A Multidisciplinary Algorithm for the Evaluation of Acute Neurologic Deficits Improves Management of Cryptogenic Stroke or Transient Ischemic Attack.

The appropriate diagnosis and management of cryptogenic stroke and transient ischemic attack (TIA) is challenging and requires multidisciplinary involvement. Joint societal guidelines exist to guide the comprehensive evaluation of these entities. This study aimed to implement a standardized multidisciplinary diagnostic algorithm for cryptogenic stroke/TIA. We performed a retrospective analysis of patients admitted to the largest regional military healthcare center with stroke or TIA considered to be cryptogenic at the time of discharge. We abstracted baseline demographics and rates of extra- and intracranial imaging, transthoracic and transesophageal echocardiography, and event monitor orders at the time of discharge. The incidence of event monitor results at 30 days and six months were included. A diagnostic algorithm for evaluation of cryptogenic stroke/TIA was created and disseminated hospital-wide using increased compliance with neuroimaging, echocardiography, and cardiac rhythm monitoring as primary endpoints for our intervention. Post-intervention data abstraction revealed similar rates of extra- and intracranial imaging, but significantly greater rates of transthoracic echocardiography (70% vs. 87%, p 0.0073), inclusion of agitated saline study (41% vs. 65%, p 0.0024), and event monitors ordered at discharge (18% vs. 35%, p 0.0045). At six months there was a higher rate of event monitors obtained (24% vs. 45%, p 0.001). Our study showed implementation of an evidence-based diagnostic algorithm for evaluation of cryptogenic stroke/TIA increases appropriate use of echocardiography and event monitoring.

Surface-water/groundwater boundaries affect seasonal PFAS concentrations and PFAA precursor transformations.

Elevated concentrations of per- and polyfluoroalkyl substances (PFAS) in drinking-water supplies are a major concern for human health. It is therefore essential to understand factors that affect PFAS concentrations in surface water and groundwater and the transformation of perfluoroalkyl acid (PFAA) precursors that degrade into terminal compounds. Surface-water/groundwater exchange can occur along the flow path downgradient from PFAS point sources and biogeochemical conditions can change rapidly at these exchange boundaries. Here, we investigate the influence of surface-water/groundwater boundaries on PFAS transport and transformation. To do this, we conducted an extensive field-based analysis of PFAS concentrations in water and sediment from a flow-through lake fed by contaminated groundwater and its downgradient surface-water/groundwater boundary (defined as ≤100 cm below the lake bottom). PFAA precursors comprised 45 ± 4.6% of PFAS (PFAA precursors + 18 targeted PFAA) in the predominantly oxic lake impacted by a former fire-training area and historical wastewater discharges. In shallow porewater downgradient from the lake, this percentage decreased significantly to 25 ± 11%. PFAA precursor concentrations decreased by 85% between the lake and 84-100 cm below the lake bottom. PFAA concentrations increased significantly within the surface-water/groundwater boundary and in downgradient groundwater during the winter months despite lower stable concentrations in the lake water source. These results suggest that natural biogeochemical fluctuations associated with surface-water/groundwater boundaries may lead to PFAA precursor loss and seasonal variations in PFAA concentrations. Results of this work highlight the importance of dynamic biogeochemical conditions along the hydrological flow path from PFAS point sources to potentially affected drinking water supplies.

Contribution of HCN1 variant to sinus bradycardia: A case report.

BACKGROUND: Missense mutations in the hyperpolarization-activated cyclic nucleotide-modulated (HCN) channel 4 (HCN4) are one of the genetic causes of cardiac sinus bradycardia. OBJECTIVE: To investigate possible HCN4 channel mutation in a young patient with profound sinus bradycardia. METHODS: Direct sequencing of HCN4 and whole-exome sequencing were performed on DNA samples from the indexed patient (P), the patient's son (PS), and a family unrelated healthy long-distance running volunteer (V). Resting heart rate was 31 bpm for P, 67 bpm for PS, and 50 bpm for V. Immunoblots, flow cytometry, and immunocytofluorescence confocal imaging were used to study cellular distribution of channel variants. Patch-clamp electrophysiology was used to investigate the properties of mutant HCN1 channels. RESULTS: In P no missense mutations were found in the HCN4 gene; instead, we found two heterozygous variants in the HCN1 gene: deletion of an N-terminal glycine triplet (72GGG74, "N-del") and a novel missense variant, P851A, in the C-terminal region. N-del variant was found before and shared by PS. These two variations were not found in V. Compared to wild type, N-del and P851A reduced cell surface expression and negatively shifted voltage-activation with slower activation kinetics. CONCLUSION: Decreased channel activity HCN1 mutant channel makes it unable to contribute to early depolarization of sinus node action potential, thus likely a main cause of the profound sinus bradycardia in this patient.

Investigating Habitability with an Integrated Rock-Climbing Robot and Astrobiology Instrument Suite.

A prototype rover carrying an astrobiology payload was developed and deployed at analog field sites to mature generalized system architectures capable of searching for biosignatures in extreme terrain across the Solar System. Specifically, the four-legged Limbed Excursion Mechanical Utility Robot (LEMUR) 3 climbing robot with microspine grippers carried three instruments: a micro-X-ray fluorescence instrument based on the Mars 2020 mission's Planetary Instrument for X-ray Lithochemistry provided elemental chemistry; a deep-ultraviolet fluorescence instrument based on Mars 2020's Scanning Habitable Environments with Raman and Luminescence for Organics and Chemicals mapped organics in bacterial communities on opaque substrates; and a near-infrared acousto-optic tunable filter-based point spectrometer identified minerals and organics in the 1.6-3.6 µm range. The rover also carried a light detection and ranging and a color camera for both science and navigation. Combined, this payload detects astrobiologically important classes of rock components (elements, minerals, and organics) in extreme terrain, which, as demonstrated in this work, can reveal a correlation between textural biosignatures and the organics or elements expected to preserve them in a habitable environment. Across >10 field tests, milestones were achieved in instrument operations, autonomous mobility in extreme terrain, and system integration that can inform future planetary science mission architectures. Contributions include (1) system-level demonstration of mock missions to the vertical exposures of Mars lava tube caves and Mars canyon walls, (2) demonstration of multi-instrument integration into a confocal arrangement with surface scanning capabilities, and (3) demonstration of automated focus stacking algorithms for improved signal-to-noise ratios and reduced operation time.

Adoption of coronary artery disease - Reporting and Data System (CAD-RADS™) and observed impact on medical therapy and systolic blood pressure control.

BACKGROUND: CAD-RADS was developed to standardize communication of per-patient maximal stenosis on coronary CT angiography (CCTA) and provide treatment recommendations and may impact primary prevention care and resource utilization. The authors sought to evaluate CAD-RADS adoption on preventive medical therapy and risk factor control amongst a mixed provider population. METHODS: Statins, aspirin (ASA), systolic blood pressure and, when available, lipid panel changes were abstracted for 1796 total patients undergoing CCTA in the 12 months before (non-standard reporting, NSR, cohort) and after adoption of the CAD-RADS reporting template. Only initiation of a medication in a treatment naïve patient, escalation from baseline dose, or transition to a higher potency was considered an escalation/initiation in lipid therapy. RESULTS: The CAD-RADS reporting template was utilized in 83.7% (751/897) of CCTAs after the CAD-RADS adoption period. After adjusting for any coronary artery disease (CAD) on CCTA, statin initiation/escalation was more commonly observed in the CAD-RADS cohort (aOR 1.46; 95%CI 1.12-1.90, p = 0.005), driven by higher rates of new statin initiation (aOR 1.79; 95%CI 1.23-2.58, p = 0.002). This resulted in a higher observed rates of total cholesterol improvement in the CAD-RADS cohort (58% vs 49%, p = 0.016). New ASA initiation was similar between reporting templates after adjustment for CAD on CCTA (aOR 1.40; 95%CI 0.97-2.02, p = 0.069). The ordering provider's specialty (cardiology vs non-cardiology) did not significantly impact the observed differences in initiation/escalation of statins and ASA (pinteraction = NS). CONCLUSIONS: Adoption of CAD-RADS reporting was associated with increased utilization of preventive medications, regardless of ordering provider specialty.

Right Ventricular Failure Following Patent Foramen Ovale Closure on Venovenous-Extracorporeal Membrane Oxygenation.

We present a patient with pulmonary arterial hypertension requiring venovenous-extracorporeal membrane oxygenation for acute respiratory distress syndrome. Refractory hypoxemia secondary to right-to-left interatrial shunting via a patent foramen ovale was discovered. Right heart catheterization with invasive occlusion test heralded worsening right heart failure so closure was aborted. (Level of Difficulty: Intermediate.).

Adoption of the Coronary Artery Disease-reporting and Data System: Reduced Downstream Testing and Cardiology Referral Rates in Patients with Non-obstructive Coronary Artery Disease.

Introduction The coronary artery disease-reporting and data system (CAD-RADS) was developed to standardize communication of per-patient maximal stenosis and provide treatment recommendations that may affect downstream testing. Methods Downstream testing, cardiology referral, and cost were abstracted for 1,796 consecutive patients undergoing coronary CT angiography (CCTA) before and after the adoption of the CAD-RADS reporting template at a single-center closed referral hospital system. Cost analysis was based on direct invasive and non-invasive testing utilizing the Center for Medicare & Medicaid Services (CMS) outpatient prospective payment system (OPPS) final rule for 2018. Results Baseline cardiovascular risk factors were balanced between the groups. Overall, referrals for downstream testing were similar between cohorts (10.7% vs 10.8%; p = 0.939). Referral for downstream testing was reduced in the CAD-RADS 1 & 2 cohort compared to non-obstructive coronary artery disease (CAD) by non-standardized reporting (NSR; 5.1% vs 14.4%, p < 0.001). This was offset by more non-diagnostic scans in the CAD-RADS cohort (9.7% vs 4.2%, p < 0.001), resulting in increased downstream testing (28.8% vs 11.4%, p = 0.038). Overall, cardiology referral rates by primary care providers (PCPs) were similar between the groups (12.2% vs 15.8%, p = 0.197). Cardiology referral rates were increased among patients with non-obstructive CAD in the NSR cohort compared with CAD-RADS 1 & 2 patients (20.5% vs 8.6%, p = 0.021). Referrals for invasive coronary angiography were low in both groups overall (3.5% vs 3.2%, p = 0.726). Median downstream testing costs were similar between the groups (p = 0.554). Conclusions Adoption of the CAD-RADS reporting template was associated with a reduction in downstream testing and cardiology referral rates among non-obstructive CAD (CAD-RADS 1 & 2) patients. Thus, CAD-RADS may impact downstream testing in patients in whom further testing can typically be deferred.

Cardiac Tamponade from Purulent Pericarditis due to Cutibacterium acnes.

Purulent pericarditis is a potentially fatal disease with high mortality rates if untreated. Cutibacterium acnes (formerly Propionibacterium acnes) is an anaerobic bacteria that is ubiquitous in skin flora and is commonly thought of as a culture contaminant; however, it does have pathogenic potential. We present a case of purulent pericarditis secondary to C. acnes leading to cardiac tamponade. Initial stabilization and diagnosis were made via pericardiocentesis; afterward the patient underwent a pericardial window. Due to a severe penicillin allergy, he was successfully treated with a 14-day course of vancomycin. To our knowledge, this represents only the third published case of purulent pericarditis with cardiac tamponade caused by C. acnes and the first case treated with a 14-day course of vancomycin.

Stochasticity in materials structure, properties, and processing-A review.

We review the concept of stochasticity-i.e., unpredictable or uncontrolled fluctuations in structure, chemistry, or kinetic processes-in materials. We first define six broad classes of stochasticity: equilibrium (thermodynamic) fluctuations; structural/compositional fluctuations; kinetic fluctuations; frustration and degeneracy; imprecision in measurements; and stochasticity in modeling and simulation. In this review, we focus on the first four classes that are inherent to materials phenomena. We next develop a mathematical framework for describing materials stochasticity and then show how it can be broadly applied to these four materials-related stochastic classes. In subsequent sections, we describe structural and compositional fluctuations at small length scales that modify material properties and behavior at larger length scales; systems with engineered fluctuations, concentrating primarily on composite materials; systems in which stochasticity is developed through nucleation and kinetic phenomena; and configurations in which constraints in a given system prevent it from attaining its ground state and cause it to attain several, equally likely (degenerate) states. We next describe how stochasticity in these processes results in variations in physical properties and how these variations are then accentuated by-or amplify-stochasticity in processing and manufacturing procedures. In summary, the origins of materials stochasticity, the degree to which it can be predicted and/or controlled, and the possibility of using stochastic descriptions of materials structure, properties, and processing as a new degree of freedom in materials design are described.

Directed Self-Assembly of Ge Quantum Dots Using Focused Si2+ Ion Beam Patterning.

We show that templating a Si surface with a focused beam of Si2+ or Si+ ions can create suitable nucleation sites for the subsequent growth of self-assembled Ge quantum dots by chemical vapor deposition. To determine the mechanism of patterning we use atomic force microscopy to show that, similar to Ga+ patterning, the formation of a surface pit is required to enable control over Ge quantum dot locations. We find that relatively high implantation doses are required to achieve patterning, and these doses lead to amorphization of the substrate. We assess the degree to which the substrate crystallinity can be recovered by subsequent processing. Using in situ transmission electron microscopy heating experiments we find that recrystallization is possible at the growth temperature of the Ge quantum dots, but defects remain that follow the pattern of the initial implantation. We discuss the formation mechanism of the defects and the benefits of using Si ions for patterning both defects and quantum dots on Si substrates.

Transcriptome profiling reveals novel BMI- and sex-specific gene expression signatures for human cardiac hypertrophy.

How obesity or sex may affect the gene expression profiles of human cardiac hypertrophy is unknown. We hypothesized that body-mass index (BMI) and sex can affect gene expression profiles of cardiac hypertrophy. Human heart tissues were grouped according to sex (male, female), BMI (lean<25 kg/m2, obese>30 kg/m2), or left ventricular hypertrophy (LVH) and non-LVH nonfailed controls (NF). We identified 24 differentially expressed (DE) genes comparing female with male samples. In obese subgroup, there were 236 DE genes comparing LVH with NF; in lean subgroup, there were seven DE genes comparing LVH with NF. In female subgroup, we identified 1,320 significant genes comparing LVH with NF; in male subgroup, there were 1,383 significant genes comparing LVH with NF. There were seven significant genes comparing obese LVH with lean NF; comparing male obese LVH with male lean NF samples we found 106 significant genes; comparing female obese LVH with male lean NF, we found no significant genes. Using absolute value of log2 fold-change > 2 or extremely small P value (10-20) as a criterion, we identified nine significant genes (HBA1, HBB, HIST1H2AC, GSTT1, MYL7, NPPA, NPPB, PDK4, PLA2G2A) in LVH, also found in published data set for ischemic and dilated cardiomyopathy in heart failure. We identified a potential gene expression signature that distinguishes between patients with high BMI or between men and women with cardiac hypertrophy. Expression of established biomarkers atrial natriuretic peptide A (NPPA) and B (NPPB) were already significantly increased in hypertrophy compared with controls.

Visualization of structural evolution and phase distribution of a lithium vanadium oxide (Li1.1V3O8) electrode via an operando and in situ energy dispersive X-ray diffraction technique.

Li1+nV3O8 (n = 0-0.2) has been extensively investigated as a cathode material for Li ion batteries because of its superior electrochemical properties including high specific energy and good rate capability. In this paper, a synchrotron based energy dispersive X-ray diffraction (EDXRD) technique was employed to profile the phase transitions and the spatial phase distribution of a Li1.1V3O8 electrode during electrochemical (de)lithiation in situ and operando. As annealing temperature during the preparation of the Li1.1V3O8 material has a strong influence on the morphology and crystallinity, and consequently influences the electrochemical outcomes of the material, Li1.1V3O8 materials prepared at two different temperatures, 500 and 300 °C (LVO500 and LVO300), were employed in this study. The EDXRD spectra of LVO500 and LVO300 cells pre-discharged at C/18, C/40 and C/150 were recorded in situ, and phase localization and relative intensity of the peaks were compared. For cells discharged at the C/18 rate, although α and ß phases were distributed uniformly within the LVO500 electrode, they were localized on two sides of the LVO300 electrode. Discharging rates of C/40 and C/150 led to homogeneous ß phase formation in both LVO500 and LVO300 electrodes. Furthermore, the phase distribution as a function of position and (de)lithiation extent was mapped operando as the LVO500 cell was (de)lithiated. The operando data indicate that (1) the lithiation reaction initiated from the side of the electrode facing the Li anode and proceeded towards the side facing the steel can, (2) during discharge the phase transformation from a Li-poor to a Li-rich α phase and the formation of a ß phase can proceed simultaneously in the electrode after the first formation of a ß phase, and (3) the structural evolution occurring during charging is not the reverse of that during discharge and takes place homogenously throughout the electrode.

Left atrial appendage morphology and risk of stroke following pulmonary vein isolation for drug-refractory atrial fibrillation in low CHA2DS2Vasc risk patients.

BACKGROUND: Cardiac CT angiography (CCTA) has become an important adjunct in the structural assessment of the pulmonary veins (PV) prior to pulmonary vein isolation (PVI). Published data is conflicting regarding a relationship between left atrial appendage (LAA) and the risk of ischemic stroke (CVA) following PVI. We investigated the associations of volumetric and morphologic left atrial (LA) and LAA measurements for CVA following PVI. METHODS: We retrospectively reviewed 332 consecutive patients with drug refractory atrial fibrillation who obtained cardiac CT angiogram (CCTA) prior to PVI. Baseline demographic data, procedural and lab details, and outcomes were obtained from abstraction of an electronic medical records system. LA, LAA, and PV volumes were measured using CCTA datasets utilizing a semi-automated 3D workstation application. LAA morphology was assigned utilizing volume rendered images as previously described. RESULTS: The study cohort was 55 ± 13 years-old, 83.7% male, low CVA risk (median CHA2DS2Vasc 1; IQR 1, 3), and 30.4% were treated with novel oral anticoagulants. Chicken wing (CW) was the most common morphology (52%), followed by windsock (WS), cauliflower (CF), and cactus (CS) at 18, 9, and 2%, respectively. CVAs occurred in 4 patients following PVI with median time to CVA of 170.5 days. All CVAs were observed in CW morphology patients. When comparing CW morphology with non-CW morphology, CVAs occurred more frequently with the CW morphology (2.1% vs 0%, p = 0.03). This difference was not significant, though, after adjusting for CHA2DS2Vasc risk factors (p = 0.14). CONCLUSION: The CW morphology was observed more commonly in patients who experienced post-PVI CVA. After adjusting for CHA2DS2Vasc risk factors, CW morphology was not an independent predictor of post-PVI CVA. These findings should be interpreted in the setting of a low CVA event rate amongst a low risk population that was highly compliant with indicated anticoagulation therapy.

Ambulatory Hemodynamic Monitoring Reduces Heart Failure Hospitalizations in "Real-World" Clinical Practice.

BACKGROUND: In the CHAMPION (CardioMEMS Heart Sensor Allows Monitoring of Pressure to Improve Outcomes in New York Heart Association [NYHA] Functional Class III Heart Failure Patients) trial, heart failure hospitalization (HFH) rates were lower in patients managed with guidance from an implantable pulmonary artery pressure sensor compared with usual care. OBJECTIVES: This study examined the effectiveness of ambulatory hemodynamic monitoring in reducing HFH outside of the clinical trial setting. METHODS: We conducted a retrospective cohort study using U.S. Medicare claims data from patients undergoing pulmonary artery pressure sensor implantation between June 1, 2014, and December 31, 2015. Rates of HFH during pre-defined periods before and after implantation were compared using the Andersen-Gill extension to the Cox proportional hazards model while accounting for the competing risk of death, ventricular assist device implantation, or cardiac transplantation. Comprehensive heart failure (HF)-related costs were compared over the same periods. RESULTS: Among 1,114 patients receiving implants, there were 1,020 HFHs in the 6 months before, compared with 381 HFHs, 139 deaths, and 17 ventricular assist device implantations and/or transplants in the 6 months after implantation (hazard ratio [HR]: 0.55; 95% confidence interval [CI]: 0.49 to 0.61; p < 0.001). This lower rate of HFH was associated with a 6-month comprehensive HF cost reduction of $7,433 per patient (IQR: $7,000 to $7,884), and was robust in analyses restricted to 6-month survivors. Similar reductions in HFH and costs were noted in the subset of 480 patients with complete data available for 12 months before and after implantation (HR: 0.66; 95% CI: 0.57 to 0.76; p < 0.001). CONCLUSIONS: As in clinical trials, use of ambulatory hemodynamic monitoring in clinical practice is associated with lower HFH and comprehensive HF costs. These benefits are sustained to 1 year and support the "real-world" effectiveness of this approach to HF management.

Leptin decreases heart rate associated with increased ventricular repolarization via its receptor.

Leptin has been proposed to modulate cardiac electrical properties via ß-adrenergic receptor activation. The presence of leptin receptors and adipocytes in myocardium raised a question as to whether leptin can directly modulate cardiac electrical properties such as heart rate and QT interval via its receptor. In this work, the role of local direct actions of leptin on heart rate and ventricular repolarization was investigated. We identified the protein expression of leptin receptors at cell surface of sinus node, atrial, and ventricular myocytes isolated from rat heart. Leptin at low doses (0.1-30 µg/kg) decreased resting heart rate; at high doses (150-300 µg/kg), leptin induced a biphasic effect (decrease and then increase) on heart rate. In the presence of high-dose propranolol (30 mg/kg), high-dose leptin only reduced heart rate and sometimes caused sinus pauses and ventricular tachycardia. The leptin-induced inhibition of resting heart rate was fully reversed by leptin antagonist. Leptin also increased heart rate-corrected QT interval (QTc), and leptin antagonist did not. In isolated ventricular myocytes, leptin (0.03-0.3 µg/ml) reversibly increased the action potential duration. These results supported our hypothesis that in addition to indirect pathway via sympathetic tone, leptin can directly decrease heart rate and increase QT interval via its receptor independent of ß-adrenergic receptor stimulation. During inhibition of ß-adrenergic receptor activity, high concentration of leptin in myocardium can cause deep bradycardia, prolonged QT interval, and ventricular arrhythmias.

PP2 prevents isoproterenol stimulation of cardiac pacemaker activity.

Increasing evidence has demonstrated the potential risks of cardiac arrhythmias (such as prolonged QT interval) using tyrosine kinase inhibitors for cancer therapy. We report here that a widely used selective inhibitor of Src tyrosine kinases, PP2, can inhibit and prevent isoproterenol stimulation of cardiac pacemaker activity. In dissected rat sinus node, PP2 inhibited and prevented isoproterenol stimulation of spontaneous beating rate. In isolated sinus node myocytes, PP2 suppressed the hyperpolarization-activated "funny" current (If) by negatively shifting the activation curve and decelerating activation kinetics, associated with decreased cell surface expression and reduced tyrosine phosphorylation of hyperpolarization-activated cyclic nucleotide-modulated channel 4 (HCN4) channel proteins. In human embryonic kidney 293 cells overexpressing recombinant human HCN4 channels, PP2 reversed isoproterenol stimulation of HCN4 and inhibited HCN4-573x, a cAMP-insensitive human HCN4 mutant. Isoprotenrenol had little effects on HCN4-573x. These results demonstrated that inhibition of presumably tyrosine Src kinase activity in heart by PP2 decreased and prevented the potential ß-adrenergic stimulation of cardiac pacemaker activity. These effects are mediated, at least partially, by a cAMP-independent attenuation of channel activity and cell surface expression of HCN4, the key channel protein that controls the heart rate.

Studying localized corrosion using liquid cell transmission electron microscopy.

Localized corrosion of Cu and Al thin films exposed to aqueous NaCl solutions was studied using liquid cell transmission electron microscopy (LCTEM). We demonstrate that potentiostatic control can be used to initiate pitting and that local compositional changes, due to focused ion beam implantation of Au(+) ions, can modify the corrosion susceptibility of Al films.

Metastable structures in Al thin films before the onset of corrosion pitting as observed using liquid cell transmission electron microscopy.

One of the fundamental challenges in understanding the early stages of corrosion pitting in metals protected with an oxide film is that there are relatively few techniques that can probe microstructure with sufficient resolution while maintaining a wet environment. Here, we demonstrate that microstructural changes in Al thin films caused by aqueous NaCl solutions of varying chloride concentrations can be directly observed using a liquid flow cell enclosed within a transmission electron microscope (TEM) holder. In the absence of chloride, Al thin films did not exhibit significant corrosion when immersed in de-ionized water for 2 days. However, introducing 0.01 M NaCl solutions led to extensive random formation of blisters over the sample surface, while 0.1 M NaCl solutions formed anomalous structures that were larger than the typical grain size. Immersion in 1.0 M NaCl solutions led to fractal corrosion consistent with previously reported studies of Al thin films using optical microscopy. These results show the potential of in situ liquid cell electron microscopy for probing the processes that take place before the onset of pitting and for correlating pit locations with the underlying microstructure of the material.

PP2 prevents ß-adrenergic stimulation of cardiac pacemaker activity.

One of the main strategies for cancer therapy is to use tyrosine kinase inhibitors for inhibiting tumor proliferation. Increasing evidence has demonstrated the potential risks of cardiac arrhythmias (such as prolonged QT interval) of these drugs. We report here that a widely used selective inhibitor of Src tyrosine kinases, 4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo[3,4-d]pyrimidine (PP2), can inhibit and prevent ß-adrenergic stimulation of cardiac pacemaker activity. First, in dissected rat sinus node, PP2 inhibited and prevented the isoproterenol-induced increase of spontaneous beating rate. Second, in isolated rat sinus node myocytes, PP2 suppressed the hyperpolarization-activated "funny" current (traditionally called cardiac pacemaker current, I(f)) by negatively shifting the activation curve and decelerating activation kinetics. Third, in isolated rat sinus node myocytes, PP2 decreased the Src kinase activity, the cell surface expression, and tyrosine phosphorylation of hyperpolarization-activated, cyclic nucleotide-modulated channel 4 (HCN4) channel proteins. Finally, in human embryonic kidney 293 cells overexpressing recombinant human HCN4 channels, PP2 reversed the enhancement of HCN4 channels by isoproterenol and inhibited 573x, a cyclic adenosine momophosphate-insensitive human HCN4 mutant. These results demonstrated that inhibition of Src kinase activity in heart by PP2 decreased and prevented ß-adrenergic stimulation of cardiac pacemaker activity. These effects are mediated, at least partially, by a cAMP-independent attenuation of channel activity and cell surface expression of HCN4, the main channel protein that controls the heart rate.