Publications by authors named "Katalin Török"

47 Publications

Effects of Exercise Dose and Detraining Duration on Mobility at Late Midlife: A Randomized Clinical Trial.

Gerontology 2021 Mar 3:1-12. Epub 2021 Mar 3.

Somogy County Móricz Kaposi Teaching Hospital, Kaposvár, Hungary.

Background: Office workers near retirement tend to be sedentary and can be prone to mobility limitations and diseases. We examined the dose effects of exergaming volume and duration of detraining on motor and cognitive function in office workers at late midlife to reduce sedentariness and mobility limitations.

Methods: In an assessor-blinded randomized trial, 160 workers aged 55-65 years performed physically active video games in a nonimmersive form of virtual reality (exergaming) in small, supervised groups for 1 h, 1×, 2×, or 3×/week for 8 weeks followed by detraining for 8 and 16 weeks. Exergaming comprises high-intensity, full-body sensorimotor coordination, balance, endurance, and strengthening exercises. The primary outcome was the 6-minute walk test (6MWT), and secondary outcomes were body mass, self-reported physical activity, sleep quality, Berg Balance Scale, Short Physical Performance Battery, fast gait speed, dynamic balance, heart rate recovery after step test, and 6 cognitive tests.

Results: The 3 groups were not different in any of the outcomes at baseline (all p > 0.05). The outcomes were stable and had acceptable reliability (intraclass correlation coefficients ≥0.334) over an 8-week control period. Training produced an inverted U-shaped dose response of no (1×), most (2×), and medium (3×/week) effects of exergaming volume in most motor and selected cognitive outcomes. The distance walked in the 6MWT (primary outcome) increased most (94 m, 19%, p < 0.05), medium (57 m, 12%, p < 0.05), and least (4 m, 1%) after exergaming 2×, 3×, or 0× (control) (all different p < 0.05). The highest responders tended to retain the exercise effects over 8 weeks of detraining, independent of training volume. This maintenance effect was less consistent after 16 weeks of detraining.

Conclusion: Less was more during training and lasted longer after detraining. A medium dose volume of exergaming produced the largest clinically meaningful improvements in mobility and selected cognitive tests in 60-year-old office workers with mild mobility limitations and intact cognition.
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http://dx.doi.org/10.1159/000513505DOI Listing
March 2021

Long-term outcomes in children with absent pulmonary valve syndrome: it is not just fixing the heart.

Arch Dis Child 2021 Feb 25. Epub 2021 Feb 25.

Department of Cardiology and Cardiovascular Surgery, Birmingham Women's and Children's Hospitals NHS Foundation Trust, Birmingham, UK

Objective: Absent pulmonary valve syndrome (APV) is a rare condition usually associated with tetralogy of Fallot (TOF). Some infants develop respiratory failure from bronchial compression and the long-term neurodevelopmental outcome is unknown. We aimed to investigate the outcomes of APV and the need for long-term ventilation (LTV).

Design, Patients And Setting: Retrospective single-centre review of patients diagnosed with APV between 2007 and 2017.

Outcome Measures: Survival, neurological disability and postoperative LTV (≥3 months of non-invasive or invasive respiratory support).

Results: Thirty patients were identified, 22 (73%) of whom were prenatally diagnosed. Pregnancy was discontinued in one patient, while in utero death occurred in three. One was lost to follow-up. Of the remaining 25 liveborn, 21 had the classic TOF/APV. One baby died immediately after birth, while two patients had palliative care due to severe airway compression and inability to wean ventilation support. Surgical repair was performed in 21 of the 25 (84%) liveborn, with one awaiting surgery. Of those undergoing surgery, two patients died: one during surgery and the other due to severe airway malacia 5 months postsurgery. In the surgical group survival from birth at 1 and 5 years was 89% (95% CI 75% to 100%). Six (30%) patients required LTV postoperatively; all had surgery within the first 6 months of life. Learning and/or other physical difficulties were evident in 63%.

Conclusions: Majority of patients with APV are diagnosed antenatally. A third of those operated required LTV and over half had learning and/or other physical difficulties. Prospective studies are needed to identify prenatal factors that predict postnatal outcomes so parents can be counselled appropriately.
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http://dx.doi.org/10.1136/archdischild-2020-320219DOI Listing
February 2021

Jobb oldali videoasszisztált thoracoscopos thymectomia a thymoma nélküli, felnőttkori myasthenia gravis sebészi kezelésében.

Magy Seb 2020 Dec 12;73(4):125-139. Epub 2020 Dec 12.

1 Országos Korányi Pulmonológiai Intézet, 1121 Budapest, Korányi Frigyes út 1.

Összefoglaló. Bevezetés: A myasthenia gravis javallatával végzett csecsemőmirigy-eltávolítás sebésztechnikai szempontból lényegesen megváltozott az elmúlt közel 30 évben. A standard műtétnek számító transsternalis és transcervicalis thymectomia mellett elterjedt a videoasszisztált thoracoscopos sebészeti (VATS), később pedig a robot sebészeti megoldás is. Két intézetünkben 2011-2012-ben vezettük be a VATS thymectomiát. Módszer: A többféle technikai megoldás közül a mediastinumot a jobb mellüreg felől megközelítő utat választottuk. Eleinte 3, később 2 pontos perimammaris portot készítettünk a thymus elérésére a beteg háton fekvő helyzetében. Minden esetben ultrahangos vágóeszközt alkalmaztunk. Kiterjesztett thymectomiára törekedve a perithymikus zsírszövetet is eltávolítottuk, szélesen megnyitva a bal oldali mellüreget is. A betegek kiválasztásában az átlagos testsúlyú vagy soványabb betegeket részesítettük előnyben. Eredmények: 8 év és 4 hónap alatt 92 beteget műtöttünk a fenti módszerrel thymoma nélküli myasthenia gravis alapbetegséggel. 20 férfi és 72 nő. Átlagéletkor 33,1 év (19-75 év). A műtéti idő 35-160 percig terjedt, átlagosan 82,3 perc volt. A tömegesebb mediastinalis zsírszövet néhány betegnél nehezítette a tájékozódást és a maradéktalan eltávolítást. Műtét alatt 4 esetben érsérülés és 3 ellenoldali tüdősérülés következett be. Két konverziót végeztünk (1-1 sternotomia és thoracotomia). Idegsérülés nem történt. Tíz beteg igényelt néhány órás művi lélegeztetést a műtét után, a többi beteget a műtőasztalon extubáltuk. Reintubáció, tracheostomia, légzési elégtelenség, műtéti halálozás nem volt. Az intenzív ápolási idő átlaga: 1,1 (0-11) nap. A teljes kórházi ápolási idő átlaga: 4,8 (3-15) nap. A drenázsidő 1-4 nap, átlagosan 1,16 nap. Két beteg (2,41%) halt meg a műtétet követően 1 és 5 éven belül. További 81 beteg 12-108 (átlag: 48) hónapos követése során a myastheniás állapotban 21 (25,3%) betegnél komplett, 4 (4,82%) betegnél gyógyszeres remisszió, 20 (24,1%) betegnél minimális manifesztáció, 28 (33,73%) betegnél egyéb javulás volt megállapítható. 4 (4,82%) beteg állapota változatlan maradt, 4 (4,82%) betegé pedig romlott. Következtetés: A VATS thymectomia teljesen új utat jelent a transsternalis módszerben járatos sebészek számára. A tömegesebb mediastinalis zsírszövet nagyon megnehezíti a műtétet. A perioperatív szak nagyon kedvező a betegek számára, és a késői eredmények is elfogadhatóak. Kérdéses, hogy a thymus minden esetben maradéktalanul eltávolítható-e ezzel a módszerrel.

Summary:

Introduction: Surgical technique of thymectomy performed for treatment of myasthenia gravis has considerably changed in the last almost 30 years. In addition to standard interventions - transsternal and transcervical thymectomy -, video-assisted thoracoscopic interventions (VATS), later on robotic surgery came into general use. In our two institutions, we apply VATS thymectomy since 2011.

Methods: There are several different surgical techniques for this purpose; we approached the mediastinum through the right thoracic cavity. We prepared initially 3, later on 2 perimammal ports for the access of the thymus; the patients were in supine position during surgery. We used an ultrasonic cutting device in all cases. In order to perform extended thymectomy, we removed the fatty tissue around the thymus and opened widely the left thoracic cavity, too. During patient enrollment, we preferred patients with normal or lower body weight.

Results: During 8 years and 4 months, we operated on 92 patients using this method for myasthenia gravis without thymoma; there were 20 male and 72 female patients at the age of 33 years on average (19-75 years). Duration of surgery was 35-160 minutes, 82.3 minutes on average. The bulky fatty tissue around the thymus made the orientation and the complete removal more difficult in a few patients. We experienced vascular injury in 4 cases and injury of the contralateral lung in 3 cases. Conversion was necessary in 2 cases (1 sternotomy and 1 thoracotomy), there were no nerve injuries. Assisted ventilation was necessary in case of ten patients in the postoperative period for a few hours; all other patients were extubated on the operating table. There was no need for repeated intubation and tracheostomy; there was no respiratory insufficiency and perioperative mortality. Duration of ICU care was 1.1 days on the average (0-11 days), that of the total hospital care 4.8 days on average (3-15 days). Duration of thoracic drainage was 1.16 days on average (1-4 days). Two patients (2.41%) died within one and five years after surgery. During 12-108 months (48 months on average) follow-up of 81 patients, 21 patients (25.3%) suffering from myasthenia total recovery was observed, pharmacologic remission was achieved in 4 patients (5.3%), minimal manifestation remained in 23 patients (24.1%), while in 28 patients (33.73%) other improvement was observed. The status of 4 patients (4.82%) remained unchanged and that of 4 patients (5.3%) worsened.

Conclusion: VATS thymectomy represents a completely new surgical method for surgeons having experience in transsternal surgical technique. Bulky mediastinal fatty tissue makes surgery very difficult. The perioperative period is advantageous for the patients and also the long term follow-up results are acceptable. It is questionable that the thymus can be completely removed with this method in all cases.
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http://dx.doi.org/10.1556/1046.73.2020.4.1DOI Listing
December 2020

High Frequency and Intensity Rehabilitation in 641 Subacute Ischemic Stroke Patients.

Arch Phys Med Rehabil 2021 01 27;102(1):9-18. Epub 2020 Aug 27.

University of Groningen, University Medical Center, Groningen, The Netherlands.

Objectives: To determine the effects of exergaming on quality of life (QoL), motor, and clinical symptoms in subacute stroke patients.

Design: A pseudorandomized controlled trial, using a before-after test design.

Setting: University hospital.

Participants: Subacute, ischemic stroke outpatients (N=3857), 680 of whom were randomized and 641 completed the study.

Interventions: We determined the effects of 5 times a week twice daily (EX2; 50 sessions; n=286) and once daily (EX1; 25 sessions; n=272) exergaming and low-intensity standard care (control [CON]; 25 sessions; n=83) on clinical, mobility, blood pressure (BP), and QoL outcomes.

Main Outcome Measures: The primary outcome was Modified Rankin Scale. Secondary outcomes were activities of daily living, 5 aspects of health-related QoL, Beck Depression Inventory, 6-minute walk test (6MWT), Berg Balance Scale (BBS), and static balance (center of pressure).

Results: During exercise, the peak heart rate was 134, 134, and 126 beats per minute in the EX2, EX1, and CON groups, respectively. mRS improved similarly in the EX2 (-1.8; effect size, d=-4.0) and EX1 (-1.4; d=-2.6) groups, but more than in the CON group (-0.7; d=-0.6). QoL, Barthel Index, BBS, 6MWT, and standing posturography improved more in the EX2 group and the same in the EX1 and CON groups. Systolic and diastolic resting BP decreased more in the EX2 and EX1 groups than in the CON group. The intervention effects did not differ between men (n=349) and women (n=292).

Conclusions: Twice daily compared with once daily high-intensity exergaming or once daily lower intensity standard care produced superior effects on clinical and motor symptoms, BP, and QoL in male and female subacute ischemic stroke participants.
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http://dx.doi.org/10.1016/j.apmr.2020.07.012DOI Listing
January 2021

Exercise Effects on Multiple Sclerosis Quality of Life and Clinical-Motor Symptoms.

Med Sci Sports Exerc 2020 05;52(5):1007-1014

University Medical Center Groningen, University of Groningen, Groningen, THE NETHERLANDS.

Introduction: Different therapies can improve clinical and motor symptoms of multiple sclerosis (MS) similarly, but studies comparing the effects of different exercise therapies on clinical and motor outcomes are scant. We compared the effects of exergaming (EXE), balance (BAL), cycling (CYC), proprioceptive neuromuscular facilitation (PNF), and a standard care wait-listed control group (CON) on clinical and motor symptoms and quality of life (QoL) in people with MS (PwMS).

Methods: PwMS (n = 68, 90% female; age, 47.0 yr; Expanded Disability Status Scale score 5-6) were randomized into five groups. Before and after the interventions (five times a week for 5 wk), PwMS were tested for MS-related clinical and motor symptoms (Multiple Sclerosis Impact Scale-29 (MSIS-29), primary outcome), QoL (EuroQol Five Dimensions Questionnaire), symptoms of depression, gait and balance ability (Tinetti Assessment Tool), static and dynamic balance and fall risk (Berg Balance Scale), walking capacity (6-min walk test), and standing posturography on a force platform.

Results: EXE, BAL, and CYC improved the MSIS-29 scores similarly. EXE and CYC improved QoL and walking capacity similarly but more than BAL. Only EXE improved gait and balance scores (Tinetti Assessment Tool). EXE and BAL improved fall risk and standing balance similarly but more than CYC. PNF and CON revealed no changes. The EuroQol Five Dimensions Questionnaire moderated the exercise effects on the MSIS-29 scores only in EXE. Changes in QoL and changes in the MSIS-29 scores correlated (R = 0.73) only in EXE.

Conclusion: In conclusion, BAL and CYC but EXE in particular, but not PNF, can improve clinical and motor symptoms and QoL in PwMS (Expanded Disability Status Scale score 5 to 6), expanding the evidence-based exercise options to reduce mobility limitations in PwMS.
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http://dx.doi.org/10.1249/MSS.0000000000002228DOI Listing
May 2020

[The occurrence of neuroinvasive symptoms caused by the West Nile virus at an emergency center].

Orv Hetil 2019 Dec;160(51):2026-2035

Sürgősségi Betegellátó Centrum,Somogy Megyei Kaposi Mór Oktató Kórház Kaposvár, Tallián Gyula u. 20-32., 7400.

According to the European Centre for Disease Prevention and Control, the prevalence of neuroinvasive symptoms caused by the West Nile virus (WNV) has significantly increased in the past years throughout Europe, including Hungary. The rise may be attributed to changes in precipitation and climate. The WNV zoonosis is spread by mosquitoes. It is mostly asymptomatic, flu-like symptoms occur in 20% of the cases and in less than 1% a neuroinvasive disease with a lethal outcome may develop. Our aim was to demonstrate the neuroinvasive symptomatology and the diagnosis and treatment of WNV infections by describing our patient cases as well as to resolve differential diagnostic dilemmas. We report the cases of 4 patients treated at the "Moritz Kaposi" Somogy County Hospital between the 31st July and 4th September, 2018, with WNV, whose diagnoses were confirmed by serological and molecular biological methods. An epidemiological overview of WNV infections was also given. Four patients were confirmed to have had WNV infection in the given time period. A wide range of neurological symptoms were observed in each patient and death occurred in one case. The patients were elderly with a number of comorbidities. The appearance of more severe, neuroinvasive symptoms following WNV infections is also characteristic of Hungary. The treatment of the infection is supportive, including giving pain relievers and the management of secondary infections. It is important to consider the possibility of a WNV infection in the case of a neurological disease of unknown origin, particularly if the symptoms indicate encephalitis. Orv Hetil. 2019; 160(51): 2026-2035.
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http://dx.doi.org/10.1556/650.2019.31575DOI Listing
December 2019

Kinetic Mechanisms of Fast Glutamate Sensing by Fluorescent Protein Probes.

Biophys J 2020 01 14;118(1):117-127. Epub 2019 Nov 14.

Molecular and Clinical Sciences Research Institute, St. George's University of London, London, United Kingdom. Electronic address:

We have developed probes based on the bacterial periplasmic glutamate/aspartate binding protein with either an endogenously fluorescent protein or a synthetic fluorophore as the indicator of glutamate binding for studying the kinetic mechanism of glutamate binding. iGluSnFR variants termed iGlu, iGlu, and iGlu cover a broad range of K-s (5.8 μM and 2.1 and 50 mM, respectively), and a novel fluorescently labeled indicator, Fl-GluBP, has a K of 9.7 μM. The fluorescence response kinetics of all the probes are consistent with a two-step mechanism involving ligand binding and isomerization either of the apo or the ligand-bound binding protein. Although the previously characterized ultrafast indicators iGlu and iGlu had monophasic fluorescence enhancement that occurred in the rate limiting isomerization step, the sensors described here all have biphasic binding kinetics with fluorescence increases occurring both in the glutamate binding and the isomerization steps. For iGlu and iGlu, the data indicate prebinding conformational change followed by ligand binding. In contrast, for iGlu and Fl-GluBP, glutamate binding is followed by isomerization. Thus, the effects of structural heterogeneity introduced by single amino acid changes around the binding site on the kinetic path of interactions with glutamate are revealed. Remarkably, glutamate binding with a diffusion-limited rate constant to iGlu and Fl-GluBP is detected for the first time, hinting at the underlying mechanism of the supremely rapid activation of the highly homologous α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor by glutamate binding.
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http://dx.doi.org/10.1016/j.bpj.2019.11.006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6950763PMC
January 2020

High-speed imaging of glutamate release with genetically encoded sensors.

Nat Protoc 2019 05 15;14(5):1401-1424. Epub 2019 Apr 15.

Institute for Synaptic Physiology, Center for Molecular Neurobiology Hamburg, Hamburg, Germany.

The strength of an excitatory synapse depends on its ability to release glutamate and on the density of postsynaptic receptors. Genetically encoded glutamate indicators (GEGIs) allow eavesdropping on synaptic transmission at the level of cleft glutamate to investigate properties of the release machinery in detail. Based on the sensor iGluSnFR, we recently developed accelerated versions of GEGIs that allow investigation of synaptic release during 100-Hz trains. Here, we describe the detailed procedures for design and characterization of fast iGluSnFR variants in vitro, transfection of pyramidal cells in organotypic hippocampal cultures, and imaging of evoked glutamate transients with two-photon laser-scanning microscopy. As the released glutamate spreads from a point source-the fusing vesicle-it is possible to localize the vesicle fusion site with a precision exceeding the optical resolution of the microscope. By using a spiral scan path, the temporal resolution can be increased to 1 kHz to capture the peak amplitude of fast iGluSnFR transients. The typical time frame for these experiments is 30 min per synapse.
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http://dx.doi.org/10.1038/s41596-019-0143-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6751072PMC
May 2019

Single Synapse Indicators of Impaired Glutamate Clearance Derived from Fast iGlu Imaging of Cortical Afferents in the Striatum of Normal and Huntington (Q175) Mice.

J Neurosci 2019 05 28;39(20):3970-3982. Epub 2019 Feb 28.

Cluster of Excellence Neurocure, Charité-Universitätsmedizin Berlin, 10117 Berlin, Germany,

Changes in the balance between glutamate (Glu) release and uptake may stimulate synaptic reorganization and even synapse loss. In the case of neurodegeneration, a mismatch between astroglial Glu uptake and presynaptic Glu release could be detected if both parameters were assessed independently and at a single-synapse level. This has now become possible due to a new imaging assay with the genetically encoded ultrafast Glu sensor iGlu We report findings from individual corticostriatal synapses in acute slices prepared from mice of either sex that were >1 year of age. Contrasting patterns of short-term plasticity and a size criterion identified two classes of terminals, presumably corresponding to the previously defined IT (intratelencephalic) and PT (pyramidal tract) synapses. The latter exhibited a higher degree of frequency potentiation/residual Glu accumulation and were selected for our first iGlu single-synapse study in Q175 mice, a model of Huntington's disease (HD). In HD mice, the decay time constant of the perisynaptic Glu concentration (TauD), as an indicator of uptake, and the peak iGlu amplitude, as an indicator of release, were prolonged and reduced, respectively. Treatment of WT preparations with the astrocytic Glu uptake blocker TFB-TBOA (100 nm) mimicked the TauD changes in homozygotes. Considering the largest TauD values encountered in WT, ∼40% of PT synapses tested in Q175 heterozygotes can be classified as dysfunctional. Moreover, HD but not WT synapses exhibited a positive correlation between TauD and the peak amplitude of iGlu Finally, EAAT2 (excitatory amino acid transport protein 2) immunoreactivity was reduced next to corticostriatal terminals. Thus, astrocytic Glu transport remains a promising target for therapeutic intervention. Alterations in astrocytic Glu uptake can play a role in synaptic plasticity and neurodegeneration. Until now, the sensitivity of synaptic responses to pharmacological transport block and the resulting activation of NMDA receptors were regarded as reliable evidence for a mismatch between synaptic uptake and release. But the latter parameters are interdependent. Using a new genetically encoded sensor to monitor extracellular glutamate concentration ([Glu]) at individual corticostriatal synapses, we can now quantify the time constant of perisynaptic [Glu] decay (as an indicator of uptake) and the maximal [Glu] elevation next to the active zone (as an indicator of Glu release). The results provide a positive answer to the hitherto unresolved question of whether neurodegeneration (e.g., Huntington's disease) associates with a glutamate uptake deficit at tripartite excitatory synapses.
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http://dx.doi.org/10.1523/JNEUROSCI.2865-18.2019DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6520508PMC
May 2019

The kinetic mechanisms of fast-decay red-fluorescent genetically encoded calcium indicators.

J Biol Chem 2019 03 16;294(11):3934-3946. Epub 2019 Jan 16.

From the Molecular and Clinical Sciences Research Institute, St. George's, University of London, London SW17 0RE, United Kingdom and

Genetically encoded calcium indicators (GECIs) are useful reporters of cell-signaling, neuronal, and network activities. We have generated novel fast variants and investigated the kinetic mechanisms of two recently developed red-fluorescent GECIs (RGECIs), mApple-based jRGECO1a and mRuby-based jRCaMP1a. In the formation of fluorescent jRGECO1a and jRCaMP1a complexes, calcium binding is followed by rate-limiting isomerization. However, fluorescence decay of calcium-bound jRGECO1a follows a different pathway from its formation: dissociation of calcium occurs first, followed by the peptide, similarly to GCaMP-s. In contrast, fluorescence decay of calcium-bound jRCaMP1a occurs by the reversal of the on-pathway: peptide dissociation is followed by calcium. The mechanistic differences explain the generally slower off-kinetics of jRCaMP1a-type indicators compared with GCaMP-s and jRGECO1a-type GECI: the fluorescence decay rate of f-RCaMP1 was 21 s, compared with 109 s for f-RGECO1 and f-RGECO2 (37 °C). Thus, the CaM-peptide interface is an important determinant of the kinetic responses of GECIs; however, the topology of the structural link to the fluorescent protein demonstrably affects the internal dynamics of the CaM-peptide complex. In the dendrites of hippocampal CA3 neurons, f-RGECO1 indicates calcium elevation in response to a 100 action potential train in a linear fashion, making the probe particularly useful for monitoring large-amplitude, fast signals, those in dendrites, muscle cells, and immune cells.
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http://dx.doi.org/10.1074/jbc.RA118.004543DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6422079PMC
March 2019

Ultrafast glutamate sensors resolve high-frequency release at Schaffer collateral synapses.

Proc Natl Acad Sci U S A 2018 05 7;115(21):5594-5599. Epub 2018 May 7.

Molecular and Clinical Sciences Research Institute, St George's, University of London, SW17 0RE London, United Kingdom;

Glutamatergic synapses display a rich repertoire of plasticity mechanisms on many different time scales, involving dynamic changes in the efficacy of transmitter release as well as changes in the number and function of postsynaptic glutamate receptors. The genetically encoded glutamate sensor iGluSnFR enables visualization of glutamate release from presynaptic terminals at frequencies up to ∼10 Hz. However, to resolve glutamate dynamics during high-frequency bursts, faster indicators are required. Here, we report the development of fast (iGlu ) and ultrafast (iGlu ) variants with comparable brightness but increased for glutamate (137 μM and 600 μM, respectively). Compared with iGluSnFR, iGlu has a sixfold faster dissociation rate in vitro and fivefold faster kinetics in synapses. Fitting a three-state model to kinetic data, we identify the large conformational change after glutamate binding as the rate-limiting step. In rat hippocampal slice culture stimulated at 100 Hz, we find that iGlu is sufficiently fast to resolve individual glutamate release events, revealing that glutamate is rapidly cleared from the synaptic cleft. Depression of iGlu responses during 100-Hz trains correlates with depression of postsynaptic EPSPs, indicating that depression during high-frequency stimulation is purely presynaptic in origin. At individual boutons, the recovery from depression could be predicted from the amount of glutamate released on the second pulse (paired pulse facilitation/depression), demonstrating differential frequency-dependent filtering of spike trains at Schaffer collateral boutons.
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http://dx.doi.org/10.1073/pnas.1720648115DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6003469PMC
May 2018

Relaxed chromatin induced by histone deacetylase inhibitors improves the oligonucleotide-directed gene editing in plant cells.

J Plant Res 2018 Jan 23;131(1):179-189. Epub 2017 Aug 23.

Institute of Plant Biology, Biological Research Centre, Hungarian Academy of Sciences, Szeged, Hungary.

Improving efficiency of oligonucleotide-directed mutagenesis (ODM) is a prerequisite for wide application of this gene-editing approach in plant science and breeding. Here we have tested histone deacetylase inhibitor treatments for induction of relaxed chromatin and for increasing the efficiency of ODM in cultured maize cells. For phenotypic assay we produced transgenic maize cell lines expressing the non-functional Green Fluorescent Protein (mGFP) gene carrying a TAG stop codon. These transgenic cells were bombarded with corrective oligonucleotide as editing reagent to recover GFP expression. Repair of green fluorescent protein function was monitored by confocal fluorescence microscopy and flow cytometry was used for quantification of correction events. Sequencing PCR fragments of the GFP gene from corrected cells indicated a nucleotide exchange in the stop codon (TAG) from T to G nucleotide that resulted in the restoration of GFP function. We show that pretreatment of maize cells with sodium butyrate (5-10 mM) and nicotinamide (1-5 mM) as known inhibitors of histone deacetylases can cause elevated chromatin sensitivity to DNase I that was visualized in agarose gels and confirmed by the reduced presence of intact PCR template for the inserted exogenous mGFP gene. Maize cells with more relaxed chromatin could serve as an improved recipient for targeted nucleotide exchange as indicated by an average of 2.67- to 3.62-fold increase in GFP-positive cells. Our results stimulate further studies on the role of the condition of the recipient cells in ODM and testing the application of chromatin modifying agents in other, programmable nuclease-based genome-editing techniques in higher plants.
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http://dx.doi.org/10.1007/s10265-017-0975-8DOI Listing
January 2018

Functional studies of chronic lymphocytic leukemia B cells expressing β-integrin type complement receptors CR3 and CR4.

Immunol Lett 2017 09 31;189:73-81. Epub 2017 May 31.

MTA-ELTE Immunology Research Group, Department of Immunology, Eötvös Loránd University, Budapest, Hungary; Department of Immunology, Eötvös Loránd University, Budapest, Hungary. Electronic address:

The expression and role of CR3 (CD11b/CD18) and CR4 (CD11c/CD18) in B cells are not yet explored in contrast to myeloid cells, where these β-integrin type receptors are known to participate in various cellular functions, including phagocytosis, adherence and migration. Here we aimed to reveal the expression and role of CR3 and CR4 in human B cells. In B cells of healthy donors CR3 and CR4 are scarcely expressed. However, two patients with chronic lymphocytic leukemia (CLL) characterized by a peculiar immune-phenotype containing both CD5-positive and CD5-negative B cell populations made possible to study these molecules in distinct B cell subsets. We found that CD11b and CD11c were expressed on both CD5-positive and CD5-negative B cells, albeit to different extents. Our data suggest that these receptors are involved in spreading, since this activity of CpG-activated B cells on fibrinogen could be partially blocked by monoclonal antibodies specific for CD11b or CD11c. CpG-stimulation lead to proliferation of both CD5-positive and CD5-negative B cells of the patients with a less pronounced effect on the CD5-positive cells. In contrast to normal B cells, CLL B cells of both patients reacted to CpG-stimulation with robust IL-10 production. The concomitant, suboptimal stimulus via the BCR and TLR9 exerted either a synergistic enhancing effect or resulted in inhibition of proliferation and IL-10 production of patients' B cells. Our data obtained studying B cells of leukemic patients point to the role of CR3 and probably CR4 in the interaction of tumor cells with the microenvironment and suggest the involvement of IL-10 producing B cells in the pathologic process.
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http://dx.doi.org/10.1016/j.imlet.2017.05.016DOI Listing
September 2017

High affinity binding of amyloid β-peptide to calmodulin: Structural and functional implications.

Biochem Biophys Res Commun 2017 05 29;486(4):992-997. Epub 2017 Mar 29.

Dept. Biochemistry and Molecular Biology, Faculty of Sciences, University of Extremadura, Avda. de Elvas, s/n, 06006 Badajoz, Spain. Electronic address:

Amyloid β-peptides (Aβ) are a major hallmark of Alzheimer's disease (AD) and their neurotoxicity develop with cytosolic calcium dysregulation. On the other hand, calmodulin (CaM), a protein which plays a major multifunctional role in neuronal calcium signaling, has been shown to be involved in the regulation of non-amyloidogenic processing of amyloid β precursor protein (APP). Using fluorescent 6-bromoacetyl-2-dimethylaminonaphthalene derivatives of CaM, Badan-CaM, and human amyloid β(1-42) HiLyte™-Fluor555, we show in this work that Aβ binds with high affinity to CaM through the neurotoxic Aβ25-35 domain. In addition, the affinity of Aβ for calcium-saturated CaM conformation is approximately 20-fold higher than for CaM conformation in the absence of calcium (apo-CaM). Moreover, the value of K of 0.98 ± 0.11 nM obtained for Aβ1-42 dissociation from CaM saturated by calcium points out that CaM is one of the cellular targets with highest affinity for neurotoxic Aβ peptides. A major functional consequence of Aβ-CaM interaction is that it slowdowns Aβ fibrillation. The novel and high affinity interaction between calmodulin and Aβ shown in this work opens a yet-unexplored gateway to further understand the neurotoxic effect of Aβ in different neural cells and also to address the potential of calmodulin and calmodulin-derived peptides as therapeutic agents in AD.
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http://dx.doi.org/10.1016/j.bbrc.2017.03.151DOI Listing
May 2017

Design and mechanistic insight into ultrafast calcium indicators for monitoring intracellular calcium dynamics.

Sci Rep 2016 12 6;6:38276. Epub 2016 Dec 6.

Molecular and Clinical Sciences Research Institute, St George's, University of London, Cranmer Terrace, London SW17 0RE, UK.

Calmodulin-based genetically encoded fluorescent calcium indicators (GCaMP-s) are powerful tools of imaging calcium dynamics from cells to freely moving animals. High affinity indicators with slow kinetics however distort the temporal profile of calcium transients. Here we report the development of reduced affinity ultrafast variants of GCaMP6s and GCaMP6f. We hypothesized that GCaMP-s have a common kinetic mechanism with a rate-limiting process in the interaction of the RS20 peptide and calcium-calmodulin. Therefore we targeted specific residues in the binding interface by rational design generating improved indicators with GCaMP6f displaying fluorescence rise and decay times (t) of 1 and 3 ms (37 °C) in vitro, 9 and 22-fold faster than GCaMP6f respectively. In HEK293T cells, GCaMP6f revealed a 4-fold faster decay of ATP-evoked intracellular calcium transients than GCaMP6f. Stimulation of hippocampal CA1 pyramidal neurons with five action potentials fired at 100 Hz resulted in a single dendritic calcium transient with a 2-fold faster rise and 7-fold faster decay time (t of 40 ms) than GCaMP6f, indicating that tracking high frequency action potentials may be limited by calcium dynamics. We propose that the design strategy used for generating GCaMP6f is applicable for the acceleration of the response kinetics of GCaMP-type calcium indicators.
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http://dx.doi.org/10.1038/srep38276DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5138832PMC
December 2016

Response of Organ Structure and Physiology to Autotetraploidization in Early Development of Energy Willow Salix viminalis.

Plant Physiol 2016 Mar 4;170(3):1504-23. Epub 2016 Jan 4.

Institute of Plant Biology, Biological Research Centre, Hungarian Academy of Sciences, 6726 Szeged, Hungary (D.D., K.T., A.C., K.P., A.V.N., B.N., L.S., G.F., I.V., F.A.); andInstitute of Experimental Botany, Academy of Sciences of the Czech Republic, Prague, Czech Republic (R.V., P.D.).

The biomass productivity of the energy willow Salix viminalis as a short-rotation woody crop depends on organ structure and functions that are under the control of genome size. Colchicine treatment of axillary buds resulted in a set of autotetraploid S. viminalis var. Energo genotypes (polyploid Energo [PP-E]; 2n = 4x = 76) with variation in the green pixel-based shoot surface area. In cases where increased shoot biomass was observed, it was primarily derived from larger leaf size and wider stem diameter. Autotetraploidy slowed primary growth and increased shoot diameter (a parameter of secondary growth). The duplicated genome size enlarged bark and wood layers in twigs sampled in the field. The PP-E plants developed wider leaves with thicker midrib and enlarged palisade parenchyma cells. Autotetraploid leaves contained significantly increased amounts of active gibberellins, cytokinins, salicylic acid, and jasmonate compared with diploid individuals. Greater net photosynthetic CO2 uptake was detected in leaves of PP-E plants with increased chlorophyll and carotenoid contents. Improved photosynthetic functions in tetraploids were also shown by more efficient electron transport rates of photosystems I and II. Autotetraploidization increased the biomass of the root system of PP-E plants relative to diploids. Sections of tetraploid roots showed thickening with enlarged cortex cells. Elevated amounts of indole acetic acid, active cytokinins, active gibberellin, and salicylic acid were detected in the root tips of these plants. The presented variation in traits of tetraploid willow genotypes provides a basis to use autopolyploidization as a chromosome engineering technique to alter the organ development of energy plants in order to improve biomass productivity.
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http://dx.doi.org/10.1104/pp.15.01679DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4775130PMC
March 2016

Fast-Response Calmodulin-Based Fluorescent Indicators Reveal Rapid Intracellular Calcium Dynamics.

Sci Rep 2015 Nov 3;5:15978. Epub 2015 Nov 3.

Institute of Cardiovascular and Cell Science, St George's, University of London, London SW17 0RE, UK.

Faithful reporting of temporal patterns of intracellular Ca(2+) dynamics requires the working range of indicators to match the signals. Current genetically encoded calmodulin-based fluorescent indicators are likely to distort fast Ca(2+) signals by apparent saturation and integration due to their limiting fluorescence rise and decay kinetics. A series of probes was engineered with a range of Ca(2+) affinities and accelerated kinetics by weakening the Ca(2+)-calmodulin-peptide interactions. At 37 °C, the GCaMP3-derived probe termed GCaMP3fast is 40-fold faster than GCaMP3 with Ca(2+) decay and rise times, t1/2, of 3.3 ms and 0.9 ms, respectively, making it the fastest to-date. GCaMP3fast revealed discreet transients with significantly faster Ca(2+) dynamics in neonatal cardiac myocytes than GCaMP6f. With 5-fold increased two-photon fluorescence cross-section for Ca(2+) at 940 nm, GCaMP3fast is suitable for deep tissue studies. The green fluorescent protein serves as a reporter providing important novel insights into the kinetic mechanism of target recognition by calmodulin. Our strategy to match the probe to the signal by tuning the affinity and hence the Ca(2+) kinetics of the indicator is applicable to the emerging new generations of calmodulin-based probes.
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http://dx.doi.org/10.1038/srep15978DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4630588PMC
November 2015

Resting Heart Rate Is Not a Good Predictor of a Clustered Cardiovascular Risk Score in Adolescents: The HELENA Study.

PLoS One 2015 26;10(5):e0127530. Epub 2015 May 26.

GENUD (Growth, Exercise, Nutrition and Development) Research Group, Facultad de Ciencias de la Salud de la Universidad de Zaragoza, Zaragoza, Spain; Department of Preventive Medicine, School of Medicine of the University of São Paulo, São Paulo, Brazil.

Background: Resting heart rate (RHR) reflects sympathetic nerve activity a significant association between RHR and all-cause and cardiovascular mortality has been reported in some epidemiologic studies.

Methods: To analyze the predictive power and accuracy of RHR as a screening measure for individual and clustered cardiovascular risk in adolescents. The study comprised 769 European adolescents (376 boys) participating in the HELENA cross-sectional study (2006-2008) were included in this study. Measurements on systolic blood pressure, HOMA index, triglycerides, TC/HDL-c, VO2máx and the sum of four skinfolds were obtained, and a clustered cardiovascular disease (CVD) risk index was computed. The receiver operating characteristics curve was applied to calculate the power and accuracy of RHR to predict individual and clustered CVD risk factors.

Results: RHR showed low accuracy for screening CVD risk factors in both sexes (range 38.5%-54.4% in boys and 45.5%-54.3% in girls). Low specificity's (15.6%-19.7% in boys; 18.1%-20.0% in girls) were also found. Nevertheless, the sensitivities were moderate-to-high (61.4%-89.1% in boys; 72.9%-90.3% in girls).

Conclusion: RHR is a poor predictor of individual CVD risk factors and of clustered CVD and the estimates based on RHR are not accurate. The use of RHR as an indicator of CVD risk in adolescents may produce a biased screening of cardiovascular health in both sexes.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0127530PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4444318PMC
February 2016

Complement receptor type 1 (CR1/CD35) expressed on activated human CD4+ T cells contributes to generation of regulatory T cells.

Immunol Lett 2015 Apr 2;164(2):117-24. Epub 2015 Mar 2.

MTA-ELTE Immunology Research Group, Budapest, Hungary; Department of Immunology, Eötvös Loránd University, Budapest, Hungary. Electronic address:

The role of complement in the regulation of T cell immunity has been highlighted recently by several groups. We were prompted to reinvestigate the role of complement receptor type 1 (CR1, CD35) [corrected] in human T cells based on our earlier data showing that activated human T cells produce C3 (Torok et al. (2012) [48]) and also by results demonstrating that engagement of Membrane Cofactor Protein (MCP, CD46) induces a switch of anti-CD35-activated [corrected] helper T cells into regulatory T cells (Kemper et al. (2003) [17]). We demonstrate here that co-ligation of CD46 and CD35, [corrected] the two C3b-binding structures present on activated CD4+ human T cells significantly enhances CD25 expression, elevates granzyme B production and synergistically augments cell proliferation. The role of CR1 in the development of the Treg phenotype was further confirmed by demonstrating that its engagement enhances IL-10 production and reduces IFNγ release by the activated CD4+ T cells in the presence of excess IL-2. The functional in vivo relevance of our findings was highlighted by the immunohistochemical staining of tonsils, revealing the presence of CD4/CD35 [corrected] double positive lymphocytes mainly in the inter-follicular regions where direct contact between CD4+ T cells and B lymphocytes occurs. Regarding the in vivo relevance of the complement-dependent generation of regulatory T cells in secondary lymphoid organs we propose a scenario shown in the figure. The depicted process involves the sequential binding of locally produced C3 fragments to CD46 and CD35 [corrected] expressed on activated T cells, which - in the presence of excess IL-2 - leads to the development of Treg cells.
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http://dx.doi.org/10.1016/j.imlet.2015.02.009DOI Listing
April 2015

Dispersal pathways and genetic differentiation among worldwide populations of the invasive weed Centaurea solstitialis L. (Asteraceae).

PLoS One 2014 31;9(12):e114786. Epub 2014 Dec 31.

Department of Biology, University of Massachusetts Boston, Boston, Massachusetts, United States of America.

The natural history of introduced species is often unclear due to a lack of historical records. Even when historical information is readily available, important factors of the invasions such as genetic bottlenecks, hybridization, historical relationships among populations and adaptive changes are left unknown. In this study, we developed a set of nuclear, simple sequence repeat markers and used these to characterize the genetic diversity and population structure among native (Eurasian) and non-native (North and South American) populations of Centaurea solstitialis L., (yellow starthistle). We used these data to test hypotheses about the invasion pathways of the species that were based on historical and geographical records, and we make inferences about historical relationships among populations and demographic processes following invasion. We confirm that the center of diversity and the native range of the species is likely the eastern Mediterranean region in the vicinity of Turkey. From this region, the species likely proceeded to colonize other parts of Europe and Asia via a slow, stepwise range expansion. Spanish populations were the primary source of seed to invade South America via human-mediated events, as was evident from historical records, but populations from the eastern Mediterranean region were also important. North American populations were largely derived from South America, but had secondary contributors. We suggest that the introduction history of non-native populations from disparate parts of the native range have allowed not just one, but multiple opportunities first in South America then again in North America for the creation of novel genotypes via intraspecific hybridization. We propose that multiple intraspecific hybridization events may have created especially potent conditions for the selection of a noxious invader, and may explain differences in genetic patterns among North and South America populations, inferred differences in demographic processes, as well as morphological differences previously reported from common garden experiments.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0114786PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4281129PMC
September 2015

Deletion of 4q28.3-31.23 in the background of multiple malformations with pulmonary hypertension.

Mol Cytogenet 2014 5;7:36. Epub 2014 Jun 5.

Department of Medical Genetics, Clinical Centre, University of Pecs, Szigeti 12, Pecs H-7624, Hungary ; Szentágothai Research Centre, University of Pecs, Ifjusag 20, Pecs H-7624, Hungary.

The 4q deletion syndrome shows a broad spectrum of clinical manifestations consisting of key features comprising growth failure, developmental delay, craniofacial dysmorphism, digital anomalies, and cardiac and skeletal defects. We have identified a de novo interstitial distal deletion in a 9 month-old girl with growth failure, developmental delay, ventricular septum defect in the subaortic region, patent foramen ovale and patent ductus arteriosus, vascular malformation of the lung, dysgenesis of the corpus callosum and craniofacial dysmorphism using array-comparative genomic hybridization. This de novo deletion is located at 4q28.3-31.23 (136,127,048 - 150,690,325), its size is 14.56 Mb, and contains 8 relevant genes (PCDH18, SETD7, ELMOD2, IL15, GAB1, HHIP, SMAD1, NR3C2) with possible contributions to the phenotype. Among other functions, a role in lung morphogenesis and tubulogenesis can be attributed to the deleted genes in our patient, which may explain the unique feature of vascular malformation of the lung leading to pulmonary hypertension. With the detailed molecular characterization of our case with 4q- syndrome we hope to contribute to the elucidation of the genetic spectrum of this disorder.
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http://dx.doi.org/10.1186/1755-8166-7-36DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4066825PMC
June 2014

Human T cell derived, cell-bound complement iC3b is integrally involved in T cell activation.

Immunol Lett 2012 Mar;143(1):131-6

Department of Immunology, Eötvös Loránd University, Budapest, Hungary.

Although the complement system is thought to be mainly involved in innate immunity and in the humoral arm of adaptive responses, evidence implicating that complement impacts T cell responses are accumulating recently. The role of the various activation products of the major complement component C3 were mainly studied so far in animal systems, and investigations regarding the effect of different C3-fragments on human T cells are sparse. Here we show that anti-CD3 activated human T lymphocytes derived from the blood and tonsil of healthy individuals produce C3, and the major cleavage fragment that appears on the T cell surface is iC3b. Based on studies carried out in allogenic system we demonstrate that the T cell membrane bound iC3b binds to the CR3 and probably to CR4 receptors expressed on monocyte-derived dendritic cells, and this interaction leads to significantly enhanced T-cell proliferation. Since neither C3aR and nor C3a binding could be detected on the membrane of anti-CD3 activated T cells, our findings indicate that in humans – in contrast to mice – the C3a peptide is most probably not involved directly in the T cell activation process.
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http://dx.doi.org/10.1016/j.imlet.2012.02.003DOI Listing
March 2012

Lobe-specific functions of Ca2+·calmodulin in alphaCa2+·calmodulin-dependent protein kinase II activation.

J Biol Chem 2011 Apr 7;286(14):12308-16. Epub 2011 Feb 7.

Division of Basic Medical Sciences, St. George's, University of London, Cranmer Terrace, London SW17 0RE, UK.

N-methyl-D-aspartic acid receptor-dependent long term potentiation (LTP), a model of memory formation, requires Ca2+·calmodulin-dependent protein kinase II (αCaMKII) activity and Thr286 autophosphorylation via both global and local Ca2+ signaling, but the mechanisms of signal transduction are not understood. We tested the hypothesis that the Ca2+-binding activator protein calmodulin (CaM) is the primary decoder of Ca2+ signals, thereby determining the output, e.g. LTP. Thus, we investigated the function of CaM mutants, deficient in Ca2+ binding at sites 1 and 2 of the N-terminal lobe or sites 3 and 4 of the C-terminal CaM lobe, in the activation of αCaMKII. Occupancy of CaM Ca2+ binding sites 1, 3, and 4 is necessary and sufficient for full activation. Moreover, the N- and C-terminal CaM lobes have distinct functions. Ca2+ binding to N lobe Ca2+ binding site 1 increases the turnover rate of the enzyme 5-fold, whereas the C lobe plays a dual role; it is required for full activity, but in addition, via Ca2+ binding site 3, it stabilizes ATP binding to αCaMKII 4-fold. Thr286 autophosphorylation is also dependent on Ca2+ binding sites on both the N and the C lobes of CaM. As the CaM C lobe sites are populated by low amplitude/low frequency (global) Ca2+ signals, but occupancy of N lobe site 1 and thus activation of αCaMKII requires high amplitude/high frequency (local) Ca2+ signals, lobe-specific sensing of Ca2+-signaling patterns by CaM is proposed to explain the requirement for both global and local Ca2+ signaling in the induction of LTP via αCaMKII.
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http://dx.doi.org/10.1074/jbc.M110.157057DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3069434PMC
April 2011

Overproduction of a rice aldo-keto reductase increases oxidative and heat stress tolerance by malondialdehyde and methylglyoxal detoxification.

Plant Mol Biol 2011 Mar 19;75(4-5):399-412. Epub 2011 Jan 19.

Institute of Plant Biology, Biological Research Center, Hungarian Academy of Sciences, Temesvári krt. 62, 6726, Szeged, Hungary.

The accumulation of toxic compounds generated by the interaction between reactive oxygen species and polyunsaturated fatty acids of membrane lipids can significantly damage plant cells. A plethora of enzymes act on these reactive carbonyls, reducing their toxicity. Based on the chromosomal localization and on their homology with other stress-induced aldo-keto reductases (AKRs) we have selected three rice AKR genes. The transcription level of OsAKR1 was greatly induced by abscisic acid and various stress treatments; the other two AKR genes tested were moderately stress-inducible. The OsAKR1 recombinant protein exhibited a high nicotinamide adenine dinucleotide phosphate-dependent catalytic activity to reduce toxic aldehydes including glycolysis-derived methylglyoxal (MG) and lipid peroxidation-originated malondialdehyde (MDA). The function of this enzyme in MG detoxification was demonstrated in vivo in E. coli and in transgenic plants overproducing the OsAKR1 protein. Heterologous synthesis of the OsAKR1 enzyme in transgenic tobacco plants resulted in increased tolerance against oxidative stress generated by methylviologen (MV) and improved resistance to high temperature. In these plants lower levels of MDA were detected both following MV and heat treatment due to the activity of the OsAKR1 enzyme. The transgenic tobaccos also exhibited higher AKR activity and accumulated less MG in their leaves than the wild type plants; both in the presence and absence of heat stress. These results support the positive role of OsAKR1 in abiotic stress-related reactive aldehyde detoxification pathways and its use for improvement of stress tolerance in plants.
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http://dx.doi.org/10.1007/s11103-011-9735-7DOI Listing
March 2011

A closer look into the GL7 antigen: its spatio-temporally selective differential expression and localization in lymphoid cells and organs in human.

Immunol Lett 2010 May 6;130(1-2):89-96. Epub 2010 Jan 6.

Immunology Research Group of Hungarian Academy of Sciences at Eotvos Lorand University, Budapest, Hungary.

The GL7 epitope was originally described as part of a late lymphocyte activation antigen expressed in mouse and widely used since then as a marker of germinal center. Here we report on its differential expression by rat and human immune cells and lymphoid organs. Expression pattern of the GL7 epitope in rats is similar to that described earlier in mice, namely that GL7 antigen appears only on lymphocytes after 48h activation. In humans lymphocytes, but not the differentiated cells of myeloid origin, express this epitope. The GL7 epitope is up-regulated upon in vitro activation of primary T cells, while a slightly decreased expression is found on B lymphocytes. Fluorescent immunohistochemistry shows discrete location of GL7(hi) cells in human tonsil. GL7 antibody intensely stains CD19(+), IgD(+), IgM(low) B lymphocytes found at the margin of B cell follicles. The GL7 epitope is constitutively and highly raft-associated in human lymphoid cells. Strong neuraminidase- and partial papain-sensitivity of the GL7 epitope on human lymphocytes indicates a sialic acid-containing epitope linked either to one (or more) membrane protein(s) or to lipids. The lymphocyte-restricted GL7 epitope expression and the activation-dependent bi-directional change in the amount of the epitope suggest a functional role for GL7 epitope linked to carbohydrate-based immunoregulation.
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http://dx.doi.org/10.1016/j.imlet.2009.12.008DOI Listing
May 2010

Time-dependent autoinactivation of phospho-Thr286-alphaCa2+/calmodulin-dependent protein kinase II.

J Biol Chem 2009 Oct 4;284(41):28146-28155. Epub 2009 Aug 4.

Division of Basic Medical Sciences, St George's, University of London, Cranmer Terrace, London SW17 0RE, United Kingdom. Electronic address:

Ca(2+)/calmodulin-dependent protein kinase II (alphaCaMKII) is thought to exert its role in memory formation by autonomous Ca(2+)-independent persistent activity conferred by Thr(286) autophosphorylation, allowing the enzyme to remain active even when intracellular [Ca(2+)] has returned to resting levels. Ca(2+) sequestration-induced inhibition, caused by a burst of Thr(305/306) autophosphorylation via calmodulin (CaM) dissociation from the Thr(305/306) sites, is in conflict with this view. The processes of CaM binding, autophosphorylation, and inactivation are dissected to resolve this conflict. Upon Ca(2+) withdrawal, CaM sequential domain dissociation is observed, starting with the rapid release of the first (presumed N-terminal) CaM lobe, thought to be bound at the Thr(305/306) sites. The time courses of Thr(305/306) autophosphorylation and inactivation, however, correlate with the slow dissociation of the second (presumed C-terminal) CaM lobe. Exposure of the Thr(305/306) sites is thus not sufficient for their autophosphorylation. Moreover, Thr(305/306) autophosphorylation and autoinactivation are shown to occur in the continuous presence of Ca(2+) and bound Ca(2+)/CaM by time courses similar to those seen following Ca(2+) sequestration. Our investigation of the activity and mechanisms of phospho-Thr(286)-alphaCaMKII thus shows time-dependent autoinactivation, irrespective of the continued presence of Ca(2+) and CaM, allowing a very short, if any, time window for Ca(2+)/CaM-free phospho-Thr(286)-alphaCaMKII activity. Physiologically, the time-dependent autoinactivation mechanisms of phospho-Thr(286)-alphaCaMKII (t(1/2) of approximately 50 s at 37 degrees C) suggest a transient kinase activity of approximately 1 min duration in the induction of long term potentiation and thus memory formation.
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http://dx.doi.org/10.1074/jbc.M109.005900DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2788865PMC
October 2009

Expression and role of CR1 and CR2 on B and T lymphocytes under physiological and autoimmune conditions.

Mol Immunol 2009 Sep 25;46(14):2767-73. Epub 2009 Jun 25.

Department of Immunology, Biological Institute, Eötvös Loránd University, Budapest, Hungary.

The involvement of complement in the development and regulation of antibody responses under both healthy and pathological conditions is known for long. Unravelling the molecular mechanisms underlying the events however is still in progress. This review focuses on the role of complement receptors CR1 (CD35) and CR2 (CD21) expressed on T and B cells. Alteration in the expression and function of these receptors may contribute to the initiation and maintenance of immune complex mediated autoimmune diseases such as systemic lupus erythematosus and rheumatoid arthritis. Recent data regarding complement receptor expression on T lymphocytes and on memory B cells are also discussed.
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http://dx.doi.org/10.1016/j.molimm.2009.05.181DOI Listing
September 2009

Calmodulin association with connexin32-derived peptides suggests trans-domain interaction in chemical gating of gap junction channels.

J Biol Chem 2008 Oct 1;283(40):26911-20. Epub 2008 Aug 1.

Division of Basic Medical Sciences, St George's, University of London, London, SW17 0RE United Kingdom.

Calmodulin plays a key role in the chemical gating of gap junction channels. Two calmodulin-binding regions have previously been identified in connexin32 gap junction protein, one in the N-terminal and another in the C-terminal cytoplasmic tail of the molecule. The aim of this study was to better understand how calmodulin interacts with the connexin32-binding domains. Lobe-specific interactions of calmodulin with connexin32 peptides were studied by stopped flow kinetics, using Ca(2+) binding-deficient mutants. Peptides corresponding to the N-terminal tail (residues 1-22) of connexin32 engaged both the N- and C-terminal lobes (N- and C-lobes) of calmodulin, binding with higher affinity to the C-lobe of calmodulin (Ca(2+) dissociation rate constants k(3,4), 1.7+/-0.5 s(-1)) than to the N-lobe (k(1,2), 10.8+/-1.3 s(-1)). In contrast, peptides representing the C-terminal tail domain (residues 208-227) of connexin32 bound either the C- or the N-lobe but only one calmodulin lobe at a time (k(3,4), 2.6+/-0.1 s(-1) or k(1), 13.8+/-0.5 s(-1) and k(2), 1000 s(-1)). The calmodulin-binding domains of the N- and C-terminal tails of connexin32 were best defined as residues 1-21 and 216-227, respectively. Our data, showing separate functions of the N- and C-lobes of calmodulin in the interactions with connexin32, suggest trans-domain or trans-subunit bridging by calmodulin as a possible mechanism of gap junction gating.
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http://dx.doi.org/10.1074/jbc.M801434200DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2555998PMC
October 2008

A two-state model for Ca2+/CaM-dependent protein kinase II (alphaCaMKII) in response to persistent Ca2+ stimulation in hippocampal neurons.

Cell Calcium 2008 Nov 23;44(5):465-78. Epub 2008 Apr 23.

Division of Basic Medical Sciences, St George's, University of London, Cranmer Terrace, London SW17 0RE, UK.

Persistent elevation of the intracellular free Ca(2+) concentration [Ca(2+)](i) is neurotoxic and therefore it is important to understand how it affects downstream components of the Ca(2+) signaling pathway. The response of calmodulin (CaM) and alphaCa(2+)/CaM-dependent protein kinase II (alphaCaMKII), to intracellular Ca(2+) overload in hippocampal neurons is studied by confocal imaging of fluorescently tagged proteins. Transient and persistent redistribution of CaM and alphaCaMKII together is seen from the cytosol to dendritic and somatic punctae. Typical persistent redistribution occurs following a lag of 138+/-(S.E.M.) 12 s and is complete at 460+/-(S.E.M.) 34 s (n=18), lack of Thr(286)-autophosphorylation of alphaCaMKII however promotes the formation of early transient punctae (peak at 40 s). In contrast, the T286D-mimick of phospho-Thr(286)-alphaCaMKII forms punctae with a delay >10 min, indicating that Thr(286)-autophosphorylation is antagonistic to CaMKII clustering. A two-state model is proposed in which phospho-Thr(286)-alphaCaMKII, formed immediately upon Ca(2+) stimulation, is primarily responsible for target interactions and memory functions of alphaCaMKII. However, a distinct clustering form denoted alphaCaMKII(c), generated upon persistent intracellular free Ca(2+) elevation, is deposited in the punctae which are made of self-interacting CaM/CaMKII complexes. Punctate deposition disables both the interactions and the activity of CaMKII.
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http://dx.doi.org/10.1016/j.ceca.2008.03.003DOI Listing
November 2008