Functional characterization of QT interval associated SCN5A enhancer variants identify combined additive effects.

Several empirical and theoretical studies suggest presence of multiple enhancers per gene that collectively regulate gene expression, and that common sequence variation impacting on the activities of these enhancers is a major source of inter-individual variability in gene expression. However, for vast majority of genes, enhancers and the underlying regulatory variation remains unknown. Even for the genes with well-characterized enhancers, the nature of the combined effects from multiple enhancers and their variants, when known, on gene expression regulation remains unexplored. Here, we have evaluated the combined effects from five SCN5A enhancers and their regulatory variants that are known to collectively correlate with SCN5A cardiac expression and underlie QT interval association in the general population. Using small deletions centered at the regulatory variants in episomal reporter assays in a mouse cardiomyocyte cell line we demonstrate that the variants and their flanking sequences play critical role in individual enhancer activities, likely being a transcription factor (TF) binding site. By performing oligonucleotide-based pulldown assays on predicted TFs we identify the TFs likely driving allele-specific enhancer activities. Using all 32 possible allelic synthetic constructs in reporter assays, representing the five biallelic enhancers in tandem in their genomic order, we demonstrate combined additive effects on overall enhancer activities. Using transient enhancer assays in developing zebrafish embryos we demonstrate the four out the five enhancer elements act as enhancers in vivo . Together, these studies extend the previous findings to uncover the TFs driving the enhancer activities of QT interval associated SCN5A regulatory variants, reveal the additive effects from allelic combinations of these regulatory variants, and prove their potential to act as enhancers in vivo .

Cardiac muscle-restricted partial loss of Nos1ap expression has limited but significant impact on electrocardiographic features.

Genome-wide association studies have identified sequence polymorphisms in a functional enhancer of the NOS1AP gene as the most common genetic regulator of QT interval and human cardiac NOS1AP gene expression in the general population. Functional studies based on in vitro overexpression in murine cardiomyocytes and ex vivo knockdown in zebrafish embryonic hearts, by us and others, have also demonstrated that NOS1AP expression levels can alter cellular electrophysiology. Here, to explore the role of NOS1AP in cardiac electrophysiology at an organismal level, we generated and characterized constitutive and heart muscle-restricted Nos1ap knockout mice to assess whether NOS1AP disruption alters the QT interval in vivo. Constitutive loss of Nos1ap led to genetic background-dependent variable lethality at or right before birth. Heart muscle-restricted Nos1ap knockout, generated using cardiac-specific alpha-myosin heavy chain promoter-driven tamoxifen-inducible Cre, resulted in tissue-level Nos1ap expression reduced by half. This partial loss of expression had no detectable effect on the QT interval or other electrocardiographic and echocardiographic parameters, except for a small but significant reduction in the QRS interval. Given that challenges associated with defining the end of the T wave on murine electrocardiogram can limit identification of subtle effects on the QT interval and that common noncoding NOS1AP variants are also associated with the QRS interval, our findings support the role of NOS1AP in regulation of the cardiac electrical cycle.

Spontaneously ruptured endometriomas presenting with symptoms and imaging findings worrisome for carcinomatosis: A case report.

INTRODUCTION AND IMPORTANCE: Endometriomas are the most common presenting subtype of endometriosis. Although most endometriomas are asymptomatic, patients can rarely present acutely with spontaneous rupture causing diffuse peritonitis and severe systemic inflammatory response. CASE PRESENTATION: Here we describe a case of ruptured endometriomas in a 26-year-old nulligravid female with a history of heavy menses, progressive abdominal distension, and a recent urinary tract infection. The patient presented to the emergency department with upper abdominal pain radiating to her back with associated nausea. Computed tomography (CT) scan demonstrated diffuse ascites with a large, multilobulated, and multicystic septated mass arising in the right pelvis and extending into the lower abdomen. Findings were concerning for peritoneal carcinomatosis and the patient was admitted for evaluation. She developed progressive signs of sepsis and was emergently brought to the operating room for surgical exploration on hospital day (HD) number two. She was found to have ruptured pelvic cysts arising from both ovaries with diffuse contamination of the abdomen by cyst contents and bilateral salpingo-oophorectomy (BSO) was performed. Final pathology demonstrated benign bilateral endometriomas. CLINICAL DISCUSSION: Endometrioma rupture is extremely rare and imaging findings may appear to represent disseminated peritoneal malignancy. CT findings demonstrating a pelvic mass with concurrent ascites should raise clinical suspicion for ruptured endometrioma, particularly in younger patients. CONCLUSION: Prompt surgical exploration and complete resection of pathologic tissue may be necessary for diagnosis and treatment in some patients with clinical deterioration related to perforated endometriomas. Combined oral contraceptives are recommended in the postoperative period.

Ruptured granulosa cell tumor of the ovary presenting with catastrophic intra-abdominal hemorrhage: A case report.

INTRODUCTION AND IMPORTANCE: Adult granulosa cell tumor (GCT) is a rare stromal cell neoplasm that most often arises from the ovary. Presenting symptoms are related to external compression of adjacent structures (mass effect) or secretion of hormones such as estrogen. Patients most commonly present with irregular menstruation, postmenopausal bleeding, and abdominal pain. Prolonged estrogen exposure can contribute to endometrial adenocarcinoma development in untreated patients. The highly vascular nature of GCTs can lead to hemorrhagic rupture in rare cases. PRESENTATION OF CASE: We describe a case of adult GCT in a 44-year-old female with a history of irregular menstrual bleeding and anemia. The patient presented with shortness of breath and abdominal pain. Computed tomography (CT) scan demonstrated possible hemorrhagic ascites of unclear etiology and a pelvic mass. The patient was brought to the operating room in hemorrhagic shock for surgical exploration where she was found to have active bleeding of a ruptured ovarian tumor for which she underwent left salpingo-oophorectomy. Postoperative course was unremarkable, and pathology demonstrated ruptured GCT. CLINICAL DISCUSSION: Although rare, ovarian tumors can present with massive bleeding following rupture. Granulosa cell tumors are surreptitious as they grow slowly, and symptoms such as distention, abdominal pain, and irregular vaginal bleeding are nonspecific. CONCLUSION: CT findings demonstrating a pelvic mass in the setting of spontaneous intra-abdominal bleeding should raise clinical suspicion, particularly in patients with histories of menstrual abnormalities. Patients with suspected intra-abdominal hemorrhage due to any cause are best treated by prompt surgical exploration and aggressive resuscitation.

Paediatric pneumonia research priorities in the context of COVID-19: A eDelphi study.

Background: Pneumonia remains the leading cause of infectious deaths in children under-five globally. We update the research priorities for childhood pneumonia in the context of the COVID-19 pandemic and explore whether previous priorities have been addressed. Methods: We conducted an eDelphi study from November 2019 to June 2021. Experts were invited to take part, targeting balance by: gender, profession, and high (HIC) and low- and middle-income countries (LMIC). We followed a three-stage approach: 1. Collating questions, using a list published in 2011 and adding newly posed topics; 2. Narrowing down, through participant scoring on importance and whether they had been answered; 3. Ranking of retained topics. Topics were categorized into: prevent and protect, diagnosis, treatment and cross-cutting. Results: Overall 379 experts were identified, and 108 took part. We started with 83 topics, and 81 further general and 40 COVID-19 specific topics were proposed. In the final ranking 101 topics were retained, and the highest ranked was to "explore interventions to prevent neonatal pneumonia". Among the top 20 topics, epidemiological research and intervention evaluation was commonly prioritized, followed by the operational and implementation research. Two COVID-19 related questions were ranked within the top 20. There were clear differences in priorities between HIC and LMIC respondents, and academics vs non-academics. Conclusions: Operational research on health system capacities, and evaluating optimized delivery of existing treatments, diagnostics and case management approaches are needed. This list should act as a catalyst for collaborative research, especially to meet the top priority in preventing neonatal pneumonia, and encourage multi-disciplinary partnerships.

Paediatric pneumonia research priorities in the context of COVID-19: An eDelphi study.

Background: Pneumonia remains the leading cause of infectious deaths in children under-five globally. We update the research priorities for childhood pneumonia in the context of the COVID-19 pandemic and explore whether previous priorities have been addressed. Methods: We conducted an eDelphi study from November 2019 to June 2021. Experts were invited to take part, targeting balance by: gender, profession, and high (HIC) and low- and middle-income countries (LMIC). We followed a three-stage approach: 1. Collating questions, using a list published in 2011 and adding newly posed topics; 2. Narrowing down, through participant scoring on importance and whether they had been answered; 3. Ranking of retained topics. Topics were categorized into: prevent and protect, diagnosis, treatment and cross-cutting. Results: Overall 379 experts were identified, and 108 took part. We started with 83 topics, and 81 further general and 40 COVID-19 specific topics were proposed. In the final ranking 101 topics were retained, and the highest ranked was to "explore interventions to prevent neonatal pneumonia". Among the top 20 topics, epidemiological research and intervention evaluation was commonly prioritized, followed by the operational and implementation research. Two COVID-19 related questions were ranked within the top 20. There were clear differences in priorities between HIC and LMIC respondents, and academics vs non-academics. Conclusions: Operational research on health system capacities, and evaluating optimized delivery of existing treatments, diagnostics and case management approaches are needed. This list should act as a catalyst for collaborative research, especially to meet the top priority in preventing neonatal pneumonia, and encourage multi-disciplinary partnerships.

Takeoff diversity in Diptera.

The order Diptera (true flies) are named for their two wings because their hindwings have evolved into specialized mechanosensory organs called halteres. Flies use halteres to detect body rotations and maintain stability during flight and other behaviours. The most recently diverged dipteran monophyletic subsection, the Calyptratae, is highly successful, accounting for approximately 12% of dipteran diversity, and includes common families like house flies. These flies move their halteres independently from their wings and oscillate their halteres during walking. Here, we demonstrate that this subsection of flies uses their halteres to stabilize their bodies during takeoff, whereas non-Calyptratae flies do not. We find that flies of the Calyptratae are able to take off more rapidly than non-Calyptratae flies without sacrificing stability. Haltere removal decreased both velocity and stability in the takeoffs of Calyptratae, but not other flies. The loss of takeoff velocity following haltere removal in Calyptratae (but not other flies) is a direct result of a decrease in leg extension speed. A closely related non-Calyptratae species (D. melanogaster) also has a rapid takeoff, but takeoff duration and stability are unaffected by haltere removal. Haltere use thus allows for greater speed and stability during fast escapes, but only in the Calyptratae clade.

Upfront boost Gamma Knife "leading-edge" radiosurgery to FLAIR MRI-defined tumor migration pathways in 174 patients with glioblastoma multiforme: a 15-year assessment of a novel therapy.

OBJECTIVE Glioblastoma multiforme (GBM) is composed of cells that migrate through the brain along predictable white matter pathways. Targeting white matter pathways adjacent to, and leading away from, the original contrast-enhancing tumor site (termed leading-edge radiosurgery [LERS]) with single-fraction stereotactic radiosurgery as a boost to standard therapy could limit the spread of glioma cells and improve clinical outcomes. METHODS Between December 2000 and May 2016, after an initial diagnosis of GBM and prior to or during standard radiation therapy and carmustine or temozolomide chemotherapy, 174 patients treated with radiosurgery to the leading edge (LE) of tumor cell migration were reviewed. The LE was defined as a region outside the contrast-enhancing tumor nidus, defined by FLAIR MRI. The median age of patients was 59 years (range 22-87 years). Patients underwent LERS a median of 18 days from original diagnosis. The median target volume of 48.5 cm3 (range 2.5-220.0 cm3) of LE tissue was targeted using a median dose of 8 Gy (range 6-14 Gy) at the 50% isodose line. RESULTS The median overall survival was 23 months (mean 43 months) from diagnosis. The 2-, 3-, 5-, 7-, and 10-year actual overall survival rates after LERS were 39%, 26%, 16%, 10%, and 4%, respectively. Nine percent of patients developed treatment-related imaging-documented changes due to LERS. Nineteen percent of patients were hospitalized for management of edema, 22% for resection of a tumor cyst or new tumor bulk, and 2% for shunting to treat hydrocephalus throughout the course of their disease. Of the patients still alive, Karnofsky Performance Scale scores remained stable in 90% of patients and decreased by 1-3 grades in 10% due to symptomatic treatment-related imaging changes. CONCLUSIONS LERS is a safe and effective upfront adjunctive therapy for patients with newly diagnosed GBM. Limitations of this study include a single-center experience and single-institution determination of the LE tumor target. Use of a leading-edge calculation algorithm will be described to achieve a consistent approach to defining the LE target for general use. A multicenter trial will further elucidate its value in the treatment of GBM.

Development of Cardiovascular Quality Indicators for Rheumatoid Arthritis: Results from an International Expert Panel Using a Novel Online Process.

OBJECTIVE: Patients with rheumatoid arthritis (RA) have a high risk of premature cardiovascular disease (CVD). We developed CVD quality indicators (QI) for screening and use in rheumatology clinics. METHODS: A systematic review was conducted of the literature on CVD risk reduction in RA and the general population. Based on the best practices identified from this review, a draft set of 12 candidate QI were presented to a Canadian panel of rheumatologists and cardiologists (n = 6) from 3 academic centers to achieve consensus on the QI specifications. The resulting 11 QI were then evaluated by an online modified-Delphi panel of multidisciplinary health professionals and patients (n = 43) to determine their relevance, validity, and feasibility in 3 rounds of online voting and threaded discussion using a modified RAND/University of California, Los Angeles Appropriateness Methodology. RESULTS: Response rates for the online panel were 86%. All 11 QI were rated as highly relevant, valid, and feasible (median rating ≥ 7 on a 1-9 scale), with no significant disagreement. The final QI set addresses the following themes: communication to primary care about increased CV risk in RA; CV risk assessment; defining smoking status and providing cessation counseling; screening and addressing hypertension, dyslipidemia, and diabetes; exercise recommendations; body mass index screening and lifestyle counseling; minimizing corticosteroid use; and communicating to patients at high risk of CVD about the risks/benefits of nonsteroidal antiinflammatory drugs. CONCLUSION: Eleven QI for CVD care in patients with RA have been developed and are rated as highly relevant, valid, and feasible by an international multidisciplinary panel.

Best practices for cardiovascular disease prevention in rheumatoid arthritis: a systematic review of guideline recommendations and quality indicators.

OBJECTIVE: Cardiovascular disease (CVD) is a leading cause of mortality in rheumatoid arthritis (RA). This study systematically reviewed and appraised guidelines and quality indicators (QIs) pertaining to CVD risk management in patients with RA. METHODS: Four electronic medical databases (Medline, Embase, CINAHL, and Web of Science) and gray literature publications were searched using terms and keywords pertaining to guidelines, QIs, RA, and CVD (RA and general population literature searched). Abstracts were screened for inclusion and rated using the Appraisal of Guidelines for Research and Evaluation II instrument independently by 2 of 3 reviewers. RESULTS: In total, 16,064 abstracts were screened and 808 underwent full-text review. A total of 17 guidelines and 3 QI sets published between 2008 and 2013 were included. A number of consistent themes emerged, including the increased CV risk faced by RA patients and the need to address modifiable risk factors on a regular basis. The role of the multidisciplinary team in risk optimization was also highlighted. Ten guidelines provided recommendations for CVD prevention in patients with RA. Unfortunately, most recommendations lacked the specificity required to determine adherence to the recommendation. Only 4 RA-specific CVD QIs were identified (1 general comorbidity screening, formal CVD risk estimation, exercise, and minimizing steroid use). CONCLUSION: Regular screening for CVD risk factors is an important part of care in patients with RA. Unfortunately, existing RA-specific CVD QIs do not adequately address risk factor management, and existing guideline recommendations lack specificity for measurement and use in quality improvement initiatives.