OPPOSITE PSYCHOLOGICAL STATES ASSOCIATED WITH RUNNING IN SOLITUDE AND STREET RACE PARTICIPATION

Running is associated with positive acute psychological effects. However, the context of running, which has received little empirical attention to date, could be expected to mediate the related subjective experiences. In this in-situ (real life) cross-sectional study, we compared the subjective psychological states before and after running in solitude and in street race running. Seventy males (n = 31 running alone and n = 39 running in a street race) completed the short version of the Positive Affect and Negative Affect Schedule (PANAS), rated their core affect (conceptualized as the momentarily perceived overall physical and psychological feeling state), and their appraised satisfaction with the completed run. Although the two groups did not differ in satisfaction with their run, except for negative affect which did not change in either of the groups, the results revealed opposite trends in psychological experiences in all measures. Positive affect, mental-, and physical core affect increased after running in solitude while they all decreased after the street race. The current results suggest that peak affective experience occurs after the run when people plan and perform their run alone, while the comparable top experience occurs before the run, likely due to the excitement of participation and social interaction, during street race running. The current work sheds light on the strong impact of the running situation on the acute psychological states associated running.  Article visualizations

Drug and dye binding induced folding of the intrinsically disordered antimicrobial peptide CM15

The rapid increase of antimicrobial resistance against conventional antibiotics has resulted in a significant focus on the use of peptides as antimicrobial agents. Understanding the structure and function relationships of these compounds is thus highly important, however, their in vivo actions are a complex issue, including interactions with small molecule agents. Here we report the folding inducing capability of some pharmaceutical substances and synthetic dyes on the intrinsically disordered (ID) cationic antimicrobial peptide CM15 (KWKLFKKIGAVLKVL). By employing circular dichroism (CD) spectroscopy, it is shown that some therapeutic drugs (suramin, pamoic acid, cromolyn) and polysulfonated dyes (Congo red, trypan blue) trigger the disorder-to-order conformational transition of CM15. The cooperative binding of 2-4 acidic molecules per peptide chain provokes its folding in a concentration dependent manner. Secondary structure analysis indicated the sharp and moderate rise of the [small alpha]-helical and [small beta]-sheet content, respectively. According to semi-empirical quantum chemical calculations, these organic molecules may induce folding by forming multiple salt-bridges with lysine residues from both N- and C-terminals as well as from the middle of the CM15 sequence. Due to the mutual neutralization of the positive and negative charges, the water solubility of the resulting complexes decreases which favours their aggregation as detected by dynamic light scattering measurements. Our findings suggest that small molecules can dramatically affect the structure of antimicrobial peptides, which may potentially alter, either enhancing or attenuating, their efficiency. It is proposed that CM15 or similar ID peptides could be useful for preliminary screening of folding inducer effect of anionic drugs and biomolecules. The data presented herein may stimulate further studies on the structural and functional impacts of related compounds on ID peptides

ÉLŐ SEJTEK ADHÉZIÓJÁNAK MONITOROZÁSA OPTIKAI BIOSZENZOROKKAL

Egy újszerű, nagy áteresztőképességű optikai bioszenzor, az Epic BenchTop (BT) segítségével azt vizsgáltuk, hogyan függ a rákos sejtek kiterülésének kinetikája a integrin-ligandum RGD-motívumok átlagos felületi sűrűségétől (νRGD) [1]. A biologiallag inaktív PLL-g-PEG kopolimer, és az RGD-funkcionalizált PLL-g-PEG-RGD vegyített oldatait használtuk a felületi bevonatok elkészítéséhez, νRGD-t négy nagységrenden keresztül hangoltuk. Modell sejtvonalként az erősen adherens HeLa rákos sejtvonalat használtuk, a kiterülés kinetikáját az Epic BT bioszenzorral egyedülálló minőségben rögzítettük. A kapott görbéket kinetikai elemzésnek vetettük alá: az adatokat a logisztikus egyenlettel illesztettük meg, hogy meghatározhassuk, hogyan függ a felületi ligandsúrúségtől a sejtkiterülés sebességi állandója (r), illetve a maximális bioszenzor jel (∆λmax). Eredményeink szerint r nem függ νRGD-től, átlagos értéke 0.062±0.004 min-1. Ezzel szemben ∆λmax, ami egyenesen arányos a kiterült sejt átlagos kontaktterületével, növekedett, ahogy νRGD-t növeltük. Ezt a viselkedést egy egyszerű monovalens kötési reakcióval leírva meghatároztuk a PLL-PEG-RGD molekula RGD-motívuma és a sejt integrinjei közötti kölcsönhatás két dimenziós disszociációs állandóját, mely 1753 μm-2-nek adódott. Ebből egyszerűen származtatható a kapcsolat 3D-s disszociációs állandója, melynek értéke ~30 μM. Mindezen eredményehez egyedülló módon teljesen noninvazív és jelölésmentes kísérleteken keresztül jutottunk

The mapping of linear B-cell epitope regions in desmoglein 1 and 3 proteins : Recognition of immobilized peptides by pemphigus patients’ serum autoantibodies

Desmosomal transmembrane glycoproteins desmoglein 1and desmoglein 3 are targets of life-threatening autoimmune blistering disorders such as Pemphigus vulgaris (PV) and Pemphigus foliaceus(PF). In these diseases pemphigus autoantibodies are produced against Dsg1 and Dsg3 proteins. The autoantibodies bind to these transmembrane elements leading to a loss of desmosomal cell-cell adhesion, and clinically, to the presence of blisters and erosions. Identification, characterization and detailed analysis of the binding sites of autoantibodies have an outstanding importance in understanding theimmunopathology of the disease and also in the design of novel diagnostics. Here we describe the localization of the B-cell epitope regions of Dsg1 and Dsg3 proteins extracellular parts recognized by IgG-type serum autoantibodies of patients with PV andPF. In our study overlapping pentadecapeptides were synthesized on hydroxypropylmethacrylate pins based on the results of in silicopredictions. To detect the interaction between theserum autoantibodies and the immobilized synthetic peptides, modified ELISA (Enzyme Linked Immunosorbent Assay) was performed with pin-attached peptides testing the serum samples of ten patients and four healthy donors. We identified five possible epitope regions (aa86-110, aa196-220, aa226-250, aa326-340, and aa486-520) within the Dsg1 protein sequence and four possible epitope regions (aa64-78, aa330-344, aa375-399, aa446-460) within the Dsg3 protein sequence using these methods. Our data showed that serum autoantibodies of patients, previously identified as Dsg1 and Dsg3 positive, are able to recognize continuous linear epitope regions of both Dsg1 and Dsg3 proteins using pin-bound overlapping peptides in modified ELISAs

Drug conjugation induced modulation of structural and membrane interaction features of cationic cell-permeable peptides

Cell-penetrating peptides might have great potential for enhancing the therapeutic effect of drug molecules against such dangerous pathogens as Mycobacterium tuberculosis (Mtb), which causes a major health problem worldwide. A set of cationic cell-penetration peptides with various hydrophobicity were selected and synthesized as drug carrier of isoniazid (INH), a first-line antibacterial agent against tuberculosis. Molecular interactions between the peptides and their INH-conjugates with cell-membrane-forming lipid layers composed of DPPC and mycolic acid (a characteristic component of Mtb cell wall) were evaluated, using the Langmuir balance technique. Secondary structure of the INH conjugates was analyzed and compared to that of the native peptides by circular dichroism spectroscopic experiments performed in aqueous and membrane mimetic environment. A correlation was found between the conjugation induced conformational and membrane affinity changes of the INH–peptide conjugates. The degree and mode of interaction were also characterized by AFM imaging of penetrated lipid layers. In vitro biological evaluation was performed with Penetratin and Transportan conjugates. Results showed similar internalization rate into EBC-1 human squamous cell carcinoma, but markedly different subcellular localization and activity on intracellular Mtb

Membrane affinity and fluorescent labelling: comparative study of monolayer interaction, cellular uptake and cytotoxicity profile of carboxyfluorescein-conjugated cationic peptides

Fluorescent labelling is a common approach to reveal the molecular details of cellular uptake, internalisation, transport, distribution processes in biological systems. The conjugation with a fluorescent moiety might affect relevant physico-chemical and in vitro transport properties of the bioactive component. A representative set of seven cationic peptides-including cell-penetrating peptides as well as antimicrobial peptides and synthetic derivatives-was selected for our comparative study. Membrane affinity of the peptides and their 5(6)-carboxyfluorescein (Cf) derivatives was determined quantitatively and compared applying Langmuir monolayer of zwitterionic (DPPC) and negatively charged (DPPC + DPPG) lipids as cell membrane models. The interaction with neutral lipid layer is mainly governed by the overall hydrophobicity of the molecule which is remarkably increased by Cf-conjugation for the most hydrophobic Magainin, Melittin and Transportan. A significantly enhanced membrane affinity was detected in negatively charged lipid model monolayer for all of the peptides since the combination of electrostatic and hydrophobic interaction is active in that case. The Cf-conjugation improved the penetration ability of Penetratin and Dhvar4 suggesting that both the highly charged character (Z/n) and the increased hydrophobicity by Cf-conjugation present important contribution to membrane interaction. This effect might also responsible for the observed high in vitro internalisation rate of Penetratin and Dhvar4, while according to in vitro studies they did not cause damage of cell membrane. From the experiments with the given seven cationic peptides, it can be concluded that the Cf-conjugation alters the degree of membrane interaction of such peptides which are moderately hydrophobic and highly charged

Tailoring Uptake Efficacy of HSV-1 gD Tailoring Uptake Efficacy of Hsv-1 GD Derived Carrier Peptides

Regions of the Herpes simplex virus-1 (HSV-1) glycoprotein D (gD) were chosen to design carrier peptides based on the known tertiary structure of the virus entry receptor complexes. These complexes consist of the following: HSV-1 gD–nectin-1 and HSV-1 gD–herpesvirus entry mediator (HVEM). Three sets of peptides were synthesised with sequences covering the (i) N-terminal HVEM- and nectin-1 binding region -5–42, (ii) the 181–216 medium region containing nectin-1 binding sequences and (iii) the C-terminal nectin-1 binding region 214–255. The carrier candidates were prepared with acetylated and 5(6)-carboxyfluorescein labelled N-termini. The peptides were chemically characterised and their conformational features in solution were also determined. In vitro internalisation profile and intracellular localisation were evaluated on SH-SY5Y neuroblastoma cells. Peptide originated from the C-terminal region 224–247 of the HSV-1 gD showed remarkable internalisation compared to the other peptides with low to moderate entry. Electronic circular dichroism secondary structure studies of the peptides revealed that the most effectively internalised peptides exhibit high helical propensity at increasing TFE concentrations. We proved that oligopeptides derived from the nectin-1 binding region are promising candidates—with possibility of Lys237Arg and/or Trp241Phe substitutions—for side-reaction free conjugation of bioactive compounds—drugs or gene therapy agents—as cargos

Autoimmun betegségekben szerepet játszó új, diagnosztikus epitópok, valamint Fc receptorhoz kötődő és effektor funkciót kiváltó IgG Fc peptidkimérák azonosítása, szintézise és funkcionális jellemzése = Identification, synthesis and functional characterisation of new, diagnostic epitopes in autoimmune diseases, and of IgG Fc peptide chimeras binding to Fc receptor and triggering effector function

Autoimmun hólyagos bőrbetegségekre jellemző B- és T-sejt epitóppeptideket szintetizáltunk szilárdfázisú peptidszintézissel, Fmoc/tBu stratégiával. A peptideket tisztítottuk, azonosítottuk. Autoantigén B-sejt epitópok azonosítására módosított ELISA módszerrel tűhegyen korábban szintetizált átlapoló desmoglein 1 és 3 peptideken hat pemphigus foliaceusban (PF), egy pemphigus vulgarisban (PV) szenvedő beteg és egészséges donor szérumát vizsgáltuk. Megállapítottuk, hogy az egészséges kontrollhoz képest a betegszérumok desmoglein specifikus autoellenanyag-tartalma általában kicsit magasabb, ezen belül öt olyan epitóprégiót találtunk, amelyeket hét beteg közül legalább öt szérum-ellenanyagai szignifikánsan erősebben ismertek fel. Megkezdtük ezen epitóprégiók szintézisét egyedi peptidek formájában, Cys-nel meghosszabbítva, későbbi, makromolekulás hordozóhoz vagy funkcionalizált ELISA lemezhez való konjugálás céljából. Desmoglein 3 autoantigén T-sejt epitóp vizsgálata pemphigus vulgarisban Két PV-ben szenvedő és két egészséges donor perifériás vér monomorfonukleáris sejtjeit (PBMC) izoláltuk, majd egy desmoglein 3 T-sejt epitóppeptid rövidített, átlapoló változataival inkubáltuk. A sejtek felülúszóiból szendvics ELISÁ-val mutattuk ki a termelődött interferon-gammát. A betegek PBMC-i szignifikánsan nagyobb mértékben termeltek IFN-gamma citokint, mint az egészséges donorok sejtjei. Leghatékonyabban a T-sejt epitóppeptid középső szakasza stimulálta a sejteket. | B- and T-cell epitope peptides characteristic for autoimmune bullous skin diseases were prepared by solid phase synthesis, Fmoc/tBu strategy. The peptides were purified and characterised. To determine autoantigenic B-cell epitopes, modified ELISA was performed on formerly synthesised pin-bound desmoglein 1 and 3 peptides with the sera of six pemphigus foliaceus (PF), a pemphigus vulgaris (PV) patients and healthy donor. The serum autoantibody binding to the peptides was generally somewhat higher in case of the patients, and we could identify five epitope regions which were recognised by at least 5 out of 7 patients significantly more strongly by patients’ sera than by those of the control. We have started the synthesis of the selected regions as individual peptides, elongated by Cys, to facilitate future conjugation to ELISA plates or macromolecular carriers. To study a desmoglein 3 autoantigen T-cell epitope in PV, we have isolated the PBMC of two PV patients and two healthy donors. The cells were incubated with truncated overlapping derivatives of T-cell epitope peptide. Interferon gamma cytokine produced by the cells was determined from the supernatants by sandwich ELISA. PBMC of patients produce

Amino acid based hydrogels with enzymatically degradable cross-links

The synthesis of a chemically cross-linked polymer hydrogel consisting exclusively of amino acids is described in this paper. A natural amino acid, aspartic acid was polymerized to polysuccinimide which was cross-linked by a tetrapeptide sequence designed for proteolytic degradation, and then the corresponding poly(aspartic acid) hydrogel was obtained by alkaline hydrolysis. The hydrogel dissolved in the presence of trypsin. According to in vitro cellular assays, the degradation products of the hydrogel cross-linked with the peptide were non-cytotoxic and non-cytostatic. The sustained release of an encapsulated macromolecular model drug, FITC-dextran, was triggered by the degradation of the hydrogel induced by trypsin. These results suggest the potential application of the trypsin-responsive hydrogel for drug delivery in the small intestine