IN BRIEF: Understanding the establishment of post-fertilization totipotency has broad implications for modern biotechnologies. This review summarizes the current knowledge of putative egg components governing this process following natural fertilization and after somatic cell nuclear transfer. ABSTRACT: The mammalian oocyte is a unique cell, and comprehending its physiology and biology is essential for understanding fertilization, totipotency and early events of embryogenesis. Consequently, research in these areas influences the outcomes of various technologies, for example, the production and conservation of laboratory and large animals with rare and valuable genotypes, the rescue of the species near extinction, as well as success in human assisted reproduction. Nevertheless, even the most advanced and sophisticated reproductive technologies of today do not always guarantee a favorable outcome. Elucidating the interactions of oocyte components with its natural partner cell - the sperm or an 'unnatural' somatic nucleus, when the somatic cell nucleus transfer is used is essential for understanding how totipotency is established and thus defining the requirements for normal development. One of the crucial aspects is the stoichiometry of different reprogramming and remodeling factors present in the oocyte and their balance. Here, we discuss how these factors, in combination, may lead to the formation of a new organism. We focus on the laboratory mouse and its genetic models, as this species has been instrumental in shaping our understanding of early post-fertilization events.

• MeSH
• Cell Nucleus * physiology   MeSH
• Embryonic Development MeSH
• Humans MeSH
• Mice MeSH
• Oocytes physiology   MeSH
• Mammals MeSH
• Semen * MeSH
• Spermatozoa physiology   MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Male MeSH
• Mice MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Review MeSH

Spermiogenesis in progenetic and adult stages of Archigetes sieboldi Leuckart, 1878, a tapeworm parasitic in oligochaetes and fish respectively, has been examined using transmission electron microscopy and cytochemical staining for glycogen. General pattern of spermiogenesis is essentially like that of other caryophyllideans, i.e., apical dense material in the zone of differentiation in the early stages of spermiogenesis, rotation of free flagellum and a flagellar bud, and proximo-distal fusion. Interestingly, rotation of a free flagellum and flagellar bud to the median cytoplasmic process (MCP) has been observed unconventionally at > 90° only in progenetic stages. Typical striated roots associated with the centrioles occur rarely in A. sieboldi, and only in form of faint structures in advanced stages of spermiogenesis. In contrast to most caryophyllideans studied to date, penetration of the nucleus into the spermatid body has started before the fusion of the free flagellum with the MCP. This feature has been reported rarely but exclusively in the family Caryophyllaeidae. The unipartite mature spermatozoon of A. sieboldi is composed of one axoneme of the 9 + '1' trepaxonematan pattern with its centriole, parallel nucleus, and parallel cortical microtubules which are situated in a moderately electron-dense cytoplasm with glycogen particles. An unusual arrangement of cortical microtubules in the two parallel rows in region I of the spermatozoon is described here for the first time in the Caryophyllidea. Ultrastructural data on spermiogenesis and the spermatozoon in A. sieboldi from tubuficids and carp are compared and discussed with those in other caryophyllideans and/or Neodermata.

• MeSH
• Axoneme ultrastructure   MeSH
• Staining and Labeling MeSH
• Cell Nucleus physiology   MeSH
• Cestoda ultrastructure   MeSH
• Cestode Infections veterinary   MeSH
• Flagella physiology   MeSH
• Glycogen analysis   MeSH
• Carps parasitology   MeSH
• Fish Diseases parasitology   MeSH
• Spermatids cytology   ultrastructure   MeSH
• Spermatogenesis physiology   MeSH
• Microscopy, Electron, Transmission MeSH
• Animals MeSH
• Check Tag
• Male MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH

The dynamics of nuclear morphology changes during apoptosis remains poorly investigated and understood. Using 3D time-lapse confocal microscopy we performed a study of early-stage apoptotic nuclear morphological changes induced by etoposide in single living HepG2 cells. These observations provide a definitive evidence that nuclear apoptotic volume decrease (AVD) is occurring simultaneously with peripheral chromatin condensation (so called "apoptotic ring"). In order to describe quantitatively the dynamics of nuclear morphological changes in the early stage of apoptosis we suggest a general molecular kinetic model, which fits well the obtained experimental data in our study. Results of this work may clarify molecular mechanisms of nuclear morphology changes during apoptosis.

• MeSH
• Single-Cell Analysis methods   MeSH
• Apoptosis physiology   MeSH
• Cell Nucleus physiology   ultrastructure   MeSH
• Hep G2 Cells MeSH
• Time-Lapse Imaging methods   MeSH
• Chromatin chemistry   metabolism   ultrastructure   MeSH
• Kinetics MeSH
• Microscopy, Confocal MeSH
• Humans MeSH
• DNA Packaging MeSH
• Models, Theoretical * MeSH
• Organelle Size physiology   MeSH
• Imaging, Three-Dimensional MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

In polarized motile cells, stress fibers display specific three-dimensional organization. Ventral stress fibers, attached to focal adhesions at both ends, are restricted to the basal side of the cell and nonprotruding cell sides. Dorsal fibers, transverse actin arcs, and perinuclear actin fibers emanate from protruding cell front toward the nucleus and toward apical side of the cell. Perinuclear cap fibers further extend above the nucleus, associate with nuclear envelope through LINC (linker of nucleoskeleton and cytoskeleton) complex and terminate in focal adhesions at cell rear. How are perinuclear actin fibers formed is poorly understood. We show that the formation of perinuclear actin fibers requires dorsal stress fibers that polymerize from focal adhesions at leading edge, and transverse actin arcs that are interconnected with dorsal fibers in spots rich in α-actinin-1. During cell polarization, the interconnected dorsal fibers and transverse arcs move from leading edge toward dorsal side of the cell. As they move, transverse arcs associate with one end of stress fibers present at nonprotruding cell sides, move them above the nucleus thus forming perinuclear actin fibers. Furthermore, the formation of perinuclear actin fibers induces temporal rotational movement of the nucleus resulting in nuclear reorientation to the direction of migration. These results suggest that the network of dorsal fibers, transverse arcs, and perinuclear fibers transfers mechanical signal between the focal adhesions and nuclear envelope that regulates the nuclear reorientation in polarizing cells.

• MeSH
• Actinin physiology   MeSH
• Actins physiology   MeSH
• Cell Nucleus physiology   MeSH
• Cell Line MeSH
• Mechanotransduction, Cellular physiology   MeSH
• Fibroblasts physiology   MeSH
• Focal Adhesions physiology   MeSH
• Stress Fibers physiology   MeSH
• Rats MeSH
• Humans MeSH
• Cell Movement physiology   MeSH
• Movement physiology   MeSH
• Cell Polarity physiology   MeSH
• Animals MeSH
• Check Tag
• Rats MeSH
• Humans MeSH
• Animals MeSH
• Publication type
• Editorial MeSH

Intracellular and extracellular mechanical forces play a crucial role during tissue growth, modulating nuclear shape and function and resulting in complex collective cell behaviour. However, the mechanistic understanding of how the orientation, shape, symmetry and homogeneity of cells are affected by environmental geometry is still lacking. Here we investigate cooperative cell behaviour and patterns under geometric constraints created by topographically patterned substrates. We show how cells cooperatively adopt their geometry, shape, positioning of the nucleus and subsequent proliferation activity. Our findings indicate that geometric constraints induce significant squeezing of cells and nuclei, cytoskeleton reorganization, drastic condensation of chromatin resulting in a change in the cell proliferation rate and the anisotropic growth of cultures. Altogether, this work not only demonstrates complex non-trivial collective cellular responses to geometrical constraints but also provides a tentative explanation of the observed cell culture patterns grown on different topographically patterned substrates. These findings provide important fundamental knowledge, which could serve as a basis for better controlled tissue growth and cell-engineering applications.

• MeSH
• Models, Biological * MeSH
• Cell Nucleus physiology   ultrastructure   MeSH
• Mechanotransduction, Cellular physiology   MeSH
• Hep G2 Cells MeSH
• Humans MeSH
• Cell Communication physiology   MeSH
• Computer Simulation MeSH
• Cell Proliferation physiology   MeSH
• Cell Size * MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

We have explored the potential relationship between ploidy level, DNA content (pg DNA nucleus(-1)), and dimensional characteristics, such as volume (μm(3)), surface area (μm(2)), and 3-D structure of erythrocyte nuclei in a series of fish ploidy level models using Feulgen image analysis densitometry, flow cytometry, and confocal laser scanning microscopy. The species were diploid tench (Tinca tinca) (2n), Cuban gar (Atractosteus tristoechus) (2n), triploid tench (3n), evolutionary tetraploid sterlet (Acipenser ruthenus) (4n), evolutionary octaploid Siberian sturgeon (A. baerii) (8n), triploid Siberian sturgeon exhibiting dodecaploidy (12n), evolutionary 12n shortnose sturgeon (A. brevirostrum), and experimentally obtained sturgeon hybrids that were tetraploid, hexaploid (6n), heptaploid (7n), octaploid, decaploid (10n), dodecaploid and/or tetradecaploid (14n). Increase in ploidy was accompanied by growth of the nucleus and an increase in the number of flattened ellipsoid nuclei with increased transverse diameter. The volume (Vvoxel ) of erythrocyte nuclei, as the sum of voxels calculated from live cells, seems more accurate than volume (Vaxis ) calculated from measuring the major and minor axis, especially at higher and odd ploidy levels. Data of absolute and relative DNA content were in agreement with previously published reports. Species of the same ploidy level, but differing in DNA content, had a similar mean erythrocyte nuclear volume (Vvoxel ), as demonstrated in sterlet and a hybrid of sterlet and beluga (48.3 and 48.9 μm(3), respectively), with a respective mean DNA content of 3.74 and 3.10 pg DNA nucleus(-1). A similar relationship was found for the ploidy 6n, 10n, 12n. The surface-to-volume ratio decreased non-linearly with increasing ploidy. The DNA in erythrocyte nuclei appeared to be more densely packed with increase in ploidy level.

• MeSH
• Cell Nucleus genetics   physiology   MeSH
• Cyprinidae genetics   MeSH
• DNA analysis   MeSH
• Erythrocytes cytology   MeSH
• Polyploidy MeSH
• Imaging, Three-Dimensional MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

• MeSH
• Cell Nucleus physiology   chemistry   MeSH
• Financing, Organized MeSH
• Histone Deacetylases physiology   genetics   metabolism   MeSH
• Ion Channels physiology   physiopathology   MeSH
• Calcineurin physiology   genetics   metabolism   MeSH
• Calmodulin physiology   genetics   metabolism   MeSH
• Cardiomegaly etiology   metabolism   physiopathology   MeSH
• Myocytes, Cardiac enzymology   physiology   MeSH
• Humans MeSH
• Sarcolemma enzymology   physiology   metabolism   MeSH
• Cardiac Electrophysiology MeSH
• Heart Failure etiology   metabolism   physiopathology   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Review MeSH

Increased cytosolic calcium ([Ca2+]i) and nitric oxide (NO) are suggested to be associated with apoptosis that is a main feature of many liver diseases and is characterized by biochemical and morphological features. We sought to investigate the events of increase in [Ca2+]i and endoplasmic reticulum (ER) calcium depletion by thapsigargin (TG), a selective inhibitor of sarco-ER-Ca2+ -ATPases, in relation to NO production and apoptotic and necrotic markers of cell death in primary rat hepatocyte culture. Cultured hepatocytes were treated with TG (1 and 5 micromol/L) for 0-24 or 24-48 h. NO production and inducible NO synthase (iNOS) expression were determined as nitrite levels and by iNOS-specific antibody, respectively. Hepatocyte apoptosis was estimated by caspase-3 activity, cytosolic cytochrome c content and DNA fragmentation, and morphologically using Annexin-V/propidium iodide staining. Hepatocyte viability and mitochondrial activity were evaluated by ALT leakage and MTT test. Increasing basal [Ca2+]i by TG, NO production and apoptotic/necrotic parameters were altered in different ways, depending on TG concentration and incubation time. During 0-24 h, TG dose-dependently decreased iNOS-mediated spontaneous NO production and simultaneously enhanced hepatocyte apoptosis. In addition, TG 5 micromol/L produced secondary necrosis. During 24-48 h, TG dose-dependently enhanced basal NO production and rate of necrosis. TG 5 micromol/L further promoted mitochondrial damage as demonstrated by cytochrome c release. A selective iNOS inhibitor, aminoguanidine, suppressed TG-stimulated NO production and ALT leakage from hepatocytes after 24-48 h. Our data suggest that the extent of the [Ca2+]i increase and the modulation of NO production due to TG treatment contribute to hepatocyte apoptotic and/or necrotic events.

• MeSH
• Apoptosis physiology   drug effects   MeSH
• Cell Death drug effects   MeSH
• Cell Nucleus drug effects   physiology   MeSH
• Financing, Organized MeSH
• Guanidines pharmacology   MeSH
• Hepatocytes physiology   drug effects   MeSH
• Caspase 3 metabolism   MeSH
• Rats MeSH
• Cells, Cultured MeSH
• Mitochondria physiology   drug effects   MeSH
• Nitric Oxide biosynthesis   MeSH
• Sarcoplasmic Reticulum Calcium-Transporting ATPases antagonists & inhibitors   metabolism   MeSH
• Nitric Oxide Synthase Type II antagonists & inhibitors   metabolism   MeSH
• Thapsigargin pharmacology   MeSH
• Calcium physiology   MeSH
• Animals MeSH
• Check Tag
• Rats MeSH
• Animals MeSH

• MeSH
• Cell Biology trends   MeSH
• Biology history   trends   MeSH
• Cell Membrane physiology   genetics   metabolism   MeSH
• Cell Nucleus physiology   genetics   MeSH
• Cellular Structures physiology   metabolism   MeSH
• Centrosome physiology   MeSH
• Chloroplasts genetics   metabolism   MeSH
• Golgi Apparatus physiology   MeSH
• Genetics, Medical methods   trends   MeSH
• Mitochondria genetics   metabolism   MeSH
• Peroxisomes physiology   MeSH

Movement of labelled plasmid DNA relative to heterochromatin foci in nuclei, visualized with HP1-GFP, was studied using live-cell imaging and object tracking. In addition to Brownian motion of plasmid DNA we found a pronounced, non-random movement of plasmid DNA towards the nearest HP1 focus, while time-lapse microscopy showed that HP1 foci are relatively immobile and positionally stable. The movement of plasmid DNA was much faster than that of the HP1 foci. Contact of transgene DNA with an HP1 focus usually resulted in cessation of the directional motion. Moreover, the motion of plasmid DNA inside the heterochromatin compartment was more restricted (limited to 0.25 microm) than when the plasmid DNA was outside heterochromatin (R = 0.7 microm). Three days after transfection most of the foreign labelled DNA colocalized with centromeric heterochromatin.

• MeSH
• Biological Transport physiology   genetics   MeSH
• Cell Nucleus physiology   MeSH
• Chromosomal Proteins, Non-Histone physiology   genetics   MeSH
• DNA physiology   genetics   MeSH
• Financing, Organized MeSH
• Heterochromatin MeSH
• Humans MeSH
• Microscopy MeSH
• Cell Line, Tumor MeSH
• Plasmids physiology   genetics   MeSH
• Transfection MeSH
• Check Tag
• Humans MeSH