The fundamental biochemical processes of 5-methylcytosine (5-mC) synthesis, maintenance, conversion and removal determine the time and spatial pattern of DNA methylation. This has a strong effect on a plethora of physiological aspects of cellular metabolism. While the presence of 5-mC within the promoter region can silence gene expression, its derivative - 5-hydroxymethylcytosine exerts an opposite effect. Dysregulations in the metabolism of 5-mC lead to an altered DNA methylation pattern which is linked with a disrupted epigenome, and are considered to play a significant part in the etiology of several human diseases. A summary of recent knowledge about the molecular processes participating in DNA methylation pattern shaping is provided here., R. Murín, M. Abdalla, N. Murínová, J. Hatok, D. Dobrota., and Obsahuje bibliografii
Recent studies have demonstrated that some microRNAs (miRNAs) inhibit bone formation by inhibiting the translation of specific genes. Several in vitro studies have suggested that miR - 23a inhibits osteogenic differentiation by suppressing the translation of Runx2, a transcription factor essential for osteoblastogenesis, and of Sa tb2, a member of the special AT-rich binding protein family. In the pr esent study, we used a gain -of-function approach to determine the roles of miR -23a in bone formation and homeostasis in vivo . The miR -23a transgenic (Tg) mice grew normally and their body size and weight were similar to those of wild -type (WT) littermates. Bone structure and morphology were similar in Tg and WT mice. Furthermore, the numbers of osteoblasts and osteoclasts, as well as their activities in bone were similar between Tg and WT mice. Our results indicate that miR -23 has limited roles in bone form ation and maintenance in vivo in mice., J. Park, S. Wada, T. Ushida, T. Akimoto., and Obsahuje bibliografii
Familial hypercholesterolemia (FH) is most frequently caused by LDLR or APOB mutations. Therefore, the aim of our study was to examine the genetic background of Slovak patients suspected of FH. Patients with clinical suspicion of FH (235 unrelated probands and 124 family relatives) were recruited throughout Slovakia during the years 2011-2015. The order of DNA analyses in probands was as follows: 1. APOB mutation p.Arg3527Gln by real-time PCR method, 2. direct sequencing of the LDLR gene 3. MLPA analysis of the LDLR gene. We have identified 14 probands and 2 relatives with an APOB mutation p.Arg3527Gln, and 89 probands and 75 relatives with 54 different LDLR mutations. Nine of LDLR mutations were novel (i.e. p .Asp90Glu, c.314-2A>G, p.Asp136Tyr, p.Ser177Pro, p.Lys225_Glu228delinsCysLys, p.Gly478Glu, p.Gly675Trpfs*42, p.Leu680Pro, p.Thr832Argfs*3). This is the first study on molecular genetics of FH in Slovakia encompassing the analysis of whole LDLR gene. Geneti c etiology of FH was confirmed in 103 probands (43.8 %). Out of them, 86.4 % of probands carried the LDLR gene mutation and remaining 13.6 % probands carried the p.Arg3527Gln APOB mutation., D. Gabčová, B. Vohnout, D. Staníková, M. Hučkova, M. Kadurová, M. Debreová, M. Kozárová, Ľ. Fábryová, Slovak FH Study Group, J. Staník, I. Klimeš, K. Rašlová, D. Gašperiková., and Obsahuje bibliografii
Interest surrounds the role of an NADPH oxidase-like enzyme in hypoxic pulmonary vasoconstriction (HPV). We have studied the effects of the NADPH oxidase inhibitors iodonium diphenyl (ID) and cadmium sulphate (CdSO4) upon HPV of isolated rat pulmonary arteries (n = 73, internal diameter 545± 23 mm). Vessels were preconstricted with prostaglandin F2a (PGF2a, 0.5 or 5 mM) prior to a hypoxic challenge. ID (10 or 50 mM), CdSO4 (100 mM) or vehicle (50 ml) was added for 30 min before re-exposure to PGF2a and hypoxia. ID and CdSO4 significantly inhibited HPV. In vessels preconstricted with 5 mM PGF2a, ID (10 and 50 mM) reduced HPV from 37.4± 5.6 % to 9.67± 4.4 % of the contractile response elicited by 80 mM KCl (P<0.05) and from 30.1± 5.0 % to 0.63± 0.6% 80 mM KCl response (P<0.01), respectively. CdSO4 (100 mM) reduced HPV from 29.4±4.0 % to 17.1±2.2% 80 mM KCl response (P<0.05). In vessels preconstricted with 0.5 mM PGF2a, ID (10 and 50 mM) reduced HPV from 16.0± 3.15% to 3.36± 1.44 % 80 mM KCl response (P<0.01) and from 15.0± 1.67 % to 2.82± 1.40 % 80 mM KCl response (P<0.001), respectively. Constriction to PGF2a was potentiated by ID. ID and CdSO4, at concentrations previously shown to inhibit neutrophil NADPH oxidase, attenuate HPV in isolated rat pulmonary arteries. This suggests that an NADPH oxidase-like enzyme is involved in HPV and could act as the pulmonary oxygen sensor., R. D. Jones, J. S. Thompson, A. H. Morice., and Obsahuje bibliografii
a1_We investigated the potential neuroprotective effect of transient hypertension on neuronal cell death induced by ischemia-reperfusion. Recovery of neurons, terminally differentiated cells, is almost entirely dependent upon active transcription and repair of DNA damage. We focused on the histochemical detection of distribution of NOR (argyrophylic nucleolar proteins) reflecting nucleolar integrity, immunohistochemical detection of PARP-1 (poly(ADP-ribose) polymerase-1), MADD (mitogen-activated death domain), a protein accumulated in nucleoli upon stimulation by ischemia, the active form of caspase-3, a universal proteolytic enzyme of apoptosis. The terminal deoxynucleotidyl-transferase (TdT)-mediated dUTP-biotin nick-end-labeling method (TUNEL) proved the presence of in situ DNA fragmentation. We used the model of transient focal cerebral ischemia in rats with occlusion of middle cerebral artery. In experimental group of rats, the transient hypertension was induced by constriction of the abdominal aorta. The period of ischemia lasted 15, 30, 60 and 120 min followed by 48 h of reperfusion. We examined the frontal lobe of the ipsilateral hemisphere for apoptosis of neurons and compared it with the intact brain tissue. In normotensive rats with transient focal cerebral ischemia, we found disintegrated nucleoli of cortical as well as subcortical neurons at all investigated periods of ischemia, whereas the neurons of intact animals showed compact nucleoli with a few satellites. Nuclear positivity for MADD and PARP-1 was apparent in the neocortex after 15 min and peaked after 30 min of ischemia. On the other hand, the subcortical neurons showed nuclear positivity after 60 and 120 min. The immunohistochemical reaction for active caspase 3 was apparent after 30 min onwards predominantly in the cortex. The TUNEL staining was distinct after 60 and 120 min., a2_In hypertensive rats, we found nucleolar disintegration, positivity for MADD, PARP-1 and caspase 3 after 30 min cortically and subcortically, followed by TUNEL positive staining of cortical neurons after 60 and 120 min. In summary, we detected delayed activation of neuronal apoptosis in transiently hypertensive rats with focal cerebral ischemia compared to normotensive animals. The apoptotic phenotype was confirmed by a panel of complementary methods showing rapid proteolysis-nucleolar segregation, MADD, PARP-1 and caspase-3 positivity as well as ultimate DNA fragmentation proved by the TUNEL assay., M. Smrčka, M. Horký, F. Otevřel, Š. Kuchtíčková, V. Kotala, J. Mužík., and Obsahuje bibliografii
Nearly 60 years has elapsed since the first isolation and identification of 7α-hydroxy-dehydroepiandrosterone, and in that time much information has been gained on its occurrence, metabolism, ontogeny, immunomodulatory activity, cell proliferation, cortisol control in local tissues and neuroactivity. Additional knowledge about this steroid may elucidate its role in obesity, neurodegenerative disturbances such as Alzheimer’s disease, or psychiatric disorders such as schizophrenia or depression. This review aims to provide a comprehensive summary of the available literature on 7α-hydroxydehydroepiandrosterone., L. Stárka., and Obsahuje bibliografii
There is considerable evidence linking alcohol consumption and sedation and TRH in the brain septum. Moreover, innate septal TRH concentration is inversely related to the degree of ethanol preference. Recently we demonstrated in rats that four-week ethanol drinking increased the septal TRH content by 50 %. We had shown previously that ethanol induces neuronal swelling, which is known to evoke the secretion of hormones, peptides and amino acids from various types of cells. We have therefore explored the effect of hyposmotic medium and of 80 and 160 mM ethanol and 80 mM urea (both permeant molecules) in isosmotic and hyperosmotic (preventing cell swelling) media on the in vitro release of TRH by the rat septum. Lowering medium osmolarity resulted in a hyposmolarity-related increase in TRH secretion. Both ethanol and urea stimulated TRH release only in isosmolar solution. Our data indicate that ethanol in clinically relevant concentrations can induce TRH release from the septum by a mechanism involving neuronal swelling., J. Kučeorvá, V. Štrbák., and Obsahuje bibliografii
Interstitial cells of Cajal (ICC) are the pacemaker cells in the gut. They have special properties that make them unique in their ability to generate and propagate slow waves in gastrointestinal muscles. The electrical slow wave activity determines the characteristic frequency of phasic contractions of the stomach, intestine and colon. Slow waves also determine the direction and velocity of propagation of peristaltic activity, in concert with the enteric nervous system. Characterization of receptors and ion channels in the ICC membrane is under way, and manipulation of slow wave activity markedly alters the movement of contents through the gut. Gastric myoelectrical slow wave activity produced by pacemaker cells (ICC) can be reflected by electrogastrography (EGG). Electrogastrography is a perspective non-invasive method that can detect gastric dysrhythmias associated with symptoms of nausea or delayed gastric emptying., P. Čamborová, P. Hubka, I. Sulková, I. Hulín., and Obsahuje bibliografii
Cytarabine is one of the most efficient drugs in the treatment of hematological malignancies. In this work, we describe the synthesis and characterization of two different polymer conjugates of cytarabine that were designed for the controlled release of cytarabine within the leukemia cells. Reactive copolymers of N-(2-hydroxypropyl)methacrylamide (HPMA) and 3-(3-methacrylamidopropanoyl)thiazolidine-2-thione) or 3-(Nmethacryloylglycyl- phenylalanylleucylglycyl)thiazolidine-2-thione were used in the study as reactive polymer precursors for reaction with cytarabine. The enzymatic release of cytarabine from the conjugate containing a GFLG spacer utilizing cathepsin B was verified. In addition to enzymolysis, the pH-dependent hydrolysis of cytarabine from both copolymers was also confirmed. Approximately 40 % and 20 % of the drug was released by spontaneous hydrolysis at pH 7.4 within 72 h from the polymer conjugates with the GFLG and β-Ala spacers, respectively. At pH 6.0, the spontaneous hydrolysis slowed down, and less than 10 % of the drug was liberated within 72 h. The results of the cytotoxicity evaluation of the polymer conjugates in vitro against various cell lines showed that the cytotoxicity of the polymer conjugates is approximately three times lower in comparison to free cytarabine., R. Pola, O. Janoušková, T. Etrych., and Obsahuje bibliografii
a1_The genes that cause or increase susceptibility to essential hypertension (EH) and related an imal models remain unknown. Their identification is unlikely to be realized with current genetic approaches, because of ambiguities in the genotype-phenotype relationships in these polygenic disorders. In turn, the phenotype is not just an aggregate of traits, but needs to be related to specific components of the circulatory control system at different stages of EH. Hence, clues about important genes must come through the phenotype, reversing the order of current approaches. A recent systems analysis has highlighted major differences in circulatory control in the two main syndromes of EH: 1) stress-and-salt-related EH (SSR-EH) - a constrictor hypertension with low blood volume; 2) hypertensive obesity - SSR-EH plus obesity. Each is initiated through sensitization of central synapses linking the cerebral cortex to the hypothalamic defense area. Several mechanis ms are probably involved, including cerebellar effects on baroreflexes. The result is a sustained increase in sympathetic neural activity at stimulus levels that have no effect in normal subjects. Subsequent progression of EH is largely thro ugh interactions with non-neural mechanisms, including changes in concentration of vascular autacoids (e.g. nitric oxide) and the amplifying effect of structural changes in large resistance vessels. The rising vasoconstriction increases heterogeneity of blood flow, causing rarefaction (decreased microvascular densit y) and deterioration of vital organs. SSR-EH also increases food intake in response to stress, but only 40% of these individual s develop hypertensive obesity. Their brain ignores the adiposity signals that normally reduce eating., a2_Hyperinsulinemia masks the sympathetic vasoconstriction through its dilator action, rais es blood volume, whilst renal nephropathy and other diabetic complications are common. In each syndrome the neural and non-neural determinants of hypertension provide targets fo r identifying high BP genes. Reading the genome from the phenotype will require new approaches, such as those used in developmental genetics. In addition, transgenic technology may help verify hypotheses and examine whether an observed effect is through single or multiple mechanisms. To obtain answers will require substantial collaborative efforts between physiologists and geneticists., P. I. Korner., and Obsahuje bibliografii