Дисертації з теми "HIFU therapy"

The rising incidence of small, asymptomatic renal tumours discovered usmg abdominal imaging during the investigation of unrelated symptoms has fuelled the desire for new therapies which avoid surgical excision. Extracorporeal High Intensity Focused Ultrasound (HIFU) was proposed as one of these modalities but so far clinical research has been ,~." inconclusive. The present work was designed to improve these clii teal outcomes through the conduct of further clinical trials, laboratory based research and the translation of new technology into existing HIFU devices. A Phase II clinical trial of patients (n=13) with newly diagnosed <4cm renal tumours (clinical stage T1a) was designed, peer reviewed and received ethical approval (Ox REC 09/H0606104). Ten of 13 patients underwent renal HIFU using a clinical HIFU device (Model JCIJC200, HAIFU, China). One patient could not be treated due to poor tumour visualisation after anaesthesia and two patients could not be treated as they became unwell before or during anaesthesia. Histological evidence of HIFU ablation in either tumour or normal renal parenchyma was seen in all ten patients. Evidence of sub-total tumour ablation was seen in 8/10 of patients. Grade 1 «50%), 2 (50-90%) & 3 (90-99%) ablation was achieved in 4/10, 3/1 0 & 3/1 0 patients respectively but complete (100%) tumour ablation was not possible. HIFU treatment caused minimal morbidity - no Grade III- V (Clavien-Dindo) complications related to HIFU treatment occurred. Grade I skin pain and induration was seen in 9/1 0 patients; Grade II skin pain occurred in a single patient. Patient demographics, imaging and tumour characteristics were used to design parameters to improve patient selection for renal HIFU. The tumour location, thickness of peri-nephric fat and renal nephrometry score were useful predictors of successful screening for treatment. Page /ii Dr R. W Ritchie Nutiield Department of Surgical Sciences - TT 2012 Abstract Diligent use of these factors could limit unnecessary treatments and Improve ablation outcomes. , It is well known that ultrasound imaging of small renal masses can be challenging. Ultrasound imaging often deteriorates further during HIFU as the abdominal wall and fat tissues swell and cause increased attenuation. This loss of imaging quality was clearly demonstrated in this clinical trial and resulted in the early termination of treatment, before ,#,J' ... ~ .•.. endpoints were reached, in a number of cases. The current clinical method for monitoring the success of HIFU ablation using hyperecho analysis of B-mode ultrasound images is also questionable. Laboratory based studies using ex-vivo bovine liver subjected to HIFU confirmed that hyperecho monitoring had low sensitivity, predictive values and overall accuracy. A novel method of HIFU monitoring - passive mapping of the emissions received from acoustic cavitation activity and other sources of non-linearity during HIFU treatment - is believed to represent a significant opportunity to improve feedback. This technique uses the passively received signature of cavity activity which, when time-reversed, gives high- resolution images of the precise location of the activity. Laboratory-based ex-vivo work, using a commercially available ultrasound system (z.one, Zonare, USA), demonstrates its superiority over hyperecho monitoring. Indeed, thresholds could be applied to successfully predict HIFU ablation with high sensitivity and specificity. This technique was successfully translated into the clinical setting through the design of a Passive Acoustic Mapping (P AM) device. Custom-built receiving elements were applied without limiting the function of the existing HIFU devices. Both pre-clinical and ethically- Page [iii Dr R. W Ritchie Nuffield Department of Surgical Sciences - TT 2012 Abstract approved clinical studies demonstrated its safe integration without significant impact on the device energy output or treatment accuracy. Using similar passive beamfonning algorithms, acoustic cavitation activity was successfully mapped and corresponded with the location of thermal ablation in both ex-vivo tissue phantoms and during clinical HIFU therapy. ,~-' It is believed that the development of new patient selection paral~~tel's will elimil?ate target those patients who are most suitable for renal HIFU - small tumours, minimal peri-nephric fat & low nephrometry score .. The use of P AM will lead to a significant improvement in the efficacy of treatment. It can be successfully applied to existing devices and predicts the location and extent ofHIFU ablation with greater accuracy that existing techniques.

High Intensity Focused Ultrasound (HIFU) is a new cancer thermal therapy method which has achieved encouraging results in clinics recently. However, the lack of a temperature monitoring makes it hard to apply widely, safely and efficiently. Conventional ultrasonic temperature estimation based on echo strain suffers from artifacts caused by signal distortion over time, leading to poor estimation and visualization of the 2D temperature map. This thesis presents a novel model-based stochastic framework for ultrasonic temperature estimation, which combines the temperature information from the ultrasound images and a theoretical model of the heat diffusion. Consequently the temperature estimation is more consistent over time and its visualisation is improved. There are 3 main contributions of this thesis related to: improving the conventional echo strain method to estimate temperature, developing and applying approximate heat models to model temperature, and finally combining the estimation and the models. First in the echo strain based temperature estimation, a robust displacement estimator is first introduced to remove displacement outliers caused by the signal distortion over time due to the thermo-acoustic lens effect. To transfer the echo strain to temperature more accurately, an experimental method is designed to model their relationship using polynomials. Experimental results on a gelatine phantom show that the accuracy of the temperature estimation is of the order of 0.1 ◦C. This is better than results reported previously of 0.5 ◦C in a rubber phantom. Second in the temperature modelling, heat models are derived approximately as Gaussian functions which are mathematically simple. Simulated results demonstrate that the approximate heat models are reasonable. The simulated temperature result is analytical and hence computed in much less than 1 second, while the conventional simulation of using finite element methods requires about 25 minutes under the same conditions. Finally, combining the estimation and the heat models is the main contribution of this thesis. A 2D spatial adaptive Kalman filter with the predictive step defined by the shape model from the heat models is applied to the temperature map estimated from ultrasound images. It is shown that use of the temperature shape model enables more reliable temperature estimation in the presence of distorted or blurred strain measurements which are typically found in practice. The experimental results on in-vitro bovine liver show that the visualisation on the temperature map over time is more consistent and the iso-temperature contours are clearly visualised.

Cavitation is a phenomenon present during many ultrasound therapies, including the thermal ablation of malignant tissue using high intensity focused ultrasound (HIFU). Inertial cavitation, in particular, has been previously shown to result in increased heat deposition and to be associated with broadband noise emissions that can be readily monitored using a passive receiver without interference from the main ultrasound signal. The present work demonstrates how an array of passive receivers can be used to generate maps of cavitation distribution during HIFU exposure, uncovering a new potential method of monitoring HIFU treatment. Using a commercially available ultrasound system (z.one, Zonare, USA), pulse transmission can be switched off and data from 64 elements of an array can be simultaneously acquired to generate passive maps of acoustic source power. For the present work, a 38 mm aperture 5-10 MHz linear array was used, with the 64 elements chosen to span the entire aperture. Theory and simulations were used to show the spatial resolution of the system, the latter showing that the broadband nature of inertial cavitation makes passive maps robust to interference between cavitating bubbles. Passive source mapping was first applied to wire scatterers, demonstrating the ability of the system to resolve broadband sources. With the array transversely placed to the HIFU axis, high-resolution passive maps are generated, and emissions from several cavitating bubbles are resolved. The sensitivity of passive mapping during HIFU exposure is compared with that of an active cavitation detector following exposure. The array was then placed within a rectangular opening in the centre of the HIFU transducer, providing a geometric setup that could be used clinically to monitor HIFU treatment. Cavitation was instigated in continuous and disjoint regions in agar tissue mimicking gel, with the expected regions of cavitation validating the passive maps obtained. Finally, passive maps were generated for samples of ox liver exposed to HIFU. The onset of inertial cavitation as detected by the passive mapping approach was found to provide a much more robust indicator of lesioning than post-exposure B-mode hyperecho, which is in current clinical use. Passive maps based on the broadband component of the received signal were able to localize the lesions both transversely and axially, however cavitation is generally indicated 5 mm prefocal to the lesions. Further work is needed to establish the source of this discrepancy. It is believed that with use of an appropriately designed cavitation detection array, passive mapping will represent a major advance in ultrasound-guided HIFU therapy. Not only can it be utilized in real-time during HIFU exposure, without the need to turn the therapeutic ultrasound field off, but it has also been shown in the context of the present work to provide a strong indicator of successful lesioning and high signal-to-noise compared to conventional B-mode ultrasound techniques.

High Intensity Focused Ultrasound (HIFU) is a promising technique for the treatment of localized cancers. The ability to focus ultrasound precisely on a predetermined volume allows the possibility of selective tissue destruction at this position without damage to surrounding tissues. However, many difficulties remain in the treatment of deep-seated tumors. In this thesis, new therapeutic and monitoring techniques are proposed to address these problems, by using phased arrays of ultrasound transducers. Two monitoring techniques based on the detection of the displacements of the ultrasonic speckle are developed, and allowed us to image the changes in the temperature and the shear modulus during HIFU therapy. In-vitro ultrasound-guided experiments are performed. Secondly, the problem of organs motion during the treatment is addressed. A method for real-time tracking the 3D motion of tissues is combined with a 2D High Intensity Focused Ultrasound multi-channel system in order to correct the respiratory motion during HIFU therapies. In the last section of this thesis, a high power ultrasonic system is developed for transcranial HIFU brain therapy. The skulls aberrations are corrected using a time reversal mirror thanks to an implanted hydrophone. In-vivo experiments are conducted on 22 sheep with minimally invasive surgery. Finally, a non-invasive protocol based on CT scans of the entire skull is developed and allows the prediction of the skulls aberrations and the skull overheating.

High intensity focused ultrasound (HIFU) is an emerging non-invasive thermal ablative modality that can be utilised for the treatment of solid organ tumours, including liver cancer. Acoustic cavitation is a phenomenon that can occur during HIFU and its presence can enhance heating rates. One major limitation of thermal ablative techniques in general, such as radiofrequency and microwave ablation, is the heat sink effect imparted by large vasculature. Thermal advection from blood flow in vessels ≥ 3 - 4 mm in diameter has been shown to significantly reduce heating rates and peak temperatures in the target tissue, potentially leading to treatment failure. With regards to HIFU therapy, a clearer understanding is required of the effects of blood flow on heating, cavitation and thermal tissue necrosis, which is the treatment endpoint in clinical thermal ablation. Therefore, the overall aim of this thesis project was to elucidate the effects of blood flow on HIFU-induced heating, cavitation and histological assessment of thermal ablation. A unique isolated, perfused porcine liver model was used in order to provide a relevant test bed, with physiological and anatomical characteristics similar to the in vivo human liver. The normothermic liver perfusion device used in all studies presented in this work can keep an organ alive in a functional state ex vivo for in excess of 72 hours. A further advantage of the liver perfusion device was that it allowed blood flow to be stopped completely and resumed rapidly, allowing studies to be conducted under zero flow conditions. A therapeutic HIFU system was used in order to deliver HIFU therapy to regions of hepatic parenchyma adjacent (≤ 3 mm) to large (≥ 5 mm) blood vessels or away from vasculature (≥ 1 cm) at either 1.06 MHz or at 3.18 MHz. Cavitation events during HIFU therapy were spatio-temporally monitored using a previously developed passive acoustic mapping (PAM) technique. The cavitation threshold at each frequency was determined through assessment of acoustic emissions acquired through PAM during HIFU exposure at a range of acoustic pressures. Real time thermal data during HIFU therapy were obtained using an implantable 400 μm thermocouple, aligned with the HIFU focus, in order to assess the effect of large vessel blood flow on peak tissue temperatures. Thermal data were obtained at 1.06 MHz, in the presence of acoustic cavitation and at 3.18 MHz, in the absence of cavitation, both in the presence and complete absence of blood flow. Finally, histological assessment of cell viability and cell death was performed in order to determine whether any heat sink effect could be overcome, with the achievement of complete tissue necrosis in treatment regions directly adjacent to large vasculature. This work demonstrated for the first time that in perfused, functional liver tissue, the presence of large vasculature and physiological blood flow does not significantly affect ablative HIFU therapy, both in terms of peak focal tissue temperatures attained and histological evidence of complete tissue necrosis. Therefore, HIFU may be superior to other ablative modalities in treating tumours in tissue regions adjacent to major vascular structures, but further work needs to be performed to correlate the experimental findings with clinical outcomes.

Thèse doctorat : Images et Systèmes : Villeurbanne, INSA : 2004.
Thèse rédigée en anglais. Introduction et conclusion en français et en anglais. Titre provenant de l'écran-titre. Bibliogr. p. 113-118. Publications de l'auteur, 3 p.

La fibrillation atriale (FA) est l'arythmie cardiaque la plus fréquente. Elle touche près de 750 000 personnes en France. La technique de traitement la plus courante est l'ablation intracardiaque par radiofréquence (RF). Son principe consiste à isoler électriquement les veines pulmonaires du reste de l'oreillette. Cependant cette technique est invasive et a une efficacité limitée. Les Ultrasons Focalisés de Haute Intensité (HIFU) permettent de léser à distance de façon précise les tissus biologiques. Un traitement de la FA par HIFU transoesophagiens aurait l'avantage d'être mini-invasif et plus efficace qu'un traitement par RF intracardiaque de par la possibilité de générer des lésions transmurales sans nécessiter de contact entre la sonde et la zone à traiter. Un applicateur HIFU transoesophagien à guidage échographique intégré a donc été développé pour le traitement de la FA. Le transducteur peut focaliser le faisceau ultrasonore de 17 mm à 55 mm de profondeur avec une intensité acoustique maximale à sa surface de 12 W*cm-2. Une procédure de traitement HIFU préservant les tissus adjacents a été simulée numériquement sur un modèle anatomique réaliste. Des lésions HIFU transoesophagiennes ont été obtenues ex vivo dans du myocarde dans des conditions anatomiques et physiologiques proches de l'in vivo. Des essais préliminaires d'élastographie par ondes de cisaillement ont permis d'évaluer la faisabilité d'un contrôle de la formation des lésions à l'aide du transducteur d'imagerie intégré. Une première série d'expérimentations in vivo sur le modèle porcin a finalement permis de valider la procédure de traitement et d'induire des dommages biologiques dans le tissu cardiaque

Le carcinome hépatocellulaire est le principal cancer primaire du foie. Les procédures d’ablation thermique par voie interstitielle constituent un type de traitement curatif de ce cancer. Ces méthodes ne permettent pas toujours, de par leur nature physique (radio fréquence, micro-onde, laser, cryothérapie), de générer une ablation conformationnelle pour un volume tumoral donné. Dans certains cas, cela peut entraîner l’ablation d’un volume important de tissus non tumoraux. L'utilisation d'une sonde interstitielle ultrasonore disposant d’un transducteur avec de nombreux éléments indépendants, capable de générer des ultrasons focalisés de haute intensité (HIFU), permettrait théoriquement de lever cette limitation. D’autre part, le nombre élevé d’éléments permettrait également de disposer de capacités d’imagerie. Les travaux présentés dans cette thèse ont donc d’abord porté sur la conception du transducteur associé à ce type de sonde. Un design particulier a été proposé spécifiquement pour le traitement d’une tumeur de 4 cm de diamètre. S’est ensuite posé la question de la stratégie de planification du traitement à adopter pour obtenir une lésion la plus conformationnelle possible. Différentes stratégies ont été évaluées par simulations numériques. Toutes ont présentés des caractéristiques semblables tant en termes de conformation que de temps de traitement total. La focalisation ultrasonore s’est avérée en elle-même suffisante pour générer un traitement conformationnel. Finalement, une plateforme robotique a été développée pour le pilotage de prototypes de sondes ultrasonores interstitielles bimodales, aussi bien en mode imagerie qu’en mode thérapie. Cette plateforme a permis de réaliser in vitro, les planifications de traitement automatiques de plusieurs fantômes de tumeurs, en se basant sur la reconstruction ultrasonore 3D issue de l’imagerie échographique obtenue in situ par les sondes ultrasonores. En revanche, en mode thérapie les prototypes de sondes n’ont pas atteint leurs spécifications et n’ont pas réussi à générer des lésions thermiques dans des tissus hépatiques in vitro. La modularité de la plateforme robotique a rendu possible son utilisation avec un système de thérapie ultrasonore différent, à la fiabilité éprouvé. Avec ce système, la planification automatique du traitement, ainsi que l’exécution du traitement HIFU associé, ont pu être effectué in vitro avec succès par la plateforme
Hepatocellular carcinoma is the most common primary cancer of the liver. Interstitial thermal ablation procedures constitute a type of curative treatments for this cancer. Given the physical nature of the phenomenon used to modify temperature (radio frequency, micro wave, laser, cryotherapy), those methods may not be able to generate a conformal treatment for a given tumor shape. In some cases, this limitation may induce the thermal ablation of a large volume of non-tumor tissues. The use of an ultrasound interstitial probe mounted with a multi-element transducer capable of generating high intensity focused ultrasound (HIFU) may theoretically help to overcome this limitation. Also a transducer with an important number of elements may also provide in situ imaging. As a first step, the design of a transducer for interstitial ultrasound probe was studied. A specific configuration has been proposed for the treatment of tumors with a diameter of 4 cm. The question of the treatment planning method to adopt to reach an optimal conformal treatment has been then addressed by comparing numerical simulations of different strategies. All strategies were sufficiently conformal and none presented real assets compared to the others. Ultrasound focusing in itself provided the desired conformal thermal ablation. Finally, a robotic platform was developed for driving interstitial dual mode ultrasound probes, both in imaging and in therapy mode. This platform allowed the automatic treatment planning of in vitro tumor mimic phantoms, based on 3D ultrasound reconstruction from the B mode images obtained in situ by the interstitial probe. However, in therapy mode, the probes did not reach their specifications and did not manage to create thermal lesions in in vitro liver tissue sample. The modularity of the robotic platform allowed driving a different HIFU system, which was more robust. With this system, the platform managed to perform with success an automatic treatment planning and then the associated HIFU treatment in in vitro tissue sample

High Intensity Focussed Ultrasound (HIFU) ablation is a promising technology for the non-invasive, targeted treatment of certain types of cancer. The technique functions by subjecting tumours to a cytotoxic level of intense, localised heating, while leaving the surrounding tissue unharmed. However, a number of limitations in the available HIFU treatment monitoring methods are currently hampering the effectiveness and clinical adoption of the therapy. This work aims to develop improved metrics of HIFU-induced biological damage that are specifically suited to monitoring and controlling HIFU ablation. Firstly, an optical method that enables straightforward quantification of thermal damage in protein-embedding hydrogels is developed. Secondly, hydrogels embedded with different cell lines are used to assess the performance of common temperature-based metrics of cell death across a range of HIFU-relevant conditions. Finally, a novel, passive acoustic detector designed for the real-time monitoring of HIFU-induced tissue damage is proposed. The detector is shown to predict lesioning with over 80% accuracy in regimes that are very likely to create lesions (60 J of acoustic energy or more), with an error rate of less than 6% for exposures that are too short to cause lesioning (up to 1 s long). The proposed detector could therefore provide a low-cost means of effectively monitoring clinical HIFU treatments passively and in real time.

La fibrillation atriale (FA) est l’arythmie cardiaque la plus fréquente. Elle touche près de 750 000 personnes en France. La technique de traitement la plus courante est l’ablation intracardiaque par radiofréquence (RF). Son principe consiste à isoler électriquement les veines pulmonaires du reste de l’oreillette. Cependant cette technique est invasive et a une efficacité limitée. Les Ultrasons Focalisés de Haute Intensité (HIFU) permettent de léser à distance de façon précise les tissus biologiques. Un traitement de la FA par HIFU transoesophagiens aurait l’avantage d’être mini-invasif et plus efficace qu’un traitement par RF intracardiaque de par la possibilité de générer des lésions transmurales sans nécessiter de contact entre la sonde et la zone à traiter. Un applicateur HIFU transoesophagien à guidage échographique intégré a donc été développé pour le traitement de la FA. Le transducteur peut focaliser le faisceau ultrasonore de 17 mm à 55 mm de profondeur avec une intensité acoustique maximale à sa surface de 12 W•cm-2. Une procédure de traitement HIFU préservant les tissus adjacents a été simulée numériquement sur un modèle anatomique réaliste. Des lésions HIFU transoesophagiennes ont été obtenues ex vivo dans du myocarde dans des conditions anatomiques et physiologiques proches de l’in vivo. Des essais préliminaires d’élastographie par ondes de cisaillement ont permis d’évaluer la faisabilité d’un contrôle de la formation des lésions à l’aide du transducteur d’imagerie intégré. Une première série d’expérimentations in vivo sur le modèle porcin a finalement permis de valider la procédure de traitement et d’induire des dommages biologiques dans le tissu cardiaque
Atrial fibrillation (AF) is the most frequent cardiac arrhythmia. This pathology affects more than 750,000 persons in France. Radiofrequency (RF) endocardial ablation is performed to treat this disease and involves the generation of transmural thermal lesions, to isolate electrically the pulmonary veins (PV) from the left atrium. The technique is, however, invasive and has a limited efficiency, especially for ensuring transmurality which requires a perfect contact between the RF probe and cardiac tissues. High Intensity Focused Ultrasound (HIFU) allows the creation of precise thermal lesions, deep within biological tissues. A transesophageal HIFU approach could provide a minimally-invasive alternative for AF treatment, since deep transmural lesions could be generated at distance from the HIFU probe. In this work, an ultrasound-guided transesophageal applicator has been developed for AF treatment. The HIFU transducer, embedding a transesophageal echocardiography (TEE) probe, can focus the acoustic beam from 17 to 55 mm axially and generate a surface acoustic intensity up to 12 W•cm-2. A complex treatment plan, the HIFU Mini-Maze (HIFUMM), was successfully simulated on a realistic anatomical human model. Transesophageal HIFU lesions were induced experimentally in static myocardium, under ex vivo configurations reflecting an increasing complexity in anatomical and physiological conditions. Investigations conducted on shear wave elastography confirmed the feasibility of using TEE to control the formation of HIFU lesions. Finally, in vivo experiments in a porcine model allowed validating the treatment procedure by inducing biological damages in beating heart

Murine macrophages were induced to over-express iNOS by treatment with a combination of cytokines, mixed with HT1080 and HCT116 human tumour cells and the toxicity of AQ4N was determined under normoxic or hypoxic conditions. The normoxic toxicity of AQ4N towards tumour cells was not affected through co-culturing with macrophages. However, under hypoxic conditions, the induction of iNOS activity in the macrophages was associated with an increase in AQ4N metabolism and a substantial increase in tumour cell toxicity, which was dependent upon the proportion of macrophages in the culture. This study is the first demonstration of tumour associated macrophage mediated pro-drug activation to result in bystander killing of human tumour cells. The oxygenase domain of the iNOS enzyme is responsible for the production of nitric oxide (NO), a potent biological mediator, whose complex role in tumour biology is still not fully understood as it seems to have both anti- and protumour effects. We were able to use the co-culture model of NO-producing macrophages and cancer cells to demonstrate that NO is also a potent tumour radiosensitiser under hypoxic conditions with a sparing effect on well-oxygenated tumour cells, also a characteristic of normal tissues. Additionally, NO produced by the macrophages resulted in a significant induction of hypoxia inducible factor-1 (HIF-1) driven transcriptional activity in the co-cultured tumour cells under both normoxic and hypoxic conditions. We therefore, suggest the use of HIF-1 inhibitors in combination with NO-based therapies and radiation to favour tumour regression.

Energy production is a very important function for the cells to maintain homeostasis, survive and proliferate. Cellular energy can be produced either through oxidative phosphorylation (OXPHOS) in the presence of oxygen or glycolysis in its absence. Cancer cells, even in the presence of oxygen prefer to produce energy through glycolysis and this confers them a survival advantage. Energy metabolism has recently attracted the interest of several laboratories as targeting the pathways for energy production in cancer cells could be an efficient anticancer treatment. For that purpose the role of various transcription factors in determining the pathway of energy production has been investigated extensively and there is evidence to suggest that oncogenic transcription factors promote glycolysis whereas tumour suppressors demote it. In line with this notion, the master regulator of cellular response to hypoxia, the Hypoxia Inducible Factor 1 (HIF-1) has been shown to induce the expression of a variety of genes encoding enzymes involved in glucose metabolism as well as OXPHOS favouring energy production through glucose metabolism in hypoxic cells. The tumour suppressor p53 on the other hand inhibits glycolysis and stimulates OXPHOS. One of the pathways through which p53 exerts these effects, is by inducing the inhibitor of glycolysis TIGAR and the cytochrome c oxidase assembly factor SCO2 gene expression under DNA damage conditions. However, the regulation of the expression of these genes in hypoxic conditions has been only partially elucidated. We hypothesised that under hypoxic conditions, TIGAR and SCO2 gene expression might be differentially regulated in cells bearing mutated p53 and in these cells the involvement of HIF-1 could be crucial. Indeed under hypoxia mimicking conditions, the TIGAR and SCO2 protein and mRNA levels were found to be modulated differentially in p53 wild type and mutant cell lines. The bioinformatics analysis revealed the presence of hypoxia responsive elements (HREs) within the regulatory region of the promoters of TIGAR and SCO2 genes. Firefly reporter assays and chromatin immunoprecipitation (ChIP) assays have indicated that HIF-1 plays a crucial role in the regulation of TIGAR gene expression. The direct involvement of HIF-1 in the regulation of SCO2 gene expression requires further investigation. We and others have recently reported that PCAF is a common cofactor for p53 and HIF-1α, regulating the protein stability and transcription target selectivity of both transcription factors thereby orchestrating the balance between life and death in cancer cells. We hypothesised that PCAF plays a similar role in the regulation of cellular energy metabolism by differentially targeting HIF-1α and p53 to the promoter of TIGAR and SCO2 genes. In this study we present evidence to support the notion that PCAF plays an import role in the regulation of TIGAR and SCO2 gene expression under hypoxic mimicking conditions. This conclusion was supported by assessing the functional consequences of PCAFwt and PCAFΔHAT overexpression on the intracellular lactate production, cellular oxygen consumption, NAD+/NADH ratio and ROS generation in cells under hypoxia mimicking conditions.

碩士
國立中央大學
機械工程學系
106
Compared with conventional open surgery, High-intensity focused ultrasound (HIFU) thermal therapy is non-invasive and has no radiation and less bleeding. The main mechanism of HIFU ablation treatment involves mechanical and thermal effects, which can generate high temperature to heat and destroy target cells. Currently, HIFU has been applied in the treatment of prostate, liver, breast, and bone tumor. With the application of targeting microbubbles, tumors can be treated by heat or local higher concentration of drug activated by HIFU. However, the focus point of HIFU must be located accurately and precisely so that it can destroy target tumor cells completely. In this study, the focus point of a HIFU system is located. After the thermocouple tip for temperature measurement is located by ultrasound scan, a robot moves the HIFU focal point along the HIFU focusing axis to heat the thermocouple tip. The position with highest temperature is defined as the true position of the HIFU focal point. The difference between the true position of the HIFU focal point and that of the thermocouple tip detected by ultrasonic image and transferred relative to the robot coordinate system is the position offset of the focus point. The experimental results show the offset is 6mm. This study also investigates the impact of phantom shape on the focusing effect. The experimental results show that the symmetry of the phantom body was helpful to the efficiency of the acoustic focusing. Finally, the calibrated HIFU system pork tissue is applied to ablate pork tissues (muscle and fat) and the targeting microbubbles, the results show that a single tissue can be quickly heated, but the heating of multiple-layer tissues are out of prediction, which requires further study. Moreover, HIFU targeting microbubbles can be detected and destroyed easily no matter they are targeting on small spots, silicon tubes or spherical sponges.

碩士
國立臺灣科技大學
電機工程系
107
Ultrasound (US) imaging is often used for guidance of high-intensity focused ultrasound (HIFU) treatment due to its high temporal resolution and relatively low cost. However, real-time US monitoring is often hindered when the strong HIFU interference overwhelms the imaging echoes. In this study, a method of Golay-encoded pulse-inversion subtraction (PIS) is proposed to better visualize the change of imaged object during the HIFU treatment. It effectively eliminates HIFU interference patterns in real-time US imaging and also improves the signal-to-interference ratio (SIR). In PIS method, the received imaging echo of positive transmit is subtracted from that of negative transmit to cancel the HIFU component. When Golay excitation is adopted, imaging echo after PIS is further decoded by matched filters for pulse compression. The performance of the proposed method was evaluated through experiments with both PVA phantom and ex-vivo swine liver. Results show that 4-bit Golay transmit can improve the SIR from -15.0 dB of un-coded transmit to 3.5 dB. With PIS, the SIR of 4-bit Golay further increases to 15.9 dB. Moreover, the effect of tissue motion due to breathing are also shown to be negligible on HIFU suppression and decoded image resolution.

碩士
義守大學
生物醫學工程學系
104
High-intensity focused ultrasound (HIFU) thermal therapy is a non-invasive means for tumor therapy. The concept is the use of thermal effect of focused ultrasound to produce a local high temperature region at the tumor tissue and resulting in irreversible damage. To ablate a deep-seated tumor, a large scale transducer is needed to compensate the attenuation of ultrasound at the propagation pathway. Owing to the technical limitation, it is difficult to fabricate a large scale, single element HIFU transducer. In this study, a large scale, geometerical ultrasound focusing system via appropriate arrangement of plane transducers on a spherical surface has been developed. Several parameters were investigated, such as the number and the diameter of transducer elements and the distribution of these transducers. Simulation results show that the proposed geometrical focusing system could result in an appropriate focused ultrasound field for HIFU thermal therapy. According to the simulation results, a 25 cm focusing ultrasound transducer with 32 elements, operated at 1 MHz has been developed. Experimental results indicate that, geometrical focusing technique could be an alternative way to substitute for large scale, single element transducer. Furthermore, temperature controller is important during HIFU treatment. A fuzzy controller was proposed to dynamically control temperature elevation. Simulation results suggest that the proposed fuzzy controller is feasible to control the temperature elevation within a prescribed time, even if blood perfusion varies with time.

博士
臺灣大學
機械工程學研究所
96
One of the main problems encountered when using conventional B-mode ultrasound (US) for targeting and monitoring purposes during ablation therapies employing high-intensity focused US (HIFU) is the appearance of strong interference in the obtained diagnostic US images. In this study, instead of avoiding the interference noise, we demonstrate how we used it to locate the focus of the HIFU transducer in both in-vitro tissue-mimicking phantoms and an ex-vivo tissue block. We found that when the B-mode image plane coincided with the HIFU focal plane, the interference noise was maximally converged and enhanced compared with the off-focus situations. Stronger interference noise was recorded when the angle between the US image plane and the HIFU axis was less than or equal to 90. By intentionally creating a target (group of bubbles) at the 3.5-MHz HIFU focus (7.1 mm in length and 0.7 mm in diameter), the position of the maximal noise convergence coincided well with the target. The differenced between the predicted focus and the actual one (bubbles) on x and z axes (axes perpendicular to the HIFU central axis, Fig. 1) were both about 0.9 mm. For y axis (HIFU central axis), the precision was within 1.0 mm. For tissue block ablation, the interference noise concentrated at the position of maximal heating of the HIFU-induced lesions. The proposed method can also be used to predict the position of the HIFU focus by using a low intensity output scheme before permanent changes in the target tissue were made. The utilization of magnetic resonance imaging (MRI) for HIFU not only real-time monitoring of HIFU ablation but also allows the evaluation of HIFU-induced lesions after treatment. Our study proposed an interleaved dual gradient-echo technique to simultaneously estimate temperature changes and magnetization transfer (MT) contrast, reflecting respectively heating conditions and degree of tissue damage during HIFU treatment. If the target hepatocellular carcinoma (HCC) is close to the surface of the liver, HIFU may overheat intervening tissue such as the diaphragm, abdominal wall, and skin. To avoid this complication, we propose inducing artificial ascites in the abdominal cavity so as to separate the liver from the peritoneum, and to serve as a heat sink to cool overlying structures and thereby avoid inducing permanent damage. Target tissue that was 10 mm below the liver surface was ablated in 12 New Zealand White rabbits: 6 in the experimental group and 6 in the control group. Artificial ascites was established in the experimental group by injecting normal saline into the abdominal cavity until the pressure reached 150 mmH2O. Artificial ascites not only reduced the probability and extent of thermal damage to intervening structures (P<0.05), but also had no adverse affect on the efficacy of HIFU ablation (P>0.05). One of the major disadvantages of HIFU ablation was the small lesion size and thus the long treatment duration. In this study, the effect of using ultrasound contrast agent (UCA) to enlarge the lesion size was studied both in vitro and in vivo. The mechanisms of lesion formation in the presence of UCA microbubbles were studied in vitro and in vivo.

Tese de mestrado integrado em Engenharia Biomédica e Biofísica (Sinais e Imagens Médicas), apresentada à Universidade de Lisboa, através da Faculdade de Ciências, 2013
O fígado é um local frequente de ocorrência de tumores primários e secundários (do intestino, colo-rectal e cancro do estômago). A incidência de cancro do fígado a nível mundial é mais do que um milhão de casos por ano. O cancro no fígado mais comum encontrado na prática clínica são as metástases hepáticas que, juntamente com o carcinoma colo-rectal são os tipos histológicos mais predominantes. O cancro do fígado primário contribuiu com mais de 21 mil novos casos e causou 18.000 mortes nos Estados Unidos em 2008. O carcinoma hepatocelular (HCC) é o cancro primário de fígado mais comum, com uma incidência global estimada de mais de meio milhão de novos casos por ano e é a terceira maior causa de mortes por cancro em todo o mundo. Sem tratamento específico, o prognóstico é muito pobre, e as taxas de sobrevivência médias para pacientes com tumores iniciais e avançados são 6-9 meses e 1-2 meses, respectivamente. O HCC recorrente ocorre em 50% a 80% dos pacientes em 5 anos após a cirurgia de ressecção, a maioria ocorrendo no prazo de 2 anos. Após o tratamento para o HCC, 50% a 90% das mortes no pós-operatório é devido a doença recorrente. A hepatotomia (ressecção hepática) é viável apenas em 10-20% dos casos, e está associada a uma mortalidade operatória de até 5%, no entanto pode atingir taxas de sobrevivência de 5 anos em cerca de 40%. A alternativa actual à cirurgia é a combinação com quimioterapia, mas esta está associada uma taxa de resposta de 20-50%, e taxa de sobrevivência global relativamente curta (12-18 meses). Como resultado, têm sido feitos esforços consideráveis no sentido de proporcionar uma alternativa minimamente invasiva à cirurgia para estes doentes. Essas alternativas actualmente incluem a ablação por radiofrequência, crioterapia e terapia com laser. A única terapia local proposta não-invasiva até à data é High Intensity Focused Ultrasound (HIFU). HIFU é um procedimento não-invasivo que por meio de uma fonte extracorporal de ultra-sons de alta intensidade concentrada induz a necrose por coagulação total de um tecido alvo específico, sem necessidade de exposição cirúrgica ou a inserção de instrumentos no interior da lesão. A intenção do tratamento por HIFU é elevar a temperatura de uma região de tecido delimitada e isolada acima dos 55_C e manter essa temperatura durante 1 segundo ou mais. Isto, por sua vez induz a necrose por coagulação e morte celular. Os pequenos comprimentos de onda (mm) do ultra-som em frequências da ordem dos megahertz (MHz) em tecidos moles permitem que ele seja focado em pequenos volumes clinicamente relevantes. O perigo da maioria dos tratamentos ablativos, onde as células cancerígenas são destruídas pelo aumento da sua temperatura, é que o tecido saudável pode ser danificado. Com HIFU, a energia pode ser focada num volume muito pequeno e bem definido, fazendo com que a passagem do ultra-som através dos tecidos intervenientes, não cause nenhum efeito cumulativo aparente para esses tecidos. HIFU foi aprovado para terapias clínicas de tumores sólidos ou para outras doenças do fígado, rim, pâncreas, recto, próstata, mama, osso, pele e do útero. Em vários centros no mundo, o HIFU está a ser utilizado clinicamente para o tratamento de tumores sólidos (tanto malignos e benignos), incluindo os da próstata, do fígado, da mama, do rim, do osso e do pâncreas, e sarcoma de tecido mole. Tanto o cancro de fígado primário (carcinoma hepatocelular) e metástases hepáticas de cancros do cólon e estômago já foram tratados com HIFU. Apesar dos sucessos clínicos iniciais com tratamentos extracorpóreos de tumores abdominais, ainda existem alguns limites a ultrapassar – especialmente no que diz respeito ao tratamento do fígado, pâncreas e rins – antes de esta técnica poder ser utilizada de forma segura e rotineiramente no interior do corpo. Outra dificuldade importante a ter em conta é o facto de o comprimento do percurso nos tecidos que sobrepõem estes orgãos ser longo e, em geral, a sua composição ser pouco homogénea. Por exemplo, o tratamento terá que transmitir energia através da caixa torácica, o que pode aumentar o risco de queimaduras na pele e danos à superfície das costelas. Do mesmo modo, uma vez que o paciente respira, os efeitos desse movimento durante o tratamento precisa de ser explorado. Podem também ocorrer efeitos de cavitação como resultado do tratamento, e estes efeitos devem ser também considerados. Finalmente, actualmente ainda não existe uma técnica de imagem em tempo real completamente eficaz para a monitorização do tratamento, sendo este um requisito necessário para que a técnica seja utilizada e aplicada com sucesso. Para resolver estas questões, foi desenvolvido um projecto financiado pelo Engineering & Physical Sciences Research Council intitulado "THIFU - Trans-Costal High Intensity Focused Ultrasound" que tem como principal objectivo o desenvolvimento de uma nova técnica para a utilização de HIFU no tratamento de cancros na zona abdominal superior. O objectivo global do programa é desenvolver soluções práticas para esses problemas. Um dos objectivos com o projecto THIFU é a optimização da ablação do tecido no volume alvo e a minimização dos efeitos em estruturas críticas que sobrepõem o local onde queremos administrar a energia ultra-sónica, tais como a pele e a caixa torácica. O planeamento exacto do tratamento e o seu monitoramento bem como a própria administração do mesmo são fundamentais para isso. A fim de melhorar o planeamento e a administração do tratamento, é necessária proceder a uma caracterização precisa das propriedades físicas do tecido. Para isso, é indispensável uma compreensão completa dos campos acústicos, das propriedades dos tecidos no caminho de propagação do ultrassom e os mecanismos pelos quais a destruição dos tecidos é induzida pelo feixe de HIFU. Desta forma, é também importante conhecer a interacção do feixe de ultra-som com os diferentes tipos de tecido, com o fim de criar com precisão um plano de tratamento para a entrega da energia ultra-sónica no foco desejado. Assim, um dos objectivos deste projecto é obter uma estimativa precisa das propriedades acústicas dos tecidos-alvo (por exemplo, a velocidade do som e o coeficiente de atenuação). Esta informação é depois utilizada para determinar a absorção de energia e de transmissão da energia ultra-sónica nos tecidos-alvo. Estes resultados serão usados como parâmetros input para o software de planeamento de tratamento desenvolvido no projecto THIFU. O objectivo do trabalho descrito nesta dissertação foi abordar os dessafios de medir as propriedades acústicas (velocidade do som, coeficiente de atenuação e o coeficiente de não-linearidade) dos tecidos moles - mais concretamente em amostras de fígado humano obtidas logo após uma hepatotomia - que são atravessados pelo feixe HIFU durante o tratamento, com um sistema \all in one" que permite avaliar estas propriedades numa única amostra e num curto espaço de tempo, evitando assim a degradação das amostras. Foi feita uma investigação inicial com o intuito de validar o sistema all in one construído para o efeito com materiais considerados de referência (as suas propriedades acústicas são bastante conhecidas), como por exemplo phantoms em gel e óleo de rícino. Foram também recolhidos dados referentes a amostras de tecidos de origem animal e mais tarde procedeu-se à quantificação das propriedades acústicas de tecido hepático humano. Os valores obtidos com um líquido de referência { óleo de rícino { medido no sistema all-in-one a 23_C estão de acordo com valores publicados por outros estudos. Foram analisadas seis amostras de fígado humano de diferentes patologias. Um total de três tipos de tecidos foi estudado: saudável/normal, cancerígeno e cirrótico. Os resultados obtidos para as amostras de tecido de fígado humano, não mostram clara diferenciação entre o tumor e tecido normal e grande variação entre as amostras. Devido ao tamanho pequeno da amostra não é possível retirar conclusões estatisticamente significativas e precisas nesta fase. No entanto, os resultados gerais mostram que o sistema all-in-one é capaz de determinar estes parâmetros em amostras de referência - como phantoms de criogel e amostras de óleo de rícino - amostras de tecido animal ex-vivo e de tecido hepático humano. Trabalho futuro irá envolver a optimização de certos factores referentes aos materiais e métodos utilizados bem como melhoramentos nos protocolos de preparação das amostras, estudos adicionais com um tamanho de amostra maiores, estudo destas propriedades nos tecidos entre um intervalo de temperaturas relevante para a técnica de HIFU, etc. A dissertação está dividida em 5 capítulos. O primeiro capítulo consiste numa introdução ao tema abordado e a descrição dos principais conceitos inerentes ao trabalho desenvolvido. O segundo capítulo descreve os materiais e metodologias utilizadas. No terceiro e quarto capítulo encontram-se descritos os principais resultados e as conclusões, respectivamente.
High-intensity focused ultrasound (HIFU) provides a potential noninvasive alternative to conventional therapies. In spite of early clinical successes with extracorporeal treatments of abdominal tumours, there are some remaining challenges, especially related to access to the liver, pancreas and kidney. This thesis addresses the challenges of measuring the acoustic (sound speed, attenuation and non-linearity coefficients) properties of soft tissues that lie within the HIFU beam during treatment. Differences in these tissue properties may affect the delivery of thermal therapies, but may also provide a basis for their monitoring with ultrasound. Novel measurement techniques have been developed and applied in the construction of an "all-in-one" tissue characterisation system that allows for the determination of all these parameters in the same region of a single sample. The finite amplitude insertion substitution (FAIS) method was used to obtain the attenuation coefficient and speed of sound. Nonlinearity coefficients were measured using an adapted version of the FAIS method. The attenuation coefficient, sound speed and the acoustic nonlinearity parameter B/A were determined for explant human liver tissue samples with different pathological characteristics. A total of three tissue types were studied: healthy/normal, tumorous and cirrhotic. The results obtained for the human liver tissue samples show no clear differentiation between tumour and normal tissue and big variation between samples. However, as a result of the small sample size and no clear pattern observed, it is not statistically possible to draw accurate and statistically significant conclusions at this stage.The overall results show that the system is able to determine these parameters in reference samples such as cryogel phantoms and castor oil samples, animal ex-vivo tissue and explant human liver tissue. Future studies with a bigger sample size will be required in order to validate the system and also explore thoroughly the relationship between the acoustic properties and temperature variations in the tissue. in reference samples such as cryogel phantoms and castor oil samples, animal ex-vivo tissue and explant human liver tissue. Future studies with a bigger sample size will be required in order to validate the system and also explore thoroughly the relationship between the acoustic properties and temperature variations in the tissue.

Hypoxia Inducible Factors (HIFs) are very important transcription factors that can respond to low oxygen concentrations in the cellular environment. Inhibition of HIF’s transcriptional activity represents a promising approach to new anticancer compounds. Herein, we describe the design and synthesis of a series of HIF-1 inhibitors. Evaluation of these inhibitors using a cell-based luciferase assay led to the discovery compounds with sub-micromolar potency.

The degree of intratumoral hypoxia is clinically correlated to poor response to therapy and increased incidence of distal spread in various cancer subtypes. Specifically, the transcription factor Hypoxia Inducible Factor-1alpha (HIF-1alpha), which is accumulated in cells in response to a hypoxic microenvironment, is implicated in poor disease outcome associated with intratumoral hypoxia. Using novel genetically engineered mouse models of primary soft tissue sarcoma, I show that in vivo genetic deletion of HIF-1alpha specifically in tumor cells 1) decreases the incidence of lung metastases by limiting sarcoma collagen deposition, and 2) improves sarcoma response to radiation therapy by limiting the inflammatory response and metabolic adaptations. These results define HIF-1alpha as a potential target for cancer therapy.

Dissertation

博士
國防醫學院
生命科學研究所
94
Abstract We investigated the antioxidant effect of PGI2 on HIF-1 regulation in human umbilical vein endothelial cells (HUVECs) and the therapy of MCT-caused pulmonary hypertension. HIF-1α, a subunit of HIF-1, is rapidly degraded under normoxic conditions. Hypoxia for 2-4 hrs can upregulate HIF-1αby inhibiting its degradation. In my study, augmentation of PGI2 via adenovirus-mediated gene transfer of both cyclooxygenase-1 and PGI2 synthase (Ad-COP-1) activated HIF-1 by stabilizing HIF-1αin cells under prolonged hypoxia (12 hrs) or the hypoxia-normoxia transition. Exogenous H2O2 abolished PGI2- and catalase-induced HIF-1αupregulation. Moreover, PGI2 attenuated NADPH oxidase activity under hypoxia. These data demonstrate a novel function of PGI2 in downregulating ROS production by attenuating NADPH oxidase activity, which stabilizes HIF-1αin HUVECs exposed to prolonged hypoxia. Pulmonary hypertension could be caused by hypoxia and monocrotaline (MCT) treatment. In my study, I would evaluate the gene therapy effects of COP-1 on pulmonary hypertension. My data show that injection of Ad-COP-1 could significantly attenuate MCT-caused pulmonary artery resistance and right ventricle hypertrophy. Ad-COP-1 infection could also reduce MCT-caused ROS production and apoptosis of endothelial cells (ECs) on vessels. Furthermore, like Ad-COP-1 effect, the anti-apoptosis gene, Bcl-2, could also significantly attenuate MCT-caused pulmonary hypertension. These indicated, except vasodilating effect, PGI2 plays a role on anti-oxidant and anti-apoptosis, during the Ad-COP-1 therapy of MCT-caused pulmonary hypertension.

碩士
國立臺灣大學
臨床牙醫學研究所
96
In Taiwan, the problem of oral cancer in increasing, the increasing rate was the first of the cancer increasing rate. The death patient rate was over half of patient. The methods of treatment cancer were multiple various methods. The aids are elongation the life of patient and cure the disease. The main strategy of the treatment for oral cancer is surgery, but the facial defect resulting from surgery. Due to reduce the invasive treatment, photodynamic therapy (PDT) has been used in the treatment of many cancers, it is the potential therapy, and its advantage is that there is no significant facial defect. The Photodynamic therapy (PDT) is a kind of new developing treatment for cancer. 5-Aminolevulinic acid (ALA) is a novel photosensitizer for photodynamic therapy. We previously showed ALA-PDT can induce cytotoxicity in oral cancer cells. The mechanism for the ALA-PDT induced cytotoxicity is still unknown. Oxygen is the important quench of the three fundamental elements. The expression of hypoxia-inducible factor-1α (HIF-1α) is the regular of oxygen. Overexpressions of HIF-1α have induced the tumor growth and angiogenesis, and affect the treatment result of the cancer therapy, such as radiotherapy and chemotherapy. The PDT has induced the reactive oxygen species (ROS), in the procedure, the O2 was decreased and due to the hypoxia situation. We want to know if the situation was the main cause. That needs us to research. Overexpression of hypoxia-inducible factor-1α (HIF-1α) has been demonstrated in a variety of human cancers and found to be significantly associated with the tumor invasion, lymph node metastasis, clinical stage, survival rate, and prognosis of these cancers. What role is the HIF-1α in the ALA-PDT? In the past study, we found the expression of VEGF was induced with expression of HIF-1α during hypoxia. How to expression of VEGF in the ALA-PDT? In this study, the immunohistochemistry, Western blot analysis, and MTA assay will be used to investigate if the expression of HIF-1α correlates with the varied treatment results of PDT. And the expression of HIF-1α in 13 specimens of oral squamous cell carcinoma (OSCC). We were calculated and compared between the specimens. Then we treated the cell lines with ALA-PDT plus the inhibitor of the HIF-1α and the COX-2, free radical inhibitor and multiple drugs to research the expression of protein. In our study, we found the expressions of HIF-1α were increased with time follow, and induced the expressions of HO-1 and VEGF, then reduced the ALA-PDT-induced apoptosis. In CA9-22 and in SAS cell lines, we used the ALA-PDT treatment could induced the apoptosis. Apoptosis induced by ALA-PDT was both time- and dose-dependent. Pretreatment of cells with imidazole and histidine (singlet oxygen inhibitor) reduced the ALA-PDT-induced apoptosis. These results indicated that singlet oxygen was an important mechanism for ALA-PDT-induced apoptosis in cells lines. In the IHC, the expressions of HIF-1α had no significant finding with gender, age, tumor size, involved depth, tumor region and pathology. The expressions of HIF-1α in nucleus had significant different findings about the resistance to the photodynamic therapy. There is higher resistance to ALA-PDT in the lesion with over expression of HIF-1α. The HIF-1α protein maybe induce the target gene and protein expression to protect the cell survive. PDT combination with anti-HIF-1α treatment enhance the response.

碩士
中國醫藥大學
物理治療學系復健科學碩士班
99
Background Peripheral nerve injury may induce nerve inflammation and neuropathic pain,and these phenomena not only cause physical and psychological discomfort of patients, but may also indirectly affect their quality of life. Low-energy laser treatment (low level laser therapy, LLLT) is often used to reduce pain, inflammation, wound healing and to help accelerate tissue regeneration, but the literatures and studies concerned about the therapeutic effects on the peripheral nerve injury and the molecular biological mechanism of LLLT were very few. In addition, in recent years, many researches explored that hypoxia inducible factor-1α (HIF-1α) involves in the process of inflammation, but its role and mechanism in regulation of peripheral nerve injury-induced inflammation was still not clear. Therefore, the animal model of sciatic chronic constriction injury (CCI) was used in this study for investigating the effects of LLLT on neuropathic pain and functional recovery. Purpose The purposes of this study were to assess the suppressive effects of LLLT on nerve function, accumulation of hypoxia-inducible factor-1 alpha (HIF-1α), proinflammatory cytokines interleukin-1β (IL-1β), tumor necrosis factorα (TNF-α) , inducible nitric oxide syntheses (iNOS), cyclooxygenase enzymes-2 (COX-2), tumor suppressor protein p53 (p53) and macrophages for controlling neuropathic pain after peripheral injury. Moreover, the effects of LLLT on activation of vascular endothelial growth factor (VEGF), nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF) and repair of injured myelin sheath for promoting functional recovery were also assessed in CCI model. Methods Totally 120 rats were divided randomly into four groups based on the surgical procedure of CCI (operated VS sham-operated) and the treatment (LLLT VS sham-operated LLLT).Each group was further divided into three subgroups according to the duration of LLLT treatment. The experimental groups in this study included (1) CCI + LLLT (CL group) included the 1-week (n=10), 2-week (n=10) and 3-week (n=10) of LLLT; (2) CCI+ sLLLT (CsL group) included the 1-week (n=10), 2-week (n=10) and 3-week (n=10) of sLLLT; (3) sCCI + LLLT (sCL group) included the 1-week (n=10), 2-week (n=10) and 3-week (n=10) of LLLT;(4) sCCI + sLLLT(sCsL group)：included the 1-week (n=10), 2-week (n=10) and 3-week (n=10) of sLLLT. Seven days after surgery, animals started using the continuous 660-nm Ga-Al-As diode laser, and the output power of the laser irradiation was 30 mW per session with irradiation time for 60 sec/ per spot. The energy densities were 9 J/cm2. Effects of LLLT in CCI animals were determined by the measurements of mechanical pain threshold, sciatic function index, (SFI), tibial function index (TFI), peroneal function index (PFI), sciatic static index (SSI ), histopathological and immunohistochemical examination, ELISA and Western blot analysis. Histopathological and immunological assessements included accumulation of hypoxia-inducible factor-1 alpha (HIF-1α), proinflammatory cytokines interleukin-1β (IL-1β), tumor necrosis factorα (TNF-α), inducible nitric oxide syntheses (iNOS), cyclooxygenase enzymes-2 (COX-2), tumor suppressor protein p53 (p53) and macrophages for controlling neuropathic pain. Results Our results demonstrated that LLLT significantly improved mechanical pain threshold, sciatic functional index SFI, TFI, PFI, SSI (CL vs. CsL: P <0.05) after CCI. LLLT can also significantly reduce overexpressions of HIF-1α, TNF-α, IL-1β, iNOS, COX-2, p53, and the excessive macrophage accumulation (CL vs. CsL: P <0.05), and significantly promote the amounts of VEGF, NGF, and BDNF and myelin sheath (CL vs. CsL: P <0.05). But there was no significant differences among 1-, 2- and 3-week of LLLT on the expression of these protein factors (P>0.05). Conclusions After CCI nerve injury, LLLT can reduce neuropathic pain by suppressing the HIF-1α, TNF-α, IL-1β, iNOS, COX-2, p53 accumulation and macrophages infiltration and promote functional recovery by increasing the amounts of VEGF, NGF, BDNF and myelin sheath.

博士
高雄醫學大學
醫學研究所
100
Topic 1: Expression of Hypoxia Inducible Factor -1α of Breast Cancer Background: Tumor angiogenesis is essential for solid tumor development, progression and metastases. The purpose of this study is to investigate the relationship between hypoxia inducible factor 1 α (HIF-1α), vascular endothelial growth factor (VEGF) and microvessel density (MVD) as well as their prognostic impacts on patients with breast cancer. Methods and Materials: We collected the clinical data of 240 female patients with breast cancer between January, 1991 and July, 2001. The immunohistochemical expression of HIF-1 α, VEGF and CD31 endothelial cell marker of microvessel density (MVD) was evaluated in formalin-fixed paraffin-embedded breast cancer tissue samples. Data of the immunohistochemical expression were analyzed by chi-square test. The overall survival and disease-free survival were analyzed by Kaplan-Meier method and log rank test. Results: High expression of HIF- 1α, VEGF and MVD was found in 73 patients (30%), 163 patients (68%) and 112 patients (47%), respectively. Expression of HIF-1α was significantly correlated with VEGF (p < 0.001), MVD (p = 0.001). In univariate analysis, high MVD is the significant poor prognostic factor of local recurrenace (p < 0.001). Advanced stage of T and N stage，high expression of HIF-1α、VEGF and high MVD were the poor prognostic factors of distant metastasis significantly (p < 0.001). By multivariate analysis, high expression of HIF-1α，advanced tumor stage and high MVD (p < 0.001) would had significantly poor impacts on clinical outcome of disease-free survival. From Kaplan-Meier analysis, the patients characterized with HIF-1α over-expression showed worse overall survival (p < 0.001). Conclusion: These findings suggest that HIF-1α may play an important role in angiogenesis, tumor progression via regulation of VEGF and microvessel development. The high expression of HIF-1α might be considered as one poor prognostic factor in patients with breast cancer. Topic 2：Comparison of Clinical Outcomes of Breast Cancer Patients with T1-2 Tumor and 1-3 Positive Nodes with or without Post-Mastectomy Radiation Therapy Paper in print, Japanese Journal of Clinical Oncology, accepted on April 26, 2012. Background: The value of postmastectomy radiation therapy (PMRT) for breast cancer patients with T1-2 and 1-3 positive nodes remains controversial. The purpose of this retrospective study was to compare the clinical outcome of breast cancer patients with T1-2 and 1-3 positive nodes with and without PMRT. Methods: Between May 1990 and June 2008, 318 breast cancer patients with T1-2 and 1-3 positive nodes who had undergone modified radical mastectomy received PMRT (n = 163) and no PMRT (n = 155). The clinico-pathologic characteristics were analyzed for clinical outcomes including loco-regional recurrence, distant metastasis, disease-free survival and overall survival. Results: During the median follow-up period of 102 months, the clinical outcome in PMRT vs. no-PMRT groups was as follows: loco-regional recurrence rate (3.1% vs. 11.0%, p = 0.006); distant metastasis rate (20.9% vs. 27.7%, p = 0.152); 10-year disease-free survival rate (73.8% vs. 61.3%, p= 0.001) and 10-year overall survival rate (82.1% vs. 76.1%, p = 0.239). In multivariate analysis, positive nodal ratio ≥ 25% (HR = 4.571, p = 0.003) and positive lymphovascular invasion (HR = 2.738, p = 0.028) were the independent poor prognostic predictors of LRR. The PMRT had significant improvement in reducing loco-regional recurrence (HR = 0.208, p = 0.004). Conclusions: Our results provide some evidence that PMRT be strongly recommended for breast cancer patients with T1-2 and 1-3 positive nodes, especially for high-risk subgroups with positive nodal ratio ≥ 25% and positive lymphovascular invasion, not only for reducing the loco-regional recurrence but also for having benefits on improving disease-free survival. Topic 3: Comparison of Local Recurrence and Distant Metastasis between Breast Cancer Patient after PMRT with and without Immediate TRAM Flap Reconstruction Published at Plastic and Reconstruction Surgery. Vol. 118, No5, 2006, P. 1079-1086 Background：The purpose of this study was to compare the local recurrence and distant metastasis of postmastectomy radiotherapy （PMRT）for breast cancer patients with and without immediate transverse rectus abdominis musculocutaneous (TRAM) flap reconstruction. Methods：Between March of 1997 and October of 2001, 191 breast cancer patients received postmastectomy radiotherapy（PMRT）: 82 patients had TRAM flap reconstruction (TRAM flap group) and 109 patients did not (non-TRAM flap group). The mean radiation dose to the chest wall or entire TRAM flap, axillary area, and lower neck was 50 Gy (range, 48 to 54 Gy). The median follow-up period was 40 months. Results：The percentages of chest wall recurrence were 3.7 percent (three of 82) in the TRAM flap group and 1.8 percent (two of 109) in the non-TRAM flap group (p = 0.653). The percentages of distant metastases were 12.2 percent (10 of 82) in the TRAM group and 15.6 percent (17 of 109) for the non-TRAM group (p = 0.67). The percentages of acute radiation dermatitis according to Radiation Therapy Oncology Group scoring criteria (TRAM flap group versus non-TRAM flap group) were as follows: grade I, 74 of 82 (90 percent) versus 93 of 109 (85 percent); grade II, seven of 82 (9 percent) versus 13 of 109 (12 percent); grade III, one of 82 (1 percent) versus three of 109 (3 percent) (p = 0.558). In the TRAM flap group, the increased percentage of fat necrosis was 8 percent. No flap loss was detected. Conclusions：There were no significant differences in the incidences of complication, locoregional recurrence, and distant metastasis between the TRAM flap and non-TRAM flap patients. The authors’ results suggest that immediate TRAM flap reconstruction can be considered a feasible treatment for breast cancer patients requiring postmastectomy radiotherapy （PMRT）.