Changes in the ultrastructure of cytoskeleton and nuclear matrix during HaCaT keratinocyte differentiation.

The cellular scaffold that comprises nuclear matrix and cytoskeleton provides mechanical support for the cell and plays a crucial role in motility, cellular signaling, regulation of gene transcription and DNA replication. In this study we examined the structure of cytoskeleton and nuclear matrix in the keratinocyte cell line HaCaT using a recently developed technique, embedment-free electron microscopy. With this method the three-dimensional structure of cellular scaffold is visualized in the cells extracted from soluble proteins and the chromatin. In actively proliferating cells the cytoskeleton appeared to consist of a continuous meshwork of 10--15 nm filaments with a smaller amount of thin (5 nm) and ultrathin (1--2 nm) filaments. In contrast to what could be expected from earlier immunofluorescence and electron microscopy studies, the cytoskeleton in HaCaT keratinocytes did not consist of superposed autonomous networks of different filaments but was a highly integrated, continuous structure filling whole cytoplasmic territory. Moreover, cytoskeletons of adjacent cells were in a direct physical contact. Nuclear matrix consisted of globular ribonucleoprotein aggregates attached to the meshwork of 20--40 nm filaments. Nuclear envelope was firmly fastened to the cytoskeleton. In keratinocytes induced to differentiation by calcium switch both the cytoskeleton and nuclear matrix were drastically rearranged and comprised a monomorphic, dense and regular meshwork of 10--15 nm filaments. Our data underscore the fact that in HaCaT keratinocyte monolayer in vitro, and probably also in the epidermis in vivo, the nuclear matrices and the cytoskeletons of adjacent cells seemed to form a continuous, highly ordered structure which is rapidly rearranged during cell differentiation. This feature may be crucial for the understanding of how the signal initiated by, e.g. mechanical forces generated through the cell--cell and cell--matrix interaction is transmitted to the nucleus producing ultimately changes in the pattern of gene expression.

Alterations in rat's brain capillaries in a model of focal cerebral necrosis.

Focal brain compression causes cerebral tissue damage. In this study we followed alterations in capillary ultrastructure in the rat cortex and neurohypophysis caused by 40 mm Hg compression for 15 minutes. One day after experiment we observed clogging of capillaries, accumulation of collagen fibrills under the basement membrane and necrosis or apoptosis of endothelial cells. Four days after it the basement membrane was multiplicated, blurred and thickened. In the neurohypophysis the formation of vessels lined with the atypical continuous endothelium was seen. There was also evidence for the migration of pericytes through the blurred basement membrane and the differentiation of pericytes into endothelial cells. Thus, vascular injury in the compressed brain is followed by a highly ordered sequence of processes in the basement membrane and perivascular cells leading to capillary repair.

Electron microscopy studies on experimental diabetes and cerebral ischemia in the rat brain.

Male Wistar rats were subjected to streptozotocin administration (85 mg/kg i.p.) and to cerebral air embolia with common carotids ligation. Electron microscope studies showed dark neurons, degeneration of endothelial cells and changes in basement membrane of brain capillaries, and changed astroglia in diabetic rats. Our results seem to support our previous findings in light microscopy and correspond with some others authors' suggestions that diabetes leads to chronic, generalized pathologic process in diabetic-rat brain, not-only dependent on vascular pathology, but which may be related to an oxidative/metabolic stress leading to a death of neurons in necrotic or apoptotic way.

Characterization of thiamine pyrophosphatase positive phagocytic cells in the neural lobe of rat pituitary.

Here, using a histochemical staining for a microglia/phagocyte marker TPP-ase (Murabe, Sano 1981), and an electron microscopy we characterized the population of pituitary phagocytic cells activated by cerebral ischemia. An intense thiamine pyrophosphatase (TPP-ase) activity was demonstrated in glial cells and some cells of blood vessels of neural lobe, late period (12 months) after experimental ischemia. TPP-ase positive cells were ultrastructurally identified as pituicytes, microglia, pericytes and perivascular cells. The product characteristic for TPP-ase activity was seen on plasma membrane of these cells. Our electron-microscopic histochemical results provide strong support for a role of pituicytes, pericytes and perivascular cells as a phagocytic cells involved in mechanism of elimination of ischemically damaged axonal endings in neural lobe.