Plasma kallikrein mediates brain hemorrhage and edema caused by tissue plasminogen activator therapy in mice after stroke.

Thrombolytic therapy using tissue plasminogen activator (tPA) in acute stroke is associated with increased risks of cerebral hemorrhagic transformation and angioedema. Although plasma kallikrein (PKal) has been implicated in contributing to both hematoma expansion and thrombosis in stroke, its role in the complications associated with the therapeutic use of tPA in stroke is not yet available. We investigated the effects of tPA on plasma prekallikrein (PPK) activation and the role of PKal on cerebral outcomes in a murine thrombotic stroke model treated with tPA. We show that tPA increases PKal activity in vitro in both murine and human plasma, via a factor XII (FXII)-dependent mechanism. Intravenous administration of tPA increased circulating PKal activity in mice. In mice with thrombotic occlusion of the middle cerebral artery, tPA administration increased brain hemorrhage transformation, infarct volume, and edema. These adverse effects of tPA were ameliorated in PPK (Klkb1)-deficient and FXII-deficient mice and in wild-type (WT) mice pretreated with a PKal inhibitor prior to tPA. tPA-induced brain hemisphere reperfusion after photothrombolic middle cerebral artery occlusion was increased in Klkb1-/- mice compared with WT mice. In addition, PKal inhibition reduced matrix metalloproteinase-9 activity in brain following stroke and tPA therapy. These data demonstrate that tPA activates PPK in plasma and PKal inhibition reduces cerebral complications associated with tPA-mediated thrombolysis in stroke.

Huntingtin is required for normal excitatory synapse development in cortical and striatal circuits.

Huntington's disease (HD) is a neurodegenerative disease caused by the expansion of a poly-glutamine (poly-Q) stretch in the huntingtin (Htt) protein. Gain-of-function effects of mutant Htt have been extensively investigated as the major driver of neurodegeneration in HD. However, loss-of-function effects of poly-Q mutations recently emerged as potential drivers of disease pathophysiology. Early synaptic problems in the excitatory cortical and striatal connections have been reported in HD, but the role of Htt protein in synaptic connectivity was unknown. Therefore, we investigated the role of Htt in synaptic connectivity in vivo by conditionally silencing Htt in the developing mouse cortex. When cortical Htt function was silenced, cortical and striatal excitatory synapses formed and matured at an accelerated pace through postnatal day 21 (P21). This exuberant synaptic connectivity was lost over time in the cortex, resulting in the deterioration of synapses by 5 weeks. Synaptic decline in the cortex was accompanied with layer- and region-specific reactive gliosis without cell loss. To determine whether the disease-causing poly-Q mutation in Htt affects synapse development, we next investigated the synaptic connectivity in a full-length knock-in mouse model of HD, the zQ175 mouse. Similar to the cortical conditional knock-outs, we found excessive excitatory synapse formation and maturation in the cortices of P21 zQ175, which was lost by 5 weeks. Together, our findings reveal that cortical Htt is required for the correct establishment of cortical and striatal excitatory circuits, and this function of Htt is lost when the mutant Htt is present.

Thrombosis and Hemorrhage in Diabetic Retinopathy: A Perspective from an Inflammatory Standpoint.

Retinal ischemia and hemorrhage are hallmarks of worsening diabetic retinopathy, which can lead to neovascularization, macular edema, and severe vision loss. Although diabetes alters expression of clotting factors and their activities, and increases retinal microthromboses, the effects of thrombotic processes on the pathogenesis of diabetic retinopathy are not fully understood. In addition to the roles of coagulation and fibrinolytic cascades in thrombosis and hemostasis, components in these systems also mediate multiple effects on the vasculature that promote inflammation. Plasma kallikrein, thrombin, and urokinase are increased in diabetic retinopathy, and exert proinflammatory effects that contribute to retinal vascular dysfunction. The accumulation and activation of these and additional coagulation factors in the vitreous due to hemorrhage and chronic retinal injury in the diabetic retina may contribute to worsening of retinal inflammation and capillary dysfunction, which lead to retinal ischemia and edema. Further understanding of the role for specific coagulation factors in diabetic retinopathy may suggest new therapeutic opportunities for this vision-threatening disease.

Plasma Kallikrein Contributes to Intracerebral Hemorrhage and Hypertension in Stroke-Prone Spontaneously Hypertensive Rats.

Plasma kallikrein (PKa) has been implicated in contributing to hemorrhage following thrombolytic therapy; however, its role in spontaneous intracerebral hemorrhage is currently not available. This report investigates the role of PKa on hemorrhage and hypertension in stroke-prone spontaneously hypertensive rats (SHRSP). SHRSP were fed with a high salt-containing stroke-prone diet to increase blood pressure and induce intracerebral hemorrhage. The roles of PKa on blood pressure, hemorrhage, and survival in SHRSP were examined in rats receiving a PKa inhibitor or plasma prekallikrein antisense oligonucleotide (PK ASO) compared with rats receiving control ASO. Effects on PKa on the proteolytic cleavage of atrial natriuretic peptide (ANP) were analyzed by tandem mass spectrometry. We show that SHRSP on high-salt diet displayed increased levels of PKa activity compared with control rats. Cleaved kininogen was increased in plasma during stroke compared to SHRSP without stroke. Systemic administration of a PKa inhibitor or PK ASO to SHRSP reduced hemorrhage and blood pressure, and improved neurological function and survival compared with SHRSP receiving control ASO. Since PKa inhibition was associated with reduced blood pressure in hypertensive rats, we investigated the effects of PKa on the cleavage of ANP. Incubation of PKa with ANP resulted in the generation fragment ANP5-28, which displayed reduced effects on blood pressure lowering compared with full length ANP. PKa contributes to increased blood pressure in SHRSP, which is associated with hemorrhage and reduced survival. PKa-mediated cleavage of ANP reduces its blood pressure lowering effects and thereby may contribute to hypertension-induced intracerebral hemorrhage.

Clinical and procedural characteristics predicting need for chronotropic support and permanent pacing post-heart transplantation.

BACKGROUND: Postoperative bradycardia can complicate orthotopic heart transplantation (OHT). Previous studies suggested donor age and surgical technique as possible risk factors. However, risk factors in the era of bicaval anastomosis have not been elucidated. OBJECTIVE: We sought to examine the association between donor/recipient characteristics with need for chronotropic support and permanent pacemaker (PPM) implantation in patients with OHT. METHODS: All patients treated with OHT between January 2003 and January 2018 at the Hospital of the University of Pennsylvania were retrospectively evaluated until June 2018. Chronotropic support was given upon postoperative inability to increase the heart rate to patient's demands and included disproportionate bradycardia and junctional rhythm. RESULTS: A total of 820 patients (mean age 51.3 ± 12.6 years; 607, 74% men) underwent 826 OHT procedures (787 patients, 95.3% bicaval anastomosis). Patients who were exposed to amiodarone (odds ratio [OR] 2.30; 95% confidence interval [CI] 1.58-3.34; P < .001) and have older donor (OR 1.02; 95% CI 1.01-1.04; P = .001) were more likely to develop need for chronotropic support. In multivariable analysis, recipient age (OR 1.03; 95% CI 1.00-1.06; P = .04) and biatrial anastomosis (OR 6.12; 95% CI 2.48-15.09) were significantly associated with PPM implantation within 6 months of OHT. No association was found between pre-OHT amiodarone use and PPM implantation. No risk factors assessed were associated with PPM implantation 6 months post-OHT. CONCLUSION: Surgical technique and donor age were the main risk factors for the need for chronotropic support post-OHT, whereas surgical technique and recipient age were risk factors for early PPM implantation.

Appropriate and inappropriate shocks in hypertrophic cardiomyopathy patients with subcutaneous implantable cardioverter-defibrillators: An international multicenter study.

BACKGROUND: Subcutaneous implantable cardioverter-defibrillators (S-ICDs) are attractive for preventing sudden cardiac death in hypertrophic cardiomyopathy (HCM) as they mitigate risks of transvenous leads in young patients. However, S-ICDs may be associated with increased inappropriate shock (IAS) in HCM patients. OBJECTIVE: The purpose of this study was to assess the incidence and predictors of appropriate shock and IAS in a contemporary HCM S-ICD cohort. METHODS: We collected electrocardiographic and clinical data from HCM patients who underwent S-ICD implantation at 4 centers. Etiologies of all S-ICD shocks were adjudicated. We used Firth penalized logistic regression to derive adjusted odds ratios (aORs) for predictors of IAS. RESULTS: Eighty-eight HCM patients received S-ICDs (81 for primary and 7 for secondary prevention) with a mean follow-up of 2.7 years. Five patients (5.7%) had 9 IAS episodes (3.8 IAS per 100 patient-years) most often because of sinus tachycardia and/or T-wave oversensing. Independent predictors of IAS were higher 12-lead electrocardiographic R-wave amplitude (aOR 2.55 per 1 mV; 95% confidence interval 1.15-6.38) and abnormal T-wave inversions (aOR 0.16; 95% confidence interval 0.02-0.97). There were 2 appropriate shocks in 7 secondary prevention patients and none in 81 primary prevention patients, despite 96% meeting Enhanced American College of Cardiology/American Heart Association criteria and the mean European HCM Risk-SCD score predicting 5.7% 5-year risk. No patients had sudden death or untreated sustained ventricular arrhythmias. CONCLUSION: In this multicenter HCM S-ICD study, IAS were rare and appropriate shocks confined to secondary prevention patients. The R-wave amplitude increased IAS risk, whereas T-wave inversions were protective. HCM primary prevention implantable cardioverter-defibrillator guidelines overestimated the risk of appropriate shocks in our cohort.

Association of regional myocardial conduction velocity with the distribution of hypoattenuation on contrast-enhanced perfusion computed tomography in patients with postinfarct ventricular tachycardia.

BACKGROUND: Cardiac magnetic resonance imaging has been shown to be beneficial for identification of the ventricular tachycardia (VT) substrate before catheter ablation. Contrast-enhanced perfusion multidetector computed tomography (CEP-MDCT) is more generalizable to clinical practice, and wall thickness and regional hypoenhancement on CEP-MDCT can identify potential substrate sites, albeit with decreased specificity. OBJECTIVE: The purpose of this study was to evaluate the association between wall thickness and attenuation on CEP-MDCT with local conduction velocity (CV) and electrogram abnormalities in patients with postinfarct VT. METHODS: Fourteen patients with postinfarct VT underwent preprocedural CEP-MDCT followed by endocardial electroanatomic mapping (EAM) and ablation. Myocardial attenuation and wall thickness were calculated from 3-dimensional MDCT images using ADAS-VT software (Galgo Medical). EAM was registered with 3-dimensional MDCT images using the CartoMERGE module of CARTO3 software (Biosense Webster). Local CV was calculated by averaging the velocity between each point and 5 adjacent points with concordant wavefront direction. RESULTS: A total of 3689 points were included. In multivariable regression analysis clustered by patient, local CV was positively associated with myocardial attenuation, bipolar voltage, unipolar voltage, and wall thickness. Each 10-HU drop in full-thickness attenuation correlated to 2.6% decrease in CV (P <.001) and 5.5% decrease in bipolar voltage amplitude (P <.001), after adjusting for wall thickness. CONCLUSION: Myocardial attenuation distribution on CEP-MDCT is associated with regional CV and electrogram amplitude. Regions with low CV identified with low attenuation on CEP-MDCT may serve as important VT substrates in postinfarct patients.

Epicardial Conduction Speed, Electrogram Abnormality, and Computed Tomography Attenuation Associations in Arrhythmogenic Right Ventricular Cardiomyopathy.

OBJECTIVES: This study sought to evaluate the association between contrast-enhanced multidetector computed tomography (CE-MDCT) attenuation and local epicardial conduction speed (ECS) and electrographic abnormalities in patients with arrhythmogenic right ventricular cardiomyopathy (ARVC) and ventricular tachycardia (VT). BACKGROUND: CE-MDCT is a widely available and fast imaging technology with high spatial resolution that is less prone to defibrillator generator-related safety issues and image artifacts. However, the association between hypoattenuation on MDCT and VT substrates in ARVC remains unknown. METHODS: Patients with ARVC who underwent CE-MDCT followed by endocardial (n = 30) and epicardial (n = 21) electroanatomical mapping (EAM) and VT ablation were prospectively enrolled. Right ventricular (RV) mid-myocardial attenuation was calculated from 3-dimensional MDCT images and registered to EAM. Local ECS was calculated by averaging the ECS between each point and 5 adjacent points with concordant wave front direction. RESULTS: A total of 17,311 epicardial and 5,204 endocardial points were included. In multivariable regression analysis clustered by patient, RV myocardial attenuation was associated with epicardial bipolar voltage amplitude (2.5% decrease in amplitude per 10 HU decrease in attenuation; p < 0.001), with endocardial unipolar voltage amplitude (0.9% decrease in amplitude per 10 HU decrease in attenuation; p < 0.001), and with ECS (0.4% decrease in ECS per 10 HU decrease in attenuation; p = 0.001). CONCLUSIONS: CE-MDCT attenuation distribution is associated with regional ECS and electrographic amplitude in ARVC. Regions with low attenuation likely reflect fibro-fatty involvement in the RV and may serve as important VT substrates in patients with ARVC who are undergoing VT ablation.

Rapid Golgi analysis method for efficient and unbiased classification of dendritic spines.

Dendritic spines are the primary recipients of excitatory synaptic input in the brain. Spine morphology provides important information on the functional state of ongoing synaptic transmission. One of the most commonly used methods to visualize spines is Golgi-Cox staining, which is appealing both due to ease of sample preparation and wide applicability to multiple species including humans. However, the classification of spines is a time-consuming and often expensive task that yields widely varying results between individuals. Here, we present a novel approach to this analysis technique that uses the unique geometry of different spine shapes to categorize spines on a purely objective basis. This rapid Golgi spine analysis method successfully conveyed the maturational shift in spine types during development in the mouse primary visual cortex. This approach, built upon freely available software, can be utilized by researchers studying a broad range of synaptic connectivity phenotypes in both development and disease.