Effect of immune system imagery on secretory IgA

This study was an investigation of the effects of physiologically-oriented mental imagery on immune functioning. College students with normal medical histories were randomly selected to one of three groups. Subjects in Group 1 participated in short educational training on the production of secretory immunoglobulin A. They were then tested on salivary IgA, skin temperature, and the Profile of Mood States (POMS) before and after listening to a 17-minute tape of imagery instructions with specially composed background "entrainment" music designed to enhance imagery. Subjects in Group 2 (placebo controls) listened to the same music but received nor formal training on the immune system. Group 3 acted as a control and subjects were tested before and after 17 minutes of no activity. Treatment groups listened to their tapes at home on a bi-daily basis for six weeks. All groups were again tested at Weeks 3 and 6. Secretory IgA was analyzed using standard radial immunodiffusion techniques. Repeated measures analyses of variance with planned orthogonal contrasts were used to evaluate the data. Significant overall increases (p less than 0.05) were found between pre- and posttests for all three trials. Groups 1 and 2 combined (treatment groups) yielded significantly greater increases in sIgA over Group 3 (control) for all three trials. Group 1 (imagery) was significantly higher than Group 2 (music) in antibody production for Trials 2 and 3. Symptomatology, recorded by subjects at Weeks 3 and 6, was significantly lower for three symptoms (rapid heartbeat, breathing difficulty, and jaw clenching), favoring both treatment groups over the control group.     

Biofeedback,relaxation training,and music: Homeostasis for coping with stress

This study compared the efficacy of five relaxation training procedures, four of which employed EMG auditory feedback: (1) biofeedback only (BF), (2) autogenic training phrases (ATP), (3) music (MU), (4) autogenic training phrases and music (ATP & MU), and (5) a control group, in developing self-regulation of a cultivated low arousal state as a countermeasure to tensed muscular reaction to stressful imagery. Twenty subjects established a pre- and posttraining frontalis region EMG biofeedback baseline measurement. Sixteen subjects were assigned at random to the 25-minute taped relaxation training procedure. After eight training sessions (4 weeks), MU and ATP & MU groups achieved highly significant differences when compared with the control group. The ATP & MU group attained the lowest postbaseline arousal level measured by the EMG. EMG as a physiological measure for transfer of training functioned well in detecting the psychophysiological affect of stressful imagery.This report is based on a thesis submitted in partial fulfillment of the requirements for the Master of Arts in Psychology degree by the author. The author extends his gratitude to Dr. Theodore Steiner, Dr. Paul Eskildsen, and Dr. Frank Hovell, who served on the committee, and to Rosemary Kolentus, for her help with this article.     

Expression of the alpha subunit of 7S nerve growth factor in the mouse submandibular gland

-NGF is an inactive serine protease that is associated in the mouse submandibular gland with a closely related serine protease, -NGF, and the neurotrophic factor, -NGF. The heterogeneity of purified -NGF has been examined by DEAE-cellulose chromatography and SDS polyacrylamide gel electrophoresis. A possible explanation for the observed heterogeneity is presented. Antibodies have been prepared against -NGF and purified by affinity chromatography so that they do not cross-react with -NGF. This antibody preparation recognizes two very similar proteins in male mouse submandibular gland RNA-directed cell-free translation mixtures. The expression of only one of these forms is regulated by testosterone. Oligonucleotide probes specific for each of the three NGF subunits have been prepared and used for Northern blot analysis of RNA from the mouse submandibular gland. The three subunits were found to be coordinately expressed and each were 30-fold more abundant in male than in female glands.Abbreviations used NGF nerve growth factor - -, -, and -NGF -, -, and -subunits of mouse 7S NGF - PBS phosphate buffered saline - DTT dithiothreitol - PPO 2,5-Diphenyloxazole - DMSO dimethylsulfoxide - HEPES N-2-Hydroxyethylpiperazine-N-2-ethanesulfonic acid - SSC 0.15M NaCl, 15 mM sodium citrate Supported by USPHS research grant NS19964. This paper is respectfully dedicated to Profs. Eric M. Shooter and Silvio Varon in recognition of their many contributions to our understanding of the structure and function of nerve growth factor.     

Zur Frage der Beziehungen zwischen Substantia granulofilamentosa und Krinom

Zusammenfassung In dieser ersten von insgesamt drei Studien über die Beziehungen zwischen Substantia granulofilamentosa und Krinom wurden am Blut Phenylhydrazin-vergifteter Meerschweinchen 57 kationische Farbstoffe verschiedener chemischer Struktur und ausgewählte basische Stoffe ohne Farbstoffcharakter auf ihre Eignung geprüft, die Substantia granulofilamentosa auszufällen und gegebenenfalls hinsichtlich der Art der Ausfällung individuelle, vor allem morphologische Merkmale erkennen zu lassen.Von den 57 Farbstoffen waren 36, vorwiegend die hauptsächlich untersuchten Derivate von Heterocyclen des Anthracens, zur Darstellung der Substantia granulofilamentosa geeignet. Zu den erstmals für die Rc-Darstellung als brauchbar erkannten Farbstoffen gehören Methylengrün, Toluylenblau, Pseudoisocyanin, Neutralviolett und Amethystviolett.Auch in einigen Alkaloiden in hoher Konzentration, insbesondere mit Chinin, gelingt die Präzipitation einer netzförmigen, basophilen Struktur, die allerdings nicht die gleiche Dichte und Farbstoffaffinität aufweist, wie die von guten Farbstoffen ausgefällte Substantia granulofilamentosa.Bei intensiv mit Phenylhydrazin behandelten Meerschweinchen wurde Spontanentmischung der basophilen Substanz der Rc bereits in den Kontrollpräparaten beobachtet.Die Mehrzahl der positiven Farbstoffe gehört ihrer chemischen Struktur nach zu den Heterocyclen des Anthracens. — Ursachen negativer Resultate der Rc-Markierung werden besprochen.Quantitative Beziehungen zwischen Änderungen der Farbstoffkonzentration und den Rc-Zahlen wurden vergleichend am Beispiel äquimolarer Lösungen von Acridinorange, 9-Aminoacridin, Neutralrot und Chinin untersucht. In dieser Reihenfolge geht die Wirksamkeit der vier Substanzen bei abnehmender Konzentration zurück.Nach Einwirkung konzentrierter (1100) geeigneter Farbstoffe auf Blutzellen zeigt die Struktur der in Rc ausgefällten Substantia granulofilamentosa gewisse farbstoffabhängige Eigenarten.Einige dieser Eigentümlichkeiten in der Darstellung der Substantia granulofilamentosa werden beschrieben. Typisch z. B. für Derivate des Acridins ist das Bild der dichten zentralen Zusammenballung der basophilen Substanz im May-Grünwald-Giemsa-nachgefärbten Präparat.Mit einigen positiven Farbstoffen (Konzentration 12000) wird in Rc das Phänomen der Vakuolenbildung, welches direkte Beziehungen zur Krinombildung hat, hervorgerufen. Die meisten Farbstoffe werden bei dieser Konzentration in Vakuolen konzentriert. Pyronin ist nur zur Darstellung der Substantia granulofilamentosa geeignet.In Froscherythrocyten vermögen Farbstoffe der Gentianaviolett-Gruppe die Substantia granulofilamentosa nicht auszufällen.Aus Vergleichsgründen wird die gleiche Farbstoffauswahl auch hinsichtlich ihrer Affinität zu den chemisch andersartig aufgebauten Heinz-Körpern untersucht. Die Heinz-Körper werden besonders intensiv mit Farbstoffen der Gentianaviolett-Gruppe und mit Oxazin angefärbt. Sie zeigen ziemlich intensiv Primärfluoreszenz, nach Einwirkung verschiedener Fluorochrome Sekundärfluoreszenz.Gesichtspunkte der Beziehungen zwischen chemischer Struktur der untersuchten Farbstoffe und ihren Wirkungen auf Rc werden besprochen.

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Summary This is the first of three studies on the relationship between Substantia granulofilamentosa and crinoma. Using the blood of phenylhydrazine-poisened guinea pigs, 57 cationic dyes of various chemical structures and selected basic substances of nondye character, are tested for their suitability to precipitate the substantia granulofilamentosa. Attention is paid to the various types of precipitates in order to find out whether they show individual morphologic features. Out of 57 dyes, 36—most of which are derivatives of heterocyclic anthracene — are suitable for the demonstration of the substantia granulofilamentosa. The group of dyes, which for the time ever have been used successfully for the demonstration of reticulocytes, includes: methylene green, toluylene blue, pseudo-isocyanine, neutral violet, and amethyst violet.After treatment with certain alkaloids in high concentrations (especially quinine) a reticular basophil structure is precipitated. It shows, however, not the same density and affinity for dyes as the precipitates, obtained with good dyes. In guinea pigs, treated with high doses of phenylhydrazine, a spontaneous segregation of the basophil substance of the reticulocytes is already observed in the controls. According to their chemical structures, the majority of posiive dyes belong to the heterocyclic anthracenes. — The reasons for negative results in the demonstration of reticulocytes are discussed. Quantitative relationships between dye concentration and number of reticulocytes are tested with equimolar solutions of acridine-orange, 9-amino acridine, neutral red, and quinine. The efficacy of these four substances decreases with decreasing concentration in the order given above. If blood cells are treated with a concentrated solution (1100) of suitable dyes, certain dye-dependent peculiarities are observed in the precipitated substantia granulofilamentosa, some of which are described in detail. A typical picture after treatment with acridine derivatives is — for example — a dense central conglomerate of the basophil substance, found after additional staining with May-Grünwald-Giemsa. Some positive dyes (concentration 12000) cause the formation of vacuoles in reticulocytes, which stands in direct relation to the formation of the crinoma. If used at this concentration most of the dyes are concentrated in vacuoles. Pyronine is suitable only for the demonstration of the substantia granulofilamentosa.Dyes that belong to the gentian violet group, are not suitable for the precipitation of the substantia granulofilamentosa in frog erythrocytes.For comparative reasons the same group of dyes is tested for its affinity to the Heinz bodies, which chemically are very different from the substantia granulofilamentosa. Heinz bodies are intensely stained with dyes belonging to the gentian violet group and by oxazine dyes. A rather intensive primary fluorescence is observed in Heinz bodies; if several fluorochromes are used, they show secondary fluorescence. The relation between the chemical structure of the dyes used in this study and their effect on reticulocytes is discussed.

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In der Arbeit werden die folgenden Abkürzungen verwendet FS Farbstoff(e) - HK Heinz-Körper - MGG May-Grünewald-Giemsa Färbung - Rc Reticulocyt(en) - RNP Ribonucleoproteide - DNS Desoxyribonucleinsäure - SGF Substantia granulofilamentosa     

Effects of Ginkgo Biloba Extract Constituents on Glycine-Activated Strychnine-Sensitive Receptors in Hippocampal Pyramidal Neurons of the Rat

Active compounds of Ginkgo biloba extract (ginkgolides A, B, C, and J) were tested on the main ligand-operated conductances in rat neurons using a patch-clamp technique in the whole-cell configuration and a concentration-clamp technique. It was found that all ginkgolides reduced glycine-activated currents in concentration- and use-dependent manners, whereas they did not affect other tested ligand-gated receptors (NMDA- and GABA-activated ones). The IC₅₀ values calculated from dose-response curves were as follows: 1.97, 0.273, 0.267, and 2.0 M for ginkgolides A, B, C, and J, respectively (200 M Gly ).The Hill coefficient in all cases was close to 1.0, which indicates a single site of the drug binding to the glycine-activated receptor.     

Pharmacological Treatment with Annexin A1 Reduces Atherosclerotic Plaque Burden in LDLR-/- Mice on Western Type Diet

Objective

To investigate therapeutic effects of annexin A1 (anxA1) on atherogenesis in LDLR^-/- mice.

Methods

Human recombinant annexin A1 (hr-anxA1) was produced by a prokaryotic expression system, purified and analysed on phosphatidylserine (PS) binding and formyl peptide receptor (FPR) activation. Biodistribution of ^99mTechnetium-hr-anxA1 was determined in C57Bl/6J mice. 12 Weeks old LDLR^-/- mice were fed a Western Type Diet (WTD) during 6 weeks (Group I) or 12 weeks (Group P). Mice received hr-anxA1 (1 mg/kg) or vehicle by intraperitoneal injection 3 times per week for a period of 6 weeks starting at start of WTD (Group I) or 6 weeks after start of WTD (Group P). Total aortic plaque burden and phenotype were analyzed using immunohistochemistry.

Results

Hr-anxA1 bound PS in Ca²⁺-dependent manner and activated FPR2/ALX. It inhibited rolling and adherence of neutrophils but not monocytes on activated endothelial cells. Half lives of circulating ^99mTc-hr-anxA1 were <10 minutes and approximately 6 hours for intravenously (IV) and intraperitoneally (IP) administered hr-anxA1, respectively. Pharmacological treatment with hr-anxA1 had no significant effect on initiation of plaque formation (-33%; P = 0.21)(Group I) but significantly attenuated progression of existing plaques of aortic arch and subclavian artery (plaque size -50%, P = 0.005; necrotic core size -76% P = 0.015, hr-anxA1 vs vehicle) (Group P).

Conclusion

Hr-anxA1 may offer pharmacological means to treat chronic atherogenesis by reducing FPR-2 dependent neutrophil rolling and adhesion to activated endothelial cells and by reducing total plaque inflammation.     

Fabrication of Micro-tissues using Modules of Collagen Gel Containing Cells

This protocol describes the fabrication of a type of micro-tissues called modules. The module approach generates uniform, scalable and vascularized tissues. The modules can be made of collagen as well as other gelable or crosslinkable materials. They are approximately 2 mm in length and 0.7 mm in diameter upon fabrication but shrink in size with embedded cells or when the modules are coated with endothelial cells. The modules individually are small enough that the embedded cells are within the diffusion limit of oxygen and other nutrients but modules can be packed together to form larger tissues that are perfusable. These tissues are modular in construction because different cell types can be embedded in or coated on the modules before they are packed together to form complex tissues. There are three main steps to making the modules: (1) neutralizing the collagen and embedding cells in it, (2) gelling the collagen in the tube and cutting the modules and (3) coating the modules with endothelial cells.Download video file.^{(58M, mov)}     

Cellular localization of induced human interferon-β mRNA by non-radioactive in situ hybridization

Summary Induced interferon- (IFN-) mRNA was localized in human FS-4 fibroblasts by in situ hybridization using biotinylated probes. The hybridization sites were detected by incubation with a nick-translated genomic DNA probe (1.8 kb) via streptavidin-colloidal gold followed by silver contrast enhancement. The positive signals were observed by reflection-contrast light microscopy. IFN- mRNA was transiently induced by poly r(I):r(C) in fibroblasts 2–4 h after induction. Induction in the presence of cycloheximide and actinomycin D (superinduction conditions) exhibited an enhanced level of IFN- mRNA with a maximum at 4–8 h. The kinetics of the IFN- mRNA expression in the cytoplasm as revealed by in situ hybridization proved to be compatible with the results of Northern biotting experiments of total cellular RNA.     

Serum antibody responses against human papillomavirus in relation to tumor characteristics,response to treatment,and survival in carcinoma of the uterine cervix

To investigate whether the serum antibody responses to human papillomavirus (HPV) in cervical carcinoma were related to the clinical and histopathological features of the tumors and how the antibody responses were affected by treatment, pretreatment serum samples from 66 patients with carcinoma of the cervix were studied for the presence of IgA or IgG responses against six defined HPV epitopes. Posttreatment serum samples were drawn from the same patients 2–24 months after initiation of treatment. There was no significant correlation between pretreatment level of any of the investigated antibodies and clinical stage or differentiation of tumor. For the IgA responses to the epitopes 24516 and 24518 in the E2 protein there was a significant correlation between an increased pretreatment antibody level and a shortened survival. A high pretreatment value of IgA against 24516 was also associated with the absence of any complete response after therapy. The antibody levels declined dramatically after therapy for most of the antigens studied. However, this decline was seen both among the 53 patients with complete remission and among the 13 patients with remaining or progressive disease. Thus, the investigated serological responses were not useful as tumor markers, since patients with progressive, latestage disease may fail to mount an antibody response to these proteins. However, pretreatment levels of the serological responses to the HPV epitopes 24516 and 24518 were associated with prognosis in cervical cancer.     

Clinical effects of constant rate infusions of medetomidine–propofol combined with sevoflurane anesthesia in Thoroughbred racehorses undergoing arthroscopic surgery

Background

The aim of the present study was to evaluate clinical efficacy of constant rate infusions (CRIs) of medetomidine–propofol combined with sevoflurane anesthesia in Thoroughbred racehorses undergoing arthroscopic surgery. Thirty horses were sedated intravenously (IV) with medetomidine (6.0 μg/kg) and midazolam (0.02 mg/kg) and induced IV with ketamine (1.0 mg/kg) and propofol (1.0 mg/kg). These horses were randomly allocated to three groups and maintained with sevoflurane and CRI of either medetomidine (3.0 μg/kg/h) (Group M; n?=?10); or medetomidine (3.0 μg/kg/h) and propofol (3.0 mg/kg/h) (Group MP3; n?=?10); or medetomidine (3.0 μg/kg/h) and propofol (6.0 mg/kg/h) (Group MP6; n?=?10). End-tidal sevoflurane concentration (ET_SEVO), cardiovascular parameters, plasma propofol concentration, and recovery time and quality were compared among groups. Data were analyzed by using ANOVA with Tukey’s multiple comparison test, considering P?<?0.05 significant.

Results

ET_SEVO (%) was 2.4?±?0.1 in Group M, 1.7?±?0.2 in Group MP3, and 1.4?±?0.2 in Group MP6; ET_SEVO declined significantly in a propofol-dose-dependent manner. The rates of dobutamine infusion (μg/kg/min) required to keep the mean arterial blood pressure over 70 mmHg were significantly lower in Group MP3 (0.20?±?0.10) and Group MP6 (0.15?±?0.06) than in Group M (0.37?±?0.18). Recovery time and quality did not differ among groups. All horses in Group MP3 required only one attempt to stand, and recovery quality was excellent. Plasma propofol concentrations were stable throughout maintenance of anesthesia in Group MP3, whereas those in Group MP6 increased significantly with increasing duration of maintenance.

Conclusions

CRIs of medetomidine–propofol reduced the sevoflurane requirement for surgical anesthesia as the propofol dose increased, compared with a CRI of medetomidine alone. Additionally, the two propofol protocols provided good maintenance of cardiovascular function. CRIs of medetomidine (3.0 μg/kg/h) and propofol (3.0 mg/kg/h) resulted in excellent-quality recovery. This protocol could therefore be an especially useful additive to sevoflurane anesthesia in Thoroughbred racehorses undergoing arthroscopic surgery.

     

Production of an extracellular polysaccharide byAgrobacterium sp DS3 NRRL B-14297 isolated from soil

A bacterium isolated from soil and identified asAgrobacterium sp produced a water-soluble extracellular polysaccharide capable of producing highly viscous solutions. Gas chromatographic analysis revealed a sugar composition of glucose, galactose and mannose in the molar ratio of 7.52.41, together with 3.7% (w/w) pyruvic acid. Methylation analyses showed the presence of (13)-, (14)- and (16)-linked glucose, (13)- and (14, 16)-linked galactose and a small portion of (13)-linked mannose residues. Succinic acid was not present. The molecular weight of the polysaccharide was estimated by light scattering to be 2×10⁶ Da. The viscosity of solutions containing the polysaccharide remained constant from pH 3 to 11, and decreased by 50% when heated from 5 to 55°C. Maximum yield of the polysaccharide, 20 g L^–1, was reached in 48 h at 30°C incubation.     

A Janus Tale of Two Active High Mobility Group Box 1 (HMGB1) Redox States

High mobility group box 1 (HMGB1), the prototypic damage–associated molecular pattern molecule, is released at sites of inflammation and/or tissue damage. There, it promotes cytokine production and chemokine production/cell migration. New work shows that the redox status of HMGB1 distinguishes its cytokine-inducing and chemokine activity. Reduced all-thiol-HMGB1 has sole chemokine activity, whereas disulfide-HMGB1 has only cytokine activity, and oxidized, denatured HMGB1 has neither. Autophagy (programmed cell survival) and apoptosis (programmed cell death) have been implicated in controlling both innate and adaptive immune functions. Reduced HMGB1 protein promotes autophagy, whereas oxidized HMGB1 promotes apoptosis. Thus, the differential activity of HMGB1 in immunity, inflammation and cell death depends on the cellular redox status within tissues.High mobility group box 1 (HMGB1), a nonhistone nuclear factor, acts extracellularly as a damage-associated molecular pattern (DAMP) molecule to modulate inflammation, promoting autophagy and innate immune responses (1–5). HMGB1 has compartment-specific functions: nuclear, intracellular (but extranuclear) and extracellular. Its extracellular functions can now be divided further into cytokine-like or cytokine-inducing, chemokinelike and proangiogenic. Signaling pathways that induce variations on the posttranslational modification, such as phosphorylation and acetylation, have been implicated in the regulation of HMGB1 release. Importantly, HMGB1 contains three cysteines, each of which is susceptible to redox modification (6,7). The redox state of these cysteines is important for the proinflammatory cytokine-stimulating and proautophagic activity of HMGB1 (8–10). Autophagy (literally “self-eating”), a lysosome-mediated catabolic process, contributes to maintenance of intracellular homeostasis and promotes cell survival in response to environmental stress (11–13).Treatment with reduced but not oxidized HMGB1 protein increases autophagy in cancer cells (9). In contrast, oxidized HMGB1 protein activates the caspase-dependent apoptotic cell death pathway (9). Venereau et al.(14) described a new role for redox control of both the cytokine-inducing and chemokine activity of HMGB1 in the setting of sterile inflammation, regulating leukocyte recruitment and their ability to secrete inflammatory cytokines (Figure 1).Open in a separate windowFigure 1Redox control of HMGB1 activity. To act as a DAMP/danger signal and inflammatory mediator, HMGB1 is transported extracellularly by two principal means: active secretion from living inflammatory cells (for example, macrophages) or passive release from necrotic cells. The activities of extracellular HMGB1 are redox dependent. All-thiol-HMGB1 promotes chemokine production and leukocyte recruitment. Disulfide-HMGB1, originating from infiltrating leukocytes, promotes release of proinflammatory cytokines and thus participates in the inflammatory response. Reactive oxygen species produced by leukocytes induces the terminal oxidation of HMGB1, which is inactivated during resolution of inflammation.Structurally, HMGB1 is composed of three domains: two positively charged proximal DNA-binding domains (A box and B box) and a negatively charged carboxyl terminus. Three cysteines are encoded within the molecule: two vicinal cysteines in box A (C23 and C45) and a single one in box B (C106). C23 and C45 can form an intermolecular disulfide bond, whereas C106 is unpaired. Therefore, three different redox forms HMGB1 (all-thiol-HMGB1, disulfide-HMGB1 and oxidized HMGB1) were derived from bacterial expression systems (14). In addition, by using tryptic digests and liquid chromatography tandem mass spectrometric analysis, Venereau et al. observed that recombinant HMGB1 can be reversibly oxidized and reduced in the presence of electron donors (for example, dithiothreitol) or acceptors (oxygen) (14).Next, Venereau et al. assessed whether individual redox forms of HMGB1 have a differential role in cytokine-stimulating and chemoattractant activities (14). They found that disulfide-HMGB1 induced activation of the nuclear factor (NF)-κB pathway and production of proinflammatory cytokines (for example, tumor necrosis factors-α, interleukin [IL]-6 and IL-8) in fibroblasts and macrophages. Interestingly, all-thiol-HMGB1 failed to induce a proinflammatory response. In contrast, all-thiol-HMGB1, but not disulfide-HMGB1, had chemoattractant activity in fibroblasts. These findings prompted them to determine whether HMGB1 inhibitors, such as box A and monoclonal antibody PDH1.1, block the chemoattractant and/or cytokine-inducing activities of HMGB1. Unexpectedly, these inhibitors prevented cell migration but not cytokine production, although they are widely used as HMGB1-targeting agents in experimental inflammatory diseases.Reactive oxygen species oxidize the HMGB1 released from dying cells, thereby neutralizing its stimulatory activity and promoting tolerance in immune cells (15,16). In addition, oxidation of C106 or lack of a disulfide bridge between C23 and C45 then causes HMGB1 to lose its proinflammatory effects in macrophages (8). Venereau et al. found that terminal oxidation by hydrogen peroxide results in the loss of both the cytokine-stimulating and chemoattractant activities of HMBG1. Moreover, the authors found that the three HMGB1 cysteine residues were required for the cytokine-stimulating activity but not for the chemoattractant activity of HMGB1. Cysteine mutant HMGB1 promotes fibroblast migration, but not cytokine expression in macrophages (14). Collectively, these findings establish a crucial role for redox in the regulation of HMGB1 activity in inflammation and migration.What is the redox state of HMGB1 in the pathogenesis of individual diseases? The redox state of HMGB1 from the human acute monocytic leukemia cell line THP-1 was measured in the presence or absence of lipopolysaccharide (LPS) and necrotic medium in vitro. Intracellular HMGB1 was all-thiol-HMGB1, whereas secreted HMGB1 contained both all-thiol- and disulfide-HMGB1 (14). Furthermore, disulfide-HMGB1 was present later and time-dependently increased in cardiotoxin-injured muscles in vivo, confirming that the redox state of HMGB1 is altered during tissue damage and inflammation. HMGB1 protein with all three cysteines mutated to serine are resistant to oxidation and induce leukocyte recruitment without inducing cytokine production (14). The activities of HMGB1 are thus redox-dependent and can be modified within the injured tissues after HMGB1 release. Therefore, release of dynamic redox-regulated HMGB1 contributes to the orderly orchestrated recruitment of leukocytes, activation of cytokine release and subsequent resolution of inflammation.Several issues remain unresolved regarding the redox control of HMGB1 activity. First, HMGB1 is specifically recognized by several cell surface receptors (2), including Toll-like receptor (TLR)-4 and the receptor for advanced glycation end products (RAGE), but most recently was joined by T-cell immunoglobulin and mucin domain 3 (TIM-3) (17). Initial studies suggest that reduced C106 is necessary for the binding of HMGB1 to one of its receptors, TLR4, to stimulate cytokine release (8). HMGB1-induced recruitment of inflammatory cells depends on forming a complex with CXCL12 and signaling via CXCR4 (18). Moreover, RAGE is required for reduced HMGB1-mediated autophagy, but not oxidized HMGB1-induced apoptosis (9). All-thiol-HMGB1, but not disulfide-HMGB1, binds CXCL12 (14). The influence of HMGB1 receptors (for example, RAGE, TLR4, TLR2, CD24, TIM-3 and triggering receptor expressed on myeloid cells 1 [TREM1]) on biological activities of individual redox forms of HMGB1 remains to be carefully investigated. Second, HMGB1 forms highly inflammatory complexes with DNA, lipoteichoic acid, LPS, IL-1β, chemokine (C-X-C motif) ligand 12 (CXCL12)/ stromal cell–derived factor-1 (SDF-1) and nucleosomes (19). There is great interest in determining whether the individual redox forms of HMGB1 have varying affinity profiles active in inflammation and immunity. Third, HMGB1 has multiple intracellular and extracellular functions in health and disease, including cancer (1,2,6,20). Additional studies will be needed to determine whether redox is required for other functions of HMGB1, such as regeneration and cellular differentiation as well as the complex interactions between autophagy and immunity (5). One additional unanswered question is where and how the formation of the disulfide takes place and whether there is an enzyme specific for regulating this. This is important, knowing that the nuclear form is mostly all thiol. Finally, the development and performance of a simple, sensitive method for the detection of individual HMGB1 redox state isoforms in clinical specimens remains to be accomplished.     

HIV-1 Specific IgA Detected in Vaginal Secretions of HIV Uninfected Women Participating in a Microbicide Trial in Southern Africa Are Primarily Directed Toward gp120 and gp140 Specificities

Background

Many participants in microbicide trials remain uninfected despite ongoing exposure to HIV-1. Determining the emergence and nature of mucosal HIV-specific immune responses in such women is important, since these responses may contribute to protection and could provide insight for the rational design of HIV-1 vaccines.

Methods and Findings

We first conducted a pilot study to compare three sampling devices (Dacron swabs, flocked nylon swabs and Merocel sponges) for detection of HIV-1-specific IgG and IgA antibodies in vaginal secretions. IgG antibodies from HIV-1-positive women reacted broadly across the full panel of eight HIV-1 envelope (Env) antigens tested, whereas IgA antibodies only reacted to the gp41 subunit. No Env-reactive antibodies were detected in the HIV-negative women. The three sampling devices yielded equal HIV-1-specific antibody titers, as well as total IgG and IgA concentrations. We then tested vaginal Dacron swabs archived from 57 HIV seronegative women who participated in a microbicide efficacy trial in Southern Africa (HPTN 035). We detected vaginal IgA antibodies directed at HIV-1 Env gp120/gp140 in six of these women, and at gp41 in another three women, but did not detect Env-specific IgG antibodies in any women.

Conclusion

Vaginal secretions of HIV-1 infected women contained IgG reactivity to a broad range of Env antigens and IgA reactivity to gp41. In contrast, Env-binding antibodies in the vaginal secretions of HIV-1 uninfected women participating in the microbicide trial were restricted to the IgA subtype and were mostly directed at HIV-1 gp120/gp140.     

Excitation-calcium release uncoupling in aged single human skeletal muscle fibers

The biological mechanisms underlying decline in muscle power and fatigue with age are not completely understood. The contribution of alterations in the excitation-calcium release coupling in single muscle fibers was explored in this work. Single muscle fibers were voltage-clamped using the double Vaseline gap technique. The samples were obtained by needle biopsy of the vastus lateralis (quadriceps) from 9 young (25–35 years; 25.9 ± 9.1; 5 female and 4 male) and 11 old subjects (65–75 years; 70.5 ± 2.3; 6 f, 5 m). Data were obtained from 36 and 39 fibers from young and old subjects, respectively. Subjects included in this study had similar physical activity. Denervated and slow-twitch muscle fibers were excluded from this study. A significant reduction of maximum charge movement (Q_max) and DHP-sensitive Ca current were recorded in muscle fibers from the 65–75 group. Q_max values were 7.6 ± 0.9 and 3.2 ± 0.3 nC/F for young and old muscle fibers, respectively (P < 0.01). No evidences of charge inactivation or interconversion (charge 1 to charge 2) were found. The peak Ca current was (–)4.7 ± 0.08 and (–)2.15 ± 0.11 A/F for young and old fibers, respectively (P < 0.01). The peak calcium transient studied with mag-fura-2 (400 m) was 6.3 ± 0.4 m and 4.2 ± 0.3 m for young and old muscle fibers, respectively. Caffeine (0.5 mm) induced potentiation of the peak calcium transient in both groups. The decrease in the voltage-/ Ca-dependent Ca release ratio in old fibers (0.18 ± 0.02) compared to young fibers (0.47 ± 0.03) (P < 0.01), was recorded in the absence of sarcoplasmic reticulum calcium depletion. These data support a significant reduction of the amount of Ca available for triggering mechanical responses in aged skeletal muscle and, the reduction of Ca release is due to DHPR-ryanodine receptor uncoupling in fast-twitch fibers. These alterations can account, at least partially for the skeletal muscle function impairment associated with aging.This work was supported by Grant-in-Aid from the American Heart Association (National) and Muscular Dystrophy Association, and National Institutes of Health (2-P60AG18484-06)     

Progesterone antagonism of androgen-dependent marking in gerbils

The role of progesterone (P) as an androgen antagonist in Mongolian gerbil territorial marking was assessed in the present investigation. Twenty-seven mature males were observed for two consecutive weeks in a marking apparatus. On the basis of the average of the two sessions, subjects were matched and assigned to one of three groups. All animals were orchidectomized and tested three weeks after surgery. Three days after a single postoperative session a schedule of hormonal replacement was initiated. Group 1 was administered 640 μg testosterone propionate (TP). Group 2 received 640 μg TP followed immediately by 3 mg P. A third group received 640 μg TP followed by a 1 mg injection of P. Subjects were tested for marking 24 hr after the injection for a cumulative period of 6 wk.Results indicated that (a) castration drastically reduced marking; (b) TP alone and TP + 1 mg P restored the behavior; (c) TP + 3 mg P inhibited its restoration; and (d) a significant interaction was present between hormonal therapy and the 6-wk testing interval. However, all three hormonal treatments restored sebaceous gland dimensions. Results are discussed in terms of a model of hormone-gene action.     

The Rewarding Aspects of Music Listening Are Related to Degree of Emotional Arousal

Background

Listening to music is amongst the most rewarding experiences for humans. Music has no functional resemblance to other rewarding stimuli, and has no demonstrated biological value, yet individuals continue listening to music for pleasure. It has been suggested that the pleasurable aspects of music listening are related to a change in emotional arousal, although this link has not been directly investigated. In this study, using methods of high temporal sensitivity we investigated whether there is a systematic relationship between dynamic increases in pleasure states and physiological indicators of emotional arousal, including changes in heart rate, respiration, electrodermal activity, body temperature, and blood volume pulse.

Methodology

Twenty-six participants listened to self-selected intensely pleasurable music and “neutral” music that was individually selected for them based on low pleasure ratings they provided on other participants'' music. The “chills” phenomenon was used to index intensely pleasurable responses to music. During music listening, continuous real-time recordings of subjective pleasure states and simultaneous recordings of sympathetic nervous system activity, an objective measure of emotional arousal, were obtained.

Principal Findings

Results revealed a strong positive correlation between ratings of pleasure and emotional arousal. Importantly, a dissociation was revealed as individuals who did not experience pleasure also showed no significant increases in emotional arousal.

Conclusions/Significance

These results have broader implications by demonstrating that strongly felt emotions could be rewarding in themselves in the absence of a physically tangible reward or a specific functional goal.     

Reduction of Seizure Occurrence from Exposure to Auditory Stimulation in Individuals with Neurological Handicaps: A Randomized Controlled Trial

Background

The purpose of this work was to determine in a clinical trial the efficacy of reducing or preventing seizures in patients with neurological handicaps through sustained cortical activation evoked by passive exposure to a specific auditory stimulus (particular music). The specific type of stimulation had been determined in previous studies to evoke anti-epileptiform/anti-seizure brain activity.

Methods

The study was conducted at the Thad E. Saleeby Center in Harstville, South Carolina, which is a permanent residence for individuals with heterogeneous neurological impairments, many with epilepsy. We investigated the ability to reduce or prevent seizures in subjects through cortical stimulation from sustained passive nightly exposure to a specific auditory stimulus (music) in a three-year randomized controlled study. In year 1, baseline seizure rates were established. In year 2, subjects were randomly assigned to treatment and control groups. Treatment group subjects were exposed during sleeping hours to specific music at regular intervals. Control subjects received no music exposure and were maintained on regular anti-seizure medication. In year 3, music treatment was terminated and seizure rates followed. We found a significant treatment effect (p = 0.024) during the treatment phase persisting through the follow-up phase (p = 0.002). Subjects exposed to treatment exhibited a significant 24% decrease in seizures during the treatment phase, and a 33% decrease persisting through the follow-up phase. Twenty-four percent of treatment subjects exhibited a complete absence of seizures during treatment.

Conclusion/Significance

Exposure to specific auditory stimuli (i.e. music) can significantly reduce seizures in subjects with a range of epilepsy and seizure types, in some cases achieving a complete cessation of seizures. These results are consistent with previous work showing reductions in epileptiform activity from particular music exposure and offers potential for achieving a non-invasive, non-pharmacologic treatment of epilepsy.

Trial Registration

Clinicaltrials.gov {"type":"clinical-trial","attrs":{"text":"NCT01459692","term_id":"NCT01459692"}}NCT01459692     

SALIVARY ANTIMICROBIAL PROTEIN RESPONSE TO PROLONGED RUNNING

Introduction

Prolonged exercise may compromise immunity through a reduction of salivary antimicrobial proteins (AMPs). Salivary IgA (IgA) has been extensively studied, but little is known about the effect of acute, prolonged exercise on AMPs including lysozyme (Lys) and lactoferrin (Lac).

Objective

To determine the effect of a 50-km trail race on salivary cortisol (Cort), IgA, Lys, and Lac.

Methods

14 subjects: (6 females, 8 males) completed a 50km ultramarathon. Saliva was collected pre, immediately after (post) and 1.5 hrs post race (+1.5).

Results

Lac concentration was higher at +1.5 hrs post race compared to post exercise (p < 0.05). Lys was unaffected by the race (p > 0.05). IgA concentration, secretion rate, and IgA/Osm were lower +1.5 hrs post compared to pre race (p < 0.05). Cort concentration was higher at post compared to +1.5 (p < 0.05), but was unaltered from pre race levels. Subjects finished in 7.81±1.2 hrs. Saliva flow rate did not differ between time points. Saliva Osm increased at post (p < 0.05) compared to pre race.

Conclusions

The intensity could have been too low to alter Lys and Lac secretion rates and thus, may not be as sensitive as IgA to changes in response to prolonged running. Results expand our understanding of the mucosal immune system and may have implications for predicting illness after prolonged running.     

Cerebral Activations Related to Audition-Driven Performance Imagery in Professional Musicians

Functional Magnetic Resonance Imaging (fMRI) was used to study the activation of cerebral motor networks during auditory perception of music in professional keyboard musicians (n = 12). The activation paradigm implied that subjects listened to two-part polyphonic music, while either critically appraising the performance or imagining they were performing themselves. Two-part polyphonic audition and bimanual motor imagery circumvented a hemisphere bias associated with the convention of playing the melody with the right hand. Both tasks activated ventral premotor and auditory cortices, bilaterally, and the right anterior parietal cortex, when contrasted to 12 musically unskilled controls. Although left ventral premotor activation was increased during imagery (compared to judgment), bilateral dorsal premotor and right posterior-superior parietal activations were quite unique to motor imagery. The latter suggests that musicians not only recruited their manual motor repertoire but also performed a spatial transformation from the vertically perceived pitch axis (high and low sound) to the horizontal axis of the keyboard. Imagery-specific activations in controls were seen in left dorsal parietal-premotor and supplementary motor cortices. Although these activations were less strong compared to musicians, this overlapping distribution indicated the recruitment of a general ‘mirror-neuron’ circuitry. These two levels of sensori-motor transformations point towards common principles by which the brain organizes audition-driven music performance and visually guided task performance.     

Effects of biofeedback on the discrimination of electrodermal activity

Twenty-four subjects were tested on their ability to discriminate between the presence and absence of negative skin potential responses before and after training to control skin potential. Training consisted of 52 discrete 30-second trials during which subjects were asked either to increase or to inhibit palmar sweating. Subjects in groups N and P were provided with analogue feedback on their skin potential activity. Group N was correctly informed that increases in sweating were indicated by increases in the negativity of skin potential; group P was misinformed that these were indicated by increases in the positivity of skin potential. Subjects in the control (C) group received no feedback. Reliable evidence of discrimination was obtained only in groups N and P, following training. However, reliable evidence of control was obtained only in group N. Thus, training to control skin potential led to an ability to identify afferentation associated with the more common (i.e., negative) skin potential responses, even though biofeedback training appeared unsuccessful in the case of group P. These findings are discussed in the context of discrimination or awareness accounts of the process of acquiring control of internal responses.This work was supported by grants from York University and from Glendon College.