The impact of gender of the examiner on orofacial pain perception and pain reporting among healthy volunteers.

OBJECTIVES: Pain on palpation of jaw muscles is a commonly used diagnostic criterion when examining patients with orofacial pain. It is not known, however, if pain reports are affected by the gender of the examiner. Our aim was to investigate if pressure pain threshold (PPT), pressure pain tolerance (PTol), and pain intensity assessed over the masseter muscles in healthy individuals are affected by the gender of the examiner. MATERIALS AND METHODS: Healthy, pain-free individuals were recruited on a voluntary basis. PPT and PTol were assessed using pressure algometry. At the PTol level, participants also rated pain intensity on a 0-10 numeric rating scale. Assessments of PPT and PTol were conducted with six repeated measurements performed twice, separately by one female and one male examiner, on each participant. RESULTS: In total, 84 participants (43 women; median age 24, IQR 6) were included. With a female examiner, women reported higher pain intensity than men (Mann Whitney U, p = 0.005). In the multivariable analysis, significantly higher PTol was predicted by male examiner. Also, a higher ratio between PTol and reported pain intensity was predicted by male examiner. CONCLUSIONS: The gender of the examiner influences pain reporting and perception in an experimental setting. This effect on pain perception related to gender of the examiner is probably related to normative gender behaviors rather than to biological alterations within the examined individual. CLINICAL RELEVANCE: In clinical and experimental settings, gender of the examiner may affect not only pain perception but also pain reporting, with potential implications for diagnostics in patients with pain.

Remote CB1 receptor antagonist administration reveals multiple sites of tonic and phasic endocannabinoid neuroendocrine regulation.

Endogenous cannabinoids (endocannabinoids, eCB) are expressed throughout the body and contribute to regulation of the hypothalamo-pituitary-adrenal (HPA) axis and general stress reactivity. This study assessed the contributions of CB1 receptors (CB1R) in the modulation of basal and stress-induced neural and HPA axis activities. Catheterized adult male rats were placed in chambers to acclimate overnight, with their catheters connected and exteriorized from the chambers for relatively stress-free remote injections. The next morning, the CB1R antagonist AM251 (1 or 2 mg/kg) or vehicle was administered, and 30 min later, rats were exposed to loud noise stress (30 min) or no noise (basal condition). Blood, brains, pituitary and adrenal glands were collected immediately after the procedures for analysis of c-fos and CB1R mRNAs, corticosterone (CORT) and adrenocorticotropin hormone (ACTH) plasma levels. Basally, CB1R antagonism induced c-fos mRNA in the basolateral amygdala (BLA) and auditory cortex (AUD) and elevated plasma CORT, indicating disruption of eCB-mediated constitutive inhibition of activity. CB1R blockade also potentiated stress-induced hormone levels and c-fos mRNA in several regions such as the bed nucleus of the stria terminalis (BST), lateral septum (LS), and basolateral amygdala (BLA) and the paraventricular nucleus of the hypothalamus (PVN). CB1R mRNA was detected in all central tissues investigated, and the adrenal cortex, but at very low levels in the anterior pituitary gland. Interestingly, CB1R mRNA was rapidly and bidirectionally regulated in response to stress and/or antagonist treatment in some regions. eCBs therefore modulate the HPA axis by regulating both constitutive and activity-dependent inhibition at multiple levels.

Diagnostic criteria for temporomandibular disorders: Diagnostic accuracy for general dentistry procedure without mandatory commands regarding myalgia, arthralgia and headache attributed to temporomandibular disorder.

The clinical examination in diagnostic criteria for temporomandibular disorders (DC/TMD) is a strict procedure and comprises mandatory commands. However, learning and using these mandatory commands in general practice have proven to be difficult and their use of DC/TMD is minimal. To investigate whether reliability on a diagnostic level for DC/TMD diagnoses differs between examiners using the mandatory commands or not. Six examiners were divided into two groups: one using the mandatory commands in DC/TMD for the clinical examination and one who did not use the mandatory commands. A reliability assessment was performed twice, one occasion for each group of examiners. The assessment was performed according to the guidelines from the International Network for Orofacial Pain and Related Disorders Methodology. Each group of examiners thereby examined 16 subjects (11 TMD patients and 5 healthy individuals) each, and the diagnostic agreement (reliability) as compared to diagnoses derived by a reference standard examiner was calculated with Cohen' s kappa coefficient. The DC/TMD diagnoses myalgia, arthralgia and headache attributed to TMD were included in the reliability assessment. There was no significant difference regarding diagnostic agreement reliability between the examiners using or not using the mandatory DC/TMD commands. This study indicates that not using the mandatory commands in DC/TMD in general practice does not impair the diagnostic reliability regarding the diagnoses myalgia, arthralgia and headache attributed to TMD compared to including the commands.

Gravity Spy: integrating advanced LIGO detector characterization, machine learning, and citizen science.

With the first direct detection of gravitational waves, the advanced laser interferometer gravitational-wave observatory (LIGO) has initiated a new field of astronomy by providing an alternative means of sensing the universe. The extreme sensitivity required to make such detections is achieved through exquisite isolation of all sensitive components of LIGO from non-gravitational-wave disturbances. Nonetheless, LIGO is still susceptible to a variety of instrumental and environmental sources of noise that contaminate the data. Of particular concern are noise features known as glitches, which are transient and non-Gaussian in their nature, and occur at a high enough rate so that accidental coincidence between the two LIGO detectors is non-negligible. Glitches come in a wide range of time-frequency-amplitude morphologies, with new morphologies appearing as the detector evolves. Since they can obscure or mimic true gravitational-wave signals, a robust characterization of glitches is paramount in the effort to achieve the gravitational-wave detection rates that are predicted by the design sensitivity of LIGO. This proves a daunting task for members of the LIGO Scientific Collaboration alone due to the sheer amount of data. In this paper we describe an innovative project that combines crowdsourcing with machine learning to aid in the challenging task of categorizing all of the glitches recorded by the LIGO detectors. Through the Zooniverse platform, we engage and recruit volunteers from the public to categorize images of time-frequency representations of glitches into pre-identified morphological classes and to discover new classes that appear as the detectors evolve. In addition, machine learning algorithms are used to categorize images after being trained on human-classified examples of the morphological classes. Leveraging the strengths of both classification methods, we create a combined method with the aim of improving the efficiency and accuracy of each individual classifier. The resulting classification and characterization should help LIGO scientists to identify causes of glitches and subsequently eliminate them from the data or the detector entirely, thereby improving the rate and accuracy of gravitational-wave observations. We demonstrate these methods using a small subset of data from LIGO's first observing run.

Cannabinoid receptor type 1 antagonism significantly modulates basal and loud noise induced neural and hypothalamic-pituitary-adrenal axis responses in male Sprague-Dawley rats.

Altered regulation of the hypothalamic-pituitary-adrenal (HPA) axis is associated with stress-induced changes in cognitive, emotional, and physical health. Recent evidence indicates that the endogenous cannabinoid (eCB) system may modulate HPA-axis function both directly and more centrally, via regulation of limbic brain systems that control HPA-axis activity. The current study examines the contribution of cannabinoid type 1 (CB1) receptor modulation throughout the neuraxis on control and stress-induced HPA-axis activity. Adult male Sprague-Dawley rats were given intraperitoneal injections of either CB1 receptor antagonist (AM251, 2 mg/kg) or vehicle 30 min prior to a session of loud white noise stress (95 dBA for 30 min) or placement in a familiar sound-proof chamber. Immediately following stress and control treatments, rats were killed, the brains and pituitary glands were excised for subsequent immediate early gene (c-fos mRNA) measurement, and trunk blood was collected for subsequent determination of corticosterone (CORT) and adrenocorticotropic (ACTH) hormone levels. AM251 treatment resulted in a potentiated plasma ACTH response to loud noise stress. AM251 treatment also increased stress-induced plasma CORT levels, but that increase may be due to an increase in basal plasma CORT levels, as was evident in control rats. AM251 treatment produced three distinctive c-fos mRNA response patterns across the various brain regions examined. In cortical (prelimbic, infralimbic, somatosensory, and auditory) and some subcortical structures (basolateral amygdala and paraventricular nucleus of the hypothalamus), AM251 treatment produced a substantial increase in c-fos mRNA that was comparable with the elevated c-fos mRNA levels present in those brain regions of both vehicle and AM251-treated stressed rats. In some other subcortical structures (bed nucleus of the stria terminalis and medial preoptic area) and the anterior pituitary, AM251 treatment produced a c-fos mRNA response pattern that was similar to the response pattern of ACTH hormone levels, that is, no effect on no noise control levels, but an augmentation of stress-induced levels. Conversely, in the medial geniculate and ventral posterior thalamus, AM251 treatment inhibited stress-induced c-fos mRNA induction. These data indicate that disruption of eCB signaling through CB1 receptors results in potentiated neural and endocrine responses to loud noise stress, but also substantial increases in activity in various brain regions and the adrenal gland.

Tonic, but not phasic corticosterone, constrains stress activatedextracellular-regulated-kinase 1/ 2 immunoreactivity within the hypothalamic paraventricular nucleus.

The negative-feedback actions of corticosterone (CORT) depend on both phasic and tonic CORT secretion patterns to regulate hypothalamic-pituitary-adrenal (HPA) axis activity. How these two different CORT secretion pattens influence specific intracellular signal transduction pathway activity within the cellular elements of the HPA axis has not been determined. For example, it is unknown whether CORT has suppressive actions over signal transduction events within medial parvocellular paraventricular nucleus (PVN) corticotrophin-releasing hormone (CRH) neurones, nor whether these suppressive actions are responsible for alterations in PVN transcriptional processes and neurohormone secretion associated with stress. The extracellular-regulated kinase (ERK) is a stress activated intracellular signalling molecule that is potentially subject to glucocorticoid negative-feedback regulation. We tested the ability of CORT to modulate levels of the active (phosphorylated) form of ERK (pERK1/2) in the PVN of rats. Acute psychological stress (restraint) produced a rapid increase in the number of PVN pERK1/2 immunopositive cells within CRH neurones. Absence of tonic CORT via adrenalectomy (ADX) produced no change in basal pERK1/2 cell counts but augmented the increased pERK1/2 cell counts elicited by acute restraint. Treatment of ADX rats with CORT in the drinking water normalised this enhanced pERK1/2 response to stress. By contrast, treatment of ADX rats with a phasic increase in CORT 1 h before restraint had no effect on pERK1/2 cell counts, despite substantially suppressing stress-induced PVN crh gene expression and adrenonocorticotrophic hormone secretion. This tonic CORT inhibition of stress-induced activation of ERK1/2 may involve both alteration of the activity of stress-dependent neural inputs to PVN CRH neurones and alteration within those neurones of stress-dependent intracellular signalling mechanisms associated with ERK activation.

Inhibitory effects of corticosterone in the hypothalamic paraventricular nucleus (PVN) on stress-induced adrenocorticotrophic hormone secretion and gene expression in the PVN and anterior pituitary.

Endogenous glucocorticoid negative-feedback influence on the hypothalamic-pituitary-adrenal (HPA) axis depends on glucocorticoid actions exerted on multiple glucocorticoid-sensitive tissues and differential glucocorticoid effects that are expressed within several distinct temporal domains. The relative contribution and underlying molecular mechanisms of action for the effects of location and timing of glucocorticoid exposure on HPA axis activity remain to be determined. In the present study, we examined the effects of acute exposure to corticosterone (CORT) at the level of the paraventricular nucleus (PVN) on the HPA axis response to a subsequent stressor in a short-term (1 h) timeframe. Intra-PVN CORT microinjection 1 h before restraint suppressed the adrenocorticotrophic hormone (ACTH) response and blunted restraint-induced corticotrophin-releasing hormone (CRH) heterogeneous nuclear (hn)RNA expression in the PVN and pro-opiomelanocortin hnRNA expression in the anterior pituitary (AP); however, it had no effect on restraint-induced plasma prolactin levels and c-fos mRNA expression (PVN and AP). This pattern of results suggests that CORT acts locally at the level of the PVN within a short-term timeframe to suppress stress-induced excitation-exocytosis coupling within CRH neurones and CRH gene induction without altering the stress-associated trans-synaptic input and intracellular signal transduction that converges on PVN c-fos gene induction. The present study is the first to demonstrate that an acute infusion of CORT into the PVN is sufficient to suppress the ACTH response to stress initiated 1 h after CORT infusion.

The non-proteolytic house dust mite allergen Der p 2 induce NF-kappaB and MAPK dependent activation of bronchial epithelial cells.

BACKGROUND: House dust mites (HDM) are well-known as a source of indoor aeroallergens and for causing allergic airway diseases. Some proteolytic HDM allergens are known to activate respiratory epithelial cells to produce pro-inflammatory mediators, while there is limited knowledge regarding such activity among non-proteolytic HDM allergens. OBJECTIVE: To investigate whether Der p 2, a major non-proteolytic allergen of Dermatophagoides pteronyssinus, activates respiratory epithelial cells to produce mediators involved in asthma pathogenesis and to elucidate the mechanism of such activation. METHODS: The human bronchial epithelial cell line BEAS-2B, normal human bronchial epithelial (NHBE) cells and the alveolar epithelial cell line A549 were exposed to recombinant Der p 2. Following exposure, we analysed a panel of soluble mediators and cell adhesion receptors involved in asthma pathogenesis by promoting recruitment, survival and binding of inflammatory cells. The involvement of nuclear factor (NF)-kappaB and mitogen-activated protein kinases (MAPKs) was studied using specific inhibitors. RESULTS: Der p 2 activated bronchial BEAS-2B and NHBE cells, but not alveolar A549 cells. In BEAS-2B cells Der p 2 induced dose-dependent up-regulation in both mRNA level and protein secretion of granulocyte-macrophage colony-stimulating factor, IL-6, IL-8, monocyte-chemotactic protein-1 and macrophage inflammatory protein-3alpha. Secretion as well as surface expression of intercellular adhesion molecule (ICAM)-1 was also up-regulated, which was associated with increased adhesion of monocytes to the epithelial cells. The release of cytokines and chemokines was regulated by NF-kappaB and MAPK activation in different ways, while expression of ICAM-1 was solely dependent on NF-kappaB activation. CONCLUSION: These results show that Der p 2 activates respiratory epithelial cells, indicating that this non-proteolytic allergen, in addition to its immunogenic properties, can aggravate respiratory airway disease by adjuvant-like activation of the lung epithelium.

Vitamin E down-modulates mitogen-activated protein kinases, nuclear factor-kappaB and inflammatory responses in lung epithelial cells.

The airway epithelium plays an active role in acute lung inflammation by producing chemotactic factors and by expressing cell adhesion molecules involved in the migration of leucocytes to extravascular spaces. We have reported previously that neutrophil migration to airways can be down-modulated by exogenously administered vitamin E (alpha-tocopherol). The mechanism for this effect is not well understood, however. The action of alpha-tocopherol was investigated in human alveolar type II and bronchial epithelial cells stimulated with tumour necrosis factor-alpha. Treatment of alveolar epithelial cells with alpha-tocopherol resulted in down-regulated cell surface expression of intercellular adhesion molecule-1 (ICAM-1). On bronchial epithelial cells, both ICAM-1 and vascular adhesion molecule-1 were decreased, leading to diminished adherence of leucocytes to the cells. The production of the neutrophil chemoattractant interleukin-8 was attenuated in both alveolar and bronchial cells. These effects were preceded by reduced activation of the mitogen-activated protein kinases (MAPK), extracellular signal-regulated kinase (ERK1/2) and p38, as well as down-regulation of nuclear factor-kappaB. Comparing the effects of alpha-tocopherol with that of specific inhibitors of MAPK and protein kinase C (PKC) revealed that effects appear to be partly independent of PKC inhibition. These results implicate the anti-inflammatory action of alpha-tocopherol in addition to its anti-oxidant properties.

Endurance during chewing in whiplash-associated disorders and TMD.

We have previously shown an association between neck injury and disturbed jaw function. This study tested the hypothesis of a relationship between neck injury and impaired endurance during chewing. Fifty patients with whiplash-associated disorders (WAD) were compared with 50 temporomandibular disorders (TMD) patients and 50 healthy subjects. Endurance was evaluated during unilateral chewing of gum for 5 min when participants reported fatigue and pain. Whereas all healthy subjects completed the task, 1/4 of the TMD and a majority of the WAD patients discontinued the task. A majority of the WAD patients also reported fatigue and pain. These findings suggest an association between neck injury and reduced functional capacity of the jaw motor system. From the results, we propose that routine examination of WAD patients should include jaw function and that an endurance test as described in this study could also be a useful tool for non-dental professionals.

Estrogen receptor beta is expressed in human embryonic brain cells and is regulated by 17beta-estradiol.

In order to study estrogen effects on developing human neurons, we have established primary cultures of neurons and glia from 8-13-week human embryo cortex and spinal cord. The neuronal identity of the cultures was verified using the neuronal synaptic vesicle and neuronal endosomal membrane markers synaptotagmin, synapsin and synaptophysin, and the glial contribution to the mixed glial-neuronal cultures was verified using the glial marker glial fibrillary acidic protein (GFAP). We here report expression of estrogen receptor beta (ERbeta) in these cells using RT-PCR and sequencing, RNAse protection assay, immunohistochemistry and immunoblotting. We found that both neuronal and mixed glial-neuronal cultures expressed ERbeta. Treatment with 17beta-estradiol gave an increased expression of ERbeta in both types of cultures. These results suggest that ERbeta is expressed in fetal brain and thus may mediate effects of estrogen in the developing nervous system. Furthermore, the results suggest that expression of ERbeta in fetal brain may be regulated by estrogen.

Specific localization of RBM1a in the nuclei of all cell types except elongated spermatids within seminiferous tubules of the human.

Recent studies have indicated that at least three regions (AZF a-c) on the long arm of the Y-chromosome code for factors are involved in spermatogenesis. One of the candidate genes in the AZFb region is RBM1a, coding for a protein with an RNA binding motif. In this study, poly clonal antibodies raised against a 15 amino acid peptide, corresponding to residues 263-304 of the deduced amino acid sequence of RBM1a, has been used to localize the RBM1a protein in the human testis. Immunohistochemistry on normal human testis using this RBM1a antibody, localized the antigen to the nuclei of spermatogonia, primary spermatocytes, and round spermatids but not to the nuclei of elongated spermatids. The antibody also specifically identified the nuclei of Sertoli cells, although the fluorescence was not as strong as in the germ cell nuclei it identified. No specific fluorescence was seen in the nuclei of either peritubular, endothelial or Leydig cells. Western blot of normal human testicular tissue using the anti-RBM1a antibody gave rise to a single specific band of approximately 55 kDa, corresponding to the expected size of RBM1a. In view of its expression in germ cells, and because RBM1a has an RNA binding domain, RBM1a may be involved in RNA processing, such as RNA splicing or RNA export which are events necessary for normal spermatogenesis.

Low number of Y-chromosome deletions in infertile azoospermic men at a Swedish andrology centre.

Recent studies have strongly indicated that at least three regions [azoospermia factor (AZF) a-c] on the long arm of the Y-chromosome code for factors involved in spermatogenesis. In order to reveal the prevalence of microdeletions in these regions in a Swedish population, 192 men consecutively referred to our andrology unit due to infertility and showing oligozoospermia (n=53) or azoospermia (n=139) but no obstruction or hormonal disturbances, were investigated. For this study we used a multiplex polymerase chain reaction (PCR) method including 13 pairs of primers divided into five different primer mixes. It was found that four men, all with azoospermia, had deletions including part of the AZFb region and probably the entire AZFc region. Testis biopsies showed different morphology ranging from absence of germ cells to hypospermatogenisis. Of special interest was one patient that was first investigated 10 years ago due to primary infertility and oligozoospermia. Today he has developed azoospermia. It is concluded that the number of patients with microdeletions on the Y chromosome is rather low (less than 3% in highly selected azoospermic men) in our study compared to a number of other studies in which a 1-55% incidence have been reported. It is possible that ethnic differences, selection criteria and methodological aspects can contribute to the difference between the present and previous studies.

TGFbeta receptor types I and II and the substrate proteins Smad 2 and 3 are present in human oocytes.

We have recently found that values of the transforming growth factor (TGF)beta1 in human ovarian follicular fluid obtained during ovarian stimulation for IVF were higher in women who subsequently became pregnant following embryo transfer. We therefore postulated that TGFbeta1 may have a beneficial effect on the preimplantation embryo and improve the chances of a successful implantation. We have used reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemistry to investigate the presence in human oocytes and preimplantation embryos of the essential components of the TGFbeta signalling pathway, TGFbeta receptors type I and II and the substrate proteins Smad 2 and 3. We found that both receptors, as well as Smad 2 and 3, were present in the unfertilized oocyte, whereas only the type I receptor and Smad 2 and 3 were present at the blastocyst stage. At the 4-cell and 8-cell stages neither of the receptors was present, but Smad 2 and 3 were present at both stages. These findings support our hypothesis that the TGFbeta1 in follicular fluid may interact with the oocyte and preimplantation embryo via TGFbeta receptors, and that TGFbeta signalling may be important for the development of the oocyte and the preimplantation embryo.

Mage-b4, a novel melanoma antigen (MAGE) gene specifically expressed during germ cell differentiation.

The MAGE genes were initially isolated from different kinds of tumors, and based on their virtually exclusive tumor-specific expression in adult tissues, they have been used as targets for cancer immunotherapy. However, although a large number of MAGE genes have now been identified and extensively studied in tumors of various origin, their functions in normal cells remain unknown. Here we describe the isolation and characterization of a novel murine MAGE homologue, Mage-b4. mRNA expression studies in a wide variety of adult and embryonic tissues revealed that Mage-b4 is specifically expressed in fetal and adult gonads. An antibody specific to Mage-b4 was developed, and using this antibody, we found that the Mage-b4 protein was confined to the cytoplasm of germ cells. Double-labeling experiments using antibodies against the meiosis-specific SCP3 protein and the Mage-b4 protein showed that Mage-b4 is down-regulated as the germ cells enter meiosis in adult testis. In contrast, Mage-b4 was expressed in female germ cells throughout meiosis, and the protein was also found in dormant primary oocytes.

Testatin: a cystatin-related gene expressed during early testis development.

To isolate genes involved in morphogenic aspects of testis development, and which may act in cell signaling pathways downstream of the testis-determining gene Sry, we have developed a modified mRNA differential display method named signal peptide differential display. It was used to target those genes that encode proteins having a signal peptide sequence. By using this method, we isolated a gene named testatin. This gene was found to be related to a group of genes that encodes cysteine protease inhibitors known as cystatins. Cystatins and their target proteases have been associated with tumor formation and metastasis, but also are involved in natural tissue remodeling events such as bone resorption and embryo implantation. We show that testatin expression is restricted to fetal gonads and adult testis. Furthermore, testatin is expressed during testis cord formation in pre-Sertoli cells, believed to be the site of Sry action, at a time immediately after the peak of Sry expression. This finding suggests that testatin might be activated by transcription factors that are known to orchestrate the early testis development pathway. This gene therefore represents one of the putative downstream targets likely to have an essential role in tissue reorganization during early testis development.

Temporal expression of platelet-derived growth factor (PDGF)-A and its receptor in human preimplantation embryos.

In order to improve assisted fertilization in humans it is important to elucidate the mechanisms of control of growth and development in the early pre-embryo. Increasing evidence shows that growth factors are of importance for such control mechanisms. As platelet-derived growth factor (PDGF) has been shown to enhance growth in a number of tissues, it may also be important in human pre-embryo development. PDGF acts as a dimer (AA, BB or AB) through its receptors: alpha alpha, beta beta and alpha beta. In order to study the role of PDGF and its receptors, we have used reverse transcription-polymerase chain reaction (RT-PCR) to examine the presence of transcripts in human pre-embryos that were surplus from the in-vitro fertilization treatment of infertile couples. Transcripts for PDGF A were present in the oocyte, 8-cell, morula and blastocyst stages but not in the 4-cell stage. Transcripts for PDGF B were not detected at any stage. PDGF receptor (PDGFR)-alpha transcripts were found in the 4-cell, 8-cell and blastocyst stages but not in the oocyte or morula stages. Transcripts for PDGFR-beta were detected from the 8-cell, morula and blastocyst stages but not in the oocyte or 4-cell stages. These results show that mRNA synthesis of both PDGF A and the two receptor subunits alpha and beta takes place from the 8-cell stage onwards, suggesting an autostimulatory pathway as a possible mechanism for growth factors during pre-embryo development.