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2.
Circ Arrhythm Electrophysiol ; 11(10): e006740, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-30354404

RESUMO

BACKGROUND: Glucagon-like peptide-1 receptor (GLP-1R) agonists improve cardiovascular outcomes in patients with type 2 diabetes mellitus. However, systemic actions of these agents cause sympathetic activation, which is generally considered to be detrimental in cardiovascular disease. Despite significant research interest in cardiovascular biology of GLP-1, the presence of GLP-1R in ventricular cardiomyocytes remains a controversial issue, and the effects of this peptide on the electrical properties of intact ventricular myocardium are unknown. We sought to determine the effects of GLP-1R agonist exendin-4 (Ex4) on ventricular action potential duration (APD) and susceptibility to ventricular arrhythmia in the rat heart in vivo and ex vivo. METHODS: Ventricular monophasic action potentials were recorded in anaesthetized (urethane) rats in vivo and isolated perfused rat hearts during sinus rhythm and ventricular pacing. RESULTS: In vivo, systemic administration of Ex4 (5 µg/kg intravenously) increased heart rate, and this effect was abolished by ß-adrenoceptor blockade. Despite causing sympathetic activation, Ex4 increased APD at 90% repolarization during ventricular pacing by 7% ( P=0.044; n=6) and reversed the effect of ß-adrenoceptor agonist dobutamine on APD at 90% repolarization. In isolated perfused hearts, Ex4 (3 nmol/L) increased APD at 90% repolarization by 14% ( P=0.015; n=6) with no effect on heart rate. Ex4 also reduced ventricular arrhythmia inducibility in conditions of ß-adrenoceptor stimulation with isoproterenol. Ex4 effects on APD and ventricular arrhythmia susceptibility were prevented in conditions of muscarinic receptor blockade or inhibition of nitric oxide synthase. CONCLUSIONS: These data demonstrate that GLP-1R activation effectively opposes the effects of ß-adrenoceptor stimulation on cardiac ventricular excitability and reduces ventricular arrhythmic potential. The effect of GLP-1R activation on the ventricular myocardium is indirect, mediated by acetylcholine and nitric oxide and, therefore, can be explained by stimulation of cardiac parasympathetic (vagal) neurons.

3.
J Cardiovasc Dev Dis ; 5(4)2018 Sep 23.
Artigo em Inglês | MEDLINE | ID: mdl-30249045

RESUMO

Haploinsufficiency of the T-box transcription factor TBX1 is responsible for many features of 22q11.2 deletion syndrome. Tbx1 is expressed dynamically in the pharyngeal apparatus during mouse development and Tbx1 homozygous mutants display numerous severe defects including abnormal cranial ganglion formation and neural crest cell defects. These abnormalities prompted us to investigate whether parasympathetic (vagal) innervation of the heart was affected in Tbx1 mutant embryos. In this report, we used an allelic series of Tbx1 mouse mutants, embryo tissue explants and cardiac electrophysiology to characterise, in detail, the function of Tbx1 in vagal innervation of the heart. We found that total nerve branch length was significantly reduced in Tbx1+/- and Tbx1neo2/- mutant hearts expressing 50% and 15% levels of Tbx1. We also found that neural crest cells migrated normally to the heart of Tbx1+/-, but not in Tbx1neo2 mutant embryos. In addition, we showed that cranial ganglia IXth and Xth were fused in Tbx1neo2/- but neuronal differentiation appeared intact. Finally, we used telemetry to monitor heart response to carbachol, a cholinergic receptor agonist, and found that heart rate recovered more quickly in Tbx1+/- animals versus controls. We speculate that this condition of decreased parasympathetic drive could result in a pro-arrhythmic substrate in some 22q11.2DS patients.

4.
J Cereb Blood Flow Metab ; : 271678X18780602, 2018 06 04.
Artigo em Inglês | MEDLINE | ID: mdl-29862863

RESUMO

KIR6.1 (KCNJ8) is a subunit of ATP sensitive potassium channel (KATP) that plays an important role in the control of peripheral vascular tone and is highly expressed in brain contractile cells (vascular smooth muscle cells and pericytes). This study determined the effect of global deletion of the KIR6.1 subunit on cerebral blood flow, neurovascular coupling and cerebral oxygenation in mice. In KIR6.1 deficient mice resting cerebral blood flow and brain parenchymal partial pressure of oxygen ( PO2) were found to be markedly lower compared to that in their wildtype littermates. However, cortical blood oxygen level dependent responses triggered by visual stimuli were not affected in conditions of KIR6.1 deficiency. These data suggest that KATP channels containing KIR6.1 subunit are critically important for the maintenance of normal cerebral perfusion and parenchymal PO2 but play no significant role in the mechanisms underlying functional changes in brain blood flow.

5.
Lab Chip ; 18(12): 1736-1749, 2018 06 12.
Artigo em Inglês | MEDLINE | ID: mdl-29762619

RESUMO

Circulating tumor cells (CTCs) are malignant cells released into the bloodstream with the potential to form metastases in secondary sites. These cells, acquired non-invasively, represent a sample of highly relevant tumor tissue that is an alternative to difficult and low-yield tumor biopsies. In recent years, there has been growing interest in genomic profiling of CTCs to enable longitudinal monitoring of the tumor's adaptive response to therapy. However, due to their extreme rarity, genotyping CTCs has proved challenging. Relevant mutations can be masked by leukocyte contamination in isolates. Heterogeneity between subpopulations of tumor cells poses an additional obstacle. Recent advances in single-cell sequencing can overcome these limitations but isolation of single CTCs is prone to cell loss and is prohibitively difficult and time consuming. To address these limitations, we developed a single cell sample preparation and genome sequencing pipeline that combines biophysical enrichment and single cell isolation using laser capture microdissection (LCM). A key component of this process is the encapsulation of enriched CTC sample in a hydrogel matrix, which enhances the efficiency of single-cell isolation by LCM, and is compatible with downstream sequencing. We validated this process by sequencing of single CTCs and cell free DNA (cfDNA) from a single patient with castration resistant prostate cancer. Identical mutations were observed in prostate cancer driver genes (TP53, PTEN, FOXA1) in both single CTCs and cfDNA. However, two independently isolated CTCs also had identical missense mutations in the genes for ATR serine/threonine kinase, KMT2C histone methyltransferase, and FANCC DNA damage repair gene. These mutations may be missed by bulk sequencing libraries, whereas single cell sequencing could potentially enable the characterization of key CTC subpopulations that arise during metastasis.


Assuntos
Separação Celular/métodos , Microdissecção e Captura a Laser/métodos , Células Neoplásicas Circulantes , Análise de Sequência de DNA/métodos , Análise de Célula Única/métodos , Cápsulas , Linhagem Celular Tumoral , Genômica , Humanos , Hidrogéis , Dispositivos Lab-On-A-Chip , Masculino , Mutação , Neoplasias da Próstata/genética
6.
Int J Cardiol ; 260: 82-87, 2018 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-29622458

RESUMO

BACKGROUND: Almost 1/3 of heart failure patients fail to respond to cardiac resynchronization therapy (CRT). A simple clinical score to predict who these patients are at the moment of referral or at time of implant may be of importance for early optimization of their management. METHODS: Observational study. A risk score was derived from factors associated to CRT response. The derivation cohort was composed of 1301 patients implanted with a CRT defibrillator in a multi-center French cohort-study. External validation of this score and assessment of its association with CRT response and all-cause mortality and/or heart transplant was performed in 1959 CRT patients implanted in 4 high-volume European centers. RESULTS: Independent predictors of CRT response in the derivation cohort were: female gender (OR = 2.08, 95% CI 1.26-3.45), NYHA class ≤ III (OR = 2.71, 95% CI 1.63-4.52), left ventricular ejection fraction ≥ 25% (OR = 1.75, 95% CI 1.27-2.41), QRS duration ≥ 150 ms (OR = 1.70, 95% CI 1.25-2.30) and estimated glomerular filtration rate ≥ 60 mL/min (OR = 2.01, 95% CI 1.48-2.72). Each was assigned 1 point. External validation showed good calibration (Hosmer-Lemeshow test-P = 0.95), accuracy (Brier score = 0.19) and discrimination (c-statistic = 0.67), with CRT response increasing progressively from 37.5% in patients with a score of 0 to 91.9% among those with score of 5 (Gamma for trend = 0.44, P < 0.001). Similar results were observed regarding all-cause mortality or heart transplant. CONCLUSION: The ScREEN score (Sex category, Renal function, ECG/QRS width, Ejection fraction and NYHA class) is composed of widely validated, easy to obtain predictors of CRT response, and predicts CRT response and overall mortality. It should be helpful in facilitating early consideration of alternative therapies for predicted non-responders to CRT therapy.


Assuntos
Terapia de Ressincronização Cardíaca/tendências , Insuficiência Cardíaca/fisiopatologia , Insuficiência Cardíaca/terapia , Idoso , Feminino , Seguimentos , Taxa de Filtração Glomerular/fisiologia , Insuficiência Cardíaca/diagnóstico , Humanos , Masculino , Pessoa de Meia-Idade , Valor Preditivo dos Testes , Fatores de Risco , Resultado do Tratamento
7.
Glia ; 66(6): 1185-1199, 2018 06.
Artigo em Inglês | MEDLINE | ID: mdl-29274121

RESUMO

Astrocytes support neuronal function by providing essential structural and nutritional support, neurotransmitter trafficking and recycling and may also contribute to brain information processing. In this article we review published results and report new data suggesting that astrocytes function as versatile metabolic sensors of central nervous system (CNS) milieu and play an important role in the maintenance of brain metabolic homeostasis. We discuss anatomical and functional features of astrocytes that allow them to detect and respond to changes in the brain parenchymal levels of metabolic substrates (oxygen and glucose), and metabolic waste products (carbon dioxide). We report data suggesting that astrocytes are also sensitive to circulating endocrine signals-hormones like ghrelin, glucagon-like peptide-1 and leptin, that have a major impact on the CNS mechanisms controlling food intake and energy balance. We discuss signaling mechanisms that mediate communication between astrocytes and neurons and consider how these mechanisms are recruited by astrocytes activated in response to various metabolic challenges. We review experimental data suggesting that astrocytes modulate the activities of the respiratory and autonomic neuronal networks that ensure adaptive changes in breathing and sympathetic drive in order to support the physiological and behavioral demands of the organism in ever-changing environmental conditions. Finally, we discuss evidence suggesting that altered astroglial function may contribute to the pathogenesis of disparate neurological, respiratory and cardiovascular disorders such as Rett syndrome and systemic arterial hypertension.


Assuntos
Astrócitos/metabolismo , Encéfalo/metabolismo , Animais , Humanos
8.
Integr Biol (Camb) ; 9(6): 519-528, 2017 06 19.
Artigo em Inglês | MEDLINE | ID: mdl-28524208

RESUMO

Hemolytic anemia is one of the hallmarks of malaria and leads to an increase in oxidized heme (hemin) within the plasma of infected individuals. While scavenger proteins sequester much of the circulating heme, it has been hypothesized that extracellular heme may play a central role in malaria pathogenesis. We have previously developed the multiplex fluidic plunger (MFP) device for the measurement of red blood cell (RBC) deformability. Here, we demonstrate that the measurement of changes in RBC deformability is a sensitive method for inferring heme-induced oxidative stress. We further show that extracellular hemin concentration correlates closely with changes in RBC deformability and we confirm that this biophysical change correlates with other indicators of cell stress. Finally, we show that reduced erythrocyte deformability corresponds with both erythrophagocytosis and RBC osmotic fragility. The MFP microfluidic device presents a simple and potentially inexpensive alternative to existing methods for measuring hemolytic cell stress that could ultimately be used to perform clinical assessment of disease progression in severe malaria.


Assuntos
Deformação Eritrocítica/fisiologia , Eritrócitos/parasitologia , Hemina/metabolismo , Plasmodium falciparum/patogenicidade , Adulto , Anemia Hemolítica/sangue , Anemia Hemolítica/etiologia , Anemia Hemolítica/parasitologia , Fenômenos Biofísicos , Desenho de Equipamento , Hemólise/fisiologia , Humanos , Técnicas In Vitro , Dispositivos Lab-On-A-Chip , Malária Falciparum/sangue , Malária Falciparum/complicações , Malária Falciparum/parasitologia , Técnicas Analíticas Microfluídicas , Fragilidade Osmótica/fisiologia , Estresse Oxidativo , Fagocitose/fisiologia , Fosfatidilserinas/sangue , Adulto Jovem
9.
Physiol Rep ; 4(15)2016 08.
Artigo em Inglês | MEDLINE | ID: mdl-27528004

RESUMO

The heart is controlled by the sympathetic and parasympathetic limbs of the autonomic nervous system with inhibitory signaling mechanisms recruited in both limbs. The aim of this study was to determine the role of inhibitory heterotrimeric G proteins in the central nervous mechanisms underlying autonomic control of the heart and its potential role in arrhythmogenesis. Mice with conditional deletion of the inhibitory heterotrimeric G protein GαO in the presympathetic area of the rostral ventral lateral medulla (RVLM) were generated to determine the role of GαO-mediated signalling in autonomic control and electrophysiological properties of the heart. GαO deletion within the RVLM was not associated with changes in heart rate (HR) or the arterial blood pressure at rest (home cage, normal behavior). However, exposure to stressful conditions (novel environment, hypoxia, or hypercapnia) in these mice was associated with abnormal HR responses and an increased baroreflex gain when assessed under urethane anesthesia. This was associated with shortening of the ventricular effective refractory period. This phenotype was reversed by systemic beta-adrenoceptor blockade, suggesting that GαO depletion in the RVLM increases central sympathetic drive. The data obtained support the hypothesis that GαO-mediated signaling within the presympathetic circuits of the RVLM contributes to the autonomic control of the heart. GαO deficiency in the RVLM has a significant impact on cardiovascular responses to stress, cardiovascular reflexes and electrical properties of the heart.


Assuntos
Sistema Nervoso Autônomo/fisiologia , Subunidades alfa Gi-Go de Proteínas de Ligação ao GTP/fisiologia , Coração/fisiologia , Bulbo/fisiologia , Animais , Pressão Sanguínea , Subunidades alfa Gi-Go de Proteínas de Ligação ao GTP/genética , Frequência Cardíaca , Hemodinâmica , Hipercapnia/fisiopatologia , Hipóxia/fisiopatologia , Camundongos , Camundongos Transgênicos , Respiração , Transdução de Sinais , Função Ventricular
10.
Clin Med (Lond) ; 16(3): 267-71, 2016 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-27251918

RESUMO

Atrial fibrillation is driven by spontaneous electrical activation emerging from the pulmonary veins. Catheter ablation using either radiofrequency or cryothermal energy electrically isolates these veins from the left atrium, both reducing the burden of atrial fibrillation episodes and improving the patient's symptoms. Catheter ablation is superior to antiarryhthmic drugs when patients are carefully selected. Underlying medical problems - including obesity, hypertension and obstructive sleep apnoea - should be optimally treated before considering ablation. Although this treatment has the potential to cure patients of their symptoms, they should be aware of the important associated procedural complications.


Assuntos
Fibrilação Atrial , Ablação por Cateter , Fibrilação Atrial/fisiopatologia , Fibrilação Atrial/cirurgia , Ablação por Cateter/efeitos adversos , Ablação por Cateter/métodos , Humanos , Complicações Pós-Operatórias
11.
Small ; 12(14): 1909-19, 2016 Apr 13.
Artigo em Inglês | MEDLINE | ID: mdl-26917414

RESUMO

Circulating tumor cells (CTCs) offer tremendous potential for the detection and characterization of cancer. A key challenge for their isolation and subsequent analysis is the extreme rarity of these cells in circulation. Here, a novel label-free method is described to enrich viable CTCs directly from whole blood based on their distinct deformability relative to hematological cells. This mechanism leverages the deformation of single cells through tapered micrometer scale constrictions using oscillatory flow in order to generate a ratcheting effect that produces distinct flow paths for CTCs, leukocytes, and erythrocytes. A label-free separation of circulating tumor cells from whole blood is demonstrated, where target cells can be separated from background cells based on deformability despite their nearly identical size. In doping experiments, this microfluidic device is able to capture >90% of cancer cells from unprocessed whole blood to achieve 10(4) -fold enrichment of target cells relative to leukocytes. In patients with metastatic castration-resistant prostate cancer, where CTCs are not significantly larger than leukocytes, CTCs can be captured based on deformability at 25× greater yield than with the conventional CellSearch system. Finally, the CTCs separated using this approach are collected in suspension and are available for downstream molecular characterization.


Assuntos
Microfluídica/instrumentação , Células Neoplásicas Circulantes , Humanos
12.
J Biomech ; 48(15): 4065-72, 2015 Nov 26.
Artigo em Inglês | MEDLINE | ID: mdl-26477408

RESUMO

A key challenge in transfusion medicine research and clinical hematology is to develop a simple and non-destructive method to measure the quality of each blood unit prior to use. RBC deformability has long been proposed as an indicator of blood quality. We measured RBC deformability using the pressure required for single cells to transit through a micrometer scale constriction to examine longitudinal changes in RBC deformability, as well as the variability in blood quality and storage capacity across donors. We used a microfluidic device to monitor deformability changes in RBCs stored in plastic tubes and in blood bags over 14 and 56 days respectively. We found consistent storage based degradation of RBC deformability with statistically significant variability in both the initial RBC deformability and storage capacity among donors. Furthermore, all samples exhibited a transient recovery phenomenon. Deformability profiling of stored RBCs using transiting pressure showed significant donor variability in initial quality and storage capacity. This measurement approach shows promise as a rapid method to individually assess the quality of stored RBC units.


Assuntos
Criopreservação , Deformação Eritrocítica , Eritrócitos/fisiologia , Coleta de Amostras Sanguíneas , Forma Celular , Eritrócitos/ultraestrutura , Feminino , Humanos , Masculino , Técnicas Analíticas Microfluídicas , Pressão , Caracteres Sexuais
13.
Curr Cardiol Rep ; 17(9): 631, 2015 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-26266757

RESUMO

Catheter ablation is superior to antiarrhythmic drugs in maintaining sinus rhythm for patients with atrial fibrillation (AF). Pulmonary vein (PV) isolation is the cornerstone of any AF ablation procedure. Conventionally, this is achieved by performing point by point lesions using radiofrequency (RF) energy. However, this is technically challenging, time consuming and is associated with a number of complications. Long-term durability of PV isolation is also a concern. To address these issues, 'one-shot' energy delivery systems and alternative energy sources have been developed. The cryoballoon system has emerged as the most commonly used alternative to point by point RF technology. In this paper, we compare the technology, biophysics and clinical data of cryoballoon to conventional RF ablation for AF. The safety and efficacy of cryoballoon compared to RF ablation is critically reviewed. We conclude by looking at future applications of this technology.


Assuntos
Antiarrítmicos/uso terapêutico , Fibrilação Atrial/terapia , Ablação por Cateter , Criocirurgia , Veias Pulmonares/patologia , Fibrilação Atrial/fisiopatologia , Humanos , Veias Pulmonares/inervação , Resultado do Tratamento
14.
Int J Cardiol ; 195: 253-8, 2015 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-26048388

RESUMO

OBJECTIVE: We tested the hypothesis that pulmonary vein (PV) measurements on pre-procedural CT/MR imaging can predict difficulty in isolation and phrenic nerve (PN) injury during cryoballoon ablation for paroxysmal atrial fibrillation (AF). METHODS: Consecutive patients with paroxysmal AF who had pre-procedural CT/MRI and underwent cryoballoon ablation as part of a randomized trial were studied. Imaging was anonymized for blinded analysis of: (1) maximum ostial diameter, (2) minimum ostial diameter, (3) ostial area and (4) ratio of maximum over minimum ostial diameter (eccentricity index). Veins that required more than 2 freezes of at least 200 s duration to isolate or not isolated were defined as difficult to isolate. Loss of PN pacing during right-sided ablation was defined as PN injury. Logistic regression was used to analyze the predictive effect of the measurements on the 2 outcomes. RESULTS: 148 PVs in 38 patients (aged 60 ± 11 years, 76% male) were analyzed. Left inferior PV (LIPV) was most difficult to isolate with 23 out of 37 PVs (62%), and PN injury occurred in 3 of 38 (8%) right superior PV (RSPV). Greater eccentricity index predicted difficulty in isolating LIPV, OR 40.33 (95% CI 1.40 to 1160, p = 0.03) and smaller eccentricity index predicted PN injury in RSPV, OR 0.01 (95% CI 0.01-0.16, p = 0.001). CONCLUSIONS: Eccentricity index measured from pre-procedural CT/MR imaging can predict difficulty of PV isolation and PN injury during cryoballoon ablation for paroxysmal AF.


Assuntos
Fibrilação Atrial/cirurgia , Criocirurgia , Complicações Intraoperatórias/prevenção & controle , Traumatismos dos Nervos Periféricos , Nervo Frênico/lesões , Veias Pulmonares , Idoso , Criocirurgia/efeitos adversos , Criocirurgia/métodos , Feminino , Humanos , Modelos Logísticos , Imagem por Ressonância Magnética/métodos , Masculino , Pessoa de Meia-Idade , Traumatismos dos Nervos Periféricos/etiologia , Traumatismos dos Nervos Periféricos/prevenção & controle , Cuidados Pré-Operatórios/métodos , Veias Pulmonares/diagnóstico por imagem , Veias Pulmonares/cirurgia , Tomografia Computadorizada por Raios X/métodos , Resultado do Tratamento
15.
Heart Rhythm ; 12(11): 2285-93, 2015 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-26051529

RESUMO

BACKGROUND: The central nervous origins of functional parasympathetic innervation of cardiac ventricles remain controversial. OBJECTIVE: This study aimed to identify a population of vagal preganglionic neurons that contribute to the control of ventricular excitability. An animal model of synuclein pathology relevant to Parkinson's disease was used to determine whether age-related loss of the activity of the identified group of neurons is associated with changes in ventricular electrophysiology. METHODS: In vivo cardiac electrophysiology was performed in anesthetized rats in conditions of selective inhibition of the dorsal vagal motor nucleus (DVMN) neurons by pharmacogenetic approach and in mice with global genetic deletion of all family members of the synuclein protein. RESULTS: In rats anesthetized with urethane (in conditions of systemic beta-adrenoceptor blockade), muscarinic and neuronal nitric oxide synthase blockade confirmed the existence of a tonic parasympathetic control of cardiac excitability mediated by the actions of acetylcholine and nitric oxide. Acute DVMN silencing led to shortening of the ventricular effective refractory period (vERP), a lowering of the threshold for triggered ventricular tachycardia, and prolongation of the corrected QT (QTc) interval. Lower resting activity of the DVMN neurons in aging synuclein-deficient mice was found to be associated with vERP shortening and QTc interval prolongation. CONCLUSION: Activity of the DVMN vagal preganglionic neurons is responsible for tonic parasympathetic control of ventricular excitability, likely to be mediated by nitric oxide. These findings provide the first insight into the central nervous substrate that underlies functional parasympathetic innervation of the ventricles and highlight its vulnerability in neurodegenerative diseases.


Assuntos
Ventrículos do Coração/inervação , Óxido Nítrico Sintase Tipo I/metabolismo , Estimulação do Nervo Vago , Nervo Vago/fisiologia , Animais , Eletrofisiologia Cardíaca , Modelos Animais de Doenças , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Parassimpatectomia/métodos , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley , Estatísticas não Paramétricas
17.
Lab Chip ; 15(10): 2278-86, 2015 May 21.
Artigo em Inglês | MEDLINE | ID: mdl-25876237

RESUMO

The enumeration and capture of circulating tumor cells (CTCs) are potentially of great clinical value as they offer a non-invasive means to access tumor materials to diagnose disease and monitor treatment efficacy. Conventional immunoenrichment of CTCs may fail to capture cells with low surface antigen expression. Micropore filtration presents a compelling label-free alternative that enriches CTCs using their biophysical rather than biochemical characteristics. However, this strategy is prone to clogging of the filter microstructure, which dramatically reduces the selectivity after processing large numbers of cells. Here, we use the resettable cell trap (RCT) mechanism to separate cells based on their size and deformability using an adjustable aperture that can be periodically cleared to prevent clogging. After separation, the output sample is stained and analyzed using multi-spectral analysis, which provides a more sensitive and unambiguous method to identify CTC biomarkers than traditional immunofluorescence. We tested the RCT device using blood samples obtained from 22 patients with metastatic castrate-resistant prostate cancer while comparing the results with the established CellSearch® system. The RCT mechanism was able to capture ≥5 CTCs in 18/22 (82%) patients with a mean count of 257 in 7.5 ml of whole blood, while the CellSearch system found ≥5 CTCs in 9/22 (41%) patients with a mean count of 25. The ~10× improvement in the CTC capture rate provides significantly more materials for subsequent analysis of these cells such as immunofluorescence, propagation by tissue culture, and genetic profiling.


Assuntos
Separação Celular , Dispositivos Lab-On-A-Chip , Células Neoplásicas Circulantes , Neoplasias de Próstata Resistentes à Castração , Separação Celular/instrumentação , Separação Celular/métodos , Tamanho Celular , Humanos , Masculino , Células Neoplásicas Circulantes/metabolismo , Células Neoplásicas Circulantes/patologia , Neoplasias de Próstata Resistentes à Castração/metabolismo , Neoplasias de Próstata Resistentes à Castração/patologia
18.
Hypertension ; 65(4): 775-83, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25712724

RESUMO

Systemic arterial hypertension has been previously suggested to develop as a compensatory condition when central nervous perfusion/oxygenation is compromised. Principal sympathoexcitatory C1 neurons of the rostral ventrolateral medulla oblongata (whose activation increases sympathetic drive and the arterial blood pressure) are highly sensitive to hypoxia, but the mechanisms of this O2 sensitivity remain unknown. Here, we investigated potential mechanisms linking brainstem hypoxia and high systemic arterial blood pressure in the spontaneously hypertensive rat. Brainstem parenchymal PO2 in the spontaneously hypertensive rat was found to be ≈15 mm Hg lower than in the normotensive Wistar rat at the same level of arterial oxygenation and systemic arterial blood pressure. Hypoxia-induced activation of rostral ventrolateral medulla oblongata neurons was suppressed in the presence of either an ATP receptor antagonist MRS2179 or a glycogenolysis inhibitor 1,4-dideoxy-1,4-imino-d-arabinitol, suggesting that sensitivity of these neurons to low PO2 is mediated by actions of extracellular ATP and lactate. Brainstem hypoxia triggers release of lactate and ATP which produce excitation of C1 neurons in vitro and increases sympathetic nerve activity and arterial blood pressure in vivo. Facilitated breakdown of extracellular ATP in the rostral ventrolateral medulla oblongata by virally-driven overexpression of a potent ectonucleotidase transmembrane prostatic acid phosphatase results in a significant reduction in the arterial blood pressure in the spontaneously hypertensive rats (but not in normotensive animals). These results suggest that in the spontaneously hypertensive rat, lower PO2 of brainstem parenchyma may be associated with higher levels of ambient ATP and l-lactate within the presympathetic circuits, leading to increased central sympathetic drive and concomitant sustained increases in systemic arterial blood pressure.


Assuntos
Pressão Sanguínea/fisiologia , Tronco Encefálico/irrigação sanguínea , Hipertensão/etiologia , Hipóxia-Isquemia Encefálica/complicações , Trifosfato de Adenosina/sangue , Animais , Tronco Encefálico/metabolismo , Modelos Animais de Doenças , Progressão da Doença , Feminino , Hipertensão/metabolismo , Hipertensão/fisiopatologia , Hipóxia-Isquemia Encefálica/metabolismo , Hipóxia-Isquemia Encefálica/fisiopatologia , Ácido Láctico/metabolismo , Masculino , Ratos , Ratos Endogâmicos SHR , Ratos Wistar , Sistema Nervoso Simpático/metabolismo , Sistema Nervoso Simpático/fisiopatologia
19.
Lab Chip ; 15(1): 159-67, 2015 Jan 07.
Artigo em Inglês | MEDLINE | ID: mdl-25325848

RESUMO

The extraordinary deformability of red blood cells gives them the ability to repeatedly transit through the microvasculature of the human body. The loss of this capability is part of the pathology of a wide range of diseases including malaria, hemoglobinopathies, and micronutrient deficiencies. We report on a technique for multiplexed measurements of the pressure required to deform individual red blood cell through micrometer-scale constrictions. This measurement is performed by first infusing single red blood cells into a parallel array of ~1.7 µm funnel-shaped constrictions. Next, a saw-tooth pressure waveform is applied across the constrictions to squeeze each cell through its constriction. The threshold deformation pressure is then determined by relating the pressure-time data with the video of the deformation process. Our key innovation is a self-compensating fluidic network that ensures identical pressures are applied to each cell regardless of its position, as well as the presence of cells in neighboring constrictions. These characteristics ensure the consistency of the measurement process and robustness against blockages of the constrictions by rigid cells and debris. We evaluate this technique using in vitro cultures of RBCs infected with P. falciparum, the parasite that causes malaria, to demonstrate the ability to profile the deformability signature of a heterogeneous sample.


Assuntos
Deformação Eritrocítica/fisiologia , Eritrócitos , Técnicas Analíticas Microfluídicas/instrumentação , Desenho de Equipamento , Eritrócitos/citologia , Eritrócitos/parasitologia , Eritrócitos/fisiologia , Humanos , Malária Falciparum/fisiopatologia , Técnicas Analíticas Microfluídicas/métodos , Plasmodium falciparum
20.
Hypertension ; 64(3): 523-9, 2014 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-24914196

RESUMO

ATP-sensitive potassium channels (KATP) regulate a range of biological activities by coupling membrane excitability to the cellular metabolic state. In particular, it has been proposed that KATP channels and specifically, the channel subunits Kir6.1 and SUR2B, play an important role in the regulation of vascular tone. However, recent experiments have suggested that KATP channels outside the vascular smooth muscle compartment are the key determinant of the observed behavior. Thus, we address the importance of the vascular smooth muscle KATP channel, using a novel murine model in which it is possible to conditionally delete the Kir6.1 subunit. Using a combination of molecular, electrophysiological, in vitro, and in vivo techniques, we confirmed the absence of Kir6.1 and KATP currents and responses specifically in smooth muscle. Mice with conditional deletion of Kir6.1 showed no obvious arrhythmic phenotype even after provocation with ergonovine. However, these mice were hypertensive and vascular smooth muscle cells failed to respond to vasodilators in a normal fashion. Thus, Kir6.1 underlies the vascular smooth muscle KATP channel and has a key role in vascular reactivity and blood pressure control.


Assuntos
Pressão Sanguínea/fisiologia , Canais KATP/fisiologia , Músculo Liso Vascular/fisiologia , Animais , Peptídeo Relacionado com Gene de Calcitonina/farmacologia , Modelos Animais de Doenças , Hipertensão/fisiopatologia , Técnicas In Vitro , Canais KATP/deficiência , Canais KATP/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Músculo Liso Vascular/citologia , Músculo Liso Vascular/efeitos dos fármacos , Técnicas de Patch-Clamp , Pinacidil/farmacologia , Vasodilatadores/farmacologia
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