La circulation artérielle coronaire est un système complexe dont les méthodes de mesures invasivespermettent une évaluation en pratique clinique.Matériels et méthodes Nous développons, dans deux revues, les principes et méthodes des différentes techniques invasivesde mesure du flux coronaire en pratique clinique. Puis nous étudions l’impact clinique de l’utilisationde la Fractional Flow Reserve (FFR) dans l’évaluation des sténoses coronaires intermédiaires, lesmoyens pharmacologiques pour mesurer la FFR et sa corrélation avec l’évaluation angiographique enfonction des facteurs de risque cardiovasculaires. Enfin, nous décrivons les principes et méthodesd’une technique de mesure du flux coronaire segmentaire permettant d’obtenir la FFR, le flux et lesrésistances absolues avec un microcathéter de perfusion qui, sur un principe de thermodilutionpermet d’évaluer distinctement la macro et la microcirculation coronaire.Résultats Nous recommandons une valeur seuil de FFR de 0,80 pour guider la revascularisation car le nombred’événements cardiovasculaires et la mortalité sont spontanément supérieurs chez les patients avecune FFR <0,80 comparativement à une FFR ≥0,80 (9,4 vs. 4,8%, P=0,06 et 7,5 vs. 3,2%, P=0,06;respectivement). Nous avons étudié différents agents hyperémiants permettant de mesurer la FFR:l’adénosine (100 μg à 200 μg) permettant d’obtenir une hyperémie maximale, et le produit decontraste permettant d’atteindre 65% de cette hyperémie maximale. La mesure de la FFR avec duproduit de contraste permet de meilleures performances diagnostiques que les indices de reposcomparé à la FFR sous adénosine. Nous avons établi que la corrélation entre la FFR et le degré desténose angiographique est faible et inversement proportionnel au nombre de facteurs de risquecardiovasculaires, particulièrement chez les patients diabétiques. Enfin, nous avons décrit dans troisétudes, le principe de thermodilution coronaire et la méthode de mesures du flux coronaire et desrésistances microvasculaires avec un microcathéter de perfusion intracoronaire spécifique. Nousavons montré que cette technique est précise (R=0,98), qu’elle induit une hyperémie maximale etlocale sans agent hyperémiant et quelle est reproductible chez l’homme (R=0,91).Conclusion La compréhension de la circulation coronaire et l’application chez l’homme des techniques demesure du flux coronaire segmentaire sont essentielles tant en pratique clinique courante qu’enrecherche. / Coronary circulation is complex and highly regulated while invasive coronary flow measurements techniques allow the assessment of coronary physiology in clinical practice. Material et methods We describe in two reviews the principles and methods of different invasive coronary flowmeasurements techniques in clinical practice. We study the clinical impact of fractional flow reserve(FFR) in intermediate coronary stenosis, the hyperemic agents and dosage to measure FFR and FFRcorrelation with angiographic indices according to risk factors accumulation. Finally, we describe the principle and method of coronary flow and microvascular resistances measurements with a dedicated infusion microcatheter for coronary thermodilution to obtain assessment of macro and microvascular components of coronary circulation. Results We recommend the FFR cut off value of 0.80 to guide revascularization based on our study showing higher myocardial infarction and death rate in patients treated with medical therapy and FFR<0.80compared to those with FFR>0.80, respectively 9.4 versus 4.8%, P=0.06 and 7,5 versus 3,2%, P=0.06. We studied different hyperemic agents and dosages and showed that intracoronary adenosine at 100μg to 200 μg induce maximal hyperemia while contrast medium induce 65% of maximal hyperemia. Therefore, FFR measurements with contrast medium is feasible and has better accuracy than restindices compared to FFR. We establish the weak correlation between FFR and angiographic indicesand weakens correlation as risk factors accumulates, especially in diabetic patients. Finally, we described in three studies the method of absolute coronary flow and microvascular resistancesmeasurements based on thermodilution principle with a dedicated infusion catheter. We showed anaccurate measurement with this technique (R=0.98), which induces maximal hyperemia without theneed of hyperemic agent with reproducible measurements in humans (R=0,91).Conclusion The use of invasive coronary flow measurements to study the coronary circulation is essential inclinical practice and in research.

Call number: LD2668 .T4 1985 T47 / Master of Science

Retina is a unique site in the human body where the microcirculation can be imaged directly and non-invasively, allowing us to study in vivo the structure and pathology of the human microcirculation. Retinal images can be quantitatively assessed with computerized imaging techniques, enabling us to measure several different quantitative traits derived from the retinal vasculature. Arterial and venular calibres are the most extensively studied traits of the retinal microvasculature and numerous epidemiological studies demonstrated promising associations with systemic and ocular diseases as well as with disease markers. However, there has been a lack of research into pathophysiological processes leading to retinal vascular signs, and how they link retinal microcirculation with coronary and cerebral microvasculature change. Information about genetic determinants underlying retinal vascular structure is therefore important for understanding the processes leading to microvascular pathophysiology. Two genome wide association studies have been published so far revealing four loci associated with retinal venular calibre and one locus with arteriolar calibre. Here the results from the genome-wide association analysis of 10 different retinal vessel traits in two population based cohorts are presented. Retinal images were measured in non-mydriatic fundus images from 808 subjects in the Orkney Complex Disease Study (ORCADES) and 390 subjects from the Croatian island of Korcula, using the semi-automated retinal vasculature measurement programme SIVA and VAMPIRE. Using pairwise estimates of kinship based on genomic sharing, heritability was calculated for each trait. Estimates of tortuosity measure and fractal dimensions present first published reports of heritability estimates for those traits. In addition correlation analysis with systemic risk factor was also completed, confirming already published results as well as revealing some new associations. A genome wide association analysis of retinal arteriolar width revealed a genome wide significant hit (1.8x10-7) in a region of chromosome 2q32 (within TTN gene). Replication was sought in a further independent Scottish population (LBC) and additional 400 retinal images were graded. The result did not replicate, however the direction of the effect was consistent and a larger sample size is required. Analysis of the remaining traits did not yield genome wide significant result,s and will also require larger sample sizes. Genetic analysis of a binary retinal trait was also explored in a case control study of retinal detachment, which is an important cause of vision loss. A two-stage genetic association discovery phase followed by a replication phase in a combined total of 2,833 RRD cases and 7,871 controls was carried out. None of the SNPs tested in the discovery phase reached the threshold for association. Further testing was carried out in independent case-control series from London (846 cases) and Croatia (120 cases). The combined meta-analysis identified one association reaching genome-wide significance for rs267738 (OR=1.29, p=2.11x10-8), a missense coding SNP and eQTL for CERS2 encoding the protein ceramide synthase 2. Additional genetic risk score, pathway analysis and genetic liability analysis were also carried out.

Coronary artery bypass graft (CABG) surgery is a common treatment for patients with coronary artery disease. A potential complication of CABG is myocardial ischemia or infarction. In this thesis, a method - based on laser Doppler flowmetry (LDF) - for detection of intra- and postoperative ischemia by myocardial perfusion monitoring is evaluated. LDF is sensitive to motion artifacts. In previous studies, a method for reduction of motion artifacts when measuring on the beating heart has been developed. By using the ECG as a reference, the perfusion signal is measured in intervals during the cardiac cycle where the cardiac motion is at a minimum, thus minimizing the artifacts in the perfusion signal. The aim of this thesis was to investigate the possibilities to use the ECG-triggered laser Doppler system for continuous monitoring of myocardial perfusion in humans during and after CABG surgery. Two studies were performed. In the first study, changes in myocardial perfusion during CABG surgery were investigated (n = 13), while the second study focused on postoperative measurements (n = 13). In addition, an ECG-triggering method was implemented and evaluated. It was found that the large variations in myocardial perfusion during CABG surgery could be monitored with the ECG-triggered laser Doppler system. Furthermore, a perfusion signal of good quality could be registered postoperatively from the closed chest in ten out of thirteen patients. In eight out of ten patients, a proper signal was obtained also the following morning, i.e., about 20 hours after probe insertion. The results show that respiration and blood pressure can have an influence on the perfusion signal. In conclusion, the results indicate that the method is able to detect fluctuations in myocardial perfusion under favourable circumstances. However, high heart rate, abnormal cardiac motion, improper probe attachment and limitations in the ECG-triggering method may result in variations in the perfusion signal that are not related to tissue perfusion. / Varje år utförs omkring 4500 kranskärlsoperationer i Sverige. En allvarlig komplikation som kan uppstå efter operationen är otillräcklig blodförsörjning till hjärtmuskeln. Den här licentiatavhandlingen handlar om utveckling och utvärdering av en metod, baserad på laserdopplerteknik, för att kunna upptäcka nedsatt blodperfusion i hjärtmuskeln på ett tidigt stadium. Laserdopplertekniken är känslig för rörelsestörningar. I tidigare studier har en metod för reducering av rörelsestörningar vid mätning på slående hjärta tagits fram. Med EKG:t som referens mäts blodperfusionen i de faser under hjärtcykeln då hjärtats rörelse är som minst, vilket minskar bidraget av rörelsestörningar i blodperfusionssignalen. I den här avhandlingen undersöks om metoden kan användas för kontinuerlig övervakning av hjärtmuskelns blodperfusion på patienter under och efter hjärtoperationer. Två studier har genomförts: en där hjärtmuskelns perfusion mättes i olika faser under kranskärlsoperationer och en där mätproben lades in i hjärtmuskeln under operationen och mätningar gjordes under det första dygnet efter operationen. Det visade sig vara möjligt att följa förändringar i hjärtmuskelns blodperfusion under operation. Det var även möjligt att registrera en perfusionssignal av god kvalitet efter operationen då bröstkorgen var stängd. Hos åtta av tio patienter erhölls en bra signal även morgonen efter operationen, dvs. ca 20 timmar efter att proben lades in. Resultaten visar också att andning och blodtryck kan ha en påverkan på blodperfusionssignalen. Slutsatsen av arbetet är att det går att se variationer i hjärtmuskelns blodperfusion med EKG-triggad laserdoppler under vissa förutsättningar. Signalen är dock i många fall svårtolkad på grund av att t ex hög hjärtfrekvens, onormal hjärtväggsrörelse eller ändrad probposition sannolikt kan ge variationer i perfusionssignalen som inte är relaterade till blodflödesförändringar. / Report code: LIU-TEK-LIC-2007:35.

This study explored hydrostatic pressure as a mechanobiological parameter to control in vitro endothelial cell tubulogenesis in 3-D hydrogels as a model microvascular tissue engineering approach. For this purpose, the present investigation used an endothelial spheroid model, which we believe is an adaptable microvascularization strategy for many tissue engineering construct designs. We also aimed to identify the operating magnitudes and exposure times for hydrostatic pressure-sensitive sprout formation as well as verify the involvement of VEGFR-3 signaling. For this purpose, we used a custom-designed pressure system and a 3-D endothelial cell spheroid model of sprouting tubulogenesis. We report that an exposure time of 3 days is the minimum duration required to increase endothelial sprout formation in response to 20 mmHg. Notably, exposure to 5 mmHg for 3 days was inhibitory for endothelial spheroid lengths without affecting sprout numbers. Moreover, endothelial spheroids exposed to 40 mmHg also inhibited sprouting activity by reducing sprout numbers without affecting sprout lengths. Finally, blockade of VEGFR-3 signaling abolished the effects of the 20-mmHg stimuli on sprout formation. Based on these results, VEGFR-3 dependent endothelial sprouting appears to exhibit a complex pressure dependence that one may exploit to control microvessel formation.

In normal tissues, blood supply is closely matched to tissue demand for wide ranges of oxygen demand and arterial pressure. This suggests that multiple mechanisms regulate blood flow. Theoretical models can be used to analyze these interacting mechanisms. One proposed mechanism for metabolic flow regulation involves the saturation-dependent release of ATP by red blood cells, which triggers an upstream conducted response signal and arteriolar vasodilation. To analyze this mechanism, oxygen and ATP levels are calculated along a flow pathway of seven representative segments, including two vasoactive arteriolar segments. The conducted response signal is dependent on ATP concentration. Arteriolar tone depends on the conducted response signal, local wall shear stress and wall tension. Arteriolar diameters are calculated based on vascular smooth muscle mechanics. The model can account for increases in perfusion consistent with experimental findings at low and moderate oxygen consumption rates despite the opposing effects of the myogenic and shear-dependent responses. Autoregulation, the maintenance of nearly constant blood flow as arterial pressure varies, is assessed in the presence or absence of the myogenic, shear-dependent and/or metabolic responses. The model results indicate that the combined effects of myogenic and metabolic regulation overcome the vasodilatory effect of the shear-dependent response to generate autoregulatory behavior. Capillary recruitment has been shown to increase the capacity for oxygen delivery during exercise. In the model, capillary density is assumed to depend on small arteriole diameter. The model predicts a significant increase in the range over which perfusion can be regulated when recruitment is included. Oscillations in diameter and tone are predicted under certain conditions, suggesting a novel mechanism for vasomotion. The conditions that give rise to oscillations are analyzed. It is shown that the appearance of oscillations depends in a complex way on a number of system parameters. In summary, the theoretical model provides a quantitative assessment of the myogenic, shear-dependent and metabolic responses that affect blood flow regulation and identifies a role for capillary recruitment and vasomotion in the control of blood flow.

Knowledge of the microcirculatory system has added significant value to the analysis of tissue oxygenation and perfusion. While developments in videomicroscopy technology have enabled medical researchers and physicians to observe the microvascular system, the available software tools are limited in their capabilities to determine quantitative features of microcirculation, either automatically or accurately. In particular, microvessel density has been a critical diagnostic measure in evaluating disease progression and a prognostic indicator in various clinical conditions. As a result, automated analysis of the microcirculatory system can be substantially beneficial in various real-time and off-line therapeutic medical applications, such as optimization of resuscitation. This study focuses on the development of an algorithm to automatically segment microvessels, calculate the density of capillaries in microcirculatory videos, and determine the distribution of blood circulation. The proposed technique is divided into four major steps: video stabilization, video enhancement, segmentation and post-processing. The stabilization step estimates motion and corrects for the motion artifacts using an appropriate motion model. Video enhancement improves the visual quality of video frames through preprocessing, vessel enhancement and edge enhancement. The resulting frames are combined through an adjusted weighted median filter and the resulting frame is then thresholded using an entropic thresholding technique. Finally, a region growing technique is utilized to correct for the discontinuity of blood vessels. Using the final binary results, the most commonly used measure for the assessment of microcirculation, i.e. Functional Capillary Density (FCD), is calculated. The designed technique is applied to video recordings of healthy and diseased human and animal samples obtained by MicroScan device based on Sidestream Dark Field (SDF) imaging modality. To validate the final results, the calculated FCD results are compared with the results obtained by blind detailed inspection of three medical experts, who have used AVA (Automated Vascular Analysis) semi-automated microcirculation analysis software. Since there is neither a fully automated accurate microcirculation analysis program, nor a publicly available annotated database of microcirculation videos, the results acquired by the experts are considered the gold standard. Bland-Altman plots show that there is ``Good Agreement" between the results of the algorithm and that of gold standard. In summary, the main objective of this study is to eliminate the need for human interaction to edit/ correct results, to improve the accuracy of stabilization and segmentation, and to reduce the overall computation time. The proposed methodology impacts the field of computer science through development of image processing techniques to discover the knowledge in grayscale video frames. The broad impact of this work is to assist physicians, medical researchers and caregivers in making diagnostic and therapeutic decisions for microcirculatory abnormalities and in studying of the human microcirculation.

In resting muscle, blood flow is regulated to meet the demand for O2 by the tissue. A modified ischemia (I)/reperfusion(R) investigation was systematically run and PISFO2, PaO2, Q and VO2 were observed. Twenty-nine spinotrapezius muscles from male Sprague-Dawley rats (284±20 grams) were surgically exteriorized for intravital microscopy to test a model relating blood flow, O2 supply and O2 demand. The model can aid in the understanding of the regulation of tissue PO2. The interstitial PO2 (PISFO2) and perivascular PO2 (PaO2) measurements were made using phosphorescence quenching microscopy (PQM). O2 consumption (VO2) values were obtained with a quasi-continuous, flash-synchronized, pressurized airbag to initiate ischemia and sample the rate of O¬2 change (dPO2/dt). Centerline red blood cell velocity was measured with an Optical Doppler Velocimeter and converted to flow using vessel diameter. 5-, 15-, 30-, 60-, 300- and 600-second ischemic durations were used to observe changes in PISFO2, Q, and VO2. A critical point was observed following 30 seconds of (I) where dPISFO2/dt during recovery was the fastest (4.25±0.72 mmHg/s) and was 1.00±0.16 mmHg/s following 600 seconds. Flow recovery, dQ/dt, peaked to 3.88±0.64 (µl•min-1)/s after 60 seconds of (I) but significantly dropped to 2.83±0.55 (µl•min-1)/s following 300 seconds of (I) but increased to 2.92±0.45 (µl•min-1)/s following 600 seconds. This gives evidence to a no-reflow phenomenon occurring in the extended periods of ischemia. A peak in VO¬2 to 309.2±45.0 nl O2/cm3•s with a time course of 160 seconds occurred following 600 seconds of ischemia. As the ischemic duration decreased, the time course and peak VO2 also decreased. VO2 following 300 seconds of (I) was significantly higher than 5-60 seconds of (I) (p <0.05) but was not significantly different from 600 seconds of (I). The information collected during the Q and VO2 studies can be incorporated into a factor, M, that relates VO2, Q and ∆PO2. M calculated for the recovery of 5- through 60-second (I) groups reasonably relates the three variables due to consistency and little variability. However, recovery in 600- and especially 300-second (I) groups showed higher variability in M which requires more consideration.

Doctor of Philosophy / Department of Anatomy and Physiology / David C. Poole / The overall aim of this dissertation is to better understand the mechanisms determining skeletal muscle oxygen delivery-utilization matching in health and disease. Emphasis is directed toward the role of nitric oxide (NO) bioavailability in modulating muscle microvascular oxygenation (PO2mv; the sole driving force for blood-myocyte oxygen flux) during transitions in metabolic demand. The first investigation of this dissertation (Chapter 2) demonstrates that alterations in NO bioavailability have a major impact on skeletal muscle PO2mv kinetics following both the onset and cessation of contractions. Specifically, increased NO levels (via the NO donor sodium nitroprusside; SNP) elevates whereas reduced NO levels (non-specific NOS inhibition with NG-nitro-L-arginine methyl ester; L-NAME) diminishes muscle PO2mv at the onset and during recovery from contractions in the spinotrapezius muscle of healthy young rats. Consistent with these results, inhibition of the neuronal NO synthase isoform (S-methyl-L-thiocitrulline; SMTC; Chapter 3) reveals alterations in NO-mediated regulation of skeletal muscle PO2mv with advanced age that likely contribute to exercise intolerance in this population. In Chapter 4 we observed that pronounced oxidative stress is implicated in these pathological responses seen in aged and diseased states. Transient elevations in the oxidant hydrogen peroxide to levels seen in the early stages of senescence and cardiovascular diseases promote detrimental effects on skeletal muscle contractile function (i.e., augmented oxygen cost of force production). Chapter 5 demonstrates that endurance exercise training improves skeletal muscle microvascular oxygenation (i.e., greater PO2mv and slower PO2mv kinetics) across the metabolic transient partly via enhanced NO-mediated function in healthy young individuals. These data carry important clinical implications given that exercise training may ameliorate NO-mediated function, muscle microvascular oxygenation deficits and consequently exercise intolerance in aged and diseased populations. In conclusion, alterations in NO bioavailability have a major impact on the dynamic balance between skeletal muscle oxygen delivery and utilization (i.e., PO2mv kinetics) in health and disease. While advanced age or the predations of disease impair considerably skeletal muscle microvascular oxygenation, exercise training-induced adaptations on the oxygen transport system constitute a non-pharmacological therapeutic intervention potentially capable of mitigating these microcirculatory deficits.

Soluble form of Junctional adhesion molecule C (sJAM-C) has been identified to cause angiogenesis as well as chemotaxis in endothelial cells. However, the role of sJAM-C in the context of cancer has not been elucidated. Our atomic force microscopy (AFM) stiffness measurements of normal mammary epithelial cells (MCF 10A) have shown a two-fold decrease in cell's stiffness in response to sJAM-C. Changes in cell stiffness are indicative of modulations in a cell's mechanical properties. Our results indicated that sJAM-C increased the MCF 10A cell migration about two-fold and also promoted a three-fold increase in chemotaxis. Additionally, sJAM-C treatment resulted in considerable filamentous-actin loss and peripheral actin ring breakage. We also found activation of Rho signaling pathway to be the main mechanism behind sJAM-C mediated alterations in MCF 10A cell cytoskeleton and motility. Our data present for the first time that sJAM-C is a pro metastatic mediator for normal mammary epithelial cells. / by Anila Qureshi. / Thesis (M.S.)--Florida Atlantic University, 2012. / Includes bibliography. / Mode of access: World Wide Web. / System requirements: Adobe Reader.