Acetaminophen (APAP) overdose is the most common cause of acute liver failure in humans. Non-alcoholic fatty liver disease is the most frequent chronic liver disease in developed countries. The aim of our work was to compare the effect of APAP on intact rat hepatocytes and hepatocytes isolated from steatotic liver in primary cultures. Male Wistar rats were fed with standard diet (10 % energy from fat) and high-fat diet (71 % energy from fat) for 6 weeks and then hepatocytes were isolated. After cell attachment, APAP (1; 2.5; 3.75 and 5 mM) was added to culture media (William´s E medium) and hepatocytes were cultured for up to 24 hours. APAP caused more severe dose-dependent damage of steatotic hepatocytes as documented by increased release of lactate dehydrogenase (LDH) and LDH leakage, decreased activity of cellular dehydrogenases (WST-1 test) and reduced albumin production. Intact steatotic hepatocytes contained lower amount of reduced glutathione (GSH). Treatment with APAP (1 and 2.5 mmol/l) caused more pronounced decrease in GSH in steatotic hepatocytes. ROS (reactive oxygen species) formation after 24-hour incubation was significantly higher in fatty hepatocytes using APAP at concentration of 3.75 and 5 mmol/l. Interleukin 6 (IL-6) production was elevated in 2.5 mM APAP-treated nonsteatotic and steatotic hepatocyte cultures at 8 hours, compared to appropriate controls. In conclusions, our results indicate that steatotic hepatocytes exert higher sensitivity to the toxic action of APAP. This sensitivity may be caused by lower content of GSH in intact steatotic hepatocytes and by more pronounced APAPinduced decrease in intracellular concentration of GSH., O. Kučera, ... [et al.]., and Obsahuje seznam literatury
Stem cells biology is one of the most frequent topic of physiological research of today. Spinal fusion represents common bone biology challenge. It is the indicator of osteoinduction and new bone formation on ectopic model. The purpose of this study was to establish a simple model of spinal fusion based on a rat model including verification of the possible use of titanium microplates with hydroxyapatite scaffold combined with human bone marrow-derived mesenchymal stem cells (MSCs). Spinous processes of two adjacent vertebrae were fixed in 15 Wistar rats. The space between bony vertebral arches and spinous processes was either filled with augmentation material only and covered with a resorbable collagen membrane (Group 1), or filled with augmentation material loaded with 5 × 10 6 MSCs and covered with a resorbable collagen membrane (Group 2). The rats were sacrificed 8 weeks after the surgery. Histology, histomorphometry and micro-CT were performed. The new model of interspinous fusion was safe, easy, inexpensive, with zero mortality. We did not detect any substantial pathological changes or tumor formation after graft implantation. We observed a nonsignificant effect on the formation of new bone tissue between Group 1 and Group 2. In the group with MSCs (Group 2) we described mino r inflamatory response which indicates the imunomodulational and antiinflamatory role of MSCs. In conclusion, this new model proved to be easy to use in small animals like rats., K. Klíma, V. Vaněček, A. Kohout, O. Jiroušek, R. Foltán, J. Štulík, V. Machoň, G. Pavlíková, P. Jendelová, E. Syková, J. Šedý., and Obsahuje bibliografii
Normal pressure hydrocephalus (NPH) is one of a few treatable conditions of cognitive decline affecting predominately elderly people. Treatment, commonly based on the ventriculoperitoneal shunt insertion, leads to a partial or complete correction of patient's state, although its effect does not unfortunately always last. The aim of our study was to observe the changes of homocysteine and selected steroids and neurosteroids and follow-up the patients with respect to the duration of the NPH-related dementia improvement. The cerebrospinal fluid and plasma levels of cortisol, cortisone, dehydroepiandrosterone (DHEA), 7α-hydroxy-DHEA, 7β-hydroxy-DHEA, 7-oxo-DHEA, 16α-hydroxy-DHEA (all LC-MS/MS), DHEA-sulphate (DHEAS) (radioimmunoassay) and homocysteine (gas chromatography) were determined in NPH-diagnosed subjects before, during and 6, 12 and 24 months after shunt insertion. The cognitive functions ameliorated after shunt insertion and remain improved within 2 years. Changes in cerebrospinal fluid DHEAS, DHEA and its ratio, cortisone/cortisol and 16α-hydroxy-DHEA and plasma DHEAS, 7β-hydroxy-DHEA, cortisone/cortisol and homocysteine were found. Mentioned changes may contribute to the clarification of NPH pathogenesis. Altered neurosteroids levels are possible indicators to be utilized in the follow-up of NPH subjects. Moreover, plasma homocysteine may serve as an early indicator of NPH-related dementia., L. Sosvorova, M. Mohapl, M. Hill, L. Starka, M. Bicikova, J. Vitku, R. Kanceva, J. Bestak, R. Hampl., and Obsahuje bibliografii
Postpartum depression affects 10-15 % women after childbirth. There is no currently generally accepted theory about the causes and mechanisms of postpartum mental disorders. The principal hypothesis concerns the association with sudden changes in the production of hormones affecting the nervous system of the mother and, on the other hand, with the ability of receptor systems to adapt to these changes. We observed changes in steroidogenesis in the period ar ound spontaneous delivery. We collected three samples of maternal blood. The first sampling was 4 weeks prior to term; the second sampling was after the onset of uterine contractions (the beginning of spontaneous labour); the third sampling was during the third stage of labour (immediately after childbirth). Additionally, we collected mixed umbilical cord blood. The almost complete steroid metabolome was analyzed by gas chroma tography-mass spectrometry followed by RIA for some steroids. Mental changes in women in the peripartum period were observed using the Hamilton Depression Rating Scale. The local Ethics Committee approved the study. We found already th e changes in androgens levels correlating with postpartum mood disorders four weeks prior to childbirth. The strongest correlations between steroid and postpartum mood change were found in venous blood samples collected from mothers after childbirth and from umbilical cord blood. The main role played testosterone, possibly of maternal origin, and estrogens originating from the fetal compartment. These results suggest that change s in both maternal and fetal steroidogenesis are involved in the development of mental changes in the postpartum period. Descriptions of changes in steroidogenesis in relation to po stpartum depression could help clarify the causes of this disease, and changes in some steroid hormones are a promising marker of mental changes in the postpartum period., A. Pařízek, M. Mikešová, R: Jirák, M. Hill, M. Koucký, A. Pašková, M. Velíková, K. Adamcová, M. Šrámková, H. Jandíková, M. Dušková, L. Stárka., and Obsahuje bibliografii
The local concentration of glucocorticoids is intensively regulated by 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD 1). Human 11β-HSD 1 also reversibly catalyzes the inter-conversion of 7α-hydroxy- and 7β-hydroxy-dehydroepiandrosterone (DHEA) into 7-oxo-DHEA. The cohort of 282 obese adolescents, 154 girls (median age 15.31 years, range 14.17-16.68 years) and 128 boys (median age 14.95 years, range 13.87-16.16 years), BMI (Body Mass Index) >90th percentile was examined. In samples collected before and after one month of reductive diet therapy, circulating levels of steroids were analyzed by liquid chromatography-tandem mass spectrometry and radioimmunoassay methods. The model of the treatment efficacy prediction was calculated. A significant reduction in circulating levels of cortisone, E2 and increased levels of 7β-hydroxy-DHEA after the reductive treatment was observed. Levels of cortisol, DHEA, DHT sustained without any significant change. The predictive Orthogonal Projections to Latent Structures (OPLS) model explained 20.1 % of variability of BMI, z-score change by the basal levels of 7α-hydroxy-DHEA, DHEA, cortisol and E2 as the strongest predictors. Reduced levels of circulating cortisone and reduced ratios of oxygenated/reduced metabolites reflect increased reductase activity of 11β-HSD 1 with reduced BMI, z-score. We hypothesize whether these changes can be attributed to the altered activity of 11β-HSD 1 in the liver., L. Máčová, L. Sosvorová, J. Vítků, M. Bičíková, M. Hill, H. Zamrazilová, B. Sedláčková, L. Stárka /., and Obsahuje bibliografii
Steroids are important marker s in pregnancy. Although estimating their levels separately in umbilical arterial (UA) and venous blood (UV) enable more precise insights into the functioning fetoplacental unit compared to using mixed umbilical blood (UM), selective aspiration of UA and UV is technically more demanding than collecting UM. We measured the levels of 67 unconjugated steroids and steroid polar conjugates in UA and UV using GC-MS in 80 women giving birth within weeks 28 to 42 of gestation. The samples were sorted into three groups: women entering labor within weeks 28- 32 (group A, n=19), weeks 33-37 (group B, n=19), and weeks 38-42 (group C, n=42) of gestation, respectively. The preterm labo rs were due to pathologies unrelated to steroid status. Most unconjugated steroids exhibited pronounced arteriovenous differences (AVD). The AVD were less distinct in more stable steroid conjugates. Most steroids positively correlate with gestational age, but unconjugated 5 β -reduced pregnanes show negative correlations, as do testosterone and androstenediol, substrates for the placental synthesis of estrogens. Tight correlations be tween steroids in UA and UV indicate that steroid measurements in UA, UV and UM can be accurately derived from each other, which is important for the diagnostics of steroid related diseases in newborns., A. Pašková ... [et al.]., and Obsahuje bibliografii a bibliografické odkazy
It has been reported that the major function of the sterol regulatory element binding protein 2 (SREBP-2) is to activate preferentially cholesterol biosynthesis in liver and adipose tissue rather than fatty acid synthesis. In the current study, we analyzed the effects of overexpression of human dominantpositive SREBP-2 transgene under control of PEPCK promoter in the spontaneously hypertensive rat (SHR) on lipid and glucose metabolism. Transgenic overexpression of SREBP-2 was associated with significantly higher hepatic triglycerides (20.4±0.9 vs. 17.0±0.05 μmol/g, P<0.05) but not cholesterol (10.6±0.4 vs. 10.9±0.4 μmol/g) and decreased relative weight of epididymal fat pad (0.73±0.03 vs. 0.830.03, P<0.05). In addition, muscle triglyceride (15.8±3.7 vs. 8.5±1.2 μmol/g, P<0.001) and cholesterol (3.6±0.5 vs. 2.1±0.1 μmol/g, P<0.05) concentrations were significantly increased in transgenic rats when compared to SHR controls. Ectopic fat accumulation was associated with significantly increased serum glucose levels (6.4±0.1 vs. 5.9±0.1 mmol/l, P<0.005) and reduced insulin levels (1.78±0.33 vs. 2.73±0.37 nmol/l, P<0.05) in transgenic rats. These results provide evidence for important role of SREBP-2 in regulation of lipid and glucose metabolism., V. Landa, V. Zídek, P. Mlejnek, M. Šimáková, J. Šilhavý, J. Trnovská, L. Kazdová, M. Pravenec., and Obsahuje bibliografii
The STESYS2 software is a new version of the IBM PC software supporting interactive stereological measurements. In comparison with the previous STESYS, it is enhanced by a number of useful options, e.g. on-line image input via a TV camera coupled with a microscope operating under MS Windows OS. The main advantage, when compared with other such software packages, is the design of the STESYS2 as a module of the freeware image processing system Image Tool which provides a user-friendly environment including a number of image processing and preprocessing routines. Capabilities of the STESYS2 are illustrated by a practical example: estimation of the surface area of capillaries in the terminal villi of human placenta by the Sandau spatial grid method., Z. Tomori, L. Matis, P. Karen, L. Kubínová, I. Krekule., and Obsahuje bibliografii
Pharmacological preconditioning by diazoxide and a model of experimental streptozotocin-induced acute diabetes mellitus (STZ-DM) provided similar levels of cardioprotection assessed as limiting myocardial infarct size. The aim was to explore the possibility of existence of another in vitro mechanism, which could be contributory to cardioprotection mediated by diazoxide treatment. Mitochondrial membrane fluidity and ATP synthase activity in isolated heart mitochondria were determined under the influence of two factors, STZ-DM condition and treatment with diazoxide. Both factors independently increased the ATP synthase activity (p<0.05), as no interaction effect was observed upon the combination of STZ-DM with diazoxide. On the other hand, the mitochondrial membrane fluidity was significantly increased by STZ-DM only; no significant main effect for diazoxide was found. Based on the results from measurements of enzyme kinetics, we assume a direct interaction of diazoxide with the molecule of ATP synthase stimulated its activity by noncompetitive activation. Our present work revealed, for the first time, that cardioprotection induced by diazoxide may not be caused exclusively by mitochondrial KATP opening, but presumably also by a direct interaction of diazoxide with ATP synthase, although the mechanisms for achieving this activation cannot be fully delineated., M. Jašová, I. Kancirová, M. Muráriková, V. Farkašová, I. Waczulíková, T. Ravingerová, A. Ziegelhöffer, M. Ferko., and Obsahuje bibliografii
Streptozotocin (STZ) is used to induce experimental diabetes in animals and is also applied for the treatment of patients with insulinoma. The aim of the present work was to investigate the direct effect of STZ on lipolysis in isolated rat adipocytes. After the isolation, the cells were incubated in a Krebs-Ringer buffer of pH 7.4, at the temperature 37 °C for 90 min with different concentrations of STZ: 0.5, 1 or 2 mmol/l. STZ caused a significant rise in basal values (99 %, 199 %, and 377 %, respectively) and epinephrine-stimulated (1 µmol/l) lipolysis (15 %, 24 % and 46 %, respectively). Augmentation of basal lipolysis by STZ was neither restricted by insulin (1 nmol/l) nor by H-89 (an inhibitor of protein kinase A, 50 µmol/l). These results indicate the stimulatory influence of STZ on the action of hormone-sensitive lipase in isolated cells of white adipose tissue. The obtained outcomes suggest that in studies employing STZ, it is necessary to consider its direct effect upon lipolysis in adipocytes., T. Szkudelski, K. Szkudelska., and Obsahuje bibliografii