Die humane Lunge kann bei der Pharmakotherapie einer Erkrankung entweder als betroffenes Organ Ziel eines verabreichten Arzneistoffes sein oder aber auch als Portal für diesen in die systemische Zirkulation fungieren. Wird ein Arzneistoff inhaliert, ist für dessen Nutzen-Risiko-Profil von zentraler Bedeutung, in welchem Ausmaß und mit welcher Geschwindigkeit dieser resorbiert und anschließend in die systemische Zirkulation umverteilt wird. Wenn bei der Behandlung einer Lungenerkrankung dagegen ein Arzneistoff z.B. nach peroraler Gabe erst in der systemischen Zirkulation anflutet, müssen ausreichend hohe Wirkstoffkonzentrationen in den betroffenen Gewebearealen sichergestellt werden. Ziel der vorliegenden Arbeit war es daher, Möglichkeiten zu finden, diese beiden Vorgänge in vitro möglichst realitätsnah messen zu können. Für die Simulation der pulmonalen Absorption nach inhalativer Applikation eines Arzneistoffs diente Beclomethasondipropionat (BDP), freigesetzt aus den handelsüblichen FCKW-freien Dosieraerosolen Sanasthmax® und Ventolair®, als Modellsubstanz. Es wurde zunächst ein einfaches Dialysemodell als Screeningverfahren entwickelt. Hier wurden BDP-Partikel unter Verwendung der beiden Dosieraerosole auf humanem Lungenhomogenat deponiert und nachfolgend die kombinierten Prozesse aus Auflösung und Umverteilung der Substanz in eine Dialyseflüssigkeit, die sich entweder aus salinem Puffer oder humanem Blutplasma zusammensetzte, untersucht. Anschließend wurde erstmals ein etabliertes humanes Lungenperfusionsmodell dahingehend modifiziert, dass eine Inhalation von BDP nach Applikation eines handelsüblichen Dosieraerosols nachgestellt werden konnte. Auf diese Weise konnte an diesem realitätsnahen Modell die initiale Phase der pulmonalen Absorption von BDP in der Perfusionsflüssigkeit verfolgt werden. Beide Modelle zeigten Unterschiede in der Auflösungs- bzw. Umverteilungskinetik von BDP in Abhängigkeit von der verwendeten Applikationsform auf. So schienen sich von Ventolair® erzeugte BDP-Partikel schneller und in größerer Menge aufzulösen als diejenige bei den Versuchen mit Sanasthmax®, was eine vermehrte Umverteilung der Substanz sowohl in die Dialyseflüssigkeit als auch Perfusionslösung zur Folge hatte. Die am Lungenperfusionsmodell beobachteten Verläufe der initialen pulmonalen Absorption von BDP nach Freisetzung aus den Dosieraerosolen Sanasthmax® oder Ventolair® korrelierten dabei sehr gut mit Daten aus einer entsprechenden Humanstudie mit gesunden Probanden. Auch standen die ermittelten Unterschiede in sinnvoller Übereinstimmung mit Untersuchungen der in den von Sanasthmax® oder Ventolair® versprühten Aerosolen enthaltenen Partikel hinsichtlich Größenverteilung, Morphologie und Lösungsverhalten in Bronchialsekret. Um die Umverteilung eines Wirkstoffs von der systemischen Zirkulation in lungenspezifisches Gewebe am humanen Lungenperfusionsmodell zu simulieren, wurden die Gewebekonzentrationen von Thalidomid (THAL) in peripherem Lungengewebe im Vergleich zu den korrespondierenden Spiegeln in einem Bronchialkarzinom erstmals mittels Mikrodialyse verfolgt. Hierzu wurde im Vorfeld für diese Substanz unter Einsatz des Komplexbildners (2 Hydroxypropyl)-beta-cyclodextrin (HPCD) ein bezüglich der Sensitivität und der zeitlichen Auflösung optimiertes Mikrodialysesystem etabliert und dessen Eigenschaften systematisch untersucht. Am Lungenperfusionsmodell wurde dann eine an klinisch relevante Plasmaspiegel angelehnte THAL-Konzentration in der Perfusionslösung vorgelegt und anschließend das Anfluten in den oben genannten Geweben mit Hilfe des entwickelten Mikrodialysesystems beobachtet. Durch Zugabe von HPCD in das Mikrodialyseperfusat konnte eine signifikante Erhöhung der Wiederfindungsrate im Dialysat (Relative Recovery) erreicht und damit ein Mikrodialysesystem etabliert werden, das neben hoher zeitlicher Auflösung eine ausreichende analytische Sensitivität für THAL aufwies. Allerdings wurden aufgrund dieses Perfusatzusatzes die Diffusionsvorgänge während der Mikrodialyse derart beeinflusst, dass übliche Methoden zur Sondenkalibrierung wie z.B. die Retrodialyse nicht mehr angewendet werden konnten, und daher in Hinblick auf die Messungen am Lungenperfusionsmodell bestehende Kalibrierverfahren modifiziert werden mussten. Bei der Untersuchung der Gewebepenetration am Lungenperfusionsmodell flutete THAL in Tumorgewebe langsamer an als in peripherem Lungengewebe, wo schnell ähnliche Konzentrationen wie in der Perfusionslösung gefunden wurden. Auch lagen die Gewebespiegel im Tumorgewebe stets unter dem ermittelten Niveau im Lungengewebe. Die erhaltenen Konzentrationsverhältnisse zwischen Perfusionslösung, peripherem Lungengewebe und Tumorgewebe deckten sich dabei mit Kenntnissen aus Humanstudien, in denen analog Plasmakonzentrationen von antineoplastischen Substanzen ebenfalls mittels Mikrodialyse in Relation zu deren Spiegeln in gesundem Gewebe und Tumorgewebe verschiedenster Ätiologie bestimmt wurden. / In pharmacotherapy the human lung may either represent the therapeutic target site of an applied drug or be used as portal for systemic drug delivery. In case of inhalation of a drug the rate and extent of pulmonary drug absorption and subsequent distribution into systemic circulation is essential for the benefit-risk ratio. Otherwise, when a drug is systemically administered, e.g. by intravenous or oral route, to treat a lung disease and therefore first appears in the systemic circulation, sufficient drug concentrations have to be achieved in the affected tissue areals. Thus, the aim of this thesis was to find methods that allow to describe these two processes in vitro as close to reality as possible. Beclomethasone dipropionate (BDP) was chosen for the simulation of pulmonary drug absorption after administration of the two commercially available HFA-propelled metered dose inhalers (pMDI) Sanasthmax® and Ventolair®. Initially a simple dialysis model was established for screening tests. In this setting BDP particles were applied to human lung homogenate using those two inhalers and subsequently the combined processes of drug dissolution and distribution of the drug into dialysis fluid consisting of either saline buffer or human blood plasma were monitored. Then an established isolated reperfused und ventilated human lung setting was used to monitor the initial pulmonary absorption of BDP by measuring drug concentrations in the reperfusion fluid. For this purpose BDP particles containing aerosols delivered by commercially available pMDI for the first time were applied to an isolated reperfused human lung. Both models revealed differences in the combined processes of dissolution and distribution of BDP delivered by the two pMDI Sanasthmax® and Ventolair®. BDP particles delivered by Ventolair® apparently dissolved faster and to a greater extent than particles delivered by Sanasthmax®, resulting in an enhanced distribution both into dialysis fluid and into reperfusion fluid. The time course of initial pulmonary absorption of BDP delivered by the pMDI Sanasthmax® or Ventolair® observed at the isolated reperfused human lung exhibited high correlation with data from a corresponding clinical study with healthy volunteers. Furthermore, the obtained differences were consistent with results from investigations on the particles found in the aerosols produced by Sanasthmax® or Ventolair® regarding their size distribution, topology and dissolution behaviour in brochial fluid. To mimic the distribution of a drug from the systemic circulation into lung specific tissue employing the isolated reperfused and ventilated human lung setting, time course of tissue concentrations of thalidomide (THAL) in peripheral lung tissue in comparison with those in tumour tissue was determined for the first time by microdialysis. Firstly a microdialysis method optimised regarding sensitivity and time resolution by utilising the complexing agent (2 hydroxypropyl)-beta-cyclodextrin (HPCD) was developed and systematically evaluated. A THAL concentration derived from clinically relevant plasma concentrations was used in the reperfusion fluid and subsequently drug influx into tissue was monitored. By adding HPCD to the microdialysis perfusate a significant increase in the relative recovery was achieved enabling the establishment of a microdialysis method that exhibited high time resolution and appropriate analytical sensitivity. However, this perfusate additive strongly affected the diffusion processes during microdialysis so that common methods for microdialysis probe calibration, particularly the retrodialysis method, did not give accurate results. Therefore, a new calibration method suitable for the lung reperfusion experiments had to be explored. Tissue penetration evaluated in the lung reperfusion experiments revealed a slower distribution of THAL into tumour tissue than into peripheral lung tissue. In the latter concentrations similar to those detected in the reperfusion fluid were rapidly observed. Additionally, THAL concentrations achieved in tumour tissue were always lower than the corresponding levels in peripheral lung tissue. The resulting relationship between reperfusion fluid concentrations, concentrations in peripheral lung tissue, and concentrations in tumour tissue was highly correlated with data from clinical studies investigating the concentrations of antineoplastic agents in healthy and tumour tissue of various etiologies by microdialysis in relation to plasma concentrations. In conclusion, methods enabling both characterisation of the distribution of inhaled drugs from lung tissue into systemic circulation and determination of tissue penetration kinetics of systemically administered drugs into lung specific tissue were successfully established. These techniques simulating pulmonary drug distribution very closely to reality may significantly contribute to the understanding of pharmacokinetic processes in the lung.

Das Hauptziel der vorliegenden Arbeit war es, eine genauere Erkenntnis über die derzeitigen Möglichkeiten der quantitativen Messung der myokardiale Perfusion im Hinblick auf die Beschreibung verschiedener myokardialer Infarkte mittels kardialer MRT zu gewinnen. Die Untersuchungen zur Perfusion im Infarktgebiet ergaben, dass ein visuell festgestellter subendokardialer Infarkt an Hand der Bestimmung der absoluten Perfusion nicht immer nachvollzogen werden konnte. Ein Zusammenhang zwischen dem Auftreten eines no-reflow im Late Enhancement und der Höhe der absoluten Perfusion im Infarktgebiet konnte nicht gezeigt werden. Die Untersuchungen zur Perfusion im Remote Myokard ergaben keinen Zusammenhang zwischen der Perfusion im Remote Myokard und dem transmuralen Ausmaß des no-reflow-Phänomens in der First Pass Perfusion. Auch korrelierte die Perfusion im Remote Myokard nicht mit dem Auftreten eines no-reflow Phänomens im Late Enhancement. Die Perfusion im Remote Myokard unterschied sich zwischen transmuralen und nicht-transmuralen Infarkten. Eine Hyperperfusion im Remote Myokard konnte erst ab einer Infarktausdehnung von 75% im Late Enhancement beobachtet werden, während eine Hypoperfusion im Remote Myokard bei allen Infarktausdehnungen zwischen 0% und 100% auftrat. Die Untersuchungen zur Perfusion bei Vorliegen eines transmuralen Infarktes“ ergaben eine signifikante Korrelation der Perfusionen in Infarktgebiet und Remote Myokard bei transmuralem Infarkten. Die Ergebnisse zur Messung der Perfusion in Abhängigkeit von der relativen Infarktgröße wiesen keinen Zusammenhang zwischen der Perfusion im Infarktgebiet und der relativen Infarktgröße auf. Ebenso konnte keine Beziehung zwischen der Perfusion im Remote Myokard und der relativen Infarktgröße dargelegt werden. Letztendlich wurde das Verhalten der quantitativen Perfusion im Infarktgebiet und im Remote Myokard in Abhängigkeit von der Zeit nach Infarktereignis untersucht. Dabei zeigte sich kein Zusammenhang zwischen dem Auftreten eines no-reflow im Late Enhancement und der Entwicklung der Perfusion im Infarktgebiet zwischen der Erst- und der Spätuntersuchung. Ebenso war kein Zusammenhang zwischen dem Auftreten eines no-reflow im Late Enhancement und der Entwicklung der Perfusion im Remote Myokard zwischen der Erst- und der Spätuntersuchung erkennbar. Die kardiale MRT ist aufgrund der gleichzeitigen Analyse von morphologischen, funktionellen, quantitativen und metabolischen Parametern in einem Untersuchungsgang ein erfolgversprechendes Bildgebungsverfahren der Zukunft, da sie nicht invasiv ist, ohne Einsatz von Röntgenstrahlung auskommt und dabei eine gute räumliche Auflösung bei hohem Gewebekontrast bietet. Studien zeigen, dass die Kombination von Stress-Perfusion und Late-Technik in einem MRT-Protokoll eine höhere Genauigkeit als die Verwendung der SPECT-Untersuchung in der klinischen Beurteilung von Koronargefäßstenosen und im Nachweis subendokardialer Infarkte aufweist. 62 Allerdings erfährt die in vorliegender Arbeit verwendete Technik der Datenverarbeitung in dieser Form ohne Zweifel noch ihre Limitation im klinischen Alltag. Grundsätzlich ist zu sagen, dass die Messung der absoluten Perfusion im Myokard mit der MRT des Herzens zurzeit sicherlich noch nicht ausgereift ist. Dennoch lassen die in dieser Arbeit vorgestellten Ergebnisse und die viel versprechende Weiterentwicklung in der magnetresonanztomographischen Bildgebung weit reichende und interessante Möglichkeiten erahnen. / The aim of this work was to provide a more precise knowledge of the current possibilities for the quantitative measurement of myocardial perfusion in terms of the description of various myocardial infarcts using cardiac MRI. The analysis regarding the determination of a subendokardial infarction using the absolute perfusion in this area showed that the location of the infarction could not always been reproduced. A correlation between the occurrence of a no-reflow in Late Enhancement and the level of absolute perfusion in the area of infarction could not be shown. The analysis on the remote myocardial perfusion showed no coherence between myocardial perfusion in remote and the transmural extent of the no-reflow phenomenon in First-pass perfusion. Also the myocardial perfusion in remote myocardium did not correlate with the occurrence of a no-reflow phenomenon in Late Enhancement. The perfusion in remote myocardium differed between transmural and non-transmural infarctions. A hyperperfusion in remote myocardium was only seen with an extent of infarction of 75% and more in Late Enhancement while a hypoperfusion in remote myocardium occured with all infarction extents between 0% and 100%. The analysis regarding the perfusion when there was a transmural infarction showed a significant correlation of the perfusions in the area of infarcation and the remote myocardium. The results for the measurement of perfusion depending on the relative size of the infarction showed no coherence between the perfusion in the area of the infarction and relative size of it. Also there was no way of showing coherence between the myocardial perfusion in remote myocardium and the relative size of the infarction. Finally the behaviour of quantitative perfusion in the area of infarction and in the remote myocardium was investigated. The results showed no coherence between the occurrence of a no-reflow phenomenon in late enhancement and the development of the perfusion in the area of infarction between the first and the late data ascertainment. Also there was no coherence between the occurrence of a no-reflow in late enhancement and the development of the remote myocardium perfusion in between the first and the late data ascertainment. The cardiac MRI is due to the simultaneous analysis of morphological, functional, quantitative and metabolic parameters during one single investigation a promising imaging procedure of the future, because it is not invasive, without the use of X-rays and requires a good spatial resolution with high tissue contrast. Studies show that the combination of stress perfusion and late-MRI technique provide a higher accuracy in the clinical evaluation of coronary stenosis and subendocardial infarctions than the single use of the SPECT- technique. Without doubt the way of processing data used in this work still comes to its limits in everyday work and the technique of receiving data of the myocardial perfusion using cardiac MRI must still be developed. Nevertheless the results presented in this work and the promising progresses in the field of MRI imaging give a hint to the interesting and far reaching possibilities that are to come.

O mecanismo de contraste mais utilizado em imagens funcionais por ressonância magnética (functional Magnetic Resonance Imaging, fMRI), também conhecido por sinal BOLD (Blood Oxygenation Level Dependent) mede indiretamente a atividade neural, sendo sensível a mudanças no fluxo cerebral sangüíneo (Cerebral Blood Flow, CBF), na taxa cerebral metabólica do oxigênio (Cerebral Metabolic Rate of Oxygen, CMRO2) e no volume cerebral sanguíneo (Cerebral Blood Volume, CBV) e, em princípio, ele pode ser utilizado para mapear perfusão cerebral. Desse modo, o objetivo principal deste trabalho foi investigar, quantitativamente, alterações perfusionais no cérebro humano mapeadas pelas mudanças do sinal BOLD em resposta à indução transitória do estado de apnéia. Para isso, imagens por ressonância magnética foram obtidas através de um scanner de 1.5 T Siemens (Magneton Vision) com seqüências do tipo EPI-BOLD. Nesta pesquisa, foi analisada a influência da duração da apnéia no sinal BOLD. Observou-se, também, a diferença ocasionada no sinal em duas situações: apnéia iniciando-se após a inspiração ou após a expiração. Além disso, foi estudada a propagação deste sinal BOLD pelas diferentes regiões cerebrais. Por último, fazendo uso deste sinal BOLD, construiu-se mapas para obter informações a respeito do volume cerebral sangüíneo. Pelos dados obtidos, foi possível analisar o comportamento do sinal BOLD quando na presença de diferentes PaO2 e PaCO2. Observaram-se, também, diferenças regionais na sensibilidade do sinal BOLD ocasionada pelo estado de apnéia induzido. Essa diferença pode estar relacionada à reatividade das artérias que irrigam cada região ou ao volume sangüíneo basal dessas artérias. Além disso, foi possível obter informações a respeito das características temporais da mudança do CBF para diferentes regiões do cérebro em resposta a hipercapnia. Também, foi feita a identificação de áreas corticais responsáveis pelo controle voluntário da respiração. Por fim, os mapas de B-CBV obtidos utilizando o contraste BOLD em resposta à apnéia foram capazes de refletir o volume sangüíneo local, embora, estudos para análise dos outros parâmetros que influenciam o sinal devam ser realizados. / The BOLD (Blood Oxygenation Level Dependent) signal, is the most used contrast mechanism of the so called functional Magnetic Resonance Imaging (fMRI). Although it indirectly measures neuronal activity, its response is directly related to cerebral blood flow (CBF), Cerebral Metabolic Rate of Oxygen (CMRO2) and Cerebral Blood Volume (CBV) and can be, in principle, used to map cerebral perfusion. Thus, the main purpose of this study was to investigate, quantitatively, some aspects of perfusional alterations in the human brain. These changes were mapped by changes in the BOLD signal as a result of a global and uniform stimulation: hypercapnia induced by breath holding paradigms. Magnetic resonance images were acquired in a 1.5 T scanner (Siemens, Magneton Vision) with EPI-BOLD fMRI sequences. It was analyzed the BOLD dependency on breath holding duration and differences on the BOLD signal due the employed breath holding techniques: breath holding after expiration or after inspiration. The regional variability of the BOLD signal propagation was also studied. Moreover, the signal was used to construct maps based on CBV information. It was possible to gain information about the BOLD signal behavior that respond to PaO2 and PaCO2 alterations. Besides, it was demonstrated its regional variations sensibility, which can be correlated with arterial reactivity or the rest CBV of this arteries. It was also possible acquire information about the temporal characteristics of CBF changes induced by hypercapnia across brain regions as well as the identification of cortical areas that were responsible to the voluntary breathing. Finally, the B-CBV maps that used the BOLD con-trast were able to reflect CBV information, although, it is necessary the study of other parameters that can influence the signal.

The methodologies utilised by pharmaceutical companies for the toxicity screening of developmental drugs are currently based on outdated two-dimensional (2D) plate-based assay systems. Although such methods provide high-throughput analysis, limitations surrounding the biomimicry of the culture environment reduces the accuracy of testing, making the process cost and time inefficient. To significantly enhance the current methods, a screening platform that is both flexible in its design and is amenable toward physiologically representative engineered tissue is required. Incorporating a flow environment within the system elicits a variety of advantages over standard static cultures, pertinently the ability to couple the flow path with automated analytical systems via the use of intuitive software. Musculoskeletal pathological conditions account for £4.76 billion of NHS spending as of 2011 (Department of Health), affecting one in four of the UK adult population. Skeletal muscle, a highly metabolic and regenerative tissue, is involved in a wide variety of functional, genetic, metabolic and degenerative pathological conditions such as muscular dystrophy, diabetes, osteoarthritis, motor neuron disease and pertinently muscular weakness associated with aging populations. Skeletal muscle tissue engineering is centred on the in vitro creation of in vivo-like tissue within laboratory environments and seeks to aid the development of future therapies, by reliably elucidating the molecular mechanisms that regulate such conditions. However, the translation of such models toward systems amenable to pharmaceutical companies has to date been limited. Microphysiological perfusion bioreactors for in vitro cell culture are a rapidly developing research niche, although state of the art systems are currently limited due to the biologically non-representative 2D culture environment, lack of adaptability toward different experimental requirements and confinement to offline analytical methods. Advancements in additive manufacture (AM), commonly known as three-dimensional (3D) printing has provided a method of production that enables researchers to hold complete design freedom and facilitate customisation of required parts. The low cost, rapid prototyping nature of AM further lends itself toward the development of such technology, with design iterations quickly and easily printed, tested and re-designed where appropriate. Issues do however, currently persist regarding the biological compatibility of printed polymers and functional material properties of parts created. As such, this thesis investigated the use of AM as a rapid and functional prototyping technique to design and develop microphysiological perfusion bioreactors. Here, biocompatibility of candidate polymers derived from commercially available 3D printing processes; fused deposition modelling (FDM), stereolithography (SL), selective laser sintering (LS) and PolyJet modelling (PJM) were elucidated. Following the biological evaluation of these polymers, their suitability, and the applicability of each process in function and manufacture of perfusion bioreactors were assessed alongside the research and development process of system designs. Specifically, attention was afforded to the homeostatic environment within bio-perfusion systems. Once finalised, the biological optimisation of designs; biocompatibility and rates of proliferation in response to the perfusion environment, was undertaken. Protocols were then established for the automated perfusion of skeletal muscle cells in both monolayer and tissue engineered 3D hydrogels. This research outlined significant contributions to the scientific literature in 3D printed polymer biocompatibility, in addition to creating bio-perfusion systems that are adaptable, analytical and facilitate the in situ phenotypic development of physiologically representative skeletal muscle tissue. Polymer biocompatibility elucidated in this work will help to facilitate the wide-ranging use of AM in biological settings. However, advancements in the chemical properties of liquid resins for advanced photo-curable processes remain necessitated for AM to be considered as a primary manufacturing technique in the biological sciences. Furthermore, although systems developed in this work have provided a base technology from which to develop and build upon, significant challenges remain in the integration of tissue engineered perfusion devices within pharmaceutical settings. Although it is plausible that the technology created in its current guise would facilitate the automated generation of skeletal muscle tissue, systems require further development to aid their usability and scale. Furthermore, work is also required to optimise the biological environment prior to mass manufacture. As such, to truly influence the pharmaceutical industry, which has invested so heavily in more traditional screening technology, a system that is all-encompassing in biology, technology and automated analytics is required.

In den letzten Jahren hat die myokardiale MR-Perfusionsbildgebung als nichtinvasives Verfahren zur Darstellung von funktionellen Veränderungen des Myokards für die Diagnostik der KHK zunehmend an Bedeutung gewonnen. Während in den letzten 20 Jahren die kardiale MRT überwiegend bei einer Magnetfeldstärke von 1,5 T durchge-führt wurde und dies auch immer noch wird, findet aktuell eine rasante Verbreitung von MR-Systemen höherer Feldstärken statt. Von der neuen Hochfeldtechnik erhofft man sich vor allem, je nach Anwendung, eine deutliche Verbesserung der Bildqualität mit höherer räumlicher und zeitlicher Auflösung, wodurch der diagnostische Nutzen noch weiter gesteigert werden könnte. In der vorliegenden Arbeit wurden mittels First-Pass-MR-Bildgebung bei einer Magnet-feldstärke von 3 T quantitative Werte für die myokardiale Perfusion von 20 gesunden Probanden unter Ruhebedingungen bestimmt. Sowohl die erhobenen absoluten Perfusionswerte (0,859 ml/g/min im Mittel) als auch die Standardabweichung des mittleren MBF (0,298 ml/g/min) entsprechen den Messungen aus den früheren Publikationen dieser Arbeitsgruppe. In der Gesamtzusammenschau bisher veröffentlichter Perfusionsstudien zeigt sich eine relativ große Variabilität der publizierten Ruheflüsse. Dabei liegt der absolute MBF dieser Arbeit im mittleren Wertebereich dieser Streubreite. Er lässt sich auch mit den in PET-Studien ermittelten Ergebnissen in Einklang bringen, welche als Goldstandard zur Bestimmung der absoluten myokardialen Perfusion beim Menschen gelten. Die vorliegende Arbeit bestätigt die bereits in anderen 3 T-Studien untersuchten Vorteile der Hochfeld-MRT. Die höhere Magnetfeldstärke ermöglicht durch das größere SNR eine signifikant bessere räumliche Auflösung und besticht vor allem durch die hohe Bildqualität. Dies könnte bei der Erkennung kleiner, subendokardial gelegener Perfusionsdefekte sowie der Erstellung von transmuralen Perfusionsgradienten von Bedeutung sein und verspricht neben einer Reduktion von Partialvolumeneffekten auch eine Verminderung von „dark rim“-Artefakten. Um diese Vorteile entsprechend nutzen zu können, wird die Entwicklung von Methoden zur pixelweisen Bestimmung der absoluten Flüsse und farblich kodierten Darstellung derselben in Form von Perfusionskarten ein weiterer Schritt in Richtung klinisch einsetzbare Diagnostik sein. Eine Voraussetzung hierfür ist die Entwicklung einer exakten und sehr stabilen Bewegungskorrektur in weiterführenden Studien. Durch den Wechsel zu einer höheren Magnetfeldstärke von 3 T und den sich daraus ergebenden Vorteilen kann das Potential der MR-Perfusionsbildgebung, insbesondere der Bestimmung quantitativer Perfusionswerte, im Bereich der nichtinvasiven KHK-Diagnostik zukünftig weiter gesteigert werden. / Absolute Quantification of Myocardial Perfusion Using High-Resolution MRI at 3 T

Le travail présenté ici porte sur l’étude de la production et du transport du monoxyde d’azote (NO) dans le poumon humain. Le NO est une molécule dont l’implication dans des processus physiologiques n’a été mis en évidence qu’en 1987. Depuis, il a été démontré que le NO joue de nombreux rôles dans le corps humain. Le NO est un gaz labile (instable) dans les conditions physiologiques, il diffuse très facilement au travers des parois et il a une grande affinité pour l’hémoglobine. La production du NO est liée à 3 isoformes différentes de la protéine appelées synthases du NO ou NO synthases. En 1991, Gustafsson et al. ont découvert du NO endogène (produit par les poumons) dans l’air exhalé chez l’homme et le cochon d’Inde. Depuis près de 15 ans, de plus en plus de groupes de recherche travaillent sur le NO pulmonaire sans s’accorder sur ses rôles exactes. Il est cependant établit que, dans les pathologies comme l’asthme, la production accrue de NO est liée aux processus inflammatoires. Le NO peut être produit par la surface épithéliale (au niveau des conduits pulmonaires) ou alvéolaire, mais les sites exactes de production sont encore débattus. De part l’impossibilité de mesures directes au-delà des premières générations de l’arbre bronchique, les modèles mathématiques sont indispensables pour interpréter les résultats de mesures de concentrations exhalées et étudier la production et le transport du NO dans le poumon. Récemment, un modèle a été proposé par notre groupe, tenant compte de la convection, de la diffusion moléculaire et de la production du NO dans le poumon. Le but de ce travail est d’utiliser et d’adapter ce modèle pour reproduire des résultats expérimentaux soit existants, soit originaux. Dans ce travail, nous montrons que l’augmentation de concentration alvéolaire chez des sujets asthmatiques bien contrôlés est liée à une augmentation de production dans les conduits et non dans les alvéoles. Nous montrons également que, sur base de résultats expérimentaux, la production bronchique dans le poumon des sujets sains doit être très hétérogène pour reproduire des résultats expérimentaux apparemment irréconciliables. Nous montrons enfin que la localisation des conduits pulmonaires subissant une constriction influence la chute de NO exhalé mesurée après cette constriction. Nous avons également participé à 2 expériences liées à la gravité qui constitueront le matériel susceptible de faire évoluer le modèle.

Continuous monitoring of blood perfusion and oxygenation is essential in assessing the health of a transplanted organ. Particularly, monitoring the perfusion and oxygenation of the organ during the two-week period after the transplant procedure is crucial in detecting a sustained loss in perfusion or a reduction in oxygen saturation before these changes render irreversible damage to the organ or patient. Pulse oximetry is a clinically accepted method of monitoring the arterial oxygen saturation of a patient in a non-invasive manner. Pulse oximeters exploit the wavelength-dependent absorption of oxygenated and deoxygenated hemoglobin to measure a patient's arterial oxygen saturation. However, traditional pulse oximeters do not provide perfusion information and produce erroneous oxygen saturation measurements under low perfusion levels. An optical blood perfusion and oxygenation sensor, based on a modified reflectance pulse oximeter, has been developed for in situ monitoring of transplanted organs. To reduce the number of animal experiments, phantoms that mimic the optical and anatomical properties of liver parenchyma have also been developed. In this work, in vitro data was gathered from dye solutions mimicking oxygenated blood that were pumped through single and multi-layer phantoms mimicking liver parenchyma and through a phantom mimicking the portal vein. A portion of the phantom data was compared to data collected from in vivo occlusion studies performed on female swine to assess the ability of the phantoms to mimic the response observed with changes in blood perfusion through liver parenchyma. Both the single layer and multilayer phantoms showed a similar response to changes in perfusion as the in vivo case. With each phantom, the signal increased linearly with increases in perfusion, but the multilayer phantom showed a higher sensitivity (approximately 30% higher) to changes in perfusion than the single layer phantom. This higher sensitivity would provide the ability to measure smaller changes in perfusion and increase the resolution of the sensor. Also, both parenchymal phantoms showed similar trends in the oxygenation studies, with the R value decreasing with increasing oxygenation. While the observations in this research demonstrate the ability to use both phantoms for in vitro experiments, the results show the multilayer phantom is a better option for mimicking perfusion because it displays similar occlusion patterns as the liver parenchyma in vivo, a higher sensitivity to changes in perfusion than the single layer phantom, and it is only slightly more complex in design (contains only two more layers of sinusoids) than the single layer phantom.

In recent years, computer tomography (CT) has become an increasingly important tool in the clinical diagnosis, mainly because of the advent of fast scanning techniques and high spatial resolution of the vision hardware. In addition to the detailed information of morphology, functional CT also gives the physiologic information, such as perfusion. It can help doctors to make better decision. Our goal in this paper is to evaluate lung perfusion by comparing pre and post contrast-enhanced CT images. After the contrast agent is injected, it flows with blood stream and causes the temporal changes in CT values. Therefore, we can quantize perfusion values from the changes of CT values between pre and post contrast-enhanced CT images. Then guided by color -coded maps, a quantitative analysis for the assessment of lung perfusion can be performed. As a result, it is easier for observer to determinate the lung perfusion distribution. Moreover, we can use color - coded images to visualize pulmonary embolism and monitor therapeutic efficacy.

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Microvasculatures may become damaged by a variety of acute and chronic diseases. In many cases, microvessel function is irreversibly compromised, leading to the dysfunction and even death of surrounding tissues. Currently, there are few therapies that directly address the treatment of microvascular insufficiency. Responding to this need, researchers are developing methods to fabricate in vitro blood vessels. Typical strategies include; cellular sodding within polymers and/or biopolymers, the formation of cylindrical cellular monolayers around polymer mandrels, and the modification of biocompatible surfaces for cellular adhesion. Using currently available techniques, simple, individual vessel conduits have been engineered with internal diameters down to 150μm. However, no evidence has been provided illustrating the formation of patent, interconnected microvessel networks without the aid of a host circulatory system. In response to this challenge, it is hypothesized that a novel flow-based experimental system will support the in vitro development of three-dimensional microvascular tissues. Addressing this hypothesis, the presented work focused on three specific aims: Specific Aim 1. Pattern planar in vitro three-dimensional microvasculatures. Specific Aim 2. Engineer a Dynamic In vitro Perfusion Chamber (DIP Chamber) for microvascular investigation. Specific Aim 3. In vitro perfusion of microvessel fragments within the DIP Chamber. Through the supporting experiments, directed endothelial sprouting from parent isolated microvessel fragments was achieved. In addition, patent in vitro microvessel networks were successfully developed. The presented experiments are the first to achieve these experimental results. In addition, the described experimental model will provide a unique method for future investigations of microcirculatory phenomena. Since no exogenous growth factors or cell signals were introduced into the constructs, it is believed that this system presents a physiological platform for future investigations into angiogenesis, angioadaptation, and network remodeling. Moreover, this model may offer a useful platform for vascular therapeutic testing and a foundation for future tissue engineering applications.