Control of protein signaling using a computationally designed GTPase/GEF orthogonal pair.

Signaling pathways depend on regulatory protein-protein interactions; controlling these interactions in cells has important applications for reengineering biological functions. As many regulatory proteins are modular, considerable progress in engineering signaling circuits has been made by recombining commonly occurring domains. Our ability to predictably engineer cellular functions, however, is constrained by complex crosstalk observed in naturally occurring domains. Here we demonstrate a strategy for improving and simplifying protein network engineering: using computational design to create orthogonal (non-crossreacting) protein-protein interfaces. We validated the design of the interface between a key signaling protein, the GTPase Cdc42, and its activator, Intersectin, biochemically and by solving the crystal structure of the engineered complex. The designed GTPase (orthoCdc42) is activated exclusively by its engineered cognate partner (orthoIntersectin), but maintains the ability to interface with other GTPase signaling circuit components in vitro. In mammalian cells, orthoCdc42 activity can be regulated by orthoIntersectin, but not wild-type Intersectin, showing that the designed interaction can trigger complex processes. Computational design of protein interfaces thus promises to provide specific components that facilitate the predictable engineering of cellular functions.

Transpleural pulmonary-to-systemic venous collaterals in a case with obstructed scimitar vein.

Scimitar syndrome is a rare cause of left-to-right shunting. Surgery is indicated for a pulmonary-to-systemic blood flow ratio greater than 1.5:1 and not infrequently is complicated by postoperative obstruction. This report presents a case of scimitar syndrome and reviews how magnetic resonance imaging (MRI) can be used for initial and follow-up assessment of the syndrome with emphasis on suspected pulmonary venous obstruction. Given the potential high incidence of postoperative occlusion, MRI provides hemodynamic and anatomic information for both initial and follow-up assessment of scimitar syndrome. MRI clearly demonstrated transpleural pulmonary-to-systemic venous collaterals draining the obstructed scimitar vein.

Identification and Evaluation of Controlled Trials in Pediatric Cardiology: Crowdsourced Scoping Review and Creation of Accessible Searchable Database.

Cardiac disease in children is associated with significant morbidity and mortality as well as increased health resource utilisation. There is a perception that there is a paucity of high-quality studies, particularly randomized controlled trials (RCTs), in the field of pediatric cardiology. We sought to identify, examine, and map the range of RCTs conducted in children with cardiac conditions, including the development of a searchable open-access database. A literature search was conducted encompassing MEDLINE, EMBASE, and the Cochrane Central Register of Controlled Trials from inception to 2018. All English-language RCTs enrolling children (age 0-21 years) with cardiac conditions were included. Data extraction and risk of bias assessments were performed in duplicate via crowdsourcing for each eligible study and entered into an online database. A total of 933 RCTs met eligibility criteria. Median trial recruitment was 49 patients (interquartile range 30-86) with 18.9% of studies (n = 176) including > 100 patients. A wide variety of populations and interventions were encompassed with congenital heart disease (79.8% of RCTs) and medications (63.3% of RCTs) often studied. Just over one-half of the trials (53.4%) clearly identified a primary outcome, and fewer than half (46.6%) fully documented a robust randomization process. Trials were summarised in a searchable online database (https://pediatrics.knack.com/cardiology-rct-database#cardiology-rcts/). Contrary to a commonly held perception, there are nearly 1,000 published RCTs in pediatric cardiology. The open-access database created as part of this project provides a resource that facilitates an efficient comprehensive review of the literature for clinicians and researchers caring for children with cardiac issues.

Implantation of drug-eluting stents for relief of obstructed infra-cardiac totally anomalous pulmonary venous connection in isomerism of the right atrial appendages.

We describe an infant with severe obstruction of infra-cardiac totally anomalous pulmonary venous connection associated with right isomerism, atrioventricular septal defect, pulmonary atresia, and multiple aortopulmonary collateral arteries. Implantation of a stent into the obstructed descending vertical vein provided effective palliation, with a dramatic increase in saturations of oxygen obviating the need for urgent high-risk surgery.

Retroesophageal brachiocephalic artery assessed by magnetic resonance imaging: a case report and literature review.

A right aortic arch with a retroesophageal brachiocephalic artery is a very rare type of vascular ring. We present a case report along with a review of published cases to date. Twelve publications from 1968-2014 describe a retroesophageal brachiocephalic artery in a total of 13 patients. The mean age at diagnosis was 8.7 ± 16 years. Four of the 13 patients were boys. Nearly half of the patients were symptomatic, with dysphagia or respiratory difficulties. Ten patients (77%) had associated congenital heart defects. Of the 13 patients with retroesophageal brachiocephalic artery, 12 had a right aortic arch, and only 1 had a left aortic arch associated with complex congenital heart disease. Investigations used in the diagnosis of the vascular ring include angiography, esophagography, echocardiography, and computed tomography. Only 2 patients had the diagnosis confirmed with magnetic resonance imaging (MRI), but this was in the setting of complex congenital heart disease. In conclusion, a right aortic arch with a retroesophageal left brachiocephalic artery is an extremely rare form of vascular ring and is often seen in association with other forms of congenital heart disease. Cardiac MRI can be used as a primary diagnostic modality for both the assessment of the vascular ring anatomy and the hemodynamics of associated cardiac malformations in the preoperative setting.

Changes in systemic and pulmonary blood flow distribution in normal adult volunteers in response to posture and exercise: a phase contrast magnetic resonance imaging study.

Hemodynamics are usually evaluated in the supine position at rest. This is only a snapshot of an individual's daily activities. This study describes circulatory adaptation, as assessed by magnetic resonance imaging, to changes in position and exercise. Phase contrast magnetic resonance imaging of blood flow within systemic and pulmonary arteries and veins was performed in 24 healthy volunteers at rest in the prone and supine position and with bicycle exercise in the supine position. No change was seen in systemic blood flow when moving from prone to supine. Exercise resulted in an increased percentage of cardiac output towards the lower body. Changes in position resulted in a redistribution of blood flow within the left lung--supine positioning resulted in decreased blood flow to the left lower pulmonary vein. With exercise, both the right and left lower lobes received increased blood flow, while the upper lobes received less.

Effectiveness of serial increases in amino-terminal pro-B-type natriuretic peptide levels to indicate the need for mechanical circulatory support in children with acute decompensated heart failure.

We sought to determine prospectively whether serial assessment of the natriuretic peptide prohormone, amino-terminal pro-B-type natriuretic peptide (NT-pro-BNP), correlated with clinical severity and outcomes in children hospitalized for acute decompensated heart failure (ADHF). Patients (>1 month of age) admitted from 2005 to 2007 with ADHF requiring intravenous vasoactive/diuretic therapy for ADHF were eligible. Serum NT-pro-BNP levels were obtained within 24 hours of admission and at prespecified intervals, and clinical caregivers were blinded to these levels. End points included hospital discharge, death or cardiac transplantation, and care escalation including the need for mechanical circulatory support (MCS) was noted. Twenty-four patients were enrolled: 22 survived to hospital discharge and 2 died. Ten required MCS (of which 6 underwent cardiac transplantation). Two patients underwent transplantation without MCS. For the entire cohort, NT-pro-BNP levels peaked at days 2 to 3 after admission, with a subsequent gradual decrease until discharge. However, for those who did require MCS, NT-pro-BNP failed to decrease consistently until after MCS initiation. At discharge, NT-pro-BNP levels were significantly decreased from admission levels but remained well above normal for all patients. Single-point NT-pro-BNP levels on admission did not correlate with independently assessed clinical scores of heart failure severity or predict the need for MCS in this cohort. In conclusion, serial NT-pro-BNP levels demonstrated an incremental trend after 48 hours in patients who went on to require MCS but decreased in all other patients and may therefore assist the decision to initiate or avoid MCS after admission for pediatric ADHF.

Mechanism of actin network attachment to moving membranes: barbed end capture by N-WASP WH2 domains.

Actin filament networks exert protrusive and attachment forces on membranes and thereby drive membrane deformation and movement. Here, we show that N-WASP WH2 domains play a previously unanticipated role in vesicle movement by transiently attaching actin filament barbed ends to the membrane. To dissect the attachment mechanism, we reconstituted the propulsive motility of lipid-coated glass beads, using purified soluble proteins. N-WASP WH2 mutants assembled actin comet tails and initiated movement, but the comet tails catastrophically detached from the membrane. When presented on the surface of a lipid-coated bead, WH2 domains were sufficient to maintain comet tail attachment. In v-Src-transformed fibroblasts, N-WASP WH2 mutants were severely defective in the formation of circular podosome arrays. In addition to creating an attachment force, interactions between WH2 domains and barbed ends may locally amplify signals for dendritic actin nucleation.