Homologous and heterologous desensitization of guanylyl cyclase-B signaling in GH3 somatolactotropes

The guanylyl cyclases, GC-A and GC-B, are selective receptors for atrial and C-type natriuretic peptides (ANP and CNP, respectively). In the anterior pituitary, CNP and GC-B are major regulators of cGMP production in gonadotropes and yet mouse models of disrupted CNP and GC-B indicate a potential role in growth hormone secretion. In the current study, we investigate the molecular and pharmacological properties of the CNP/GC-B system in somatotrope lineage cells. Primary rat pituitary and GH3 somatolactotropes expressed functional GC-A and GC-B receptors that had similar EC50 properties in terms of cGMP production. Interestingly, GC-B signaling underwent rapid homologous desensitization in a protein phosphatase 2A (PP2A)-dependent manner. Chronic exposure to either CNP or ANP caused a significant down-regulation of both GC-A- and GC-B-dependent cGMP accumulation in a ligand-specific manner. However, this down-regulation was not accompanied by alterations in the sub-cellular localization of these receptors. Heterologous desensitization of GC-B signaling occurred in GH3 cells following exposure to either sphingosine-1-phosphate or thyrotrophin-releasing hormone (TRH). This heterologous desensitization was protein kinase C (PKC)-dependent, as pre-treatment with GF109203X prevented the effect of TRH on CNP/GC-B signaling. Collectively, these data indicate common and distinct properties of particulate guanylyl cyclase receptors in somatotropes and reveal that independent mechanisms of homologous and heterologous desensitization occur involving either PP2A or PKC. Guanylyl cyclase receptors thus represent potential novel therapeutic targets for treating growth-hormone-associated disorders

A Functional Screen Provides Evidence for a Conserved, Regulatory, Juxtamembrane Phosphorylation Site in Guanylyl Cyclase A and B

Kinase homology domain (KHD) phosphorylation is required for activation of guanylyl cyclase (GC)-A and -B. Phosphopeptide mapping identified multiple phosphorylation sites in GC-A and GC-B, but these approaches have difficulty identifying sites in poorly detected peptides. Here, a functional screen was conducted to identify novel sites. Conserved serines or threonines in the KHDs of phosphorylated receptor GCs were mutated to alanine and tested for reduced hormone to detergent activity ratios. Mutation of Ser-489 in GC-B to alanine but not glutamate reduced the activity ratio to 60% of wild type (WT) levels. Similar results were observed with Ser-473, the homologous site in GC-A. Receptors containing glutamates for previously identified phosphorylation sites (GC-A-6E and GC-B-6E) were activated to ∼20% of WT levels but the additional glutamate substitution for S473 or S489 increased activity to near WT levels. Substrate-velocity assays indicated that GC-B-WT-S489E and GC-B-6E-S489E had lower Km values and that WT-GC-B-S489A, GC-B-6E and GC-B-6E-S489A had higher Km values than WT-GC-B. Homologous desensitization was enhanced when GC-A contained the S473E substitution, and GC-B-6E-S489E was resistant to inhibition by a calcium elevating treatment or protein kinase C activation – processes that dephosphorylate GC-B. Mass spectrometric detection of a synthetic phospho-Ser-473 containing peptide was 200–1300-fold less sensitive than other phosphorylated peptides and neither mass spectrometric nor 32PO4 co-migration studies detected phospho-Ser-473 or phospho-Ser-489 in cells. We conclude that Ser-473 and Ser-489 are Km-regulating phosphorylation sites that are difficult to detect using current methods

The Implementation and Evaluation of a Media Literacy Intervention About PAES Use in Sport Science Students

With respect to both competitive and amateur/fitness sports, media may strongly influence young people’s opinions and behaviors concerning the use of PAES (Performance and Appearance Enhancing Substances). The present investigation addressed this topic by focusing on sport sciences students’ beliefs concerning the possible role of media related to the implementation and evaluation of a PAES-focus media literacy intervention conducted with sport science students. This study relied on a sample of 521 students (attrition rate 10.3%) (45.1% female, mean age = 22.6, SD = 2.20), which provided baseline data on students’ levels of media literacy concerning the use of PAES (i.e. “descriptive sample”), and a sample of 248 students, who participated in and provided data on the media literacy intervention. This latter sample included a group of 128 students (44.5% female, mean age = 23.03, SD = 3.76) who actively participated in the intervention (i.e. “intervention group”), and a group of 120 students who did not (i.e. “control group”, 53.3% female, mean age = 22.25, SD = 2.47). All students filled out media literacy questionnaires targeting students’ awareness of media influence, their views about the realism of media content, their sense of confidence in dealing with media messages, and their positive attitudes toward PAES use. Analyses of questionnaire data showed that students are relatively aware of media influence on people’s views and behaviors with respect to PAES use. At the same time, students also believed that young people do not consider media as “realistic sources” of information; nonetheless, they also did not consider themselves entirely capable of dealing effectively with media messages. With respect to the intervention, students overall appreciated and greatly welcomed the educational program on media literacy, and the analyses of intervention data across intervention and control groups showed that key media literacy variables changed over time, attesting to the overall effectiveness of the intervention.Ministry of Health, Italy CUPB86G17000660005 2017-

How Much Do Focal Infarcts Distort White Matter Lesions and Global Cerebral Atrophy Measures?

BACKGROUND: White matter lesions (WML) and brain atrophy are important biomarkers in stroke and dementia. Stroke lesions, either acute or old, symptomatic or silent, are common in older people. Such stroke lesions can have similar signals to WML and cerebrospinal fluid (CSF) on magnetic resonance (MR) images, and may be classified accidentally as WML or CSF by MR image processing algorithms, distorting WML and brain atrophy volume from the true volume. We evaluated the effect that acute or old stroke lesions at baseline, and new stroke lesions occurring during follow-up, could have on measurement of WML volume, cerebral atrophy and their longitudinal progression. METHODS: We used MR imaging data from patients who had originally presented with acute lacunar or minor cortical ischaemic stroke symptoms, recruited prospectively, who were scanned at baseline and about 3 years later. We measured WML and CSF volumes (ml) semi-automatically. We manually outlined the acute index stroke lesion (ISL), any old stroke lesions present at baseline, and new lesions appearing de novo during follow-up. We compared baseline and follow-up WML volume, cerebral atrophy and their longitudinal progression excluding and including the acute ISL, old and de novo stroke lesions. A non-parametric test (Wilcoxon's signed rank test) was used to compare the effects. RESULTS: Among 46 patients (mean age 72 years), 33 had an ISL visible on MR imaging (median volume 2.05 ml, IQR 0.88–8.88) and 7 of the 33 had old lacunes at baseline: WML volume was 8.54 ml (IQR 5.86–15.80) excluding versus 10.98 ml (IQR 6.91–24.86) including ISL (p < 0.001). At follow-up, median 39 months later (IQR 30–45), 3 patients had a de novo stroke lesion; total stroke lesion volume had decreased in 11 and increased in 22 patients: WML volume was 12.17 ml (IQR 8.54–19.86) excluding versus 14.79 ml (IQR 10.02–38.03) including total stroke lesions (p < 0.001). Including/excluding lacunes at baseline or follow-up also made small differences. Twenty-two of the 33 patients had tissue loss due to stroke lesions between baseline and follow-up, resulting in a net median brain tissue volume loss (i.e. atrophy) during follow-up of 24.49 ml (IQR 12.87–54.01) excluding versus 24.61 ml (IQR 15.54–54.04) including tissue loss due to stroke lesions (p < 0.001). Including stroke lesions in the WML volume added substantial noise, reduced statistical power, and thus increased sample size estimated for a clinical trial. CONCLUSIONS: Failure to exclude even small stroke lesions distorts WML volume, cerebral atrophy and their longitudinal progression measurements. This has important implications for design and sample size calculations for observational studies and randomised trials using WML volume, WML progression or brain atrophy as outcome measures. Improved methods of discriminating between stroke lesions and WML, and between tissue loss due to stroke lesions and true brain atrophy are required

The association between farming activities, precipitation, and the risk of acute gastrointestinal illness in rural municipalities of Quebec, Canada: a cross-sectional study

<p>Abstract</p> <p>Background</p> <p>Increasing livestock density and animal manure spreading, along with climate factors such as heavy rainfall, may increase the risk of acute gastrointestinal illness (AGI). In this study we evaluated the association between farming activities, precipitation and AGI.</p> <p>Methods</p> <p>A cross-sectional telephone survey of randomly selected residents (n = 7006) of 54 rural municipalities in Quebec, Canada, was conducted between April 2007 and April 2008. AGI symptoms and several risk factors were investigated using a phone questionnaire. We calculated the monthly prevalence of AGI, and used multivariate logistic regression, adjusting for several demographic and risk factors, to evaluate the associations between AGI and both intensive farming activities and cumulative weekly precipitation. Cumulative precipitation over each week, from the first to sixth week prior to the onset of AGI, was analyzed to account for both the delayed effect of precipitation on AGI, and the incubation period of causal pathogens. Cumulative precipitation was treated as a four-category variable: high (≥90^thpercentile), moderate (50^thto <90^thpercentile), low (10^thto <50^thpercentile), and very low (<10^thpercentile) precipitation.</p> <p>Results</p> <p>The overall monthly prevalence of AGI was 5.6% (95% CI 5.0%-6.1%), peaking in winter and spring, and in children 0-4 years old. Living in a territory with intensive farming was negatively associated with AGI: adjusted odds ratio (OR) = 0.70 (95% CI 0.51-0.96). Compared to low precipitation periods, high precipitation periods in the fall (September, October, November) increased the risk of AGI three weeks later (OR = 2.20; 95% CI 1.09-4.44) while very low precipitation periods in the summer (June, July, August) increased the risk of AGI four weeks later (OR = 2.19; 95% CI 1.02-4.71). Further analysis supports the role of water source on the risk of AGI.</p> <p>Conclusions</p> <p>AGI poses a significant burden in Quebec rural municipalities with a peak in winter. Intensive farming activities were found to be negatively associated with AGI. However, high and very low precipitation levels were positively associated with the occurrence of AGI, especially during summer and fall. Thus, preventive public health actions during such climate events may be warranted.</p

Eosinophilic esophagitis

Eosinophilic esophagitis (EoE) is an atopic condition of the esophagus that has become increasingly recognized over the last decade. Diagnosis of the disorder is dependent on the patient’s clinical manifestations and histologic findings on esophageal mucosal biopsies. Patients with eosinophilic esophagitis should be referred to both an allergist and gastroenterologist for optimal management, which may include dietary modifications, pharmacologic agents such as corticosteroids, leukotriene modifiers and biologics as well as mechanical dilatation of the esophagus. The epidemiology, pathophysiology, diagnosis, treatment, and prognosis of EoE are discussed in this review

Natriuretic peptide activation of extracellular regulated kinase 1/2 (ERK1/2) pathway by particulate guanylyl cyclases in GH3 somatolactotropes.

The natriuretic peptides, Atrial-, B-type and C-type natriuretric peptides (ANP, BNP, CNP), are regulators of many endocrine tissues and exert their effects predominantly through the activation of their specific guanylyl cyclase receptors (GC-A and GC-B) to generate cGMP. Whereas cGMP-independent signalling has been reported in response to natriuretic peptides, this is mediated via either the clearance receptor (Npr-C) or a renal-specific NPR-Bi isoform, which both lack intrinsic guanylyl cyclase activity. Here, we report evidence of GC-B-dependent cGMP-independent signalling in pituitary GH3 cells. Stimulation of GH3 cells with CNP resulted in a rapid and sustained enhancement of ERK1/2 phosphorylation (P-ERK1/2), an effect that was not mimicked by dibutryl-cGMP. Furthermore, CNP-stimulated P-ERK1/2 occurred at concentrations below that required for cGMP accumulation. The effect of CNP on P-ERK1/2 was sensitive to pharmacological blockade of MEK (U0126) and Src kinases (PP2). Silencing of the GC-B1 and GC-B2 splice variants of the GC-B receptor by using targeted short interfering RNAs completely blocked the CNP effects on P-ERK1/2. CNP failed to alter GH3 cell proliferation or cell cycle distribution but caused a concentration-dependent increase in the activity of the human glycoprotein α-subunit promoter (αGSU) in a MEK-dependent manner. Finally, CNP also activated the p38 and JNK MAPK pathways in GH3 cells. These findings reveal an additional mechanism of GC-B signalling and suggest additional biological roles for CNP in its target tissues

Specific orofacial problems experienced by musicians

Background: Patients who play musical instruments (especially wind and stringed instruments) and vocalists are prone to particular types of orofacial problems. Some problems are caused by playing and some are the result of dental treatment. This paper proposes to give an insight into these problems and practical guidance to general practice dentists. Method: Information in this paper is gathered from studies published in dental, music and occupational health journals, and from discussions with career musicians and music teachers. Results: Orthodontic problems, soft tissue trauma, focal dystonia, denture retention, herpes labialis, dry mouth and temporomandibular joint (TMJ) disorders were identified as orofacial problems of career musicians. Options available for prevention and palliative treatment as well as instrument selection are suggested to overcome these problems. Conclusions: Career musicians express reluctance to attend dentists who are not sensitive to their specific needs. General practitioner dentists who understand how the instruments impact on the orofacial structures and are aware of potential problems faced by musicians are able to offer preventive advice and supportive treatment to these patients, especially those in the early stages of their career

The role of PET/CT in detection of gastric cancer recurrence

<p>Abstract</p> <p>Background</p> <p>In the course of surveillance of gastric cancer recurrence after curative resection, contrast CT scan is used in general. However, new findings from CT scan are not always confirmatory for the recurrence. In this case, we usually use short-term follow up strategy or therapeutic intervention with clinical decision. Recently, the use of fusion Positron Emission Tomography/Computed Tomography (PET/CT) is increasing. The purpose of this study is to evaluate the efficacy and usefulness of PET/CT for detecting recurrence of gastric cancer after curative resection.</p> <p>Methods</p> <p>Fifty two patients who received curative resection of gastric cancer and had undergone PET/CT and contrast CT for surveillance of recurrence until Dec 2006 in Seoul National University Hospital were analyzed retrospectively. Recurrence of gastric cancer was validated by histologic confirmation (n = 17) or serial contrast CT follow up with at least 5 month interval (n = 35). McNemar's test and Fisher's exact test were used to evaluate sensitivity and specificity of PET/CT and contrast CT.</p> <p>Results</p> <p>Of 52 patients, 38 patients were confirmed as recurrence. The sensitivity was 68.4% (26/38) for PET/CT and 89.4% (34/38) for contrast CT (p = 0.057). The specificity was 71.4% (10/14) and 64.2% (9/14), respectively (p = 1.0). In terms of the recurred sites, the sensitivity and specificity of PET/CT were similar to those of contrast CT in all sites except peritoneum. Contrast CT was more sensitive than PET/CT (p = 0.039) for detecting peritoneal seeding. Additional PET/CT on contrast CT showed no further increase of positive predictive value regardless of sites. Among 13 patients whose image findings between two methods were discordant and tissue confirmation was difficult, the treatment decision was made in 7 patients based on PET/CT, showing the final diagnostic accuracy of 42.8% (3/7).</p> <p>Conclusion</p> <p>PET/CT was as sensitive and specific as contrast CT in detection of recurred gastric cancer except peritoneal seeding. However, additional PET/CT on contrast CT did not increase diagnostic accuracy in detection of recurred gastric cancer. Further studies are warranted to validate the role of PET/CT in detection of gastric cancer recurrence.</p