissl stain three and four days after ischemia. DNA fragmentation was examined by the terminal dUTP nick-end labeling (TUNEL) reaction. Prior to cell death, there were no TUNEL-positive neurons in area CA1b. By three days after ischemia, when neuronal degeneration was nearly complete, 14 out of 16 gerbils exhibited a positive TUNEL reaction throughout area CA1b stratum pyramidale. In 13 out of 14 gerbils treated with diazepam, no TUNEL-positive neurons were observed in this region. Imidazenil was less effective than diazepam with respect to both neuroprotection and prevention of DNA fragmentation. Three days after ischemia, six out of eight gerbils treated with imidazenil showed partial to complete neuroprotection. Imidazenil completely prevented DNA fragmentation in only one of the animals; varying degrees of TUNEL reaction persisted in the remainder. To determine whether the neurons protected by diazepam had a normal ultrastructure, gerbils were killed two to 30 days after ischemia and the hippocampal neurons in area CA1b were examined by electron microscopy. Within the first 48h after ischemia, early cytoplasmic changes of varying degrees (e.g., vacuolation, rough endoplasmic reticulum stacking, swollen mitochondria) and electron-dense dendrites were observed in gerbils not treated with diazepam. Degeneration was nearly complete by three days after ischemia. In contrast, pyramidal neuron ultrastructure appeared normal in gerbils that exhibited complete area CA1b neuroprotection (defined at the light microscope level) by diazepam when studied two, seven or 30 days after ischemia. In gerbils with partial protection of area CA1b, most of the remaining neurons exhibited varying degrees of necrosis when studied 30 days after ischemia. No apoptotic bodies were observed. We conclude that: (i) diazepam can fully protect CA1 pyramidal cells from the toxic effects of transient cerebral ischemia; (ii) when diazepam affords only partial neuroprotection, the residual CA1 pyramidal cells exhibit ultrastructural abnormalities consistent with necrotic damage; and (iii) diazepam is a more efficacious neuroprotectant than the partial benzodiazepine receptor agonist, imidazenil.

Animals↗