Improving Radiotherapy Targeting for Cancer Treatment Through Space and Time

Radiotherapy is a common medical treatment in which lethal doses of ionizing radiation are preferentially delivered to cancerous tumors. In external beam radiotherapy, radiation is delivered by a remote source which sits several feet from the patient\u27s surface. Although great effort is taken in properly aligning the target to the path of the radiation beam, positional uncertainties and other errors can compromise targeting accuracy. Such errors can lead to a failure in treating the target, and inflict significant toxicity to healthy tissues which are inadvertently exposed high radiation doses. Tracking the movement of targeted anatomy between and during treatment fractions provides valuable localization information that allows for the reduction of these positional uncertainties. Inter- and intra-fraction anatomical localization data not only allows for more accurate treatment setup, but also potentially allows for 1) retrospective treatment evaluation, 2) margin reduction and modification of the dose distribution to accommodate daily anatomical changes (called `adaptive radiotherapy\u27), and 3) targeting interventions during treatment (for example, suspending radiation delivery while the target it outside the path of the beam). The research presented here investigates the use of inter- and intra-fraction localization technologies to improve radiotherapy to targets through enhanced spatial and temporal accuracy. These technologies provide significant advancements in cancer treatment compared to standard clinical technologies. Furthermore, work is presented for the use of localization data acquired from these technologies in adaptive treatment planning, an investigational technique in which the distribution of planned dose is modified during the course of treatment based on biological and/or geometrical changes of the patient\u27s anatomy. The focus of this research is directed at abdominal sites, which has historically been central to the problem of motion management in radiation therapy

Adaptations of Escherichia coli strains to oxidative stress are reflected in properties of their structural proteomes.

BACKGROUND:The reconstruction of metabolic networks and the three-dimensional coverage of protein structures have reached the genome-scale in the widely studied Escherichia coli K-12 MG1655 strain. The combination of the two leads to the formation of a structural systems biology framework, which we have used to analyze differences between the reactive oxygen species (ROS) sensitivity of the proteomes of sequenced strains of E. coli. As proteins are one of the main targets of oxidative damage, understanding how the genetic changes of different strains of a species relates to its oxidative environment can reveal hypotheses as to why these variations arise and suggest directions of future experimental work. RESULTS:Creating a reference structural proteome for E. coli allows us to comprehensively map genetic changes in 1764 different strains to their locations on 4118 3D protein structures. We use metabolic modeling to predict basal ROS production levels (ROStype) for 695 of these strains, finding that strains with both higher and lower basal levels tend to enrich their proteomes with antioxidative properties, and speculate as to why that is. We computationally assess a strain's sensitivity to an oxidative environment, based on known chemical mechanisms of oxidative damage to protein groups, defined by their localization and functionality. Two general groups - metalloproteins and periplasmic proteins - show enrichment of their antioxidative properties between the 695 strains with a predicted ROStype as well as 116 strains with an assigned pathotype. Specifically, proteins that a) utilize a molybdenum ion as a cofactor and b) are involved in the biogenesis of fimbriae show intriguing protective properties to resist oxidative damage. Overall, these findings indicate that a strain's sensitivity to oxidative damage can be elucidated from the structural proteome, though future experimental work is needed to validate our model assumptions and findings. CONCLUSION:We thus demonstrate that structural systems biology enables a proteome-wide, computational assessment of changes to atomic-level physicochemical properties and of oxidative damage mechanisms for multiple strains in a species. This integrative approach opens new avenues to study adaptation to a particular environment based on physiological properties predicted from sequence alone

Improving Accuracy of Information Extraction from Quantitative Magnetic Resonance Imaging

Quantitative MRI offers the possibility to produce objective measurements of tissue physiology at different scales. Such measurements are highly valuable in applications such as drug development, treatment monitoring or early diagnosis of cancer. From microstructural information in diffusion weighted imaging (DWI) or local perfusion and permeability in dynamic contrast (DCE-) MRI to more macroscopic observations of the local intestinal contraction, a number of aspects of quantitative MRI are considered in this thesis. The main objective of the presented work is to provide pre-processing techniques and model modification in order to improve the reliability of image analysis in quantitative MRI. Firstly, the challenge of clinical DWI signal modelling is investigated to overcome the biasing effect due to noise in the data. Several methods with increasing level of complexity are applied to simulations and a series of clinical datasets. Secondly, a novel Robust Data Decomposition Registration technique is introduced to tackle the problem of image registration in DCE-MRI. The technique allows the separation of tissue enhancement from motion effects so that the latter can be corrected independently. It is successfully applied to DCE-MRI datasets of different organs. This application is extended to the correction of respiratory motion in small bowel motility quantification in dynamic MRI data acquired during free breathing. Finally, a new local model for the arterial input function (AIF) is proposed. The estimation of the arterial blood contrast agent concentration in DCE-MRI is augmented using prior knowledge on local tissue structure from DWI. This work explores several types of imaging using MRI. It contributes to clinical quantitative MRI analysis providing practical solutions aimed at improving the accuracy and consistency of the parameters derived from image data

Modelling, Simulation and Data Analysis in Acoustical Problems

Modelling and simulation in acoustics is currently gaining importance. In fact, with the development and improvement of innovative computational techniques and with the growing need for predictive models, an impressive boost has been observed in several research and application areas, such as noise control, indoor acoustics, and industrial applications. This led us to the proposal of a special issue about “Modelling, Simulation and Data Analysis in Acoustical Problems”, as we believe in the importance of these topics in modern acoustics’ studies. In total, 81 papers were submitted and 33 of them were published, with an acceptance rate of 37.5%. According to the number of papers submitted, it can be affirmed that this is a trending topic in the scientific and academic community and this special issue will try to provide a future reference for the research that will be developed in coming years

Magnetic resonance imaging of colonic function

The overall aim of this work was to develop MRI methods and techniques to study the physiology and the pathology of the gastrointestinal tract, with particular attention to the colon. Besides, the development of new methods was aimed in order to perform quantitative analysis using proton and fluorine MRI. In particular the first experimental chapter describes the development and the optimisation of imaging protocols for studying colonic function in undisturbed physiologically relevant conditions. In addition a texture analysis method based on Gabor filters is developed and used for the objective assessment of colonic content characteristics. The mechanisms of action of common anti-diarrhoeal and anti-constipation agents are also investigated. The last experimental chapter describes the development of methods for using markers to measure GI transit. Transit time, i.e. the time it takes for a marker to pass through the entire gut, is often affected by functional gastrointestinal disorders, therefore it is of primary importance to develop a non-invasive and effective technique for the diagnosis of such gastrointestinal diseases. The use of fluorinated agents and its many advantages compared to other techniques is outlined and the first in vivo studies at high field are presented. The use of gadolinium based compounds as an additional marker is also discussed

Developing of an organ on chip device as novel in vitro platform to study organ mechanobiology: Peristalsis on a chip.

Developing of an organ on chip device as novel in vitro platform to study organ mechanobiology: Peristalsis on a chip. Knowing the mechanical properties of the gastrointestinal (GI) tract appears to be important for understanding the molecular and cellular responses to mechanical stimuli on physiological processes such as foods, xenobiotic or drugs digestion/absorption. These processes are mediated by various intestinal cells such as epithelial cells, interstitial cells, smooth muscle cells, and neurocytes. The loss or dysfunction of specific cells or mechanical strength of cell bowel wall directly results in GI tract disease. Reversing the abnormal status of pathogenic cells has been considered crucial to treatment of gut diseases. Gut bioengineered models have been developing for the purpose to replace the damaged tissues and to provide three-dimensional platforms that mimic the in vivo environment to study drug development, absorption and toxicity. Nevertheless, the need to develop more complex models in vitro to study mechanical stress is growing. In this perspective, this project will allow us to get an automatized microfluidic gut platform to evaluate the pathophysiology of the small intestine through the study of the shear stress of the bolus on the epithelial cells layer at the lumen side of the healthy or diseased 3D intestine models. To this aim, the major goals of this project are the the design and fabrication of complex and innovative microfluidic device provided with an integrated PDMS membrane designed to mimic the crypt-villus axis in order to promote the differentiation of the intestinal epithelium and the establishment of peristaltic motion by means of an automatized and controlled elettrovalve system. The platform was used to estimate the intestinal transport properties of the bolus and the physiological condition of the shear stress under peristaltic motion. An important feature of the device, is the possibility to induce a fluid flow both at the basolateral and the lumen side of the intestinal epithelium, therefore the possibility to introduce integrated electrodes in the apical side and basoteral side in order to be enable continuous monitoring of cells behaviour and differentiation through TransEpithelial Electrical Resistance measurements. The effect of PDMS membrane morphology, peristaltic motion and shear stress on intestinal epithelial cell differentiation, mucus production and molecules adsorption process has been evaluated. The development of the Peristalsis on chip device could be reduce the poorly predictive preclinical evaluation generated by the phylogenetic distance between laboratory animals and humans, the discrepancy between current in vitro systems and the human body, and the restrictions of in silico modelling

Feeling full and being full : how gastric content relates to appetite, food properties and neural activation

Aim: This thesis aimed to further determine how gastric content relates to subjective experiences regarding appetite, how this relation is affected by food properties and whether this is visible in neural activation changes. Method: This was studied using questionnaires, MRI of the stomach and fMRI of the brain. Randomized, controlled crossover experiments with healthy men and for one experiment women were performed. Results: MRI measurements of the stomach as opposed to an indirect measurement by proxy, such as 13C breath testing are to be preferred. We show that gastric emptying is affected by energy load, and to a much smaller extent by viscosity. Additionally we show that a thick shake containing 100 kcal will yield higher fullness sensations than a thin shake containing 500 kcal. In the chapter we name this phenomenon ‘phantom fullness’, i.e., a sense of fullness and satiation caused by the taste and mouthfeel of a food which is irrespective of actual stomach fullness. A liquid meal followed by a drink of water empties about twice as fast in the first 35 minutes compared to the same amount of water incorporated within the liquid meal. Using MRI we were able to show layering within the stomach and increased emptying of this watery layer. With 300mL of increased gastric content inducing distention, appetite was lowered. Ingestion led to significant changes in activation in the right insula and parts of the left and right inferior frontal cortices over time. Women retain significantly more fluid after a carbonated drink in their stomach than men. When comparing correlations between subjective ratings and intragastric liquid and gas and total gastric volume, nausea and fullness correlated strongest with the liquid fraction within the stomach, bloating strongest with total gastric volume. Conclusion: There are marked differences betweengastric content and subjective experiences regarding appetite. Viscosity is a main driver of these differences. Combined gastric MRI and brain fMRI measurements need to be performed to understand this further.</p

The Role of Schwann Cell Mitochondrial Metabolism in Schwann Cell Biology and Axonal Survival

Mitochondrial dysfunction has emerged as a common cause of peripheral neuropathies. While the role of neuronal and axonal mitochondria in peripheral nerve disease is well appreciated, whether Schwann cell: SC) mitochondrial deficits contribute to peripheral neuropathies is unclear. Greater insight into the biology and pathology of SC mitochondrial metabolism could be relevant to the treatment of peripheral neuropathies, particularly because SCs critically support axonal stability and function as well as promote peripheral nerve regeneration. The present thesis investigates the contribution of SC mitochondrial deficits to disease progression in peripheral neuropathies as well as the gene regulatory network that drives the SC regenerative response after injury and in disease states. We describe the generation and characterization of the first mouse model useful in directly interrogating the contribution of SC mitochondrial dysfunction to peripheral neuropathy. These mice: Tfam-SCKOs) were produced through the tissue-specific deletion of the mitochondrial transcription factor A gene: Tfam), which is required for mtDNA transcription and replication. Interestingly, induction of SC-specific mitochondrial dysfunction did not affect SC survival; instead, these deficits resulted in a severe, progressive peripheral neuropathy characterized by extensive axonal degeneration that recapitulated critical features of human neuropathy. Mechanistically, we demonstrated that SC mitochondrial dysfunction activates a maladaptive integrated stress response and causes a shift in lipid metabolism away from new lipid biosynthesis towards increased lipid oxidation. These alterations in lipid metabolism caused the early depletion of key myelin lipid components as well as a dramatic accumulation of acylcarnitine lipid intermediates. Importantly, release of acylcarnitines from SCs was toxic to axons and induced their degeneration. Our results show that normal mitochondrial function in SCs is essential for maintenance of axonal survival and normal peripheral nerve function, suggesting that SC mitochondrial dysfunction contributes to human peripheral neuropathies. Moreover, our work identifies alterations in SC lipid metabolism and the accumulation of toxic lipid intermediates as novel mechanisms driving some of the pathology in peripheral neuropathies associated with mitochondrial dysfunction. Tfam-SCKO mice showed a severe deficiency in their ability to remyelinate peripheral nerve axons after injury. To gain insight into the highly orchestrated process of SC-mediated support of axonal regeneration, we also investigated the transcriptional and post-transcriptional gene regulatory program that drives the SC regenerative response. We profiled the expression of SC microRNAs: miRNAs) after peripheral nerve lesions as well as characterized the injury response of SCs with disrupted miRNA processing and showed that SC miRNAs modulated the injury response largely by targeting positive regulators of SC dedifferentiation/proliferation. SC miRNAs cooperated with transcriptional regulators to promote rapid and robust transitions between the distinct differentiation states necessary to support nerve regeneration. Moreover, we identified miR-34a and miR-140 as regulators of SC proliferation and myelination. We then used a novel computational approach to infer the gene regulatory network involved in this SC injury response and gain insight on cooperative regulation of this process by transcription factors and miRNAs. Together, the results described in the present thesis represent a significant increase in our understanding of how mitochondrial abnormalities specifically in SCs contribute to clinical impairment in patients with peripheral neuropathy. Moreover, the mechanistic characterization of lipid metabolism abnormalities in SCs following mitochondrial dysfunction elucidates potentially important therapeutic targets. Finally, our analysis of the transcriptional and post-transcriptional gene regulatory network involved in the SC regenerative response also provides valuable insight that could be harnessed to help restore normal nerve function in patients with peripheral neuropathy