Comparison of implicit and explicit FSI coupling strategies in cardiovascular system = Comparación de las estrategias implícitas y explícitas de acoplamiento FSI en el sistema cardiovascular

Comparación de los esquemas de integración temporal explícito e implícito, en la simulación del flujo sanguíneo y su interacción con la pared arterial. There are two major strategies in FSI coupling techniques: implicit and explicit. The general difference between these methodologies is how many times the data is exchanged between the fluid and solid domains at each FSI time-step. In both coupling strategies, the pressure values coming from fluid domain calculations at each time-step are exported to the solid domain, and consequently, the solid domain is analyzed with these imported forces. In contrast to the explicit coupling, in the implicit approach the fluid and solid domain’s data is exchanged several times until the convergence is achieved. Although this method may boost the numerical stabilization, it increases the computational cost due to the extra data exchanges. In cardiovascular simulations, depending on the analysis objectives, one may choose an explicit or implicit approach. In the current work, the advantage of an explicit coupling strategy is highlighted when simulation of pulsatile blood flow in elastic arteries is desired.

Pulmonary Artery-Vein Segmentation in Real and Synthetic CT Images = Segmentación Arteria-Vena Pulmonares en Imágenes de CT Reales y Sintéticas

La tomografía axial computerizada (TAC) es la modalidad de imagen médica preferente para el estudio de enfermedades pulmonares y el análisis de su vasculatura. La segmentación general de vasos en pulmón ha sido abordada en profundidad a lo largo de los últimos años por la comunidad científica que trabaja en el campo de procesamiento de imagen; sin embargo, la diferenciación entre irrigaciones arterial y venosa es aún un problema abierto. De hecho, la separación automática de arterias y venas está considerado como uno de los grandes retos futuros del procesamiento de imágenes biomédicas. La segmentación arteria-vena (AV) permitiría el estudio de ambas irrigaciones por separado, lo cual tendría importantes consecuencias en diferentes escenarios médicos y múltiples enfermedades pulmonares o estados patológicos. Características como la densidad, geometría, topología y tamaño de los vasos sanguíneos podrían ser analizados en enfermedades que conllevan remodelación de la vasculatura pulmonar, haciendo incluso posible el descubrimiento de nuevos biomarcadores específicos que aún hoy en dípermanecen ocultos. Esta diferenciación entre arterias y venas también podría ayudar a la mejora y el desarrollo de métodos de procesamiento de las distintas estructuras pulmonares. Sin embargo, el estudio del efecto de las enfermedades en los árboles arterial y venoso ha sido inviable hasta ahora a pesar de su indudable utilidad. La extrema complejidad de los árboles vasculares del pulmón hace inabordable una separación manual de ambas estructuras en un tiempo realista, fomentando aún más la necesidad de diseñar herramientas automáticas o semiautomáticas para tal objetivo. Pero la ausencia de casos correctamente segmentados y etiquetados conlleva múltiples limitaciones en el desarrollo de sistemas de separación AV, en los cuales son necesarias imágenes de referencia tanto para entrenar como para validar los algoritmos. Por ello, el diseño de imágenes sintéticas de TAC pulmonar podría superar estas dificultades ofreciendo la posibilidad de acceso a una base de datos de casos pseudoreales bajo un entorno restringido y controlado donde cada parte de la imagen (incluyendo arterias y venas) está unívocamente diferenciada. En esta Tesis Doctoral abordamos ambos problemas, los cuales están fuertemente interrelacionados. Primero se describe el diseño de una estrategia para generar, automáticamente, fantomas computacionales de TAC de pulmón en humanos. Partiendo de conocimientos a priori, tanto biológicos como de características de imagen de CT, acerca de la topología y relación entre las distintas estructuras pulmonares, el sistema desarrollado es capaz de generar vías aéreas, arterias y venas pulmonares sintéticas usando métodos de crecimiento iterativo, que posteriormente se unen para formar un pulmón simulado con características realistas. Estos casos sintéticos, junto a imágenes reales de TAC sin contraste, han sido usados en el desarrollo de un método completamente automático de segmentación/separación AV. La estrategia comprende una primera extracción genérica de vasos pulmonares usando partículas espacio-escala, y una posterior clasificación AV de tales partículas mediante el uso de Graph-Cuts (GC) basados en la similitud con arteria o vena (obtenida con algoritmos de aprendizaje automático) y la inclusión de información de conectividad entre partículas. La validación de los fantomas pulmonares se ha llevado a cabo mediante inspección visual y medidas cuantitativas relacionadas con las distribuciones de intensidad, dispersión de estructuras y relación entre arterias y vías aéreas, los cuales muestran una buena correspondencia entre los pulmones reales y los generados sintéticamente. La evaluación del algoritmo de segmentación AV está basada en distintas estrategias de comprobación de la exactitud en la clasificación de vasos, las cuales revelan una adecuada diferenciación entre arterias y venas tanto en los casos reales como en los sintéticos, abriendo así un amplio abanico de posibilidades en el estudio clínico de enfermedades cardiopulmonares y en el desarrollo de metodologías y nuevos algoritmos para el análisis de imágenes pulmonares. ABSTRACT Computed tomography (CT) is the reference image modality for the study of lung diseases and pulmonary vasculature. Lung vessel segmentation has been widely explored by the biomedical image processing community, however, differentiation of arterial from venous irrigations is still an open problem. Indeed, automatic separation of arterial and venous trees has been considered during last years as one of the main future challenges in the field. Artery-Vein (AV) segmentation would be useful in different medical scenarios and multiple pulmonary diseases or pathological states, allowing the study of arterial and venous irrigations separately. Features such as density, geometry, topology and size of vessels could be analyzed in diseases that imply vasculature remodeling, making even possible the discovery of new specific biomarkers that remain hidden nowadays. Differentiation between arteries and veins could also enhance or improve methods processing pulmonary structures. Nevertheless, AV segmentation has been unfeasible until now in clinical routine despite its objective usefulness. The huge complexity of pulmonary vascular trees makes a manual segmentation of both structures unfeasible in realistic time, encouraging the design of automatic or semiautomatic tools to perform the task. However, this lack of proper labeled cases seriously limits in the development of AV segmentation systems, where reference standards are necessary in both algorithm training and validation stages. For that reason, the design of synthetic CT images of the lung could overcome these difficulties by providing a database of pseudorealistic cases in a constrained and controlled scenario where each part of the image (including arteries and veins) is differentiated unequivocally. In this Ph.D. Thesis we address both interrelated problems. First, the design of a complete framework to automatically generate computational CT phantoms of the human lung is described. Starting from biological and imagebased knowledge about the topology and relationships between structures, the system is able to generate synthetic pulmonary arteries, veins, and airways using iterative growth methods that can be merged into a final simulated lung with realistic features. These synthetic cases, together with labeled real CT datasets, have been used as reference for the development of a fully automatic pulmonary AV segmentation/separation method. The approach comprises a vessel extraction stage using scale-space particles and their posterior artery-vein classification using Graph-Cuts (GC) based on arterial/venous similarity scores obtained with a Machine Learning (ML) pre-classification step and particle connectivity information. Validation of pulmonary phantoms from visual examination and quantitative measurements of intensity distributions, dispersion of structures and relationships between pulmonary air and blood flow systems, show good correspondence between real and synthetic lungs. The evaluation of the Artery-Vein (AV) segmentation algorithm, based on different strategies to assess the accuracy of vessel particles classification, reveal accurate differentiation between arteries and vein in both real and synthetic cases that open a huge range of possibilities in the clinical study of cardiopulmonary diseases and the development of methodological approaches for the analysis of pulmonary images.

Fluid-solid interaction in arteries incorporating the autoregulation concept in boundary conditions

In pre-surgery decisions in hospital emergency cases, fast and reliable results of the solid and fluid mechanics problems are of great interest to clinicians. In the current investigation, an iterative process based on a pressure-type boundary condition is proposed in order to reduce the computational costs of blood flow simulations in arteries, without losing control of the important clinical parameters. The incorporation of cardiovascular autoregulation, together with the well-known impedance boundary condition, forms the basis of the proposed methodology. With autoregulation, the instabilities associated with conventional pressure-type or impedance boundary conditions are avoided without an excessive increase in computational costs. The general behaviour of pulsatile blood flow in arteries, which is important from the clinical point of view, is well reproduced through this new methodology. In addition, the interaction between the blood and the arterial walls occurs via a modified weak coupling, which makes the simulation more stable and computationally efficient. Based on in vitro experiments, the hyperelastic behaviour of the wall is characterised and modelled. The applications and benefits of the proposed pressure-type boundary condition are shown in a model of an idealised aortic arch with and without an ascending aorta dissection, which is a common cardiovascular disorder.

Microvascular and tissue oxygen gradients in the rat mesentery

One of the most important functions of the blood circulation is O2 delivery to the tissue. This process occurs primarily in microvessels that also regulate blood flow and are the site of many metabolic processes that require O2. We measured the intraluminal and perivascular pO2 in rat mesenteric arterioles in vivo by using noninvasive phosphorescence quenching microscopy. From these measurements, we calculated the rate at which O2 diffuses out of microvessels from the blood. The rate of O2 efflux and the O2 gradients found in the immediate vicinity of arterioles indicate the presence of a large O2 sink at the interface between blood and tissue, a region that includes smooth muscle and endothelium. Mass balance analyses show that the loss of O2 from the arterioles in this vascular bed primarily is caused by O2 consumption in the microvascular wall. The high metabolic rate of the vessel wall relative to parenchymal tissue in the rat mesentery suggests that in addition to serving as a conduit for the delivery of O2 the microvasculature has other functions that require a significant amount of O2.

Static and dynamic lengths of neutrophil microvilli

Containing most of the L-selectin and P-selectin glycoprotein ligand-1 (PSGL-1) on their tips, microvilli are believed to promote the initial arrest of neutrophils on endothelium. At the rolling stage following arrest, the lifetimes of the involved molecular bonds depend on the pulling force imposed by the shear stress of blood flow. With two different methods, electron microscopy and micropipette manipulation, we have obtained two comparable neutrophil microvillus lengths, both ≈0.3 μm in average. We have found also that, under a pulling force, a microvillus can be extended (microvillus extension) or a long thin membrane cylinder (a tether) can be formed from it (tether formation). If the force is ≤34 pN (± 3 pN), the length of the microvillus will be extended; if the force is >61 pN (± 5 pN), a tether will be formed from the microvillus at a constant velocity, which depends linearly on the force. When the force is between 34 pN and 61 pN (transition zone), the degree of association between membrane and cytoskeleton in individual microvilli will dictate whether microvillus extension or tether formation occurs. When a microvillus is extended, it acts like a spring with a spring constant of ≈43 pN/μm. In contrast to a rigid or nonextendible microvillus, both microvillus extension and tether formation can decrease the pulling force imposed on the adhesive bonds, and thus prolonging the persistence of the bonds at high physiological shear stresses.

Force-mediated kinetics of single P-selectin/ligand complexes observed by atomic force microscopy

Leukocytes roll along the endothelium of postcapillary venules in response to inflammatory signals. Rolling under the hydrodynamic drag forces of blood flow is mediated by the interaction between selectins and their ligands across the leukocyte and endothelial cell surfaces. Here we present force-spectroscopy experiments on single complexes of P-selectin and P-selectin glycoprotein ligand-1 by atomic force microscopy to determine the intrinsic molecular properties of this dynamic adhesion process. By modeling intermolecular and intramolecular forces as well as the adhesion probability in atomic force microscopy experiments we gain information on rupture forces, elasticity, and kinetics of the P-selectin/P-selectin glycoprotein ligand-1 interaction. The complexes are able to withstand forces up to 165 pN and show a chain-like elasticity with a molecular spring constant of 5.3 pN nm−1 and a persistence length of 0.35 nm. The dissociation constant (off-rate) varies over three orders of magnitude from 0.02 s−1 under zero force up to 15 s−1 under external applied forces. Rupture force and lifetime of the complexes are not constant, but directly depend on the applied force per unit time, which is a product of the intrinsic molecular elasticity and the external pulling velocity. The high strength of binding combined with force-dependent rate constants and high molecular elasticity are tailored to support physiological leukocyte rolling.

Modulation of cognition-specific cortical activity by gonadal steroids: A positron-emission tomography study in women

There is considerable evidence from animal studies that gonadal steroid hormones modulate neuronal activity and affect behavior. To study this in humans directly, we used H215O positron-emission tomography to measure regional cerebral blood flow (rCBF) in young women during three pharmacologically controlled hormonal conditions spanning 4–5 months: ovarian suppression induced by the gonadotropin-releasing hormone agonist leuprolide acetate (Lupron), Lupron plus estradiol replacement, and Lupron plus progesterone replacement. Estradiol and progesterone were administered in a double-blind cross-over design. On each occasion positron-emission tomography scans were performed during (i) the Wisconsin Card Sorting Test, a neuropsychological test that physiologically activates prefrontal cortex (PFC) and an associated cortical network including inferior parietal lobule and posterior inferolateral temporal gyrus, and (ii) a no-delay matching-to-sample sensorimotor control task. During treatment with Lupron alone (i.e., with virtual absence of gonadal steroid hormones), there was marked attenuation of the typical Wisconsin Card Sorting Test activation pattern even though task performance did not change. Most strikingly, there was no rCBF increase in PFC. When either progesterone or estrogen was added to the Lupron regimen, there was normalization of the rCBF activation pattern with augmentation of the parietal and temporal foci and return of the dorsolateral PFC activation. These data directly demonstrate that the hormonal milieu modulates cognition-related neural activity in humans.

Potential roles of osteopontin and αVβ3 integrin in the development of coronary artery restenosis after angioplasty

Angioplasty procedures are increasingly used to reestablish blood flow in blocked atherosclerotic coronary arteries. A serious complication of these procedures is reocclusion (restenosis), which occurs in 30–50% of patients. Migration of coronary artery smooth muscle cells (CASMCs) to the site of injury caused by angioplasty and subsequent proliferation are suggested mechanisms of reocclusion. Using both cultured human CASMCs and coronary atherectomy tissues, we studied the roles of osteopontin (OPN) and one of its receptors, αvβ3 integrin, in the pathogenesis of coronary restenosis. We also measured the plasma levels of OPN before and after angioplasty and determined the effect of exogenous OPN on CASMC migration, extracellular matrix invasion, and proliferation. We found that cultured CASMCs during log phase of growth and smooth muscle cell layer of the coronary atherosclerotic tissues of patients express both OPN mRNA and protein at a significantly elevated level compared with controls. Interestingly, whereas the baseline plasma OPN levels in control samples were virtually undetectable, those in patient plasma were remarkably high. We also found that interaction of OPN with αvβ3 integrin, expressed on CASMCs, causes migration, extracellular matrix invasion, and proliferation. These effects were abolished when OPN or αvβ3 integrin gene expression in CASMCs was inhibited by specific antisense S-oligonucleotide treatment or OPN-αvβ3 interaction was blocked by treatment of CASMCs with antibodies against OPN or αvβ3 integrin. Our results demonstrate that OPN and αvβ3 integrin play critical roles in regulating cellular functions deemed essential for restenosis. In addition, these results raise the possibility that transient inhibition of OPN gene expression or blocking of OPN-αvβ3 interaction may provide a therapeutic approach to preventing restenosis.

Imaging of H217O distribution in the brain of a live rat by using proton-detected 17O MRI

Imaging of H217O has a number of important applications. Mapping the distribution of H217O produced by oxidative metabolism of 17O-enriched oxygen gas may lead to a new method of metabolic functional imaging; regional cerebral blood flow also can be measured by measuring the H217O distribution after the injection of 17O-enriched physiological saline solution. Previous studies have proposed a method for indirect detection of 17O. The method is based on the shortening of the proton T2 in H217O solutions, caused by the residual 17O-1H scalar coupling and transferred to the bulk water via fast chemical exchange. It has been shown that the proton T2 of H217O solutions can be restored to that of H216O by irradiating the resonance frequency of the 17O nucleus. The indirect 17O image thus is obtained by taking the difference between two T2-weighted spin-echo images: one acquired after irradiation of the 17O resonance and one acquired without irradiation. It also has been established that, at relatively low concentrations of H217O, the indirect method yields an image that quantitatively reflects the H217O distribution in the sample. The method is referred to as PRIMO (proton imaging of oxygen). In this work, we show in vivo proton images of the H217O distribution in a rat brain after an i.v. injection of H217O-enriched physiological saline solution. Implementing the indirect detection method in an echo-planar imaging sequence enabled obtaining H217O images with good spatial and temporal resolution of few seconds.

Positron-emission tomography imaging of long-term shape recognition challenges

Long-term visual memory performance was impaired by two types of challenges: a diazepam challenge on acquisition and a sensory challenge on recognition. Using positron-emission tomography regional cerebral blood flow imaging, we studied the effect of these challenges on regional brain activation during the delayed recognition of abstract visual shapes as compared with a baseline fixation task. Both challenges induced a significant decrease in differential activation in the left fusiform gyrus, suggesting that this region is involved in the automatic or volitional comparison of incoming and stored stimuli. In contrast, thalamic differential activation increased in response to memory challenges. This increase might reflect enhanced retrieval attempts as a compensatory mechanism for restoring recognition performance.

Suppression of protein synthesis in brain during hibernation involves inhibition of protein initiation and elongation

Protein synthesis (PS) has been considered essential to sustain mammalian life, yet was found to be virtually arrested for weeks in brain and other organs of the hibernating ground squirrel, Spermophilus tridecemlineatus. PS, in vivo, was below the limit of autoradiographic detection in brain sections and, in brain extracts, was determined to be 0.04% of the average rate from active squirrels. Further, it was reduced 3-fold in cell-free extracts from hibernating brain at 37°C, eliminating hypothermia as the only cause for protein synthesis inhibition (active, 0.47 ± 0.08 pmol/mg protein per min; hibernator, 0.16 ± 0.05 pmol/mg protein per min, P < 0.001). PS suppression involved blocks of initiation and elongation, and its onset coincided with the early transition phase into hibernation. An increased monosome peak with moderate ribosomal disaggregation in polysome profiles and the greatly increased phosphorylation of eIF2α are both consistent with an initiation block in hibernators. The elongation block was demonstrated by a 3-fold increase in ribosomal mean transit times in cell-free extracts from hibernators (active, 2.4 ± 0.7 min; hibernator, 7.1 ± 1.4 min, P < 0.001). No abnormalities of ribosomal function or mRNA levels were detected. These findings implicate suppression of PS as a component of the regulated shutdown of cellular function that permits hibernating ground squirrels to tolerate “trickle” blood flow and reduced substrate and oxygen availability. Further study of the factors that control these phenomena may lead to identification of the molecular mechanisms that regulate this state.

Impaired metabolic modulation of α-adrenergic vasoconstriction in dystrophin-deficient skeletal muscle

The neuronal isoform of nitric oxide synthase (nNOS) is highly expressed in mammalian skeletal muscle, but its functional role has not been defined. NO has been implicated in the local metabolic regulation of blood flow in contracting skeletal muscle in part by antagonizing sympathetic vasoconstriction. We therefore hypothesized that nNOS in skeletal muscle is the source of the NO mediating the inhibition of sympathetic vasoconstriction in contracting muscle. In the mdx mouse, a model of Duchenne muscular dystrophy in which dystrophin deficiency results in greatly reduced expression of nNOS in skeletal muscle, we found that the normal ability of skeletal muscle contraction to attenuate α-adrenergic vasoconstriction is defective. Similar results were obtained in mutant mice that lack the gene encoding nNOS. Together these data suggest a specific role for nNOS in the local metabolic inhibition of α-adrenergic vasoconstriction in active skeletal muscle.

Effects of in vivo adventitial expression of recombinant endothelial nitric oxide synthase gene in cerebral arteries

Nitric oxide (NO), synthesized from l-arginine by NO synthases (NOS), plays an essential role in the regulation of cerebrovascular tone. Adenoviral vectors have been widely used to transfer recombinant genes to different vascular beds. To determine whether the recombinant endothelial NOS (eNOS) gene can be delivered in vivo to the adventitia of cerebral arteries and functionally expressed, a replication-incompetent adenoviral vector encoding eNOS gene (AdCMVNOS) or β-galactosidase reporter gene (AdCMVLacZ) was injected into canine cerebrospinal fluid (CSF) via the cisterna magna (final viral titer in CSF, 109 pfu/ml). Adventitial transgene expression was demonstrated 24 h later by β-galactosidase histochemistry and quantification, eNOS immunohistochemistry, and Western blot analysis of recombinant eNOS. Electron microscopy immunogold labeling indicated that recombinant eNOS protein was expressed in adventitial fibroblasts. In AdCMVNOS-transduced arteries, basal cGMP production and bradykinin-induced relaxations were significantly augmented when compared with AdCMVLacZ-transduced vessels (P < 0.05). The increased receptor-mediated relaxations and cGMP production were inhibited by eNOS inhibitors. In addition, the increase in cGMP production was reversed in the absence of calcium, suggesting that the increased NO production did not result from inducible NOS expression. The present study demonstrates the successful in vivo transfer and functional expression of recombinant eNOS gene in large cerebral arteries. It also suggests that perivascular eNOS gene delivery via the CSF is a feasible approach that does not require interruption of cerebral blood flow.

Noninvasive measurement of gene expression in skeletal muscle

We have developed a noninvasive detection method for expression of viral-mediated gene transfer. A recombinant adenovirus was constructed by using the gene for arginine kinase (AK), which is the invertebrate correlate to the vertebrate ATP-buffering enzyme, creatine kinase. Gene expression was noninvasively monitored using 31P-magnetic resonance spectroscopy (31P-MRS). The product of the AK enzyme, phosphoarginine (PArg), served as an MRS-visible reporter of AK expression. The recombinant adenovirus coding for arginine kinase (rAdCMVAK) was injected into the right hindlimbs of neonatal mice. Two weeks after injection of rAdCMVAK, a unique 31P-MRS resonance was observed. It was observable in all rAdCMVAK injected hindlimbs and was not present in the contralateral control or the vehicle injected limb. PArg and phosphocreatine (PCr) concentrations were calculated to be 11.6 ± 0.90 and 13.6 ± 1.1 mM respectively in rAdCMVAK injected limbs. AK activity was demonstrated in vivo by monitoring the decreases in PArg and ATP resonances during prolonged ischemia. After 1 h of ischemia intracellular pH was 6.73 ± 0.06, PCr/ATP was decreased by 77 ± 8%, whereas PArg/ATP was decreased by 50 ± 15% of basal levels. PArg and PCr returned to basal levels within 5 min of the restoration of blood flow. AK activity persisted for at least 8 mo after injection, indicating that adenoviral-mediated gene transfer can produce stable expression for long periods of time. Therefore, the cDNA encoding AK provides a useful reporter gene that allows noninvasive and repeated monitoring of gene expression after viral mediated gene transfer to muscle.

Orbitofrontal cortex: A key prefrontal region for encoding information

Little is known about the specific functional contribution of the human orbitofrontal cortex with regard to memory processing, although there is strong evidence from lesion studies in monkeys that it may play an important role. The present investigation measured changes in regional cerebral blood flow with positron emission tomography in normal human subjects who were instructed to commit to memory abstract visual patterns. The results indicated that the rostral orbitofrontal region (area 11), which is primarily linked with the anterior medial temporal limbic region and lateral prefrontal cortical areas, is involved in the process of encoding of new information.