Ultrastructural changes of kidney in diabetic rats.

OBJECTIVES: To demonstrate histopathological changes in diabetic kidneys of the rat on ultrastructural level with focusing on glomerular cellular environment. METHODS: Male Wistar rats were administered streptozotocin and after 8 weeks, their kidneys were studied using qualitative transmission electron microscope analysis. RESULTS: In diabetic kidneys, the presence of irregularly thickened glomerular membrana basalis (GBM), often up to six times as compared to healthy kidney GBM, mainly around peripheral capillary loops, was striking. We also demonstrated lamination and split appearance of GBM, with presence of delicate fibrillar structure, which was absent within normal GBM. Accumulation of mesangial matrix was found only occasionally. CONCLUSION: Model may be useful to better understand the mechanisms that play role in the progression of DM-induced renal disease and to comprehend the morphological changes, which this process leaves within the tissue. The clear association between ultra-structure and functional markers may not be necessary (Fig. 4, Ref. 27).

The first imported human infestation with Furuncular myiasis in man in the Slovakia and current knowledge of myiasis.

INTRODUCTION: Furuncular myiasis is caused by the genus of botfly Dermatobia hominis. It belongs to the family Cuterebridae and is indigenous to Central and South America. OBJECTIVE: to present a case report of the first case of this disease in Slovakia. CURRENT STATE OF PROBLEM SOLUTION: The term myiasis refers to infestation of the host (animal, man) by botfly larvae. Its larvae burrow under the skin. They feed on the host's living tissues and fluids. MateriAl and methods: Patient's history analysis, parasitological examination. RESULTS: A 58-year-old woman after returning from Central America found in the skin above her m. gluteus mayor 2 indurations, which contained three botfly larvae. DISCUSSION: Infestation with botfly larvae Dermatobia hominis is for man annoying and from a health point of view dangerous. CONCLUSION: With proper diagnosis, it is possible to remove the larvae safely from furuncles. The authors point to the first case of imported infestation with Furuncular myiasis caused by botfly Dermatobia hominis in man introduced to Slovakia. They note that increasing tourism spread to the countries with the endemic occurrence of Furuncular myiasis will cause its higher prevalence also in Central European countries (Fig. 5, Ref. 45).

Iron and its relation to glycoconjugates in human globus pallidus.

Iron and eosinophilic, argyrophilic spheroid structures of glycoconjugates are observed in the pallido-nigral system of human and nonhuman primates. In the present study, we map the localization and distribution of ferritin and hemosiderin and their relation to neutral and acidic (sulphated and carboxylated) glycoconjugates in human globus pallidus. We investigated tissues of human globus pallidus without any motor abnormalities and psychiatric symptoms. Acidic (Alcian blue, AB pH 2.5) and neutral glycoconjugates (PAS reaction) showed spheroid deposits with the size of 5-15 µm. Staining reaction utilizing AB (pH 1.0) displays sulphated fibers and highly sulphated (AB pH 0.5) glycoconjugates round deposits. AF/AB pH 2.5 for separating sulphated from carboxylated glycoconjugates revealed the presence of both sulphated and carboxylated glycoconjugates. Perls' and Turnbull's positive reaction showed the presence of Fe(III) and Fe(II) (hemosiderin) and Ag-NOR reaction showed ferric phosphate (ferritin) deposits. Scanning electron microscope with microanalysis revealed the iron in glycoconjugates globular deposits. We suppose the presence of glycoconjugates in the samples is the result of elimination and inactivation of iron as inductor of reactive oxygen species. They can be a useful neuroprotective agent in CNS degradation (Tab. 2, Fig. 7, Ref. 44).

Iron-rich complexes in human spleen in hereditary spherocytosis.

OBJECTIVE: Tissue iron plays an important role in the development of certain diseases. Although it is one of biogenic elements, its excess induces the reactive oxygen species (ROS) formation. The aim of the present work is to examine the protection against free or loosely bound iron from the view of morphology and chemical composition of iron-rich complexes in human spleen tissues with hereditary spherocytosis (HS) by scanning and transmission electron microscope with energy-dispersive microanalysis (EDX). RESULTS: The examination of human spleen tissues by scanning and transmission electron microscope showed covering of iron-rich particles. EDX revealed many iron-rich complexes of multi-element composition in HS samples with sulphur and phosphorus as the major elements. Detection was negative in the reference samples. CONCLUSION: The covering of iron-rich particles can be explained by elimination and isolation of ferritin/iron complexes from surrounding environment to prevent the ROS formation. Sulphur, phosphorus and their compounds are probably the most significant elements that influence the ROS formation (Fig. 5, Ref. 16). Full Text in PDF www.elis.sk.

Iron accumulation in human spleen in autoimmune thrombocytopenia and hereditary spherocytosis.

BACKGROUND: Although the iron is an essential element for the physiological functions of cells, tissues and organs, it is also an important inductor of reactive oxygen species (ROS). MATERIAL AND METHODS: Three groups of human spleen with autoimmune thrombocytopenia (AITP), hereditary spherocytosis (HS) and reference samples stained by haematoxylin and eosin, Perls' reaction for nonheme Fe(III) iron and Alcian blue for glycoconjugates detection were studied. RESULTS: Positive Perls' reaction in both AITP and HS groups was seen. Higher positivity in the HS than in AITP group was observed. HS group showed a higher amount of acidic glycoconjugates deposits than AITP group. Iron overload in HS and AITP leads to overproduction of ROS. CONCLUSION: We suggest that acidic glycoconjugates deposits are involved in antioxidant defence by elimination and restriction of iron as a ROS inducer (Fig. 4, Ref. 19).

Perspectives and complexity of an experimental cancer study. the secrets of tumorigenesis. (To the Gupta's, Chaffer's and Weinberg's "perspectives" and to the nurse's "horizons").

We would like to add to the "mysteriousness", our observations from the application of "identical" BP6 cells either intraperitoneally or subcutaneously. In connection with the concept that tumor development is not only a portrayal of cells proliferation, we could presume that different "environment" will result in structurally different tumors. Morphological differences observed are not significant, but they are present (Fig. 2, Ref. 12). Full Text (Free, PDF) www.bmj.sk.

[Changes in membrane enzymes in warm liver ischemia]. / Zmeny membránových enzýmov pri teplej ischémii pecene.

BACKGROUND: The aim of the study is to perform a morphological analysis of certain membrane enzymes and parenchymal alterations during warm ischemia in the pig liver. METHODOLOGY: Standard hepatectomies were performed in a set of 24 pigs. Intraoperative intravenous (portal vein) hydroxyl radical scavenger Stobadine and Pentoxifylline was administered. Tissue specimens were removed from the margo acutum in interval 10, 60, 80 et 100 minutes. Applied methods of morphological analysis were concentrated on studying the activity of enzymes, which are regarded as cell membrane condition markers. First of them was the alkaline phosphatase, the second studied enzyme was the 5'-mononucleotidase. Detected findings in control group were compared to other two groups. We evaluated each sample using Image Forge Version 1.1 software (PROVER LTD., Bratislava). Applying this software we measured percentage of pre-measured activity levels of both enzymes. RESULTS: In normal pig liver, the activity of alkaline phosphatase was regularly distributed in sinusoid walls in the whole lobule. At 10 minutes after the beginning of the warm ischemia, we have found focal losses of enzyme activity, especially in cells of zone 3 of the liver lobuli. One hour after the beginning of warm ischemia were the local losses of enzyme activity even more pronounced. Decreased bile canalicular 5'-mononucleotidase activity reflects cell damage, especially in pericentral areas after 60 min.of ischemia and a further decrease in activity at all surfaces of the membrane. Total amount of enzyme activity is different. CONCLUSIONS: Morhological findings of enzyme activities showed that zone 2 and 3 of the liver lobule are essential for the organ survival. The study showed, that intravenous administration of Pentoxifylline and Stobadine probably protects the liver from warm ischemia injury.

Warm liver ischemia in experiment and lysosomal markers.

BACKGROUND: The aim of the study is to perform a morphological analysis of certain lysosomal enzymes and parenchymal alterations during warm ischemia in the pig liver. METHODS: Standard hepatectomies were performed in a set of 24 pigs. Intra-operative intravenous (portal vein) Pentoxiphylline and hydroxyl radical scavenger Stobadine was administered. Tissue specimens were removed from the margo acutum in 10 minutes interval. RESULT: In normal pig liver, the acid phosphatase (ACP) activity is in not numerous Kupffer cells and on the biliary pole of hepatocytes, diffusely in the whole parenchyma. One hour after the beginning of warm ischemia, there was an increase in ACP activity in the cytoplasm of hepatocytes. The activity in Kupffer cells could not be detected. Lactate dehydrogenase (LDH) is localized exclusively in the cytoplasmic matrix of liver cells, so only cytoplasmic enzymes leak into the blood plasma. LDH activity has remained low in areas around portal and central veins. CONCLUSION: Morphological findings of enzyme activities showed that zone 2 and 3 of the liver lobule are essential for the organ survival and signs of diffusion of lysosomal enzymes into the cytoplasm of hepatocytes indicate one of the possible explanations for the findings after liver reperfusion. The study showed that intravenous administration of Pentoxiphylline and Stobadine protects the liver from warm ischemia injury (Tab. 2, Fig. 2, Ref. 37). Full Text (Free, PDF) www.bmj.sk.

Cristobalite and Hematite Particles in Human Brain.

Foreign substances get into the internal environment of living bodies and accumulate in various organs. Cristobalite and hematite particles in the glial cells of pons cerebri of human brain with diagnosis of Behhet disease with scanning electron microscopy (SEM), energy-dispersive microanalysis (EDX), and transmission electron microscopy (TEM) with diffraction were identified. SEM with EDX revealed the matter of irregular micrometer-sized particles sometimes forming polyhedrons with fibrilar or stratified structure. It was found in some particles Ti, Fe, and Zn. Some particles contained Cu. TEM and electron diffraction showed particles of cristobalite and hematite. The presence of the particles can be a result of environmental effect, disruption of normal metabolism, and transformation of physiologically iron-ferrihydrite into more stable form-hematite. From the size of particles can be drawn the long-term accumulation of elements in glial cells.

The interaction of beta-adrenoceptor blocking drugs with platelet aggregation, calcium displacement and fluidization of the membrane.

beta-Adrenoceptor blocking drugs interfere with adenosine diphosphate-stimulated platelet aggregation. Alprenolol, exaprolol, Kö 1124 and propranolol inhibited the aggregation, metipranolol decreased the extent and rate of aggregation significantly. Atenolol potentiated the aggregation measured by amplitude significantly. The interaction of beta-adrenoceptor blocking drugs with aggregation correlated with the displacement of calcium ions from binding sites in isolated platelets and the fluidization of the whole platelets and isolated platelet membrane as measured with electron spin resonance of the spin probe. The most potent were highly liposoluble drugs alprenolol, exaprolol, metipranolol and propranolol which increased the calcium displacement and membrane fluidity, the least active was atenolol decreasing these phenomena. The inhibition by beta-adrenoceptor blocking drugs of stimulated platelet aggregation is rather a result of unspecific than specific receptor interaction.

Conformation study of the membrane models by the Maxwell displacement current technique and oxidative stress.

The role of biological membranes as a target in biological radiation damage is still unclear. Recently much attention has been paid to the dynamic behaviour of the cell membrane. Maxwell displacement current technique (MDC) provides new possibility of conformation study of the membrane models. Oxidative stress can impair macromolecules in the cell on a molecular level. MDC technique enables to study the changes in molecular orientations and/or conformations of cell membranes. The combination of different methods in structural biology can clarify membrane chemical and physical properties.

Chondroitin sulphates A, B and C, collagen types I-IV and fibronectin in venous sinus of the red pulp in human spleen.

The morphological relationship of chondroitin sulphates A, B, and C, collagen types I-IV and fibronectin in the wall of venous sinuses of the red pulp in human spleen has not been a focus of interest among morphologists. Regarding the hypothesis that the structure of the spleen lends it the function of a blood filter the substances described in our study might play a significant role in the functional morphology. Of 146 human spleen surgical specimens, groups of 12 specimens each were examined under a light microscope using the method of antibodies against fibronectin, against collagen types I-IV and against chondroitin sulphates A, B, and C. The sections of the red pulp of human spleen stained with hematoxylin and eosin provided limited information about the wall of the sinuses. Chondroitin sulphates A and B were observed on the surface of sinus-lining cells (SLC), and fibronectin was detected on the surface of the annular fibers. Collagen type 11 was observed almost in the same places as chondroitin sulphates A and B. Collagen type IV was present in annular fibers of the wall of the sinus and in the basement membrane, like fibronectin. Chondroitin sulphate was not present in the walls of sinuses. Binding of antibodies against chondroitin sulphate A and against chondroitin sulphate B indicates the presence of chondroitin sulfates on the surface of SLC, where they probably play a role in helping the human organism to recognize alien and self substances. The presence of chondroitin,sulphates A and B probably affects inhibition of binding of cells with collagen type I, but not with fibronectin.

Combined chronic toxicity/carcinogenicity study of the fungicide trimorphamide in rats.

Trimorphamide, a new fungicide, was fed over 2 years to groups of 50 male and 50 female rats at dietary levels of 0, 67, 330 and 665 mg/kg. No adverse effects on mortality, haematology, urinalysis, organ weights, incidence and severity of tumors were observed. Isolated observations of abnormalities in blood chemistry suggested the possibility of certain renal damage in the highest dosage group; however, the clinical significance of these findings is dubious. According to the results of the present study, 67 mg/kg in the diet can be considered as the "no observed effect level".

Metastasizing adenocarcinoma of the female prostate (Skene's paraurethral glands). Histological and immunohistochemical prostate markers studies and first ultrastructural observation.

The case of a 46-year-old women with well-differentiated adenocarcinoma of the female prostate (Skene's paraurethral glands and ducts) with inguinal metastases is reported. Besides adenocarcinomatous structures, also more solid parts of the tumor and anaplastic regions with dark cells were found on histological examination. Clear cancerous cells were typical for glandular and solid tumor parts. The cancerous cells showed distinct immunohistochemical positivity of prostate specific antigen (PSA) and prostate (specific) acid phosphatase [P(S)AcP]. These are the first published results of electron microscopic examination of formalin fixed tissue showing the ultrastructure of female prostate carcinoma, comparable to that of the male prostate carcinoma. In the female, similar to the male, the prostate carcinoma probably originates from the secretory (luminal) cells of the female prostatic glands.

Delayed effects of neonatal exposure to Tween 80 on female reproductive organs in rats.

Neonatal female rats were injected ip (0.1 ml/rat) with Tween 80 in 1, 5 or 10% aqueous solution on days 4-7 after birth. Treatment with Tween 80 accelerated maturation, prolonged the oestrus cycle, and induced persistent vaginal oestrus. The relative weight of the uterus and ovaries was decreased relative to the untreated controls. Squamous cell metaplasia of the epithelial lining of the uterus and cytological changes in the uterus were indicative of chronic oestrogenic stimulation. Ovaries were without corpora lutea, and had degenerative follicles.

Morphologic changes in the gastric mucosa of rats after administration of N-methyl-N'-nitro-N-nitrosoguanidine. Electronmicroscopic study.

Electron microscopic findings of the effects of exposure to the cancerogenic substance N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) upon the gastric mucosa of rats are presented. MNNG was administered in drinking water at a concentration of 83 mg/l over a period of eight weeks. The resulting findings included occurrence of myelin figures in membrane structures of epithelial cells, occasional disappearance of nuclear membrane, numerous intranuclear cytoplasmic pseudoinclusions, and increased number of perichromatin and interchromatin granules and of nuclear corpuscles. Narrowing of the layer of peripheral condensed chromatin and broadening of the layer of dispersed chromatin were frequently observed. In the cytoplasm of epithelial cells, production of autophagous vacuoles was detected in differentiated types of cells, the presence of great amount of polysomes and profiles of rough endoplasmic reticulum diversely deformed was seen in the undifferentiated neck cells. These cells presented a marked proliferative activity. Some of the changes described in the nuclei of epithelial cells were also observed in different mesenchymal elements in the lamina propria mucosae. The demonstrated changes are indicative of both regressive and progressive effects of the cancerogen used in our experiment.

Chronic lymphoproliferative disease of large granular lymphocytes.

An 80-year-old patient has been followed for hepato- and splenomegaly, hemolytic anemia, neutropenia with lymphocytosis with large granular lymphocyte predominance in his peripheral blood, with infiltration of bone marrow, liver and probably also spleen. Determination of surface markers of proliferating lymphocytes in peripheral blood showed a mixed phenotype of T suppressor/cytotoxic and natural killer cells (SIg-, E+, T3+, T8+, EAC+, Leu7-, N901+, NK9+, VIB C5 and VIB E3-, Ia-). An in vitro cytotoxic test showed the functional inactivity of the cells tested also after human leukocyte interferon stimulation. Chromosomal analysis neither of peripheral blood lymphocytes nor of bone marrow cells proved the monoclonality marker. Following long-term prednisone therapy, the improvement of anemia, later also neutropenia accompanied by the decrease of lymphocytes has been achieved. As the disease present in our patient was distinguished only in recent years and in our country has not been reported yet, the details on its clinical, morphologic, hematologic, cytogenetic and mainly immunophenotypic characteristics are given in this paper. The problems concerning classification of the disease and determination of its biological nature are discussed.

Immunohistochemical identification of mastocytes.

Antibodies to mastocytes were localized in cells and tissues following formaldehyde or paraformaldehyde fixation and embedding into paraffin or Durcupan ACM, and after chymotrypsin treatment of the sections. Immunomicroscopic examination yielded a more distinct picture of the amount and localization of mastocytes in different tissues.

Immunohistochemical staining using coupled immunofluorescence and immunocomplex technique.

An immunomicroscopic method combining the use of fluorescent antibodies and enzyme immunocomplex technique is presented. The basic steps of this procedure are: 1. Rabbit antiserum (IgG) to the antigen to be determined, 2. FITC-conjugated antirabbit globulin, 3. Peroxidase + antiperoxidase complex, 4. 3,3-diaminobenzidine + H2O2. The method renders it possible to study simultaneously in the same section antigens and antibodies that are to be localized, and that both by immunofluorescent and immunoenzymatic techniques, it provides means for further elimination of some of the deficiencies of immunofluorescent technique, and in suitable cases it also makes electronoptic examination available.

Histochemistry and cytochemistry of thiamine pyrophosphatase (TPP-ase 3.6.1.) in the rat gastric mucosa.

The paper deals with the histochemical and cytochemical distribution of thiamine pyrophosphatase (TPP-ase 3.6.1.) in the normal rat gastric mucosa using the method of NOVIKOFF and GOLDFISCHER (1961). In light microscopy, activity was demonstrated only in the GOLGI apparatus of the foveolar mucous cells; in the other specialized gastric mucosal cells no activity was established. At electron microscopic level, the reaction product was localized in the membranes of the cisternae of the GOLGI apparatus of foveolar mucous cells, undifferentiated neck cells, parietal cells, chief cells and in endocrine cells. The reaction product was not found in the endoplasmic reticulum of any of the above mentioned types of cells with the exception of parietal cells. The deposits of the reaction product in the cytoplasmic membrane, in the capillary endothelium and on the surface of the plasma membrane of foveolar mucous cells represents the sites of activity of alkaline phosphatase. The examination of the TPP-ase in the gastric mucosa inables us to estimate the activity of the GOLGI apparatus of gastric mucosal cells in the norm and in different pathological conditions connected with its changes.