Differentiation of epidermal growth factor-responsive neural precursor cells within neurospheres.

Genetically unmodified neural precursor cells (NPCs) derived from rate embryonic cortex were stimulated with Long-epidermal growth factor in vitro to form multicellular neurospheres. In our study, histology and transmission electron microscopy was for the first time utilized for analysis of three-dimensional neurospheres. Immunohistochemical phenotypization carried out on paraffin-embedded section revealed that most cells in the neurosphere expressed high levels of vimentin that is normally found in immature neural cells. Majority of glial cells expressed glial fibrillary acidic protein (GFAP) that is a specific marker for differentiated astrocytes. On the other hand, myelin basic protein (MBP), a marker of oligodendrocytes, was found only in a discrete subpopulation of glial cells. Similarly, electron microscopy recognized only rate myelin sheaths surrounding axons. Some cells displayed neuron-like morphology and were labelled with antibodies recognizing the neuronal antigens: 210 kDa neurofilaments (NF 210) and microtubule associated protein-2 (MAP-2). Surprisingly well-developed interneuronal synapses were found electronmicroscopically. The presence of synaptic vesicles was also confirmed by immunohistochemical detection of synaptophysin. Our results show that NPCs in neurospheres gradually differentiate into glial and neuronal cells.

Cerebrospinal fluid dissemination of fetal neural isografts in brain of adult rats.

Our previous studies tend to document the temporary invasiveness of intraparenchymal fetal isografts. The present study exploits a model situation in which contacted host brain surface remains undamaged. We analyzed the behavior of transplant fragments that accidentally disseminated in cerebrospinal fluid pathways. The transplant seeding occurred in 7 cases out of 85 cases of healthy solid isografts taken from various parts of fetal brain. Fragments were found either as free (floating or simply lying close to the surface) or attached. Some attached fragments invasively penetrated the brain parenchyma. This speaks in favor of invasive properties of fetal neural transplants.

Cultivation of neural EGF-responsive precursor cells.

We demonstrated that proliferation of dissociated neural E14 and E19 rat precursor cells could be induced by Long-EGF. Dividing EGF-responsive neural progenitor cells stimulated by Long-EGF formed spherical multicellular clusters (neurospheres) which may reach macroscopical size. We have studied the inner organization of cells in semithin sections and revealed that the cell population within the neurosphere is not uniform. These are original findings as other researchers did not process neurospheres histologically. Cells located in a central area possessed neuron-like morphology whereas peripheral cells where differentiated to a lesser degree. Generally, we have observed several apoptotic cells or apoptotic bodies per section. Moreover, the central portion of the neurosphere contained degenerating cells that probably die from worsened nutrition conditions in the large neurosphere. After plating the neurosphere in serum-supplemented medium, cells began to migrate radially from the edge of the neurosphere and differentiate. The cells lying in the vicinity to the cluster mimicked radial glia whereas the cells located at the periphery of the colony took morphology of astroglial cells. These observations suggest EGF-responsive neural precursor cells retained their ability to produce both neuronal and glial phenotypes after prolonged cultivation period.

Skin side-effects of amiodarone therapy.

Amiodarone is a antiarrhythmic drug with many side-effects which include also the induction of photosensitivity and the development of greyish-blue pigmentations on the skin exposed to light. The authors examined 64 patients treated with amiodarone and 32 controls by means of a phototest using a xenon lamp. In 9.4% photosensitivity was recorded, in 9.4% there were greyish-blue hyperpigmentations in the face and on the back of the hands. Histological identification of these hyperpigmentations including electron microscopy was also performed.

Liver damage induced by intrabiliary turpentine in rats.

Inflammation of the rat bile duct induced by administration of turpentine into it has been used to study the influence of the impaired duct on liver function. Turpentine was dissolved in olive oil 1:1000 and 1:500. A 2 h ligation of the bile duct was used to promote a local effect. Contemporary groups of intact, sham-operated, control rats (given 0.9% NaCl by intrabiliary injection) and animals with total chronic obstruction were compared to assess the significance of changes. Serum concentrations of total and conjugated bilirubin, cholesterol and creatinine, activities of S-alanine-aminotransferase, S-aspartate aminotransferase and alkaline phosphatase, mortality of rats, and also total body weight compared with the weight of the liver, were investigated on days 1, 4, 8, 12, 16, 32 and 64 after surgery and turpentine, or following ligation of the bile duct. An increase in bilirubin and cholesterol, an augmentation of enzymatic activity and the histological changes were indicative of hepatotoxicity or cholestasis. The turpentine concentration--effect, manifested in body-weight change, suggests some specificity of the effect. There were no changes in serum creatinine arterial blood pressure, heart rate or portal blood pressure, when turpentine was administered by the intrabiliary route. These results suggest primary liver damage.

[Ultrastructure of the epidermis in laboratory rats under normal conditions and after percutaneous administration of an organophosphate]. / Ultrastruktura epidermis laboratorního potkana v norme a po perkutánním podání organofosfátu.

For the study of the normal epidermal layers of the stratum basale, stratum spinosum, stratum granulosum and (male rats at the age of 2 years) were used. Skin samples were obtained from the spinal area and processed by the method of transmission electron microscopy to get semithin and ultrathin sections. An electron microscopical examination preceded by an evaluation of the semithin sections in the light microscope showed that the epidermis of both age groups contained keratinocytes and Langerhans cells. Keratinocytes are typically arranged in epidermis layers of the stratum basale, stratum spinosum, stratum granulosum and stratum corneum. These layers are of different ultrastructure. The Langerhans cells are localized suprabasally in the epithelium. In the course of keratinization, prominent ultrastructural changes in keratinocytes occur. The cell organelles of keratinocytes including the nucleus disappear and the formation of specific cell components--prominent bundles of tonofilaments, keratohyalin granules and keratinosomes--can be observed. Keratinized cells of the stratum corneum exhibit a homogeneous or fibrillar inner structure. Langerhans cells contain characteristic cytoplasmic organelles--Langerhans granules. The epidermis of old rats differs from that of young animals in a special arrangement of the dermo-epidermal junction. The most typical change (in comparison with the epidermis of young rats) is the presence of multiplied lamina densa. The qualitative study of the normal epidermis of the laboratory rat is completed by morphometric analysis. The measurements concern keratinocytes of the stratum basale. Cell dimensions under the light microscope as well as some cell organelles in ultrathin sections are evaluated. Another part of the study is aimed to estimate morphological changes in the epidermis of rats which were percutaneously administered a toxic organophosphate. In this experiment, the tissue processing for electron microscopy could be realized only when the exposed epidermal area had been previously decontaminated. A group of intoxicated animals was therefore compared with a control group, the epidermis of which was affected by the decontaminating agent only. The ultrastructural changes noted in the epidermis of intoxicated animals show no specificity. Damaged membrane organelles and vacuoles in keratinocytes, dense cells, dilated epidermal intercellular spaces containing the material of medium density, etc. are present. Furthermore, the above-mentioned changes proved to be manifestation of a combined effect of the proper toxic substance and the decontaminating agent.

[The effect of a diet containing long-chain fatty acids on the formation of insoluble elastin in the rat aorta]. / Vliv diety obsahujici mastné kyseliny s dlouhým retezcem na tvorbu nerozpustného elastinu v aorte potkanú.

The paper deals with the influence of long-chain fatty acids on the formation of insoluble elastin in vivo. Oil isolated from the sea fish named flounder has been chosen as the source of those long-chain fatty acids. The diet containing the flounder oil was administered to pregnant rat females. The influence of fatty acids on the formation of insoluble elastin was evaluated according to the percentage changes of insoluble elastin in the aortas of newborn rats and to the changes of the amount of desmosine cross-links in isolated elastin. The administration of the diet with long-chain fatty acids causes no changes in the content of elastin in the aortas of the new-born rats, meanwhile the amount of desmosine increases considerably. The results on the aortas of new-born rats shown here are in good agreement with the changes on the aortas of 3-month-old rats published in a previous paper.

Structural and functional changes after the administration of tetrachlormethane in the liver of rats fed on diets with different protein contents.

Morphological and biochemical changes characterizing the degree of liver damage and the development of liver repair were studied in rats fed 21 days on a low protein diet (LPD), a standard diet (SLD) and a high protein diet (HPD) and then given a single i.p. injection of tetrachlormethane (CCl4) in a dose of 0.75 ml/kg body weight. The HPD was found to increase sensitivity to CCl4, but it also promoted the liver repair process, as seen from the increment in liver DNA synthesis and the total DNA content of the liver, increased ploidy of the hepatocytes and growth of the size of their nuclei and of the hepatocytes themselves. An increase in the total surface area of the membranes of the granular endoplasmic reticulum and the inner and outer membrane of the mitochondria, but a decrease in the surface area of the membranes of the smooth endoplasmic reticulum, were also observed after the administration of CCl4. The LPD raised liver resistance to CCl4, but the development of liver repair activity differed from the process after the SLD and HPD, since polyploidy of the hepatocytes (especially the growth of octaploid cells) predominated and there was also an increase in the number of binuclear hepatocytes. Cell hypertrophy was expressed less in rats fed on the LPD than in animals given the HPD. As far as liver repair was concerned, the HPD showed no explicit advantages over the SLD.