Ultrastructural morphometry of precompacted bovine embryos produced in vivo and in vitro after activation by electric pulse AC/DC.

Early bovine precompacted embryos (at 1- to 8-blastomere stage) were analyzed by electron microscopy. The volume density of cellular components was determined by morphometric analysis to quantify the ultrastructure of early bovine embryos produced either in vivo or parthenogenetically after stimulation of oocytes by electric pulse AC/DC. In embryos obtained in vivo, most of cellular volume was occupied by cytoplasm (82.93%). The relative volume of lipids, vacuoles, mitochondria was relatively low (5.46; 5.07; 3.78%, respectively), and the relative volume of Golgi apparatus and cell inclusions was the lowest (1.51%). AC/DC-derived parthenogenotes had a relative high area occupied by vacuoles and lipids (18.68 vs. 14.33%) and a significantly lower relative volume was occupied by cytoplasm (60.63%) when compared with the control in vivo embryos. These observations demonstrated that parthenogenetic embryos had significantly altered ultrastructure, indicating extensive subcellular damages. These findings are discussed from the physiological and functional point of view.

Comparative morphometry of precompacted bovine embryos produced in vivo or in vitro after parthenogenetic activation and intracytoplasmic sperm injection (ICSI): ultrastructural analysis.

Early bovine precompacted embryos (1 to 8 blastomeres) were analysed by electron microscopy. The volume density of cellular components was determined by morphometric analysis to quantify the ultrastructure of early bovine embryos produced either in vivo or in vitro both after fertilisation by intracytoplasmic sperm injection (ICSI) or from electrically stimulated oocytes (AC/DC). In normal embryos obtained in vivo (control), most of the cellular volume was occupied by cytoplasm (82.93%). The relative volume of lipids, vacuoles, mitochondria, Golgi apparatus and inclusion bodies was minimal. In the group of embryos after parthenogenetic activation (AC/DC) a relatively high proportion of the volume was occupied by vacuoles and lipids (18.68% vs 14.33%). Early ICSI-derived embryos contained the lowest relative volume of cytoplasm (58.33%) compared with the control embryos (in vivo) and parthenogenetically AC/DC-activated embryos and a higher volume was occupied by lipids (13.25%) and vacuoles (12.92%). It is concluded that in vitro produced embryos have a significantly altered ultrastructure, indicating extensive cellular damage.

Autoradiographically detected impairment of RNA synthesis pattern in 8-16 cell bovine embryos after irradiation.

Early preimplantation bovine embryos at 8- or 16-cell stage were analysed by [5-3H]uridine autoradiography for distribution of newly synthesized RNA after 60Co irradiation with a single dose of 1 Gy, 2 Gy or 4 Gy gamma rays, respectively. Embryos irradiated with a single dose of 1 Gy showed equally decreased synthesis of RNA in nucleoplasma as well as in nucleolus. In embryos irradiated with a single dose of 2 Gy or 4 Gy, RNA synthesis was decreased and localized mostly on the periphery of the nucleus; in both cases of irradiation, the nucleus center being without labelling. In most of embryos irradiated with a dose of 4 Gy, the nucleoli were not labelled, and an increasing occurrence appeared of various nucleus chromatin segregation forms, mainly as its marginalization.

[Morphological study of gastrocnemius muscle in Japanese quail chicks hatched out in space flight]. / Morfologicheskie issledovaniia ikronozhnoi myshtsy ptentsov iaponskogo perepela, vyvedennykh v usloviiakh kosmicheskogo poleta.

Slovak experiment Quail SK-6 aboard orbital station Mir in the period of February 20-27, 1999 resulted in hatching out of viable chicks of the Japanese quail. The paper presents morphologic results of investigation of m. gastrocnemius of the chicks after 4-5-d life in space microgravity. On the whole, histologically the gastrocnemius of the space chicks was very much similar to that of the control group. Muscular fibers were, which is typical, striated; bundles of myofibrills and numerous nuclei were on the periphery of the fiber under the sarcolemma. However, despite the similarity of the muscular histology, the chicks borne and exposed in microgravity displayed some deviations in the gastrocnemius as compared to their controls. Thus, Z-lines in some of the muscular fibers had a zigzag form or were doubled. In places, mitochondria were increased in size and lined up in chains longer than a sarcomere; internal membranes of the mitochondria were thickened and vacuolized, there were expanded tubules in the sarcotubular apparatus, and muscular fibers with multiple fatty vacuoles.

[Morphological changes in the adrenal gland of Japanese quail chicken hatched out in microgravity]. / Morfologicheskie izmeneniia v nadpochechnikakh ptentsov iaponskogo perepela, vylupivshikhsia v usloviiakh nevesomosti.

The paper presents results of studying effects of spaceflight factors on the adrenal structure in Japanese quail hatchlings (Coturnix coturnix japonica) incubated and kept alive 4 to 5 days aboard the space station Mir. In comparison with the ground controls, cortical cells in the adrenals of space hatchlings contained copious fat drops, mitochondrias, and significantly dilated cisterns of the endoplasmic reticulum. Accumulation of fat drops evidences lowered cholesterol transformation and disturbed genesis of steroids. Cytoplasm of the medullar chromaffin cells was full of dark large granules; some of these cells also had enlarged mitochondrias and dilated endoplasmic reticulum cisterns.

The histopathological characterization of oral Candida leukoplakias.

Sixty four biopsy specimens of oral Candida leukoplakias were examined. Histological sections were stained with periodic acid-Schiff (PAS) reagent and by Grocott's silver method. C. albicans isolates were cultivated from all the patients but fungal hyphae were proved histologically only in 23 of them. PAS-positive and Grocott-positive elements were exclusively observed in the superficial epithelial layers, e.g., the cornified layer and the stratum granulosum. The inflammatory reaction was characterized by an early phase during which polymorphonuclear cells predominated. The skeletal muscle fascicles immediately adjacent to the infected epithelium showed striking degeneration and atrophy associated with a marked infiltrate of chronic inflammatory cells.

Functional development of small intestine of Japanese quail hatched on MIR orbital station.

The effect of microgravity on functional development of the small intestine of Japanese quails incubated for 2-3 d and hatched on the orbital station MIR was examined. After 5 d of space flight duodenal and jejunal alkaline phosphatase (AP) activity of the experimental group was compared with the AP activity in quails of the same age hatched on the Earth (laboratory controls). Short-term microgravity leading to decreased food intake resulted in significant increase of AP activity in both duodenal and jejunal enterocytes (P<0.001) of the experimental quails. The results suggest that increased AP activity probably reflects the delayed functional development of the small intestine as a consequence of inappropriate food intake during non-physiological conditions of space flight.

[The Sertoli cell nucleolus in domestic and wild animals]. / Jadierko Sertoliho buniek niektorých domácich a divozijucich zvierat.

Sertoli cells produce special microenvironment for developing germ cells; therefore it is assumed that they play primary role in the onset and control of spermatogenesis. In this connection we extended our previous study on the ultrastructure of Sertoli cells in different domestic and wild animals with special regard to nucleolus. Sertoli cells of domestic and wild ruminants possess the typical vesicular nucleolus except for fallow deer, in this species no vesicular nucleolus occurs in Sertoli cells even during the rut. In roe.buck, another wild ruminant with seasonal spermatogenesis, cyclic changes were found in the nucleolus of Sertoli cells. If no spermatogenesis is present, the Sertoli cells have a reticular nucleolus. Membranous vesicles appear in the nucleolus of Sertoli cells of roe-buck at the onset of spermatogenesis 1-2 months before rut. In domestic ruminants with continuous spermatogenesis the vesicular nucleolus in Sertoli cells is present permanently. During postnatal development of bull and ram the vesicular nucleolus appears in Sertoli cells just before the onset of spermatogenesis. In experimental cryptorchidism of bulls a vesicular nucleolus is found in the Sertoli cells. Our observations and experiments support a hypothesis that Sertoli cells have primary role at the onset and the maintenance of spermatogenesis.

[Ultrastructure of frozen and thawed ram sperm]. / Ultrastruktúra zmrazených a rozmrazených spermií barana.

With the aid of a transmission electron microscope we studied the ultrastructure of frozen and thawed ram sperm. The cryoprotective agents do not completely prevent the occurrence of structural changes in spermatozoa. The sperm membrane system is affected. The greatest and most frequent damage is to the acrosome, the cell membrane and mitochondria are less injured. We observed regional differences in the damage to the cell membrane. It is most damaged above the acrosome, least in the postacrosomal area and in the principal piece of the tail. We found considerable variability in the changes among individual spermatozoa from the same ejaculate. The changes are not in the nature of mechanical damage as a result of the occurrence of ice microcrystals. We therefore think that the occurrence of structural changes is dependent on damage at molecular level and consists of a change in the physical-chemical properties of membranes.

[Agglutination and acrosome reaction of ram spermatozoa incubated in the bovine folicular fluid]. / Aglutinácia a akrozómová reakcia spermií barana, inkubovaných v bovinnej folikulárnej tekutine.

Fresh-ejaculated sperm of ram was incubated at a temperature of 38 degrees C in the bovine follicular fluid and homologous blood serum (blood plasma). The spermatozoa were studied in native state under a microscope with phase contrast and as ultra-thin sections under a transmission electron microscope. In the follicular fluid and in the blood serum, strong agglutination of spermatozoa occurred, with its maximum after about two hours. Only the heads of spermatozoa agglutinated, the flagella being loose were arranged parallelly. Although the progressive motility of spermatozoa was not observed, the motility of flagella was not affected. The investigation under the electron microscope showed that the agglutination occurred only in acrosomes of intact spermatozoa. It was also found out that the follicular fluid induced the acrosome reaction of spermatozoa. The course of the acrosome reaction is similar to that in the other mammals: first of all plasma membrane becomes undulated, then it fuses with the outer acrosome membrane, giving origin to vesicles within the entire acrosome, except the equatorial segment. The acrosome reaction was found in about 5% spermatozoa, and therefore it may be assumed that secretions of the oviduct and uterus play their role to induce the acrosome reaction.

[Ultrastructural changesin acrosomes and mitochondria of bull spermatozoa incubated in vitro]. / Ultrastrukturálne zmeny akrozómu a mitochondrii spermii býka inkubovaných in vitro

The structural changes on the acrosomes and mitochondria of bull spermatozoa incubated at 37 degrees C for 1/2--6 hours are described in this paper. The spermatozoa were examined by means of a transmission electron microscope. The changes of the acrosome manifest as acrosome swelling, as the loss of acrosome matter, and as a disintegration of the external acrosome membrane. The internal acrosome membrane remains preserved. The cytoplasmatic membrane of many spermatozoa swells and, later, disintegrates, first over the acrosome and in the middle part of the flagellum. The changes of mitochondria manifest in a decrease of the density of the mitochondrial matrix and in a disintegration of mitochondrial septa. A fibrous material fills the mitochondria, instead of the septa. Some mitochondria lose their content entirely. The described changes are of degenerative character. Despite this, a certain amount of spermatozoa showing such changes on the acrosome and on mitochondria can be found even in fresh semen. The author described other structural changes of mitochondria similar to the changes first described by Hackenbrock (1969) in isolated mitochondria of liver cells. It is assumed that these structural changes of spermatic mitochondria are not of degenerative character and reflect the state of their metabolic activity.