The rapid dissemination of the 2009 pandemic H1N1 influenza virus emphasizes the need for universal influenza vaccines that would broadly protect against multiple mutated strains. Recent efforts have ...focused on the highly conserved hemagglutinin (HA) stem domain, which must undergo a significant conformational change for effective viral infection. Although the production of isolated domains of multimeric ectodomain proteins has proven difficult, we report a method to rapidly produce the properly folded HA stem domain protein from influenza virus A/California/05/2009 (H1N1) by using Escherichia coli -based cell-free protein synthesis and a simple refolding protocol. The T4 bacteriophage fibritin foldon placed at the C terminus of the HA stem domain induces trimer formation. Placing emphasis on newly exposed protein surfaces, several hydrophobic residues were mutated, two polypeptide segments were deleted, and the number of disulfide bonds in each monomer was reduced from four to two. High pH and Brij 35 detergent emerged as the most beneficial factors for improving the refolding yield. To stabilize the trimer of the HA stem-foldon fusion, new intermolecular disulfide bonds were finally introduced between foldon monomers and between stem domain monomers. The correct immunogenic conformation of the stabilized HA stem domain trimer was confirmed by using antibodies CR6261, C179, and FI6 that block influenza infection by binding to the HA stem domain trimer. These results suggest great promise for a broadly protective vaccine and also demonstrate a unique approach for producing individual domains of complex multimeric proteins.
IMPORTANCE: Publicly available data sets hold much potential, but their unique design may require specific analytic approaches. OBJECTIVE: To determine adherence to appropriate research practices for ...a frequently used large public database, the National Inpatient Sample (NIS) of the Agency for Healthcare Research and Quality (AHRQ). DESIGN, SETTING, AND PARTICIPANTS: In this observational study of the 1082 studies published using the NIS from January 2015 through December 2016, a representative sample of 120 studies was systematically evaluated for adherence to practices required by AHRQ for the design and conduct of research using the NIS. EXPOSURES: None. MAIN OUTCOMES AND MEASURES: All studies were evaluated on 7 required research practices based on AHRQ’s recommendations and compiled under 3 domains: (1) data interpretation (interpreting data as hospitalization records rather than unique patients); (2) research design (avoiding use in performing state-, hospital-, and physician-level assessments where inappropriate; not using nonspecific administrative secondary diagnosis codes to study in-hospital events); and (3) data analysis (accounting for complex survey design of the NIS and changes in data structure over time). RESULTS: Of 120 published studies, 85% (n = 102) did not adhere to 1 or more required practices and 62% (n = 74) did not adhere to 2 or more required practices. An estimated 925 (95% CI, 852-998) NIS publications did not adhere to 1 or more required practices and 696 (95% CI, 596-796) NIS publications did not adhere to 2 or more required practices. A total of 79 sampled studies (68.3% 95% CI, 59.3%-77.3%) among the 1082 NIS studies screened for eligibility did not account for the effects of sampling error, clustering, and stratification; 62 (54.4% 95% CI, 44.7%-64.0%) extrapolated nonspecific secondary diagnoses to infer in-hospital events; 45 (40.4% 95% CI, 30.9%-50.0%) miscategorized hospitalizations as individual patients; 10 (7.1% 95% CI, 2.1%-12.1%) performed state-level analyses; and 3 (2.9% 95% CI, 0.0%-6.2%) reported physician-level volume estimates. Of 27 studies (weighted; 218 studies 95% CI, 134-303) spanning periods of major changes in the data structure of the NIS, 21 (79.7% 95% CI, 62.5%-97.0%) did not account for the changes. Among the 24 studies published in journals with an impact factor of 10 or greater, 16 (67%) did not adhere to 1 or more practices, and 9 (38%) did not adhere to 2 or more practices. CONCLUSIONS AND RELEVANCE: In this study of 120 recent publications that used data from the NIS, the majority did not adhere to required practices. Further research is needed to identify strategies to improve the quality of research using the NIS and assess whether there are similar problems with use of other publicly available data sets.
Electrical synapses are formed by gap junctions and permit electrical coupling, which shapes the synchrony of neuronal ensembles. Here, we provide a direct demonstration of receptor-mediated ...strengthening of electrical coupling in mammalian brain. Electrical coupling in the inferior olive of rats was strengthened by activation of NMDA-type glutamate receptors (NMDARs), which were found at synaptic loci and at extrasynaptic loci 20–100 nm proximal to gap junctions. Electrical coupling was strengthened by pharmacological and synaptic activation of NMDARs, whereas costimulation of ionotropic non-NMDAR glutamate receptors transiently antagonized the effect of NMDAR activation. NMDAR-dependent strengthening (1) occurred despite increased input conductance, (2) induced Ca2+-influx microdomains near dendritic spines, (3) required activation of the Ca2+/calmodulin-dependent protein-kinase II, (4) was restricted to neurons that were weakly coupled, and (5) thus strengthened coupling, mainly between nonadjacent neurons. This provided a mechanism to expand the synchronization of rhythmic membrane potential oscillations by chemical neurotransmitter input.
•Receptor-mediated augmentation of electrical synapses was explored in mammalian brain•Extrasynaptic NMDA receptors were found adjacent to gap junctions•NMDA receptor activation strengthened weak electrical coupling via CaMKII activation•Glutamate neurotransmission can expand synchronized activity by this mechanism
Turecek et al. demonstrate NMDA receptor-mediated strengthening of electrical synapses in the inferior olive of the mammalian brain. Their work reveals synaptic and intradendritic mechanisms by which synchronized neuronal activity can be expanded by glutamatergic neurotransmission.
In this study involving patients with type 1 diabetes, sensor-augmented insulin-pump therapy plus automated insulin suspension when glucose dropped below 70 mg per deciliter reduced nocturnal ...hypoglycemia, without affecting glycated hemoglobin values.
Severe nocturnal hypoglycemia can be catastrophic,
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and hypoglycemia remains one of the most formidable barriers to improving glycemic control in patients with diabetes.
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Sensor-augmented insulin-pump therapy offers substantial glycemic benefits, as compared with multiple daily insulin injections, but has not been shown to lower the risk of severe hypoglycemia significantly.
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The automatic suspension of insulin delivery when a preset sensor glucose threshold is reached has the potential to mitigate hypoglycemia. The low-glucose suspend feature, available in the Medtronic Paradigm Veo pump outside the United States since 2009, was used in this study in the intervention group; the feature allows . . .
Aside from the established role for platelets in regulating hemostasis and thrombosis, recent research has revealed a discrete role for platelets in the separation of the blood and lymphatic vascular ...systems. Platelets are activated by interaction with lymphatic endothelial cells at the lymphovenous junction, the site in the body where the lymphatic system drains into the blood vascular system, resulting in a platelet plug that, with the lymphovenous valve, prevents blood from entering the lymphatic circulation. This process, known as “lymphovenous hemostasis,” is mediated by activation of platelet CLEC-2 receptors by the transmembrane ligand podoplanin expressed by lymphatic endothelial cells. Lymphovenous hemostasis is required for normal lymph flow, and mice deficient in lymphovenous hemostasis exhibit lymphedema and sometimes chylothorax phenotypes indicative of lymphatic insufficiency. Unexpectedly, the loss of lymph flow in these mice causes defects in maturation of collecting lymphatic vessels and lymphatic valve formation, uncovering an important role for fluid flow in driving endothelial cell signaling during development of collecting lymphatics. This article summarizes the current understanding of lymphovenous hemostasis and its effect on lymphatic vessel maturation and synthesizes the outstanding questions in the field, with relationship to human disease.
The MiniMed 640G sensor-augmented insulin pump system (Medtronic, Inc., Northridge, CA) can automatically suspend insulin delivery in advance of predicted hypoglycemia and restart it upon recovery. ...The aims of this analysis were to determine the rate at which predicted hypoglycemia was avoided with this strategy, as well as to assess user acceptance of the system and its insulin management features.
Forty subjects with type 1 diabetes used the system for 4 weeks. We retrospectively evaluated performance of the system, using downloaded pump and sensor data, and evaluated user acceptance via questionnaires.
There were 2,322 suspend before low events (2.1 per subject-day). The mean (± SD) duration of pump suspension events was 56.4 ± 9.6 min, and the mean subsequent sensor glucose (SG) nadir was 71.8 ± 5.2 mg/dL. SG values following 1,930 (83.1%) of the predictive suspensions did not reach the preset low limit. Nadir SG values of ≤50 and ≤60 mg/dL were seen in 207 (8.9%) and 356 (15.3%) of the predictive suspensions, respectively. Blood glucose (BG) and SG values before and during the study were comparable (P > 0.05). The mean absolute relative difference between paired SG and BG values was 10.9 ± 13.8%. Subjects felt confident using the system, agreed that it helped protect them from hypoglycemia, and wished to continue using it.
Automatic insulin pump suspension as implemented in the MiniMed 640G system can help patients avoid hypoglycemia, without significantly increasing hyperglycemia.
In this randomized trial, sensor-augmented pump therapy was compared with a regimen of multiple daily insulin injections in adults and children with inadequately controlled type 1 diabetes. Pump ...therapy was associated with a significant improvement in glycated hemoglobin levels, as compared with injection therapy.
Improved glycemic control can reduce the microvascular and macrovascular complications associated with type 1 diabetes mellitus,
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and diabetes practitioners are continuously challenged to optimize glucose control while minimizing severe hypoglycemia and weight gain. Insulin pumps and systems for continuous glucose monitoring represent technologies designed to assist patients with type 1 diabetes in safely reaching glycemic goals. Among adults, the use of an insulin pump has been shown to reduce glycated hemoglobin levels without an increased risk of hypoglycemia, as compared with a regimen of multiple daily insulin injections, but results in children have been inconsistent.
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Recent studies have suggested . . .
Background:
We investigated wearable components of the Dexcom G6 continuous glucose monitoring (CGM) System in simulated therapeutic and diagnostic radiologic procedures.
Methods:
G6 transmitters ...were loaded with simulated glucose data and attached to sensors. Sets of sensor/transmitter pairs were exposed to x-rays to simulate a radiotherapeutic procedure and to radiofrequency (RF) and magnetic fields to simulate diagnostic magnetic resonance imaging (MRI). The x-ray simulation provided a cumulative dose of 80 Gy. The MRI simulation used RF fields oscillating at 64 or 128 MHz and magnetic fields of 1.5 or 3 T. During the MRI simulation, displacement force, induced heating, and induced currents were measured. After the simulations, bench tests were used to assess data integrity on the transmitters and responsiveness of sensors to various concentrations of aqueous glucose.
Results:
Glucose concentrations reported by sensor/transmitter pairs after undergoing x-irradiation or a simulated MRI exam were similar to those from control (unexposed) devices. During the 3 T MRI simulation, the devices experienced a displacement force of 306 g, which was insufficient to dislodge the sensor/transmitter from the substrate, RF-induced heating of <2°C, and an induced current of <16 pA. Data stored on the transmitters prior to the MRI simulation remained intact.
Conclusion:
Wearable components of the G6 CGM System retain basic functionality and data integrity after exposure to simulated therapeutic and diagnostic radiologic procedures. The devices are unlikely to be affected by x-irradiation used in typical imaging studies. Simulated MRI procedures create displacement force, minimal heating, and current in sensor/transmitter pairs.
Background:
Excess carbohydrate intake during hypoglycemia can lead to rebound hyperglycemia (RH). We investigated associations between RH and use of real-time continuous glucose monitoring (rtCGM) ...and an rtCGM system’s predictive alert.
Methods:
RH events were series of sensor glucose values (SGVs) >180 mg/dL starting within two hours of an antecedent SGV <70 mg/dL. Events were characterized by their frequency, duration (consecutive SGVs >180 mg/dL × five minutes), and severity (area under the glucose concentration-time curve). To assess the impact of rtCGM, data gathered during the four-week baseline phase (without rtCGM) and four-week follow-up phase (with rtCGM) from 75 participants in the HypoDE clinical trial (NCT02671968) of hypoglycemia-unaware individuals were compared. To assess the impact of predictive alerts, we identified a convenience sample of 24 518 users of an rtCGM system without predictive alerts who transitioned to a system whose predictive alert signals an SGV ≤55 mg/dL within 20 minutes (Dexcom G5 and G6, respectively). RH events from periods of blinded versus unblinded rtCGM wear and from periods of G5 and G6 wear were compared with paired t tests.
Results:
Compared to RH events in the HypoDE baseline phase, the mean frequency, duration, and severity of events fell by 14%, 12%, and 23%, respectively, in the follow-up phase (all P < .05). Compared to RH events during G5 use, the mean frequency, duration, and severity of events fell by 7%, 8%, and 13%, respectively, during G6 use (all P < .001).
Conclusions:
Rebound hypreglycemia can be objectively quantified and mitigated with rtCGM and rtCGM-based predictive alerts.
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