Enhanced phosphorylation of caveolar PKC-α limits peptide internalization in lung endothelial cells.

We previously reported that the vasoactive peptide 1 (P1, "SSWRRKRKESS") modulates the tension of pulmonary artery vessels through caveolar endothelial nitric oxide synthase (eNOS) activation in intact lung endothelial cells (ECs). Since PKC-α is a caveolae resident protein and caveolae play a critical role in the peptide internalization process, we determined whether modulation of caveolae and/or caveolar PKC-α phosphorylation regulates internalization of P1 in lung ECs. Cell monolayers were incubated in culture medium containing Rhodamine red-labeled P1 (100 µM) for 0-120 min. Confocal examinations indicate that P1 internalization is time-dependent and reaches a plateau at 60 min. Caveolae disruption by methyl-ß-cyclodextrin (CD) and filipin (FIL) inhibited the internalization of P1 in ECs suggesting that P1 internalizes via caveolae. P1-stimulation also enhances phosphorylation of caveolar PKC-α and increases intracellular calcium (Ca(2+)) release in intact cells suggesting that P1 internalization is regulated by PKC-α in ECs. To confirm the roles of increased phosphorylation of PKC-α and Ca(2+) release in internalization of P1, PKC-α modulation by phorbol ester (PMA), PKC-α knockdown, and Ca(2+) scavenger BAPTA-AM model systems were used. PMA-stimulated phosphorylation of caveolar PKC-α is associated with significant reduction in P1 internalization. In contrast, PKC-α deficiency and reduced phosphorylation of PKC-α enhanced P1 internalization. P1-mediated increased phosphorylation of PKC-α appears to be associated with increased intracellular calcium (Ca(2+)) release since the Ca(2+) scavenger BAPTA-AM enhanced P1 internalization. These data indicate that caveolar integrity and P1-mediated increased phosphorylation of caveolar PKC-α play crucial roles in the regulation of P1 internalization in lung ECs.

A Special Clinical Scenario for Transcatheter Aortic Valve Replacement: "Iatrogenic Double-Outlet" Left Ventricle.

A 67-year-old man with a history of chest radiotherapy and severe aortic valve stenosis with calcification of the ascending aortic wall underwent implantation of an apicoaortic conduit from the left ventricular apex to the descending aorta. Eight years later, he presented with progressive exertional dyspnea. Imaging revealed severe native aortic valve insufficiency and calcification, with worsening left ventricular function. We decided to leave the apicoaortic conduit intact and perform transcatheter aortic valve replacement with a balloon-expandable prosthesis. Despite concerns that eliminating the obstruction across the native left ventricular outflow tract might decrease conduit flow and eventually cause graft thrombosis and peripheral embolization, we elected to move forward after a multidisciplinary discussion. The procedure resulted in angiographically and qualitatively similar forward flow across the newly implanted prosthesis and the existing apicoaortic conduit, with no hemodynamic or electrical dysfunction. The patient was discharged from the hospital the next day. At the 1-month follow-up visit, the patient felt well and reported marked functional improvement, with minimal symptoms during moderate to heavy exertion. The stroke volume index across the new bioprosthetic valve was low (13 mL/m2 at 1 mo and 18 mL/m2 at 1 y), suggesting that a substantial amount of blood was still exiting the ventricle through the left ventricle-to-aorta conduit. This report offers some guidance for treating patients with existing apicoaortic conduits and suggests that transcatheter aortic valve replacement is safe and effective if native aortic valve insufficiency develops.

Peptide-stimulation enhances compartmentalization and the catalytic activity of lung endothelial NOS.

We reported that an 11 amino acid synthetic peptide (P1) activates lung endothelial cell nitric oxide synthase (eNOS) independent of its change in expression and/or phosphorylation. Since caveolae/eNOS dissociation is known to enhance the catalytic activity of eNOS, we examined whether P1-mediated increase of eNOS activity is associated with caveolae/cholesterol modulation, increased caveolin-1 phosphorylation, and intracellular compartmentalization of eNOS in pulmonary artery endothelial cells (PAEC). PAEC were incubated with or without (control) P1 or cholesterol modulators/caveolae disruptors, cholesterol oxidase (CHOX) and methyl-beta-cyclodextrin (CD), for 1 h at 37 degrees C. After incubation cells were used for: i) immunoprecipitation, ii) isolation of plasma membrane (PM)-, Golgi complex (GC)-, and non-Golgi complex (NGC)-enriched fractions, iii) immunofluorescence confocal imaging, and iv) electron microscopy for localization and/or eNOS activity. P1, CHOX, and CD-stimulation caused dissociation of eNOS from PM with increased localization to GC and/or NGC. P1 and CHOX significantly increased eNOS activity in PM and GC and CD-stimulation increased eNOS activity localized only in GC. P1 increased phosphorylation of caveolin-1 in intact cells and GC fraction. Immunofluorescence and/or immunogold labeled imaging/electron microscopy analysis of P1-, CHOX-, and CD-stimulated intact cells confirmed eNOS/caveolae dissociation and translocation of eNOS to GC. These results suggest that: i) P1-stimulation translocates eNOS to GC and enhances the catalytic activity of eNOS in both the PM and GC fractions of PAEC, ii) CHOX- but not CD-mediated caveolae and/or cholesterol modulation mimics the effect of P1-stimulated compartmentalization and activation of eNOS in PAEC, and iii) P1-stimulated caveolae/cholesterol modulation, phosphorylation of caveolin-1, and activation of eNOS is physiologically relevant since P1 is known to enhance NO/cGMP-dependent vasorelaxation in the pulmonary circulation.

Acute Biventricular Mechanical Circulatory Support for Cardiogenic Shock.

BACKGROUND: Biventricular failure is associated with high in-hospital mortality. Limited data regarding the efficacy of biventricular Impella axial flow catheters (BiPella) support for biventricular failure exist. The aim of this study was to explore the clinical utility of percutaneously delivered BiPella as a novel acute mechanical support strategy for patients with cardiogenic shock complicated by biventricular failure. METHODS AND RESULTS: We retrospectively analyzed data from 20 patients receiving BiPella for biventricular failure from 5 tertiary-care hospitals in the United States. Left ventricular support was achieved with an Impella 5.0 (n=8), Impella CP (n=11), or Impella 2.5 (n=1). All patients received the Impella RP for right ventricular (RV) support. BiPella use was recorded in the setting of acute myocardial infarction (n=11), advanced heart failure (n=7), and myocarditis (n=2). Mean flows achieved were 3.4±1.2 and 3.5±0.5 for left ventricular and RV devices, respectively. Total in-hospital mortality was 50%. No intraprocedural mortality was observed. Major complications included limb ischemia (n=1), hemolysis (n=6), and Thrombolysis in Myocardial Infarction major bleeding (n=7). Compared with nonsurvivors, survivors were younger, had a lower number of inotropes or vasopressors used before BiPella, and were more likely to have both devices implanted simultaneously during the same procedure. Compared with nonsurvivors, survivors had lower pulmonary artery pressures and RV stroke work index before BiPella. Indices of RV afterload were quantified for 14 subjects. Among these patients, nonsurvivors had higher pulmonary vascular resistance (6.8; 95% confidence interval [95% CI], 5.5-8.1 versus 1.9; 95% CI, 0.8-3.0; P<0.01), effective pulmonary artery elastance (1129; 95% CI, 876-1383 versus 458; 95% CI, 263-653; P<0.01), and lower pulmonary artery compliance (1.5; 95% CI, 0.9-2.1 versus 2.7; 95% CI, 1.8-3.6; P<0.05). CONCLUSIONS: This is the largest, retrospective analysis of BiPella for cardiogenic shock. BiPella is feasible, reduces cardiac filling pressures and improves cardiac output across a range of causes for cardiogenic shock. Simultaneous left ventricular and RV device implantation and lower RV afterload may be associated with better outcomes with BiPella. Future prospective studies of BiPella for cardiogenic shock are required.

Understanding Why Patients Return to the Emergency Department after Mild Traumatic Brain Injury within 72 Hours.

INTRODUCTION: Although there are approximately 1.1 million case presentations of mild traumatic brain injury (mTBI) in the emergency department (ED) each year, little data is available to clinicians to identify patients who are at risk for poor outcomes, including 72-hour ED return after discharge. An understanding of patients at risk for ED return visits during the hyperacute phase following head injury would allow ED providers to develop clinical interventions that reduce its occurrence and improve outcomes. METHODS: This institutional review board-approved consecutive cohort study collected injury and outcome variables on adults with the purpose of identifying positive predictors for 72-hour ED return visits in mTBI patients. RESULTS: Of 2,787 mTBI patients, 145 (5%) returned unexpectedly to the ED within 72 hours of hospital discharge. Positive predictors for ED return visits included being male (p=0.0298), being black (p=0.0456), having a lower prehospital Glasgow Coma Score (p=0.0335), suffering the injury due to a motor vehicle collision (p=0.0065), or having a bleed on head computed tomography (CT) (p=0.0334). ED return visits were not significantly associated with age, fracture on head CT, or symptomology following head trauma. Patients with return visits most commonly reported post-concussion syndrome (43.1%), pain (18.7%), and recall for further clinical evaluation (14.6%) as the reason for return. Of the 124 patients who returned to the ED within 72 hours, one out of five were admitted to the hospital for further care, with five requiring intensive care unit stays and four undergoing neurosurgery. CONCLUSION: Approximately 5% of adult patients who present to the ED for mTBI will return within 72 hours of discharge for further care. Clinicians should identify at-risk individuals during their initial visits and attempt to provide anticipatory guidance when possible.

Impact of helmet use in traumatic brain injuries associated with recreational vehicles.

Objective. To study the impact of helmet use on outcomes after recreational vehicle accidents. Methods. This is an observational cohort of adult and pediatric patients who sustained a TBI while riding a recreational vehicle. Recreational vehicles included bicycles, motorcycles, and all-terrain vehicles (ATVs), as well as a category for other vehicles such as skateboards and scooters. Results. Lack of helmet use was significantly associated with having a more severe traumatic brain injury and being admitted to the hospital. Similarly, 25% of those who did wearing a helmet were admitted to the ICU versus 36% of those who did not (P = 0.0489). The hospital length of stay was significantly greater for patients who did not use helmets. Conclusion. Lack of helmet use is significantly correlated with abnormal neuroimaging and admission to the hospital and ICU; these data support a call for action to implement more widespread injury prevention and helmet safety education and advocacy.

TBI surveillance using the common data elements for traumatic brain injury: a population study.

BACKGROUND: To characterize the patterns of presentation of adults with head injury to the Emergency Department. METHODS: This is a cohort study that sought to collect injury and outcome variables with the goal of characterizing the very early natural history of traumatic brain injury in adults. This IRB-approved project was conducted in collaboration with our Institution's Center for Translational Science Institute. Data were entered in REDCap, a secure database. Statistical analyses were performed using JMP 10.0 pro for Windows. RESULTS: The cohort consisted of 2,394 adults, with 40% being women and 79% Caucasian. The most common mechanism was fall (47%) followed by motor vehicle collision (MVC) (36%). Patients sustaining an MVC were significantly younger than those whose head injury was secondary to a fall (P < 0.0001). Ninety-one percent had CT imaging; hemorrhage was significantly more likely with worse severity as measured by the Glasgow Coma Score (chi-square, P < 0.0001). Forty-four percent were admitted to the hospital, with half requiring ICU admission. In-hospital death was observed in 5.4%, while neurosurgical intervention was required in 8%. For all outcomes, worse TBI severity per GCS was significantly associated with worse outcomes (logistic regression, P < 0.0001, adjusted for age). CONCLUSION: These cohort data highlight the burden of TBI in the Emergency Department and provide important demographic trends for further research.