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Biomedical subjects

G Molnár

Publications and source records attributed to G Molnár.

At least 19 recordsLinked to original sources
PubMed2001

Role of prenylation in the interaction of Rho-family small GTPases with GTPase activating proteins.

The role of prenylation in the interaction of Rho-family small GTPases with their GTPase activating proteins (GAPs) was investigated. Prenylated and nonprenylated small GTPases were expressed in Sf9 insect cells and Escherichia coli, respectively. Nucleotide binding to and hydrolysis by prenylated and nonprenylated proteins were identical, but three major differences were observed in their reactions with GAPs. (1) Membrane-associated GAPs accelerate GTP hydrolysis only on prenylated Rac1 and RhoA, but they are inactive on the nonprenylated form of these proteins. The difference is independent of the presence of detergents. In contrast to Rac1 and RhoA, nonprenylated Cdc42 is able to interact with membrane-localized GAPs. (2) Full-length p50RhoGAP and p190RhoGAP react less intensely with nonprenylated Rac1 than with the prenylated protein, whereas no difference was observed in the reaction of isolated GAP domains of either p50RhoGAP or Bcr with the different types of Rac1. (3) Fluoride exerts a significant inhibitory effect only on the interaction of prenylated Rac1 with the isolated GAP domains of p50RhoGAP or Bcr. The effect of fluoride is not influenced by addition or chelation of Al(3+). This is the first detailed study demonstrating that prenylation of the small GTPase is an important factor in determining its reaction with GAPs. It is suggested that both intramolecular interactions and membrane targeting of GAP proteins represent potential mechanisms regulating Rac signaling.

Enzyme Activation↗
PubMed2001

Mechanism of NADPH oxidase activation by the Rac/Rho-GDI complex.

The low molecular weight GTP binding protein Rac is essential to the activation of the NADPH oxidase complex, involved in pathogen killing during phagocytosis. In resting cells, Rac exists as a heterodimeric complex with Rho GDP dissociation inhibitor (Rho-GDI). Two types of interactions exist between Rac and Rho-GDI: a protein-lipid interaction, implicating the polyisoprene of the GTPase, as well as protein-protein interactions. Using the two-hybrid system, we show that nonprenylated Rac1 interacts very weakly with Rho-GDI, pointing to the predominant role of protein-isoprene interaction in complex formation. In the absence of this strong interaction, we demonstrate that three sites of protein-protein interaction, Arg66(Rac)-Leu67(Rac), His103(Rac), and the C-terminal polybasic region Arg183(Rac)-Lys188(Rac), are involved and cooperate in complex formation. When Rac1 mutants are prenylated by expression in insect cells, they all interact with Rho-GDI. Rho-GDI is able to exert an inhibitory effect on the GDP/GTP exchange reaction except in the complex in which Rac1 has a deletion of the polybasic region (Arg183(Rac)-Lys188(Rac)). This complex is, most likely, held together through protein-lipid interaction only. Although able to function as GTPases, the mutants of Rac1 that failed to interact with Rho-GDI also failed to activate the NADPH oxidase in a cell-free assay after loading with GTP. Mutant Leu119(Rac)Gln could both interact with Rho-GDI and activate the NADPH oxidase. The Rac1/Rho-GDI and Rac1(Leu119Gln)/Rho-GDI complexes, in which the GTPases were bound to GDP, were found to activate the oxidase efficiently. These data suggest that Rho-GDI stabilizes Rac in an active conformation, even in the GDP-bound state, and presents it to its effector, the p67phox component of the NADPH oxidase.

Amino Acid Sequence↗
PubMed2001

A Ca(2+)-activated NADPH oxidase in testis, spleen, and lymph nodes.

Superoxide and its derivatives are increasingly implicated in the regulation of physiological functions from oxygen sensing and blood pressure regulation to lymphocyte activation and sperm-oocyte fusion. Here we describe a novel superoxide-generating NADPH oxidase referred to as NADPH oxidase 5 (NOX5). NOX5 is distantly related to the gp91(phox) subunit of the phagocyte NADPH oxidase with conserved regions crucial for the electron transport (NADPH, FAD and heme binding sites). However, NOX5 has a unique N-terminal extension that contains three EF hand motifs. The mRNA of NOX5 is expressed in pachytene spermatocytes of testis and in B- and T-lymphocyte-rich areas of spleen and lymph nodes. When heterologously expressed, NOX5 was quiescent in unstimulated cells. However, in response to elevations of the cytosolic Ca(2+) concentration it generated large amounts of superoxide. Upon Ca(2+) activation, NOX5 also displayed a second function: it became a proton channel, presumably to compensate charge and pH alterations due to electron export. In summary, we have identified a novel NADPH oxidase that generates superoxide and functions as a H(+) channel in a Ca(2+)-dependent manner. NOX5 is likely to be involved in Ca(2+)-activated, redox-dependent processes of spermatozoa and lymphocytes such as sperm-oocyte fusion, cell proliferation, and cytokine secretion.

Amino Acid Sequence↗
PubMed2001

Characterization of membrane-localized and cytosolic Rac-GTPase-activating proteins in human neutrophil granulocytes: contribution to the regulation of NADPH oxidase.

We have investigated the intracellular localization and molecular identity of Rac-GTPase-activating proteins (Rac-GAPs) in human neutrophils. Immunoblot analysis detected the presence of both p190RhoGAP and Bcr mainly in the cytosol. An overlay assay performed with [gamma-(32)P]GTP-bound Rac revealed dominant GAP activity related to a 50 kDa protein both in the membrane and cytosol. This activity could be identified by Western blotting and immunoprecipitation with specific antibody directed against the GAP domain of p50RhoGAP. Using a semirecombinant or fully purified cell-free activation assay of the Rac-activated enzyme NADPH oxidase, we demonstrated the regulatory effect of both the membrane-localized and soluble GAPs. We suggest that in neutrophil granulocytes Rac-GAPs have redundant function and represent suitable targets for both the up-regulation and down-regulation of the NADPH oxidase.

Animals↗
PubMed2001

Predictors of discharge to home during the first year after right hemisphere stroke.

OBJECTIVES: The aim of this study was to explore predictive factors of the length of hospital stay at the acute stage of right hemisphere stroke. Special attention was paid to the possible role of anosognosia for hemiparesis and anosognosia for neglect in this prediction. PATIENTS AND METHODS: A consecutive series of 57 patients having their first right hemisphere stroke were examined at the acute phase. Forty-nine patients were included in this study and followed-up for 12 months. The examinations were conducted within 2 weeks of onset. The outcome variable was the time (days) from stroke to discharge to home. The predictors were age, gender, size of infarct, neglect, hemiparesis, verbal memory, unawareness of illness, anosognosia for neglect, anosognosia for hemiparesis and presence of a relative at home. RESULTS: Hemiparesis and unawareness of illness lengthened the duration of the hospital stay, the presence of a relative reduced it. Neglect was the best single predictor of poor outcome, but it had no additional value in the combination of the three predictors above. Neither anosognosia for hemiparesis nor anosognosia for neglect were important predictors. CONCLUSION: Hemiparesis, unawareness of illness and presence of a relative at home were the best predictors of the time from right hemisphere stroke to discharge to home.

Activities of Daily Living↗
PubMed2000

Volumetric measurements of right cerebral hemisphere infarction: use of a semiautomatic MRI segmentation technique.

The applications of a new segmentation software, Anatomatic, in the evaluation of volumetric measurements of brain infarctions and the new Medimag 3D software in the evaluation of 3D image representation of infarctions are described. These programs are applied to magnetic resonance imaging. The aim of this study is to evaluate the use of these software packages in making accurate volumetric measurements in 40 patients with right cerebral infarctions, in determining the correlations between the quantitated lesions and neurological/neuropsychological dysfunctions and in creating realistic 3D views of the infarctions. Using Anatomatic, reproducible infarction volumes were achieved with ease and within a reasonably fast time. Medimag helped achieve realistic 3D representations of the infarctions. When compared, the semiautomatic segmentation proved to be much faster and yielded higher infarction volumes than the manual segmentation technique. Significantly positive correlations between the infarction volumes and neurological dysfunctions and neuropsychological deficit (neglect) helped to explain the effect of volumes on the clinical status of the patients.

Adult↗
PubMed2000

Visual neglect as a predictor of functional outcome one year after stroke.

OBJECTIVE: The aim was to study the role of visual neglect in acute right hemisphere brain infarct as a predictor of poor functional outcome during the first year after stroke. In particular, we were interested in the additional value of neglect measures besides hemiparesis, hemianopia, cognitive deficits and age. PATIENTS AND METHODS: A consecutive series of 57 patients with a neuroradiologically verified right hemisphere infarct was examined within 10 days of the stroke. Fifty patients were followed up for 1 year. Neglect was measured with the Conventional and the Behavioural subtests of the Behavioural Inattention Test (BITC and BITB, respectively). The predictors were determined at the 10-day examination. Functional outcome was assessed 3, 6 and 12 months after the onset with the Frenchay Activities Index. RESULTS: Neglect in BITB was the best single predictor, which together with high age formed the best combination of predictors for poor functional outcome at each follow-up. Hemiparesis was also included in this prediction model at the 3-month follow-up, but hemianopia, BITC, or visuoconstructional and memory deficits showed no additional predictive value. However, neglect usually recovered soon. When neurological and cognitive deficits were assessed at the same time as the outcome, hemiparesis rather than neglect was the strongest correlate of poor outcome. CONCLUSION: Neglect in acute stroke is an important predictor of poor functional recovery. Residual neglect, which could be compensated in the follow-up tests, may nevertheless restrict patients' real-life activities and hobbies.

Acute Disease↗
PubMed2000

Low plasma antioxidant activity is associated with high lesion volume and neurological impairment in stroke.

BACKGROUND AND PURPOSE: Oxidative stress is probably involved in neuronal damage induced by ischemia-reperfusion. The purpose of this study was to assess the role of antioxidant activity in cerebral ischemic stroke. METHODS: Antioxidant activity of blood plasma and cerebrospinal fluid was assessed in 22 patients with cerebral hemisphere infarction that was verified and quantified by MRI. RESULTS: Low total peroxyl radical trapping potential of plasma, but not of cerebrospinal fluid, was associated with high lesion volume and high neurological impairment assessed by scores on NIH Stroke Scale, Barthel Index, and Hand Motor Score tests. The plasma concentrations of ascorbic acid, alpha-tocopherol, and protein thiols were also associated with the degree of neurological impairment. CONCLUSIONS: These data suggest that the antioxidant activity of plasma may be an important factor providing protection from neurological damage caused by stroke-associated oxidative stress.

Adult↗
PubMed2000

Primary mono-layer cell cultures as model system for studying of environmental toxic agents: organochlorine compounds.

Organic pollution of water and soil has various harmful effects on biological systems (1). Chlorine substituted benzol compounds are one these xenobiotic substances, which are toxic to the environment (2). They can also accumulate in plant and animal tissues (3), which provides ample reason to study the effects of sublethal doses of chloro-benzols on various cell cultures. In this study the toxic effects of chloro-benzols were investigated on avian fibroblast and mammalian hepatocyte cultures. The fibroblast cultures were prepared from eggs preadapted to chloro-benzol during a fourteen-day-long incubation period. The Wistar rat hepatocyte monolayer cultures were exposed to a direct treatment of 1,2,4-tri-chloro-benzol (0.01 microgram/ml-1 microgram/ml) for 3 hours. Following the treatment with chloro-benzol, the viability of the cells was measured, together with lactic dehydrogenase activity, in both kinds of cultures. The effect of tri-chloro-benzol treatment on chicken eggs was not significant. The cells of chicken embryos were not damaged after the 1,2,4-tri-chloro-benzol treatment. The hepatocyte cultures showed the toxic effects of 1,2,4-tri-chloro-benzol after the direct and acute treatment. The cell viability decreased and the LDH activity increased significantly. These results show that the primary cell cultures are suitable for studying the effects of organochlorine compounds.

Animals↗
PubMed2000

Analysis and modulation of spike trains and oscillations in identified neural network of Lymnaea stagnalis L.

Identified neurons and members of functionally characterized clusters of the central nervous system of Lymnaea stagnalis L. were studied. Long-term spike trains (10-100 min) were collected using current clamp method. Firing patterns were analyzed by several mathematical tools e.g.: spike density function (SDF), interspike interval (ISI), Fourier-transform. Both the spike trains and oscillation of firing were modulated by 5HT (2 x 10(-5) M) and mu-opioid peptides (10(-5) M). Co-application of 5HT (2 x 10(-5) M) and DAGO (10(-5) M) turned the firing of the neurons (RPeD1 and A cells) opposite to the running pattern and eliminated the 0.3 Hz oscillation causing a new slow periodicity (0.1-0.05 Hz).

Action Potentials↗
PubMed2000

Nonlinear activity of identified Lymnaea neurons.

Two long-lasting discharges of action potentials were recorded from a buccal cell of the pond snail, respectively, before and after superfusing the preparation with low-calcium solution. The corresponding sequences of interspike intervals were then analysed by the nonlinear prediction methods. The results yield evidence of a small but clear nonlinearity only in the second of analysed tachograms. This finding is evaluated and discussed.

Action Potentials↗
PubMed1999

Regulation of cell surface expression of Fas (CD95) ligand and susceptibility to Fas (CD95)-mediated apoptosis in activation-induced T cell death involves calcineurin and protein kinase C, respectively.

We show that an influenza hemagglutinin-specific CD4+ murine T cell hybridoma (IP-12-7) enters the apoptotic suicide program via the Fas ligand (FasL)/Fas-mediated pathway upon T cell receptor (TCR) stimulation. These cells express Fas and FasL mRNA, cell surface Fas and intracellular FasL, but do not enter apoptosis upon Fas ligation prior to TCR stimulation. TCR stimulation additionally results in protein synthesis-dependent cell surface expression of the preformed FasL. Addition of phorbol dibutyrate (PBu2) alone was sufficient to induce susceptibility to Fas ligation induced apoptosis, while addition of both PBu2 and calcium ionophore A23187 were required to induce FasL cell surface expression. Addition of cyclosporin A completely inhibited TCR-mediated death and FasL cell surface up-regulation, but had no effect on apoptosis induced directly by Fas ligation following TCR stimulation. Inhibitors of protein kinase C (PKC) (Gö 6976 and GF 109203X) completely inhibited TCR-induced susceptibility to Fas ligation, but only partially inhibited TCR-induced cell surface expression of FasL. PKC isoenzymes alpha, beta, delta and zeta were expressed by this cell line and only the alpha and betaI isoforms translocated to the membrane fraction upon TCR stimulation. Our data suggest that in activation-induced T cell apoptosis PKC is involved in pathways that mediate the acquisition of Fas susceptibility, while calcineurin is required for cell surface expression of the preformed FasL.

Animals↗
PubMed1999

Periodic and oscillatory firing patterns in identified nerve cells of Lymnaea stagnalis L.

Firing patterns in identified neurons of Lymnaea stagnalis L. were analyzed by various mathematical methods including spike density function (SDF), interspike-interval histograms (ISI), Fourier transform and correlation analysis. Input-3 (IP3) events observed in most of the neurons of the respiratory regulatory system caused prominent changes in the firing frequency of the cells. Similarly, quasiperiodic firing patterns were observed in the neurons of buccal ganglia controlling feeding behavior. Apart from the known periodic patterns a fine oscillation of firing rate was observed in a large number of neurons in the visceral and parietal ganglia. The frequency of this oscillation varied between 0.2 and 0.4 Hz. The most obvious oscillatory patterns were found in the A-cells presumably resulted by periodically appearing synaptic excitation. Moderate intracellular hyperpolarizing current injection, low-Ca/high-Mg saline and application of d-tubocurarine failed to abolish the slow oscillations. Application of Ca-channel blocker cadmium, however, completely eliminated the oscillation in a reversible manner.

Action Potentials↗
PubMed1998

Cortical silent period evoked by transcranial magnetic stimulation in ischemic stroke.

OBJECTIVES: Transcranial magnetic stimulation (TMS) of the motor cortex produces motor evoked potentials (MEPs). Besides this excitatory response, TMS has inhibitory effects. When TMS is performed during voluntary muscle contraction, the MEP is followed by a pause in electromyographic activity (cortical silent period, SP). The aim of this study was to evaluate the clinical usefulness of the SP. METHODS: We studied SP changes in 50 patients with acute hemispheric brain infarction. A stimulator with a round coil and a fixed intensity of 90% of maximum was used to evoke MEPs. RESULTS: SP was elicited on the affected side in 29 of the 50 patients. The mean SP duration was markedly longer on the affected side in the patient group. There were no significant differences between left and right sides in the means of the MEP amplitude ratio (amplitude related to corresponding amplitudes to peripheral electric stimulation) and MEP latencies in the patient group. Prolonged SP was found in 25 of the 29 patients (86%) whereas only 4 (14%) had abnormalities in MEP latency or amplitude ratio. The mean SP duration was significantly prolonged also in a subgroup of 14 patients with normal hand function. CONCLUSIONS: The SP measurement is an easily performed and sensitive method to detect even subclinical disturbances in motor system function in ischemic stroke.

Adult↗
PubMed1998

Transcription of the Arabidopsis CPD gene, encoding a steroidogenic cytochrome P450, is negatively controlled by brassinosteroids.

The Arabidopsis CPD gene encodes a cytochrome P450 steroid side-chain hydroxylase (CYP90) that plays an essential role in the biosynthesis of the plant hormone brassinolide. Expression of the CPD gene is confined to cotyledons and leaf primordia in etiolated seedlings and detectable in the adaxial parenchyma of expanding leaves in light-grown plants. Transcription of the CPD gene is not affected by the plant growth factors auxin, ethylene, gibberellin, cytokinin, jasmonic acid and salicylic acid, but is specifically down-regulated by brassinolide in both dark and light. Steady-state mRNA levels of a CPD promoter-driven uidA reporter gene correlate with the expression of resident CPD gene in transgenic plants. Intermediates of the early and late C-6 oxidation pathways of brassinolide, carrying C-22 and C-23 side-chain hydroxyls, efficiently inhibit the activity of the CPD promoter. Repression of CPD transcription by brassinosteroids is sensitive to the protein synthesis inhibitor cycloheximide, indicating a requirement for de novo synthesis of a regulatory factor.

Arabidopsis↗