A 2D/3D image analysis system to track fluorescently labeled structures in rod-shaped cells: application to measure spindle pole asymmetry during mitosis.

BACKGROUND: The yeast Schizosaccharomyces pombe is frequently used as a model for studying the cell cycle. The cells are rod-shaped and divide by medial fission. The process of cell division, or cytokinesis, is controlled by a network of signaling proteins called the Septation Initiation Network (SIN); SIN proteins associate with the SPBs during nuclear division (mitosis). Some SIN proteins associate with both SPBs early in mitosis, and then display strongly asymmetric signal intensity at the SPBs in late mitosis, just before cytokinesis. This asymmetry is thought to be important for correct regulation of SIN signaling, and coordination of cytokinesis and mitosis. In order to study the dynamics of organelles or large protein complexes such as the spindle pole body (SPB), which have been labeled with a fluorescent protein tag in living cells, a number of the image analysis problems must be solved; the cell outline must be detected automatically, and the position and signal intensity associated with the structures of interest within the cell must be determined. RESULTS: We present a new 2D and 3D image analysis system that permits versatile and robust analysis of motile, fluorescently labeled structures in rod-shaped cells. We have designed an image analysis system that we have implemented as a user-friendly software package allowing the fast and robust image-analysis of large numbers of rod-shaped cells. We have developed new robust algorithms, which we combined with existing methodologies to facilitate fast and accurate analysis. Our software permits the detection and segmentation of rod-shaped cells in either static or dynamic (i.e. time lapse) multi-channel images. It enables tracking of two structures (for example SPBs) in two different image channels. For 2D or 3D static images, the locations of the structures are identified, and then intensity values are extracted together with several quantitative parameters, such as length, width, cell orientation, background fluorescence and the distance between the structures of interest. Furthermore, two kinds of kymographs of the tracked structures can be established, one representing the migration with respect to their relative position, the other representing their individual trajectories inside the cell. This software package, called "RodCellJ", allowed us to analyze a large number of S. pombe cells to understand the rules that govern SIN protein asymmetry. CONCLUSIONS: "RodCell" is freely available to the community as a package of several ImageJ plugins to simultaneously analyze the behavior of a large number of rod-shaped cells in an extensive manner. The integration of different image-processing techniques in a single package, as well as the development of novel algorithms does not only allow to speed up the analysis with respect to the usage of existing tools, but also accounts for higher accuracy. Its utility was demonstrated on both 2D and 3D static and dynamic images to study the septation initiation network of the yeast Schizosaccharomyces pombe. More generally, it can be used in any kind of biological context where fluorescent-protein labeled structures need to be analyzed in rod-shaped cells. AVAILABILITY: RodCellJ is freely available under http://bigwww.epfl.ch/algorithms.html, (after acceptance of the publication).

Direct visualization of the trimeric structure of the ASIC1a channel, using AFM imaging.

There has been confusion about the subunit stoichiometry of the degenerin family of ion channels. Recently, a crystal structure of acid-sensing ion channel (ASIC) 1a revealed that it assembles as a trimer. Here, we used atomic force microscopy (AFM) to image unprocessed ASIC1a bound to mica. We detected a mixture of subunit monomers, dimers and trimers. In some cases, triple-subunit clusters were clearly visible, confirming the trimeric structure of the channel, and indicating that the trimer sometimes disaggregated after adhesion to the mica surface. This AFM-based technique will now enable us to determine the subunit arrangement within heteromeric ASICs.

Micropapillary pattern in lung adenocarcinoma: aspect on 18F-fluorodeoxyglucose positron emission tomography/computed tomography imaging.

We diagnosed a non-small cell lung carcinoma in a 49-year-old female patient with the histopathological diagnosis of stage IIIB mixed bronchioloalveolar and papillary adenocarcinoma with extensive micropapillary feature, which was not visualized on the preoperative multimodality imaging with positron emission tomography (PET) and computed tomography (CT). The micropapillary component characterized by a unique growth pattern with particular morphological features can be observed in all subtypes of lung adenocarcinoma. Micropapillary component is increasingly recognized as a distinct entity associated with higher aggressiveness. Even the most modern multimodality PET/CT imaging technology may fail to adequately visualize this important component with highly relevant prognostic implications. Thus, the pathologist needs to consciously look for a micropapillary component in the surgical specimen or in preoperative biopsies or cytology. This may have potential future treatment implications, as adjuvant or neoadjuvant chemotherapy may be of relevance, even in the early stages of the disease.

18F-fluorodeoxyglucose positron emission tomography/computed tomography and magnetic resonance imaging in patients with liver metastases from uveal melanoma: results from a pilot study.

PURPOSE: F-fluorodeoxyglucose (FDG) positron emission tomography/computed tomography (PET/CT) and MRI are used for detecting liver metastases from uveal melanoma. The introduction of new treatment options in clinical trials might benefit from early response assessment. Here, we determine the value of FDG-PET/CT with respect to MRI at diagnosis and its potential for monitoring therapy. MATERIAL AND METHODS: Ten patients with biopsy-proven liver metastases of uveal melanoma enrolled in a randomized phase III trial (NCT00110123) underwent both FDG-PET coupled with unenhanced CT and gadolinium-diethylene triamine pentaacetic acid-enhanced liver MRI within 4 weeks. FDG-PET and MRI were evaluated blindly and then compared using the ratio of lesion to normal liver parenchyma PET-derived standardized uptake value (SUV). The influence of lesion size and response to chemotherapy were studied. RESULTS: Overall, 108 liver lesions were seen: 34 (31%) on both modalities (1-18 lesions/patient), four (4%) by PET/CT only, and 70 (65%) by MRI only. SUV correlated with MRI lesion size (r=0.81, P<0.0001). PET/CT detected 26 of 33 (79%) MRI lesions of more than or equal to 1.2 cm, whereas it detected only eight of 71 (11%) lesions of less than 1.2 cm (P<0.0001). MRI lesions without PET correspondence were small (0.6±0.2 vs. 2.1±1.1 cm, P<0.0001). During follow-up (six patients, 30 lesions), the ratio lesion-to-normal-liver SUV diminished in size-stable lesions (1.90±0.64-1.46±0.50, P<0.0001), whereas it increased in enlarging lesions (1.56±0.40-1.99±0.56, P=0.032). CONCLUSION: MRI outweighs PET/CT for detecting small liver metastases. However, PET/CT detected at least one liver metastasis per patient and changes in FDG uptake not related to size change, suggesting a role in assessing early therapy response.

Mophological analysis of the human vasculature from medical imaging. Application to cerebral and coranary arteries

In this project, we have investigated new ways of modelling and analysis of human vasculature from Medical images. The research was divided in two main areas: cerebral vasculature analysis and coronary arteries modeling. Regarding cerebral vasculature analysis, we have studed cerebral aneurysms, internal carotid and the Circle of Willis (CoW). Aneurysms are abnormal vessel enlargements that can rupture causing important cerebral damages or death. The understanding of this pathology, together with its virtual treatment, and image diagnosis and prognosis, includes identification and detailed measurement of the aneurysms. In this context, we have proposed two automatic aneurysm isolation method, to separate the abnormal part of the vessel from the healthy part, to homogenize and speed-up the processing pipeline usually employed to study this pathology, [Cardenes2011TMI, arrabide2011MedPhys]. The results obtained from both methods have been also compared and validatied in [Cardenes2012MBEC]. A second important task here the analysis of the internal carotid [Bogunovic2011Media] and the automatic labelling of the CoW, Bogunovic2011MICCAI, Bogunovic2012TMI]. The second area of research covers the study of coronary arteries, specially coronary bifurcations because there is where the formation of atherosclerotic plaque is more common, and where the intervention is more challenging. Therefore, we proposed a novel modelling method from Computed Tomography Angiography (CTA) images, combined with Conventional Coronary Angiography (CCA), to obtain realistic vascular models of coronary bifurcations, presented in [Cardenes2011MICCAI], and fully validated including phantom experiments in [Cardene2013MedPhys]. The realistic models obtained from this method are being used to simulate stenting procedures, and to investigate the hemodynamic variables in coronary bifurcations in the works submitted in [Morlachi2012, Chiastra2012]. Additionally, another preliminary work has been done to reconstruct the coronary tree from rotational angiography, and published in [Cardenes2012ISBI].

Creación de una demo 3D mediante gráficos procedurales

Aquest projecte consisteix en el desenvolupament d’una demo 3D utilitzant exclusivament gràfics procedurals per tal d’avaluar la seva viabilitat en aplicacions més complexes com els videojocs. En aquesta aplicació es genera un terreny aleatori explorable amb vegetació i textures creades proceduralment.

Reporting Guidance for Oncologic 18F-FDG PET/CT Imaging.

The written report (or its electronic counterpart) is the primary mode of communication between the physician interpreting an imaging study and the referring physician. The content of this report not only influences patient management and clinical outcomes but also serves as legal documentation of services provided and can be used to justify medical necessity, billing accuracy, and regulatory compliance. Generating a high-quality PET/CT report is perhaps more challenging than generating a report for other imaging studies because of the complexity of this hybrid imaging modality. This article discusses the essential elements of a concise and complete oncologic (18)F-FDG PET/CT report and illustrates these elements through examples taken from routine clinical practice.

Dose reconstruction in the context of tomotherapy

In vivo dosimetry is a way to verify the radiation dose delivered to the patient in measuring the dose generally during the first fraction of the treatment. It is the only dose delivery control based on a measurement performed during the treatment. In today's radiotherapy practice, the dose delivered to the patient is planned using 3D dose calculation algorithms and volumetric images representing the patient. Due to the high accuracy and precision necessary in radiation treatments, national and international organisations like ICRU and AAPM recommend the use of in vivo dosimetry. It is also mandatory in some countries like France. Various in vivo dosimetry methods have been developed during the past years. These methods are point-, line-, plane- or 3D dose controls. A 3D in vivo dosimetry provides the most information about the dose delivered to the patient, with respect to ID and 2D methods. However, to our knowledge, it is generally not routinely applied to patient treatments yet. The aim of this PhD thesis was to determine whether it is possible to reconstruct the 3D delivered dose using transmitted beam measurements in the context of narrow beams. An iterative dose reconstruction method has been described and implemented. The iterative algorithm includes a simple 3D dose calculation algorithm based on the convolution/superposition principle. The methodology was applied to narrow beams produced by a conventional 6 MV linac. The transmitted dose was measured using an array of ion chambers, as to simulate the linear nature of a tomotherapy detector. We showed that the iterative algorithm converges quickly and reconstructs the dose within a good agreement (at least 3% / 3 mm locally), which is inside the 5% recommended by the ICRU. Moreover it was demonstrated on phantom measurements that the proposed method allows us detecting some set-up errors and interfraction geometry modifications. We also have discussed the limitations of the 3D dose reconstruction for dose delivery error detection. Afterwards, stability tests of the tomotherapy MVCT built-in onboard detector was performed in order to evaluate if such a detector is suitable for 3D in-vivo dosimetry. The detector showed stability on short and long terms comparable to other imaging devices as the EPIDs, also used for in vivo dosimetry. Subsequently, a methodology for the dose reconstruction using the tomotherapy MVCT detector is proposed in the context of static irradiations. This manuscript is composed of two articles and a script providing further information related to this work. In the latter, the first chapter introduces the state-of-the-art of in vivo dosimetry and adaptive radiotherapy, and explains why we are interested in performing 3D dose reconstructions. In chapter 2 a dose calculation algorithm implemented for this work is reviewed with a detailed description of the physical parameters needed for calculating 3D absorbed dose distributions. The tomotherapy MVCT detector used for transit measurements and its characteristics are described in chapter 3. Chapter 4 contains a first article entitled '3D dose reconstruction for narrow beams using ion chamber array measurements', which describes the dose reconstruction method and presents tests of the methodology on phantoms irradiated with 6 MV narrow photon beams. Chapter 5 contains a second article 'Stability of the Helical TomoTherapy HiArt II detector for treatment beam irradiations. A dose reconstruction process specific to the use of the tomotherapy MVCT detector is presented in chapter 6. A discussion and perspectives of the PhD thesis are presented in chapter 7, followed by a conclusion in chapter 8. The tomotherapy treatment device is described in appendix 1 and an overview of 3D conformai- and intensity modulated radiotherapy is presented in appendix 2. - La dosimétrie in vivo est une technique utilisée pour vérifier la dose délivrée au patient en faisant une mesure, généralement pendant la première séance du traitement. Il s'agit de la seule technique de contrôle de la dose délivrée basée sur une mesure réalisée durant l'irradiation du patient. La dose au patient est calculée au moyen d'algorithmes 3D utilisant des images volumétriques du patient. En raison de la haute précision nécessaire lors des traitements de radiothérapie, des organismes nationaux et internationaux tels que l'ICRU et l'AAPM recommandent l'utilisation de la dosimétrie in vivo, qui est devenue obligatoire dans certains pays dont la France. Diverses méthodes de dosimétrie in vivo existent. Elles peuvent être classées en dosimétrie ponctuelle, planaire ou tridimensionnelle. La dosimétrie 3D est celle qui fournit le plus d'information sur la dose délivrée. Cependant, à notre connaissance, elle n'est généralement pas appliquée dans la routine clinique. Le but de cette recherche était de déterminer s'il est possible de reconstruire la dose 3D délivrée en se basant sur des mesures de la dose transmise, dans le contexte des faisceaux étroits. Une méthode itérative de reconstruction de la dose a été décrite et implémentée. L'algorithme itératif contient un algorithme simple basé sur le principe de convolution/superposition pour le calcul de la dose. La dose transmise a été mesurée à l'aide d'une série de chambres à ionisations alignées afin de simuler la nature linéaire du détecteur de la tomothérapie. Nous avons montré que l'algorithme itératif converge rapidement et qu'il permet de reconstruire la dose délivrée avec une bonne précision (au moins 3 % localement / 3 mm). De plus, nous avons démontré que cette méthode permet de détecter certaines erreurs de positionnement du patient, ainsi que des modifications géométriques qui peuvent subvenir entre les séances de traitement. Nous avons discuté les limites de cette méthode pour la détection de certaines erreurs d'irradiation. Par la suite, des tests de stabilité du détecteur MVCT intégré à la tomothérapie ont été effectués, dans le but de déterminer si ce dernier peut être utilisé pour la dosimétrie in vivo. Ce détecteur a démontré une stabilité à court et à long terme comparable à d'autres détecteurs tels que les EPIDs également utilisés pour l'imagerie et la dosimétrie in vivo. Pour finir, une adaptation de la méthode de reconstruction de la dose a été proposée afin de pouvoir l'implémenter sur une installation de tomothérapie. Ce manuscrit est composé de deux articles et d'un script contenant des informations supplémentaires sur ce travail. Dans ce dernier, le premier chapitre introduit l'état de l'art de la dosimétrie in vivo et de la radiothérapie adaptative, et explique pourquoi nous nous intéressons à la reconstruction 3D de la dose délivrée. Dans le chapitre 2, l'algorithme 3D de calcul de dose implémenté pour ce travail est décrit, ainsi que les paramètres physiques principaux nécessaires pour le calcul de dose. Les caractéristiques du détecteur MVCT de la tomothérapie utilisé pour les mesures de transit sont décrites dans le chapitre 3. Le chapitre 4 contient un premier article intitulé '3D dose reconstruction for narrow beams using ion chamber array measurements', qui décrit la méthode de reconstruction et présente des tests de la méthodologie sur des fantômes irradiés avec des faisceaux étroits. Le chapitre 5 contient un second article intitulé 'Stability of the Helical TomoTherapy HiArt II detector for treatment beam irradiations'. Un procédé de reconstruction de la dose spécifique pour l'utilisation du détecteur MVCT de la tomothérapie est présenté au chapitre 6. Une discussion et les perspectives de la thèse de doctorat sont présentées au chapitre 7, suivies par une conclusion au chapitre 8. Le concept de la tomothérapie est exposé dans l'annexe 1. Pour finir, la radiothérapie «informationnelle 3D et la radiothérapie par modulation d'intensité sont présentées dans l'annexe 2.

FIB-SEM tomography in biology.

Three-dimensional information is much easier to understand than a set of two-dimensional images. Therefore a layman is thrilled by the pseudo-3D image taken in a scanning electron microscope (SEM) while, when seeing a transmission electron micrograph, his imagination is challenged. First approaches to gain insight in the third dimension were to make serial microtome sections of a region of interest (ROI) and then building a model of the object. Serial microtome sectioning is a tedious and skill-demanding work and therefore seldom done. In the last two decades with the increase of computer power, sophisticated display options, and the development of new instruments, an SEM with a built-in microtome as well as a focused ion beam scanning electron microscope (FIB-SEM), serial sectioning, and 3D analysis has become far easier and faster.Due to the relief like topology of the microtome trimmed block face of resin-embedded tissue, the ROI can be searched in the secondary electron mode, and at the selected spot, the ROI is prepared with the ion beam for 3D analysis. For FIB-SEM tomography, a thin slice is removed with the ion beam and the newly exposed face is imaged with the electron beam, usually by recording the backscattered electrons. The process, also called "slice and view," is repeated until the desired volume is imaged.As FIB-SEM allows 3D imaging of biological fine structure at high resolution of only small volumes, it is crucial to perform slice and view at carefully selected spots. Finding the region of interest is therefore a prerequisite for meaningful imaging. Thin layer plastification of biofilms offers direct access to the original sample surface and allows the selection of an ROI for site-specific FIB-SEM tomography just by its pronounced topographic features.

Hybrid PET/MRI in the assessment of cardiac viability: added value compared to PET/CT

Purpose: We evaluated the potential for hybrid PET/MRI devices to provide integrated metabolic, functional and anatomic characterisation of patients with suspected coronary artery disease.Methods and Materials: Ten patients (5 with suspected hibernating myocardium and 5 healthy volunteers) performed an imaging study using a hybrid PET/MRI (Philips). Viability assessed by 18F-FDG was performed in diseased patients along with MRI anatomic and functional study and reassessed within 30 minutes by conventional PET/CT. Non-contrast right coronary artery (RCA) targeted and whole heart 3D coronary angio-MRI using ECG-gating and respiratory navigator was performed in healthy volunteers with reconstruction performed using MPR and volume rendering. The extent of metabolic defect (MD) using PET/MRI and PET/CT was compared in patients and coronary territories (LAD, CX, RCA). Assessability of coronary lumen was judged as good, sub-optimal or non-assessable using a 16-segments coronary model.Results: Metabolic assessment was successful in all patients with MD being 19.2% vs 18.3% using PET/MRI and PET/CT, respectively (P=ns). The MD was 10.2%, 6 %, and 3 % vs 9.3%, 6 % and 3 % for LAD, CX and RCA territories, respectively (P= ns). Coronary angio-MRI was successful in all volunteers with 66 coronary segments visualised overall. The RCA was fully visualised in 4/5 volunteers and the left coronary arteries in 4/5 volunteers. Assessability in visualised segments was good, sub-optimal and non-assessable in 88 %, 2 % and 10 %, respectively.Conclusion: Hybrid PET/MRI devices may enable metabolic evaluation comparable to PET/CT with additional value owing to accurate functional and anatomical information including coronary assessment.