Změnit instituci
Pokročilé nano a mikrotechnologie Pokročilé materiály Strukturní biologie Gen. a prot. rostlin. systémů Molekulární medicína Výzkum mozku a lidské mysli Molekulární vet. medicína

Selected CEITEC Publications - 3Q 2013

Full versions of all publications can be downloaded by clicking on the titles.

 

Current Drug Metabolism
Nanocarriers for anticancer drugs - New trends in nanomedicine

Drbohlavova J., Chomoucka J., Adam V., Ryvolova M., Eckschlager T., Hubalek J., Kizek R.

Research Group: Smart Nanodevices
Research Programme: Advanced Nanotechnologies and Microtechnologies

Abstract
This review provides a brief overview of the variety of carriers employed for targeted drug delivery used in cancer therapy and summarizes advantages and disadvantages of each approach. Particularly, the attention was paid to polymeric nanocarriers, liposomes, micelles, polyethylene glycol, poly(lactic-co-glycolic acid), dendrimers, gold and magnetic nanoparticles, quantum dots, silica nanoparticles, and carbon nanotubes. Further, this paper briefly focuses on several anticancer agents (paclitaxel, docetaxel, camptothecin, doxorubicin, daunorubicin, cisplatin, curcumin, and geldanamycin) and on the influence of their combination with nanoparticulate transporters to their properties such as cytotoxicity, short life time and/or solubility.

 

The Journal of Neuroscience
In vivo Stimulus-Induced Vasodilation Occurs without IP3 Receptor Activation and May Precede Astrocytic Calcium Increase

Nizar K., Uhlirova H., Tian P., Saisan P., Cheng Q., Reznichenko L., Weldy K., Steed T., Sridhar V., MacDonald C., Cui J., Gratiy S., Sakadzic S., Boas D., Beka T., Einevoll G., Chen J., Masliah E., Dale A., Silva G., Devor A.

Research Group:  Experimental Biophotonics
Research Programme: Advanced Nanotechnologies and Microtechnologies

Abstract
Calcium-dependent release of vasoactive gliotransmitters is widely assumed to trigger vasodilation associated with rapid increases in neuronal activity. Inconsistent with this hypothesis, intact stimulus-induced vasodilation was observed in inositol 1,4,5-triphosphate (IP3) type-2 receptor (R2) knock-out (KO) mice, in which the primary mechanism of astrocytic calcium increase—the release of calcium from intracellular stores following activation of an IP3-dependent pathway—is lacking. Further, our results in wild-type (WT) mice indicate that in vivo onset of astrocytic calcium increase in response to sensory stimulus could be considerably delayed relative to the simultaneously measured onset of arteriolar dilation. Delayed calcium increases in WT mice were observed in both astrocytic cell bodies and perivascular endfeet. Thus, astrocytes may not play a role in the initiation of blood flow response, at least not via calcium-dependent mechanisms. Moreover, an increase in astrocytic intracellular calcium was not required for normal vasodilation in the IP3R2-KO animals.

 

NANO LETTERS
Control and Near-Field Detection of Surface Plasmon Interference Patterns

Dvorak P., Neuman T., Brinek L., Samoril T., Kalousek R., Dub P., Varga P., Sikola T.

Research Group:  Fabrication and Characterisation of Nanostructures
Research Programme: Advanced Nanotechnologies and Microtechnologies

Abstract
The tailoring of electromagnetic near-field properties is the central task in the field of nanophotonics. In addition to 2D optics for optical nanocircuits, confined and enhanced electric fields are utilized in detection and sensing, photovoltaics, spatially localized spectroscopy (nanoimaging), as well as in nanolithography and nanomanipulation. For practical purposes, it is necessary to develop easy-to-use methods for controlling the electromagnetic near-field distribution. By imaging optical near-fields using a scanning near-field optical microscope, we demonstrate that surface plasmon polaritons propagating from slits along the metal− dielectric interface form tunable interference patterns. We present a simple way how to control the resulting interference patterns both by variation of the angle between two slits and, for a fixed slit geometry, by a proper combination of laser beam polarization and inhomogeneous far-field illumination of the structure. Thus the modulation period of interference patterns has become adjustable and new variable patterns consisting of stripelike and dotlike motifs have been achieved, respectively.

 

Journal of the European Ceramic Society
Two-stage master sintering curve applied to two-step sintering of oxide ceramics

Pouchlý V., Maca K., Shen Z.

Research Group:  Advanced Ceramic Materials
Research Programme: Advanced Materials

Abstract
Tetragonal (3 mol% Y2O3) and two cubic zirconia (8 mol% Y2O3) as well as alumina green bodies were used for the construction of the Master Sintering Curve (MSC) created from sets of constant-rate-of-heating (CRH) sintering experiments. The activation energies calculated according to the MSC theory were 770 kJ/mol for Al2O3, 1270 kJ/mol for t-ZrO2, 620 kJ/mol and 750 kJ/mol for c-ZrO2. These values were verified by an alternative approach based on an analysis of the densification rate in the intermediate sintering stage. The MSCs established from the Two-Step Sintering (TSS) experiments showed at high densities a significant deflection from those constructed from the CRH experiments. This deflection was explained by lower sintering activation energy in the closed porosity stage. A new two-stage MSC model was developed to reflect the change in sintering activation energy and to describe TSS. The efficiency of TSS of four materials under investigation was correlated with their activation energies during the final sintering stage.

 

IEEE Transactions on Industrial Electronics
AC Drive Observability Analysis

Vaclavek P., Blaha P., Herman I.

Research Group:  Cybernetics in Material Science
Research Programme: Advanced Materials

Abstract
AC induction motors and permanent magnet synchronous drives became very popular for motion control applications due to their simple and reliable construction. Sensorless drive control is required in many applications to reduce drive production costs. While many approaches to magnetic flux, rotor speed, or other quantities needed to control electrical machine were proposed, conditions under which these quantities can be estimated are not often sufficiently investigated. In this paper; induction machine and permanent-magnet-synchronous-machine drive state observability analysis is presented, together with conditions allowing reliable rotor speed and position estimation. A method based on the nonlinear dynamical system state observability theory is proposed, resulting in a unified approach to the ac drive observability analysis.

 

Journal of Alloys and Compounds
Beneficial effect of carbon on hydrogen desorption kinetics from Mg–Ni–In alloy

Cermak J., Kral L.

Research Programme: Advanced Materials
Research Group: Advanced Metallic Materials and Metal Based Composites

Abstract:
In the present paper, hydrogen desorption kinetics from hydrided Mg–Ni–In–C alloys was investigated. A chemical composition that substantially accelerates hydrogen desorption was found. It was observed that carbon improves the hydrogen desorption kinetics significantly. Its beneficial effect was found to be optimum close to the carbon concentration of about cC≈5 wt.%. With this composition, stored hydrogen can be desorbed readily at temperatures down to about 485 K, immediately after hydrogen charging. This can substantially shorten the hydrogen charging/discharging cycle of storage tanks using Mg–Ni-based alloys as hydrogen storage medium. For higher carbon concentrations, unwanted phases precipitated, likely resulting in deceleration of hydrogen desorption and lower hydrogen storage capacity.

 

Structure
Subunit Folds and Maturation Pathway of a dsRNA Virus Capsid

Němeček D., Boura E., Wu W., Cheng N., Plevka P., Qiao J., Mindich L., Heymann J.B., Hurley J.H., Steven A.C.

Research Group: CryoEM & Structural Virology
Research Programme: Structural Biology

Abstract
The cystovirus φ6 shares several distinct features with other double-stranded RNA (dsRNA) viruses, including the human pathogen, rotavirus: segmented genomes, nonequivalent packing of 120 subunits in its icosahedral capsid, and capsids as compartments for transcription and replication. φ6 assembles as a dodecahedral procapsid that undergoes major conformational changes as it matures into the spherical capsid. We determined the crystal structure of the capsid protein, P1, revealing a flattened trapezoid subunit with an α-helical fold. We also solved the procapsid with cryo-electron microscopy to comparable resolution. Fitting the crystal structure into the procapsid disclosed substantial conformational differences between the two P1 conformers. Maturation via two intermediate states involves remodeling on a similar scale, besides huge rigid-body rotations. The capsid structure and its stepwise maturation that is coupled to sequential packaging of three RNA segments sets the cystoviruses apart from other dsRNA viruses as a dynamic molecular machine.

 

ChemBioChem
Structural Study of the Partially Disordered Full-Length δ Subunit of RNA Polymerase from Bacillus subtilis

Papoušková V., Kadeřávek P., Otrusinová O., Rabatinová A., Šanderová H., Nováček J., Krásný L., Sklenář V., Žídek, L. 

Research Groups: Biomolecular NMR Spectroscopy
Research Programme: Structural Biology

Abstract
The partially disordered δ subunit of RNA polymerase was studied by various NMR techniques. The structure of the well-folded N-terminal domain was determined based on inter-proton distances in NOESY spectra. The obtained structural model was compared to the previously determined structure of a truncated construct (lacking the C-terminal domain). Only marginal differences were identified, thus indicating that the first structural model was not significantly compromised by the absence of the C-terminal domain. Various 15N relaxation experiments were employed to describe the flexibility of both domains. The relaxation data revealed that the C-terminal domain is more flexible, but its flexibility is not uniform. By using paramagnetic labels, transient contacts of the C-terminal tail with the N-terminal domain and with itself were identified. A propensity of the C-terminal domain to form β-type structures was obtained by chemical shift analysis. Comparison with the paramagnetic relaxation enhancement indicated a well-balanced interplay of repulsive and attractive electrostatic interactions governing the conformational behavior of the C-terminal domain. The results showed that the δ subunit consists of a well-ordered N-terminal domain and a flexible C-terminal domain that exhibits a complex hierarchy of partial ordering.

 

Journal of the American Chemical Society
Relative Stability of Different DNA Guanine Quadruplex Stem Topologies Derived Using Large-Scale Quantum-Chemical Computations

Šponer J., Mládek A., Špačková N., Cang X., Cheatham T.E., Grimme S.

Research Groups: Structure and Dynamics of Nucleic Acids
Research Programme: Structural Biology

Abstract
We provide theoretical predictions of the intrinsic stability of different arrangements of guanine quadruplex (G-DNA) stems. Most computational studies of nucleic acids have applied Molecular Mechanics (MM) approaches using simple pairwise-additive force fields. The principle limitation of such calculations is the highly approximate nature of the force fields. In this study, we for the first time apply accurate QM computations (DFT-D3 with large atomic orbital basis sets) to essentially complete DNA building blocks, seven different folds of the cation-stabilized twoquartet G-DNA stem, each having more than 250 atoms. The solvent effects are approximated by COSMO continuum solvent. We reveal sizable differences between MM and QM descriptions of relative energies of different G-DNA stems, which apparently reflect approximations of the DNA force field. Using the QM energy data, we propose correction to earlier free energy estimates of relative stabilities of different parallel, hybrid, and antiparallel G-stem folds based on classical simulations. The new energy ranking visibly improves the agreement between theory and experiment. We predict the 5′-anti-anti-3′ GpG dinucleotide step to be the most stable one, closely followed by the 5′-syn-anti-3′ step. The results are in good agreement with known experimental structures of 2-, 3-, and 4-quartet G-DNA stems. Besides providing specific results for G-DNA, our study highlights basic limitations of force field modeling of nucleic acids. Although QM computations have their own limitations, mainly the lack of conformational sampling and the approximate description of the solvent, they can substantially improve the quality of calculations currently relying exclusively on force fields.

 

Plant Physiology
Proteome analysis in Arabidopsis reveals shoot- and root-specific targets of cytokinin action and differential regulation of hormonal homeostasis

Zdarska M., Zatloukalova P., Benitez M., Sedo O., Potesil D., Novak O., Svacinova J., Pesek B., Malbeck J., Vasickova J., Zdrahal Z., Hejatko J.

Research Group: Functional Genomics and Proteomics of Plants & Core Facility – Genomics
Research Programme: Genomics and Proteomics of Plant Systems

Abstract
The plant hormones cytokinins (CKs) regulate multiple developmental and physiological processes in Arabidopsis (Arabidopsis thaliana). Responses to CKs vary in different organs and tissues (e.g. the response to CKs has been shown to be opposite in shoot and root samples). However, the tissue-specific targets of CKs and the mechanisms underlying such specificity remain largely unclear. Here, we show that the Arabidopsis proteome responds with strong tissue and time specificity to the aromatic CK 6-benzylaminopurine (BAP) and that fast posttranscriptional and/or posttranslational regulation of protein abundance is involved in the contrasting shoot and root proteome responses to BAP. We demonstrate that BAP predominantly regulates proteins involved in carbohydrate and energy metabolism in the shoot as well as protein synthesis and destination in the root. Furthermore, we found that BAP treatment affects endogenous hormonal homeostasis, again with strong tissue specificity. In the shoot, BAP up-regulates the abundance of proteins involved in abscisic acid (ABA) biosynthesis and the ABA response, whereas in the root, BAP rapidly and strongly up-regulates the majority of proteins in the ethylene biosynthetic pathway. This was further corroborated by direct measurements of hormone metabolites, showing that BAP increases ABA levels in the shoot and 1-aminocyclopropane-1-carboxylic acid, the rate-limiting precursor of ethylene biosynthesis, in the root. In support of the physiological importance of these findings, we identified the role of proteins mediating BAP-induced ethylene production, METHIONINE SYNTHASE1 and ACC OXIDASE2, in the early root growth response to BAP.

 

Lab on a Chip
Detection of electrochemiluminescence from floating metal platelets in suspension

Juskova P., Neuzil P., Manz A., Foret F.

Research Group: Smart Nanodevices & Bioanalytical Instrumentation
Research Programme: Advanced Nanotechnologies and Microtechnologies & Genomics and Proteomics of Plant Systems

Abstract
We present a generation of electrochemiluminescence (ECL) signal, based on square shaped gold electrodes with a size of 50 μm positioned inside a fused silica capillary. The ECL was generated using electric pulses with duration in the range from 100 ms to 5 s and an electrical field strength from 300 V cm-1 to 500 V cm-1. We have demonstrated that the electrochemical reaction with detectable optical output can be produced using freely moving and thus disposable electrodes.

 

Journal of Experimental Botany
Proteome and metabolome profiling of cytokinin action in Arabidopsis identifying both distinct and similar responses to cytokinin down- and up-regulation

Černý M., Kuklová A., Hoehenwarter W., Fragner L., Novák O., Rotková G., Jedelský P.L., Žáková K., Šmehilová M., Strnad M., Weckwerth W., Brzobohatý B.

Research Group: Developmental and Production Biology – Omics Approaches
Research Programme: Genomics and Proteomics of Plant Systems

Abstract
In plants, numerous developmental processes are controlled by cytokinin (CK) levels and their ratios to levels of other hormones. While molecular mechanisms underlying the regulatory roles of CKs have been intensely researched, proteomic and metabolomic responses to CK deficiency are unknown. Transgenic Arabidopsis seedlings carrying inducible barley cytokinin oxidase/dehydrogenase (CaMV35S>GR>HvCKX2) and agrobacterial isopentenyl transferase (CaMV35S>GR>ipt) constructs were profiled to elucidate proteome- and metabolome-wide responses to down- and up-regulation of CK levels, respectively. Proteome profiling identified >1100 proteins, 155 of which responded to HvCKX2 and/or ipt activation, mostly involved in growth, development, and/or hormone and light signalling. The metabolome profiling covered 79 metabolites, 33 of which responded to HvCKX2 and/or ipt activation, mostly amino acids, carbohydrates, and organic acids. Comparison of the data sets obtained from activated CaMV35S>GR>HvCKX2 and CaMV35S>GR>ipt plants revealed unexpectedly extensive overlaps. Integration of the proteomic and metabolomics data sets revealed: (i) novel components of molecular circuits involved in CK action (e.g. ribosomal proteins); (ii) previously unrecognized links to redox regulation and stress hormone signalling networks; and (iii) CK content markers. The striking overlaps in profiles observed in CK-deficient and CK-overproducing seedlings might explain surprising previously reported similarities between plants with down- and up-regulated CK levels.


Nature Methods
MiTCR: software for T-cell receptor sequencing data analysis

Bolotin D.A., Shugay M., Mamedov I.Z., Putintseva E.V., Turchaninova M.A., Zvyagin I.V., Britanova O.V., Chudakov D.M.

Research Group: Adaptive Immunity Group
Research Programme: Molecular Medicine

Abstract
Matrix metalloproteinases (MMPs) are involved in many physiological and pathological processes. Numerous MMPs assays were developed for both clinical and research purposes, but far more attention was turned to understanding their biological functions. In this review, enzymatic, immunochemical and fluorimetric methods as well as in vivo imaging methods are discussed. Moreover, we aimed our attention on additional methods that are now subject to investigation, such as phage display, Multiple-Enzyme/Multiple-Reagent Assay System (MEMRAS) and activity based profiling.

 

Haematologica
Multiple productive immunoglobulin heavy chain gene rearrangements in chronic lymphocytic leukemia are mostly derived from independent clones

Plevova K.,Skuhrova Francova H., Burckova K., Brychtova Y., Doubek M., Pavlova S., Malcikova J., Mayer J., Tichy B., Pospisilova S.

Research Group: Medical Genomics
Research Programme: Molecular Medicine

Abstract
In chronic lymphocytic leukemia, usually a monoclonal disease, multiple productive immunoglobulin heavy chain gene rearrangements are identified sporadically. Prognostication of such cases based on immunoglobulin heavy variable gene mutational status can be problematic, especially if the different rearrangements have discordant mutational status. To gain insight into the possible biological mechanisms underlying origin of the multiple rearrangements, we performed a comprehensive immunogenetic and immunophenotypic characterization of 31 cases with the multiple rearrangements identified in a cohort of 1147 chronic lymphocytic leukemia patients. For the majority of cases (25/31), we provide evidence for the co-existence of at least two B lymphocyte clones with chronic lymphocytic leukemia phenotype. We also identified clonal drifts in serial samples, likely driven by selection forces. More specifically, higher immunoglobulin variable gene identity to germline and longer complementarity determining region 3 were preferred in persistent or newly appearing clones, a phenomenon more pronounced in patients with stereotyped B cell receptors. Finally, we report that other factors, such as TP53 gene defects and therapy administration, influence clonal selection. Our findings are relevant to clonal evolution in the context of antigen stimulation and transition of monoclonal B-cell lymphocytosis to chronic lymphocytic leukemia.

 

Leukemia
Distinct patterns of novel gene mutations in poor-prognostic stereotyped subsets of chronic lymphocytic leukemia: the case of SF3B1 and subset #2

Strefford J.C., Sutton L.-A., Baliakas P., Agathangelidis A., Malcikova J., Plevova K., Scarfo L., Davis Z., Stalika E., Cortese D., Cahill N., Pedersen L.B., di Celle P.F., Tzenou T., Geisler C., Panagiotidis P., Langerak A.W., Chiorazzi N., Pospisilova S., Oscier D., Davi F., Belessi C., Mansouri L., Ghia P., Stamatopoulos K., Rosenquist R.

Research Group: Medical Genomics
Research Programme: Molecular Medicine

Abstract
Recent studies have revealed recurrent mutations of the NOTCH1, SF3B1 and BIRC3 genes in chronic lymphocytic leukemia (CLL), especially among aggressive, chemorefractory cases. Nevertheless, it is currently unknown whether their presence may differ in subsets of patients carrying stereotyped B-cell receptors and also exhibiting distinct prognoses. Here, we analysed the mutation status of NOTCH1, SF3B1 and BIRC3 in three subsets with particularly poor prognosis, that is, subset #1, #2 and #8, aiming to explore links between genetic aberrations and immune signalling. A remarkably higher frequency of SF3B1 mutations was revealed in subset #2 (44%) versus subset #1 and #8 (4.6% and 0%, respectively; P<0.001). In contrast, the frequency of NOTCH1 mutations in subset #2 was only 8%, lower than the frequency observed in either subset #1 or #8 (19% and 14%, respectively; P=0.04 for subset #1 versus #2). No associations were found for BIRC3 mutations that overall were rare. The apparent non-random association of certain mutations with stereotyped CLL subsets alludes to subset-biased acquisition of genomic aberrations, perhaps consistent with particular antigen/antibody interactions. These novel findings assist in unravelling specific mechanisms underlying clinical aggressiveness in poor-prognostic stereotyped subsets, with far-reaching implications for understanding their clonal evolution and implementing biologically oriented therapy.

 

Schizophrenia Research
A detailed analysis of the effect of repetitive transcranial magnetic stimulation on negative symptoms of schizophrenia: A double-blind trial
 
Prikryl R., Ustohal L., Prikrylova Kucerova H., Kasparek T., Venclikova S., Vrzalova M., Ceskova E.
 
Research Group: Applied Neuroscience
Research Programme: Brain and Mind Research
 
Abstract
Objective: The aim of the study was to assess the effect of rTMS not only on the general severity of negative schizophrenia symptoms, but also particularly on their individual domains, such as affective flattening or blunting, alogia, avolition or apathy, anhedonia, and impaired attention. 
Methods: Forty schizophrenic male patients on stable antipsychotic medication with prominent negative symptoms were included in the study. They were divided into two groups: 23 were treated with active and 17 with placebo rTMS. Both treatments were similar, but placebo rTMS was administered using a purposebuilt sham coil. Stimulation was applied to the left dorsolateral prefrontal cortex (DLPFC). The stimulation frequency was 10 Hz; stimulation intensity was 110% of the individual motor threshold intensity. Each patient received 15 rTMS sessions on 15 consecutive working days (five working days “on” and two weekend days “off” design). Each daily session consisted of 20 applications of 10-second duration with 30-second intervals between sequences. The patients and raters were blind to condition of stimulation treatment.
Results: The active rTMS led to a statistically significantly higher reduction of the Scale for the Assessment of Negative Symptoms (SANS) total score and of all domains of negative symptoms of schizophrenia. After Bonferroni adjustments for multiple testing, the statistical significance disappeared in alogia only.
Conclusion: High-frequency rTMS stimulation over the left DLPFC at a high stimulation intensity with a sufficient number of applied stimulating pulses may represent an efficient augmentation of antipsychotics in alleviating the negative symptoms of schizophrenia.

NeuroImage
Superior temporal sulcus and social cognition in dangerous drivers

Zelinkova J., Shaw D.J., Marecek R., Mikl M., Urbanek T., Peterkova L., Zamecnik P., Brazdil M.

Research Programme: Brain and Mind Research
Research Group: Multi-modal and Functional Neuroimaging & Behavioural and Social Neuroscience

Abstract
Understanding the neural systems underpinning social cognition is a primary focus of contemporary social neuroscience. Using functional magnetic resonance imaging (fMRI), the present study asked if brain activity reflecting socio-cognitive processes differs between individuals according to their social behavior; namely, between a group of drivers with frequent traffic offenses and a group with none. Socio-cognitive processing was elicited by employing videos froma traffic awareness campaign, consisting of reckless and anti-social driving behavior ending in tragic consequences, and control videos with analogous driving themes but without such catastrophic endings. We investigated whether relative increases in brain function during the observation of these campaign stimuli compared with control videos differed between these two groups. To develop the results of our previous study we focused our analyses on superior temporal sulcus/gyrus (STS/STG). This revealed a bigger increase in brain activity within this region during the campaign stimuli in safe compared with dangerous drivers. Furthermore, by thematically coding drivers' verbal descriptions of the stimuli, we also demonstrate differences in STS reactivity according to drivers' scores on two indices of socio-cognitive processing: subjects' perceived consequences of actors' actions, and their affective evaluation of the clips. Our results demonstrate the influence of social behavior and socio-cognitive processing on STS reactivity to social stimuli, developing considerably our understanding of the role of this region in social cognition.

Journal of Neuroinflammation
Bilateral elevation of interleukin-6 protein and mRNA in both lumbar and cervical dorsal root ganglia following unilateral chronic compression injury of the sciatic nerve
 
Dubovy P., Brazda V., Klusakova I., Hradilova-Svizenska I.
Research Group: Cellular and Molecular Neurobiology
Research Programme: Brain and Mind Research
 
Abstract
Background: Current research implicates interleukin (IL)-6 as a key component of the nervous-system response to injury with various effects. Methods: We used unilateral chronic constriction injury (CCI) of rat sciatic nerve as a model for neuropathic pain. Immunofluorescence, ELISA, western blotting and in situ hybridization were used to investigate bilateral changes in IL-6 protein and mRNA in both lumbar (L4-L5) and cervical (C7-C8) dorsal root ganglia (DRG) following CCI. The operated (CCI) and sham-operated (sham) rats were assessed after 1, 3, 7, and 14 days. Withdrawal thresholds for mechanical hyperalgesia and latencies for thermal hyperalgesia were measured in both ipsilateral and contralateral hind and fore paws.
Results: The ipsilateral hind paws of all CCI rats displayed a decreased threshold of mechanical hyperalgesia and withdrawal latency of thermal hyperalgesia, while the contralateral hind and fore paws of both sides exhibited no significant changes in mechanical or thermal sensitivity. No significant behavioral changes were found in the hind and fore paws on either side of the sham rats, except for thermal hypersensitivity, which was present bilaterally at 3 days. Unilateral CCI of the sciatic nerve induced a bilateral increase in IL-6 immunostaining in the neuronal bodies and satellite glial cells (SGC) surrounding neurons of both lumbar and cervical DRG, compared with those of naïve control rats. This bilateral increase in IL-6 protein levels was confirmed by ELISA and western blotting. More intense staining for IL-6 mRNA was detected in lumbar and cervical DRG from both sides of
rats following CCI. The DRG removed from sham rats displayed a similar pattern of staining for IL-6 protein and mRNA as found in naive DRG, but there was a higher staining intensity in SGC.
Conclusions: Bilateral elevation of IL-6 protein and mRNA is not limited to DRG homonymous to the injured nerve, but also extended to DRG that are heteronymous to the injured nerve. The results for IL-6 suggest that the neuroinflammatory reaction of DRG to nerve injury is propagated alongside the neuroaxis from the lumbar to the remote cervical segments. This is probably related to conditioning of cervical DRG neurons to injury.
 

FEMS Microbiology  Ecology
Extended spectrum beta-lactamase and fluoroquinolone resistance genes and plasmids among Escherichia coli isolates from zoo animals, Czech Republic

Dobiasova, H. ; Dolejska, M.; Jamborova, I.; Brhelova, E.; Blazkova, L.; Papousek, I. ; Kozlova, M.; Klimes, J.; Cizek, A. ; Literak, I.

Research Group: Molecular Bacteriology
Research Programme: Molecular Veterinary Medicine

Abstract
Commensal Escherichia coli isolates from healthy zoo animals kept in Ostrava Zoological Garden, Czech Republic, were investigated to evaluate the dissemination of extended-spectrum beta-lactamase (ESBL) and plasmid-mediated quinolone resistance (PMQR) genes. A total of 160 faecal samples of various animal species were inoculated onto MacConkey agar with cefotaxime (2 mg L-1) or ciprofloxacin (0.05 mg L-1) to obtain ESBL- or PMQR-positive E. coli isolates. Clonality of E. coli isolates was investigated by multilocus sequence typing and pulsed-field gel electrophoresis. Plasmids carrying ESBL or PMQR genes were typed by PCR-based replicon typing, plasmid multilocus sequence typing and restriction fragment length polymorphism. Forty-nine (71%, n = 69) cefotaxime-resistant and 15 (16%, n = 94) ciprofloxacin-resistant E. coli isolates harboured ESBL or PMQR genes. Isolates were assigned to 18 sequence types (ST) and 20 clusters according to their macrorestriction patterns by pulsed-field gel electrophoresis. The genes blaCTX-M-1 and qnrS1 were detected on highly related IncI1 plasmids assigned to clonal complex 3 (ST3, ST38) and on non-related IncN plasmids of ST1 and ST3, respectively. The gene qnrS1 was located on related IncX1 plasmids. Dissemination of antibiotic resistance is associated with spreading of particular E. coli clones and plasmids of specific incompatibility groups among various animal species.

 

Journal of General Virology
Preliminary epitope mapping of Torque teno sus virus 1 and 2 putative capsid protein and serological detection of infection in pigs

Jarosova V., Celer V.

Research Group: Molecular Virology
Research Programme: Molecular Veterinary Medicine

Abstract
The aim of this work is to identify antigenic regions within the ORF1 protein of Torque teno susvirus 1 (TTSuV1) and Torque teno virus sus 2(TTSuV2) that could be used as antigens to detect virus-specific antibodies following infection in pigs. Protein sequences of TTSuV ORF1 genes were analysed to predict linear antigenic epitopes. Synthesized peptides were analysed for serological reactivity with swine sera. Such an antigenic region was identified at the C terminus of the ORF1 protein of both viruses and showed serological reactivity with 78% (TTSuV1) and 88% (TTSuV2) of swine sera. An ELISA with an immunodominant peptide as antigen was used to examine the sera of piglets, aged 4–20 weeks, and adults. Results indicated that TTSuV1- and TTSuV2-specific antibodies were detectable at 4 weeks. Antibody titres increased from week 10 and peaked at week 20. A relatively high antibody titre persisted to adulthood.

 
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