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Current knowledge about the molecular properties of the crustacean ion channels is rather limited even if crustaceans have been widely used as a model in neuroscience. We cloned for the first time two different potassium channel genes from the freshwater crayfish Astacus leptodactylus (Eschscholtz, 1823), one of the genes functionally expressed in the Xenopus oocytes. The open-reading frames of the genes were 1,203 and 3,447 bp, respectively. The nucleic acid sequence of the genes and associated proteins were similar to those of a typical potassium channel. BLAST analyses indicated that one of the cloned genes had a substantial similarity to an inward-rectifier potassium channel whereas the other gene was similar to a high-conductance-KCa type potassium channel reported in related species. Transmembrane topology and three-dimensional structure of the coded proteins were calculated and functional regions of the channel proteins responsible for ion selectivity, voltage sensing, gating, and calcium binding were identified. One of the cloned channel genes has been expressed in the Xenopus oocytes. Analysis of the expressed potassium currents confirmed that the cloned gene was coding a typical Kir-type potassium channel with ATP sensitivity.

Desmin is a muscle-specific intermediate filament protein that has fundamental role in muscle structure and force transmission. Whereas human desmin protein is encoded by a single gene, two desmin paralogs (desma and desmb) exist in zebrafish. Desma and desmb show differential spatiotemporal expression during zebrafish embryonic and larval development, being similarly expressed in skeletal muscle until hatching, after which expression of desmb shifts to gut smooth muscle. We generated knockout (KO) mutant lines carrying loss-of-function mutations for each gene by using CRISPR/Cas9. Mutants are viable and fertile, and lack obvious skeletal muscle, heart or intestinal defects. In contrast to morphants, knockout of each gene did not cause any overt muscular phenotype, but did alter calcium flux in myofibres. These results point to a possible compensation mechanism in these mutant lines generated by targeting nonsense mutations to the first coding exon.

Aim: to examine the implantation of chitosan channels stuffed with mesenchyme-originated stem/progenitor cells (MSPCs) derived from adult rats in a spinal cord transection model. The level of axonal regeneration, the effect of chitosan channels on the survival of MSPCs, and the functional recovery results were also evaluated. Material and methods: chitosan channels stuffed with MSPCs were implanted at the level of T8 in a transected rat spinal cord. MSPCs were harvested from the pelvic bone marrow of adult rats, and the MSPC-chitosan channel group was compared with three control groups. The axonal regeneration capacity, the effect of chitosan channels on the survival of MSPCs, and the functional recovery results were compared among four groups. The survival of MSPCs was evaluated using histopathological techniques and electron microscopy, axonal regeneration/germination was evaluated by confocal microscopy, and locomotor function was assessed for 4 weeks using the Basso, Beattie, and Bresnahan locomotor score. Results: the MSPC-chitosan channel group exhibited enhanced survival of transplanted MSPCs compared with MSPCs transplanted directly into the lesion cavity, although no significant difference was detected in locomotor function between the treatment and control groups. The MSPC-chitosan channel group demonstrated thicker myelination of axons than the other groups. Conclusion: chitosan channels promoted the survival of transplanted MSPCs and created a tissue bridge after complete spinal cord transection. They also induced axonal regeneration and germination. No significant improvement in functional recovery was found between the groups., Hacettepe University

Background.This study compared the effect of smear layer on the penetration depth and push-out bond strength of various root canal sealers. Methods.A total of 90 extracted human mandibular premolars were assigned into 2 groups: smear layer preserved and smear layer removed. Then the roots were further divided into 3 subgroups according to the sealer tested: AH 26, BioRoot RCS and MTA Plus. Obturation was performed with gutta-percha and the relevant sealer was mixed with 0.1% rhodamine B. Three 1-mm-thick slices were obtained from the mid-third area of each root. Two slices were selected for the push-out test and the remaining slice was used to calculate the dentinal tubule penetration depth and percentage. Results.The retention of MTA Plus and BioRoot RCS was higher than that of AH 26 when the smear layer was preserved (P

WOS: 000431796100001, PubMed: 29748557, Amelogenin (AMG) is a cell adhesion molecule that has an important role in the mineralization of enamel and regulates events during dental development and root formation. The purpose of the present study was to investigate the effects of recombinant human AMG (rhAMG) on mineralized tissue-associated genes in cementoblasts. Immortalized mouse cementoblasts (OCCM-30) were treated with different concentrations (0.1, 1, 10, 100, 1000, 10,000, 100,000 ng . mL(-1)) of recombinant human AMG (rhAMG) and analyzed for proliferation, mineralization and mRNA expression of bone sialoprotein (BSP), osteocalcin (OCN), collagen type I (COL I), osteopontin (OPN), runt-related transcription factor 2 (Runx2), cementum attachment protein (CAP), and alkaline phosphatase (ALP) genes using quantitative RT-PCR. The dose response of rhAMG was evaluated using a real-time cell analyzer. Total RNA was isolated on day 3, and cell mineralization was assessed using von Kossa staining on day 8. COL I, OPN and lysosomal-associated membrane protein-1 (LAMP-1), which is a cell surface binding site for amelogenin, were evaluated using immunocytochemistry. F-actin bundles were imaged using confocal microscopy. rhAMG at a concentration of 100,000 ng . mL(-1) increased cell proliferation after 72 h compared to the other concentrations and the untreated control group. rhAMG (100,000 ng . mL(-1)) upregulated BSP and OCN mRNA expression levels eightfold and fivefold, respectively. rhAMG at a concentration of 100,000 ng . mL(-1) remarkably enhanced LAMP-1 staining in cementoblasts. Increased numbers of mineralized nodules were observed at concentrations of 10,000 and 100,000 ng . mL(-1) rhAMG. The present data suggest that rhAMG is a potent regulator of gene expression in cementoblasts and support the potential application of rhAMG in therapies aimed at fast regeneration of damaged periodontal tissue., TUBITAK (Turkey)Turkiye Bilimsel ve Teknolojik Arastirma Kurumu (TUBITAK) [SBAG108S265]; BMBF (Germany)Federal Ministry of Education & Research (BMBF) [TUR08/09], This study was supported by TUBITAK SBAG108S265 (Turkey) and BMBF TUR08/09 (Germany). We thank Johan Svensson and Leif Bulow for rhAMG production and characterization studies from Department of Pure and Applied Biochemistry, Center for Chemistry and Chemical Engineering Lund University, Lund, Sweden for rhAMG synthesis.

In order to improve the immunogenicity of the highly purified vaccine antigens, addition of an adjuvant to formulation, without affecting the safety of the vaccine, has been the key aim of the vaccine formulators. In recent years, adjuvants which are composed of a delivery system and immunopotentiators have been preferred to induce potent immune responses. In this study, we have combined Salmonella Typhi porins and chitosan to develop a new adjuvant system to enhance the immunogenicity of the highly purified antigens. Cationic gels, microparticle (1.69 +/- 0.01 mu m) and nanoparticles (337.7 +/- 1.7 mu m) based on chitosan were prepared with high loading efficiency of porins. Cellular uptake was examined by confocal laser scanning microscopy, and the macrophage activation was investigated by measuring the surface marker as well as the cytokine release in vitro in J774A.1 macrophage murine cells. Porins alone were not taken up by the macrophage cells whereas in combination with chitosan a significant uptake was obtained. Porins-chitosan combination systems were found to induce CD80, CD86 and MHC-II expressions at different levels by different formulations depending on the particle size. Similarly, TNF-alpha and IL-6 levels were found to increase with porins-chitosan combination. Our results demonstrated that combination of porins with chitosan as a particulate system exerts enhanced adjuvant effect, suggesting a promising adjuvant system for subunit vaccines with combined immunostimulating activity.

Familial Mediterranean fever (FMF), the most common of the systemic autoinflammatory disorders, is caused by mutations in the MEFV (Mediterranean Fever) gene, which encodes the protein pyrin. Neutrophils, one of the major components during inflammation, are the main cell type that expresses pyrin. In response to an inflammatory stimulus, neutrophils migration to their main active site. To date, several pyrin-interacting proteins have been demonstrated to co-localise with the cytoskeletal protein actin, which is important in the process of neutrophil migration and raises the question of whether pyrin plays a role in the actin cytoskeletal network during inflammatory cell migration. In this study, we examined the possible role of pyrin during inflammatory cell migration in neutrophils. We generated a cell migration assay with neutrophils and primary neutrophils from patients. We also knocked down pyrin expression using siRNA and then performed cell migration assay. We showed co-localisation of pyrin and F-actin at the leading edge during inflammatory cell migration. In pyrin knocked down cells, we identified a significant decrease in neutrophil migration. In addition, we demonstrated a dramatic increase in migration in the neutrophils of FMF patients compared with a healthy control group. These data together provide new insight into the cellular function of pyrin and demonstrate an important link between pyrin and polymerising actin in the process of inflammatory cell migration.

Objective: Relevancy of the reference genes have not been investigated in crayfish tissue samples. Various genes have been proposed for establishing a reference level to analyze qPCR results obtained in different tissue samples. However, majority of those genes have been investigated solely in mammalian species. There is no report related to the relevancy and the efficiency of those common reference genes in crayfish tissue samples yet. The primary aim of the present work is to compare the expression level of a set of reference genes in various crayfish tissue samples to establish a reliable reference gene for quantitative expression level comparisons.Methods: cDNA copies were synthesized by RT-PCR from equalised total RNA samples of nine different tissue samples. qPCR were performed on cDNA samples by using four different primer pairs specific for β-actin, Peptidylprolyl Isomerase B, Sarcoplasmic Calcium Binding Protein and 18S ribosomal RNA genes.Results: qActin2 amplified gene fragments for β-actin in all samples. Cycle time of the amplification reaction differed substantially between the investigated tissue types. A similar difference was observed between the melting curves of the products. The primer pair PPIB amplified gene fragments for Peptidylprolyl Isomerase B only in heart, muscle and intestine samples but not in the rest of the tissue samples. Further, both amplification and melting curves for heart, muscle and intestine were different. qSCP amplified some gene fragments for Sarcoplasmic Reticulum Binding Protein only in muscle, heart and ganglion samples. The 18S_1 amplified fragments for 18S. Amplification curves the samples superimposed on each other. Melting curves of the products were very similar. Variance of the obtained data was the lowest for 18S ribosomal RNA.Conclusion: In refer to present results; it is conceivable to propose that among all the genes investigated in the present study, expression level of ribosomal 18S RNA is the most homogeneously distributed one within the body parts of the crayfish. Further, it is minimally affected by extreme biological changes such as molting. Thus, 18S Ribosomal RNA would be suggested as a reference gene to be used when investigating tissue specific expression level of a gene in the crayfish.

The present study was designed to determine whether the properties of local Ca2+release and its related regulatory mechanisms might provide insight into the role of sex differences in heart functions of control and streptozotocin-induced diabetic adult rats. Left ventricular developed pressure, the rates of pressure development and decay (±dP/d t), basal intracellular Ca2+level ([Ca2+]i), and spatiotemporal parameters of [Ca2+]itransients were found to be similar in male and female control rats. However, spatiotemporal parameters of Ca2+sparks in cardiomyocytes isolated from control females were significantly larger and slower than those in control males. Diabetes reduced left ventricular developed pressure to a lower extent in females than in males, and the diabetes-induced depressions in both +dP/d t and −dP/d t were less in females than in males. Diabetes elicited a smaller reduction in the amplitude of [Ca2+]itransients in females than in males, a smaller reduction in sarcoplasmic reticulum-Ca2+load, and less increase in basal [Ca2+]i. Similarly, the elementary Ca2+events and their control proteins were clearly different in both sexes, and these differences were more marked in diabetes. Diabetes-induced depression of the Ca2+spark amplitude was significantly less in females than in matched males. Levels of cardiac ryanodine receptors (RyR2) and FK506-binding protein 12.6 in control females were significantly higher than those shown in control males. Diabetes induced less RyR2 phosphorylation and FK506-binding protein 12.6 unbinding in females. Moreover, total and free sulfhydryl groups were significantly less reduced, and PKC levels were less increased, in diabetic females than in diabetic males. The present data related to local Ca2+release and its related proteins describe some of the mechanisms that may underlie sex-related differences accounting for females to have less frequent development of cardiac diseases.

Objectives The aim of this study was to compare the push-out bond strength and dentinal tubule penetration of root canal sealers used with coated core materials and conventional gutta-percha. Materials and Methods A total of 72 single-rooted human mandibular incisors were instrumented with NiTi rotary files with irrigation of 2.5% NaOCl. The smear layer was removed with 17% ethylenediaminetetraacetic acid (EDTA). Specimens were assigned into four groups according to the obturation system: Group 1, EndoRez (Ultradent Product Inc.); Group 2, Activ GP (Brasseler); Group 3, SmartSeal (DFRP Ltd. Villa Farm); Group 4, AH 26 (Dentsply de Trey)/gutta-percha (GP). For push-out bond strength measurement, two horizontal slices were obtained from each specimen (n = 20). To compare dentinal tubule penetration, remaining 32 roots assigned to 4 groups as above were obturated with 0.1% Rhodamine B labeled sealers. One horizontal slice was obtained from the middle third of each specimen (n = 8) and scanned under confocal laser scanning electron microscope. Tubule penetration area, depth, and percentage were measured. Kruskall-Wallis test was used for statistical analysis. Results EndoRez showed significantly lower push-out bond strength than the others (p < 0.05). No significant difference was found amongst the groups in terms of percentage of sealer penetration. SmartSeal showed the least penetration than the others (p < 0.05). Conclusions The bond strength and sealer penetration of resin-and glass ionomer-based sealers used with coated core was not superior to resin-based sealer used with conventional GP. Dentinal tubule penetration has limited effect on bond strength. The use of conventional GP with sealer seems to be sufficient in terms of push-out bond strength.

Voltage-gated sodium channel genes and associated proteins have been cloned and studied in many mammalian and invertebrate species. However, there is no data available about the sodium channel gene(s) in the crayfish, although the animal has frequently been used as a model to investigate various aspects of neural cellular and circuit function. In the present work, by using RNA extracts from crayfish abdominal ganglia samples, the complete open reading frame of a putative sodium channel gene has firstly been cloned and molecular properties of the associated peptide have been analyzed. The open reading frame of the gene has a length of 5793 bp that encodes for the synthesis of a peptide, with 1930 amino acids, that is 82 % similar to the a-peptide of a sodium channel in a neighboring species, Cancer borealis. The transmembrane topology analysis of the crayfish peptide indicated a pattern of four folding domains with several transmembrane segments, as observed in other known voltage-gated sodium channels. Upon analysis of the obtained sequence, functional regions of the putative sodium channel responsible for the selectivity filter, inactivation gate, voltage sensor, and phosphorylation have been predicted. The expression level of the putative sodium channel gene, as defined by a qPCR method, was measured and found to be the highest in nervous tissue.

The defects identified in the mechanical activity of the hearts from type 1 diabetic animals include alteration of Ca2+ signaling via changes in critical processes that regulate intracellular Ca2+ concentration. These defects result partially from a dysfunction of cardiac ryanodine receptor calcium release channel (RyR2). The present study was designed to determine whether the properties of the Ca2+ sparks might provide insight into the role of RyR2 in the altered Ca2+ signaling in cardiomyocytes from diabetic animals when they were analyzed together with Ca2+ transients. Basal Ca2+ level as well as Ca2+-spark frequency of cardiomyoctes isolated from 5-week streptozotocin (STZ)-induced diabetic rats significantly increased with respect to aged-matched control rats. Ca2+ transients exhibited significantly reduced amplitude and prolonged time courses as well as depressed Ca2+ loading of sarcoplasmic reticulum in diabetic rats. Spatio-temporal properties of the Ca2+ sparks in cardiomyocytes isolated from diabetic rats were also significantly altered to being almost parallel to the changes of Ca2+ transients. In addition, RyR2 from diabetic rat hearts were hyperphosphorylated and protein levels of both RyR2 and FKBP12.6 depleted. These data show that STZ-induced diabetic rat hearts exhibit altered local Ca2+ signaling with increased basal Ca2+ level.

We performed genome-wide homozygosity mapping and mapped a novel myopathic phenotype to chromosomal region 1q25 in a consanguineous family with three affected individuals manifesting proximal and distal weakness and atrophy, rigid spine and contractures of the proximal and distal interphalangeal hand joints. Additionally, cardiomyopathy and respiratory involvement were noted. DNA sequencing of torsinA-interacting protein 1 (TOR1AIP1) gene encoding lamina-associated polypeptide 1B (LAP1B), showed a homozygous c.186delG mutation that causes a frameshift resulting in a premature stop codon (p.E62fsTer25). We observed that expression of LAP1B was absent in the patient skeletal muscle fibres. Ultrastructural examination showed intact sarcomeric organization but alterations of the nuclear envelope including nuclear fragmentation, chromatin bleb formation and naked chromatin. LAP1B is a type-2 integral membrane protein localized in the inner nuclear membrane that binds to both A- and B-type lamins, and is involved in the regulation of torsinA ATPase. Interestingly, luminal domain-like LAP1 (LULL1)-an endoplasmic reticulum-localized partner of torsinA-was overexpressed in the patient's muscle in the absence of LAP1B. Therefore, the findings suggest that LAP1 and LULL1 might have a compensatory effect on each other. This study expands the spectrum of genes associated with nuclear envelopathies and highlights the critical function for LAP 1B in striated muscle. (C) 2014 Elsevier B.V. All rights reserved.

Background Autosomal recessive limb girdle muscular dystrophy (LGMD2) is a heterogeneous group of myopathies characterised by progressive muscle weakness involving proximal muscles of the shoulder and pelvic girdles including at least 17 different genetic entities. Additional loci have yet to be identified as there are families which are unlinked to any of the known loci. Here we have investigated a consanguineous family with LGMD2 with two affected individuals in order to identify the causative gene defect. Methods and results We performed genome wide homozygosity mapping and mapped the LGMD2 phenotype to chromosome 2q35–q36.3. DNA sequence analysis of the highly relevant candidate gene DES revealed a homozygous splice site mutation c.1289-2A>G in the two affected family members. Immunofluorescent staining and western blot analysis showed that the expression and the cytoskeletal network formation of mutant desmin were well preserved in skeletal muscle fibres. Unlike autosomal dominant desminopathies, ultrastructural alterations such as disruption of myofibrillar organisation, formation of myofibrillar degradation products and dislocation/ aggregation of membranous organelles were not present. This novel splice site mutation results in addition of 16 amino acids within the tail domain of desmin, which has been suggested to interact with lamin B protein. We also detected a specific disruption of desmin-lamin B interaction in the skeletal muscle of the patient by confocal laser scanning microscopy. Conclusions Our study reveals that autosomal recessive mutations in DES cause LGMD2 phenotype without features of myofibrillar myopathy.

The purpose of this study was to investigate adhesion, proliferation and type I collagen (COL I) mRNA expression of gingival fibroblasts on different membranes used in periodontal applications. Collagen (C), acellular dermal matrix (ADM) and polylactic acid; polyglycolic acid; lactide/glycolide copolymer (PLGA) biodegradable membranes were combined with gingival fibroblasts in culture and incubated for 48 h. Cell adhesion was examined with scanning electron and confocal microscopy. MTT assay was used to measure proliferation. COL I mRNA expression was assessed using quantitative-polymerase chain reaction (QPCR). The PLGA group exhibited the lowest cell survival on day 5 and 10, and lowest cell proliferation on days 5, 10 and 14. While cell proliferation was similar in C and ADM groups, the C membrane showed a slightly greater increase in viable cells to day 10. Confocal and scanning electron microscopy confirmed the results of proliferation and MTT assays. The highest COL I mRNA expression was noted in the PLGA membrane group when compared to the C (p

Aim. The purpose of this study was to investigate proliferation, morphology, mineralization and mRNA expressions of mineralized tissue associated proteins of PDL cells on smooth (S), sandblasted small-grit (SSG), sandblasted large-grit (SLG) and sodium titanate (NaTi) coated titanium alloys, in vitro. Methods and materials: PDL cells were cultured with DMEM media containing 10% FBS on the S, SSG, SLG and NaTi titanium surfaces. PDL cell proliferation, mineralization and immunohistochemistry experiments for Bone Sialoprotein (BSP) were performed. The morphology of the PDL cells was examined using confocal and scanning electron microscopy (SEM). Gene expression profiles of cells were evaluated using a quantitative-polymerase chain reaction (Q-PCR) for type I collagen (COL I), Osteocalcin (OCN), osteopontin (OPN) and Runt-related transcription factor-2 (Runx2) on days 7 and 14. Results. Proliferation results on days 6 and 10 were similar in groups, while those of day 13 revealed a decrease in the NaTi group when compared to the S group. NaTi surface induced BSP mRNA expression which was correlated with mineralization tests and BSP immunostaining results. Increased Runx2 mRNA expression was also noted in the NaTi surface when compared to other surfaces. Conclusions. This study considers the NaTi surface as a potential alternative to SSG and SLG surfaces. This surface might provide a promising environment for PDL ligament-anchored implants.

The effects of antidromic potential spread were investigated in the stretch receptor neurons of the crayfish. Current and potential responses to conductance changes were recorded in the dynamic clamp condition and compared to those obtained by using some conventional clamp methods and a compartmental neuron model. An analogue circuit was used for dynamic calculation of the injected receptor current as a function of the membrane potential and the given conductance change. Alternatively, receptor current responses to a mechanical stimulus were recorded and compared when the cell was voltage clamped to a previously recorded impulse wave form and the resting potential, respectively. Under dynamic clamp, the receptor current had an oscillating waveform which contrasts with the conventional recordings. Frequency, amplitude and sign of the oscillations were dependent on the applied conductance level, reversal potential and electrotonic attenuation. Mean current amplitude and frequency of the evoked impulse responses were smaller under dynamic clamp, especially for large conductance increases. However, firing frequency was larger if plotted against the mean current response. Recorded responses were similar to those calculated in the model. It was not possible to evoke any adaptation in the slowly adapting neuron by using the dynamic clamp. Evoked potential change served as a self limiting response, preventing the depolarization block. However, impulse duration was significantly shorter in the rapidly adapting neuron when the dynamic clamp was used. It was concluded that, in the stretch receptor neurons during a conductance increase, antidromic potential spread modulates the receptor responses and contributes to adaptation.

1. The transducer properties of the rapidly adapting stretch receptor neurone of the crayfish (Pacifastacus leniusculus) were studied using a two-microelectrode voltage clamp technique. 2. The impulse response to ramp-and-hold extensions of the receptor muscle typically consisted of a high frequency burst followed by cessation of impulses within a relatively short time depending on the amplitude of extension. The type of adaptation was consistent with earlier studies. The stimulus-response relationship for the impulse frequency was non-linear and had a slope in a log-log plot of 2.9. 3. When impulse generation was blocked by tetrodotoxin (TTX), (block of Na+ channels) the receptor potential was extension dependent and similar to that found in the slowly adapting receptor. For small extensions there was an initial peak followed by a fall to a steady potential level. For large extensions the potential response during the ramp phase consisted of a peak followed by a constant potential level lasting to the end of the ramp. When the extension changed to the hold phase the potential fell towards a steady state. The relation between extension and amplitude of receptor potential was non-linear and saturated at -40 to -30 mV (extensions > 15% of zero length, lo). 4. When potassium channels were blocked by TEA (50 mM) and 4-aminopyridine (4-AP, 5 mM) (and Na+ channels blocked by TTX) the shape of the generator potential become less complex with an increased amplitude for large extensions. 5. When the receptor neurone was voltage clamped at the resting potential, extension of the receptor muscle produced an inwardly directed receptor current, the stretch-induced current (SIC). The response consisted of a fast transient phase which decayed towards a steady state. The SIC peak amplitude was dependent on extension in a sigmoidal fashion and saturated at 190 nA (extensions > 25% of lo). The slope of the steepest part of the stimulus-response relation (between 10 and 20% extension) was 4.7 +/- 0.25 (mean +/- S.E.M.) in a log-log plot. 6. The peak amplitude of the SIC increased with increasing extension speed (ramp steepness), the relation between the slope of the ramp and current amplitude being a first order (hyperbolic) function. The amplitude of the receptor current was voltage dependent and had a reversal potential of +16.2 +/- 1.8 mV (mean +/- S.E.M., 32 cells). From the reversal potential the permeability ratio, PNa/PK, of the transducer permeability system was calculated to be 1.5. The I-V curve of SIC was non-linear.(ABSTRACT TRUNCATED AT 400 WORDS)

Bovine herpesvirus 1 (BHV-1), is a major pathogen of cattle which causes serious infections, including infectious bovine rhinotracheitis (IBR)/infectious pustular vulvovaginitis (IPV). At present, BHV-1 is still a serious threat to animal health and productivity in Turkey, hence to develop a more efficient and economical vaccine system against BHV-1 is certainly an important necessity. A mucosal vaccination strategy would provide both mucosal and systemic immune responses to protect disease progression and transmission. However, vaccination through mucosal membranes requires adjuvants/delivery systems in order to enhance the immunogenicity of the antigens. Chitosan, which is a biodegradable, biocompatible and bioadhesive natural polysaccharide, has been shown to be promising both as a delivery system and an adjuvant for mucosal vaccination. In this study, microparticles with appropriate size (10μm), positive surface charge and high loading efficiency (∼95%) were prepared for mucosal delivery of BHV-1, using various types of chitosan with different molecular weight and solubility. Particles were shown to be taken up by the cells, mostly around the nucleus, whereas with aggregates which were bigger in size were adsorbed at the surface. Furthermore, gel formulations with a suitable viscosity which would provide easy application and remain on the mucosa for extended period of time were also developed with a high zeta potential indicating a stable system. Both the BHV-1 loaded microparticle and gel formulations were shown to maintain cell viability and antigen integrity. Chitosan-based formulations are suggested as promising adjuvant/delivery systems for mucosal immunization against BHV-1.

Purali, Nuhan and Bo Rydqvist. Action potential and sodium current in the slowly and rapidly adapting stretch receptor neurons of the crayfish ( Astacus astacus). J. Neurophysiol. 80: 2121–2132, 1998. Action potentials (APs) and sodium current from the slowly and the rapidly adapting stretch receptor neurons in the crayfish ( Astacus astacus) were recorded with a two microelectrode voltage- and current-clamp technique. In the rapidly adapting neuron the APs had a duration of 3.2 ± 0.2 ms (means ± SE) and an amplitude of 55.2 ± 1.5 mV. In the slowly adapting receptor neuron APs had a duration of 4.1 ± 0.2 ms and an amplitude 79.9 ± 2.0 mV. APs in the rapidly adapting neuron had a larger amplitude if they were recorded from the axon. In the rapidly adapting neuron adaptation of the impulse response was prolonged by hyperpolarization or by exposure to scorpion venom. Also, sinusoidal current stimulation added to the current steps prevented impulse adaptation. Block of the potassium currents in the slowly adapting neuron resulted in a rapid adaptation of the impulse response. The maximum sodium current amplitude was 313 ± 15 nA in slowly adapting neuron and 267 ± 11 nA in the rapidly adapting neuron. The current-voltage relationship showed a hump most marked in the slowly adapting neuron and abolished when a depolarizing prepulse was given. In the rapidly adapting neuron the inactivation starts at a more negative potential ( Eh = −45 mV) and is faster compared with the slowly adapting neuron ( Eh = −41 mV). The crude scorpion venom of Leiurus quinquestriatus (ScVLq) shifted h ∞ curve toward more positive potentials and slowed down the rate of inactivation. The results indicate the possible presence of more than one Na+ channel population and that the relative density and the spatial distribution is different in the slowly and rapidly adapting neuron. The difference contributes to the adaptive properties of the two receptor neurons.

MEFV (MEditerranien FeVer) gene mutations cause Familial Mediterranean Fever (FMF). This gene encodes a protein termed as Pyrin, which appears to play an important role in the inflammatory pathways. It is far characterized that Pyrin, which is expressed in neutrophils, interacts with PSTPIP1 and actin proteins. In previous studies PSTPIP1 has been shown to interact with cell migration proteins and actin polymerization is a main force driving neutrophil migration. Therefore, we hypothesized that Pyrin can play role in cell migration through the interaction with actin and PSTPIP1.

mRNA expressions related to osteogenic differentiation of MC3T3-E1 cells on electro-polished smooth (S), sandblasted small-grit (SSG) and sandblasted large-grit (SLG) surfaces of titanium alloys were investigated in vitro. Gene expression profiles of cells were evaluated using the RT2 Profiler PCR microarray on day 7. Mineralizing tissue-associated proteins, differentiation factors and extracellular matrix enzymes mRNA expressions were measured using Q-PCR. SLG surface upregulated 23 genes over twofolds and downregulated 3 genes when compared to the S surface. In comparison to the SSG surface, at least a twofold increase in 25 genes was observed in the SLG surface. BSP, OCN, OPN, COL I and ALP mRNA expressions increased in the SLG group when compared to the S and the SSG groups. BMP-2, BMP-6 and TGF-beta mRNA expressions increased in both the SSG and the SLG surfaces. MMP-2 and MMP-9 mRNA expressions increased as the surface roughness increased. This study demonstrated that surface roughness of titanium implants has a significant effect on cellular behavior and SLG surface apparently increased gene expressions related to osteogenesis when compared to the S and the SSG surfaces.

Familial Mediterranean fever (FMF) is one of the most common autoinflammatory disorders and is characterized by episodic attacks of fever, along with inflammation. FMF pathogenesis is associated with various mutations in the MEFV gene, which encodes pyrin. Pyrin is expressed predominantly in neutrophils that have an important role in the innate immune response. Several proteins related with actin machinery have been identified as pyrin-interacting proteins in our in vitro cell migration models. Thus, in this study, we hypothesized that pyrin may have a key role in neutrophil migration during inflammation and decided to do functional analysis on pyrin silenced cell lines and primary neutrophils.