Mathematical Models of Tumors and Their Remote Metastases

Clinical observations and indications in the literature have led us to investigate several models of tumors. For example, it has been shown that a tumor has the ability to send out anti-growth factors, or inhibitors, to keep its remote metastases from growing. Thus, we model the depleting effect of such a growth inhibitor after the removal of the primary tumor (thus removing the source) as a function of time t and distance from the original tumor r. It has also been shown clinically that oxygen and glucose are nutrients critical to the survival and growth of tumors. Thus, we model the effects of immersing a tumor into a nutrient bath. Similarly, this model could represent the addition of nutrient to the tissue surrounding a spherical tumor. In this paper, we model several problems associated with the clinical observations noted above, and draw conclusions based on the obtained results

Microfluidic device flow field characterization around tumor spheroids with tunable necrosis produced in an optimized off-chip process

Tumor spheroids are a 3-D tumor model that holds promise for testing cancer therapies in vitro using microfluidic devices. Tailoring the properties of a tumor spheroid is critical for evaluating therapies over a broad range of possible indications. Using human colon cancer cells (HCT-116), we demonstrate controlled tumor spheroid growth rates by varying the number of cells initially seeded into microwell chambers. The presence of a necrotic core in the spheroids could be controlled by changing the glucose concentration of the incubation medium. This manipulation had no effect on the size of the tumor spheroids or hypoxia in the spheroid core, which has been predicted by a mathematical model in computer simulations of spheroid growth. Control over the presence of a necrotic core while maintaining other physical parameters of the spheroid presents an opportunity to assess the impact of core necrosis on therapy efficacy. Using micro-particle imaging velocimetry (micro-PIV), we characterize the hydrodynamics and mass transport of nanoparticles in tumor spheroids in a microfluidic device. We observe a geometrical dependence on the flow rate experienced by the tumor spheroid in the device, such that the “front” of the spheroid experiences a higher flow velocity than the “back” of the spheroid. Using fluorescent nanoparticles, we demonstrate a heterogeneous accumulation of nanoparticles at the tumor interface that correlates with the observed flow velocities. The penetration depth of these nanoparticles into the tumor spheroid depends on nanoparticle diameter, consistent with reports in the literature

High throughput analysis of the penetration of iron oxide/polyethylene glycol nanoparticles into multicellular breast cancer tumor spheroids

The purpose of this study was to design and optimize a system for the high-throughput analysis of multicellular tumor spheroids (MCTS), and validate the system through the study of a complex biological model. The system was successfully created and optimized, allowing the histological recovery of MCTS at rates up to 90% for microarrays of 24-spheroids. Arrays of 96-spheroids were recovered at rates up to 86%. The system was used to study the penetration of 5k Da-polyethylene coated superparamagnetic iron-oxide nanoparticles (5k-PEG SPIONs) into HTB-126 breast cancer spheroids cultured to a mean diameter of 486 micrometer (± 25.2 micrometer). Results were compared to an identical study using 2D cultures. Positive staining for the SPION dosage of 100 microgram/milliliter in 2D culture regardless of incubation time was observed along with a lack of staining for all other concentrations in both 2D and 3D. SPION incubation led to necrosis in breast cancer spheroids after 3 days

Characterization of polysaccharide multilayered capsules for tissue engineering applications

Cell encapsulation has been widely studied as an alternative therapy for almost every human diseases and disorders. This technique enables the inclusion of various types of living cells inside spherical systems which, among other capabilities, mimic the environment provided by the extracellular matrix. This new therapeutic approach has already proved to be successful either in vitro or in vivo studies, thus becoming one of the most promising tools in tissue engineering and regenerative medicine. The main goal of this thesis was to explore some of the potential of cell encapsulation using simple and versatile techniques that can be performed in physiological and friendly conditions to the cells. In a first approach, cells were encapsulated in liquid-core capsules using a three step methodology: (i) the precipitation of a polymer solution of alginate into a bath of calcium chloride (ionotropic gelation), (ii) deposition of polyelectrolyte multilayers onto the surface of the beads, in a process called layer-by-layer. (iii) use of EDTA to liquefy the alginate core. Two different natural-based polymers were used, alginate, the most studied copolymer for cell encapsulation and chitosan, a polymer widely explored in a variety of biomedical applications. Both polymers were proved to be biocompatible, biodegradable and can be manipulated under physiological conditions. All the capsules produced exhibited spherical shape, smooth surface and liquid-core. The results shown that encapsulated cells were viable and proliferating few days after the alginate-chitosan multilayer buildup, which suggests that the develop capsules posses a semipermeable membrane which allows the correct diffusion of nutrients and metabolites. A preliminary study was started to test the feasibility of culturing anchorage-dependent cells in PLLA solid microparticles previously treated with human serum fibronectin followed by the encapsulation of the whole set in alginate-chitosan liquid-core capsules. The results are still very incipient but very promising

Engineering novel micro-scaffolds and bottom up strategies for in vitro building of vascularized hybrid tissues

There is a significant demand of de-novo engineered tissue grafts capable of replacing biological tissue and/or organ functions for clinical applications. The major obstacle to achieve this goal is the difficulty of recreating all of the complex biochemical and biomechanical functions of the tissue to regenerate, together with transport limitations into the bulk of these newly synthesized tissues. Oxygen and nutrients are supplied to cells and tissues naturally by the microvasculature, which is composed of branching, variable diameter blood vessels. Replicating the complex architecture and functionalities of native tissue vasculature is therefore one of the most important challenge in tissue engineering strategies. To date, bottom up techniques are strong powerful tools to build large viable tissue constructs by packing and sintering cell-laden scaffold-based micro-modules (μ-scaffolds) in a mould. In fact, after sintering and further μ-scaffolds degradation it is possible to achieve large viable tissues in vitro, replicating the composition and structure of native tissue and suitable for studying biological processes involved in new tissue genesis, maturation and remodelling. The aim of this work is to design and engineering novel μ-scaffolds and bottom up assembly techniques to fabricate vascularized layered tissues and to study the effect of μ-scaffolds spatial distribution and co-culture of human dermal fibroblasts (HDFs) together with human umbilical vein endothelial cells (HUVECs) on new tissue growth and vascularization in vitro. To achieve these aims, in first part of this study, we fabricated porous polycaprolactone (PCL) μ-scaffolds with bioinspired trabecular structure and we demonstrated that these newly developed μ-scaffolds supported the in vitro adhesion, growth, and biosynthesis of HDFs. The μ-scaffolds were fabricated by using a fluidic emulsion/porogen leaching/particle coagulation process and by using polyethylene oxide (PEO) as a biocompatible pore-generating agent. In particular, the effect of the composition of the polymeric solution and the flow rate of the continuous phase on μ-scaffolds size distribution, morphology and architectural properties were assessed with the aim to find the best preparation conditions for biological characterization. In vitro culture of HDFs showed that μ-scaffolds supported cells adhesion, colonization, proliferation and biosynthesis in the entire three-dimensional porosity up to 25 days. The second part of this study involved the development of a soft-lithography approach to control the spatial assembly of μ-scaffolds and to create two distinctive μ-scaffolds patterns, namely ordered and disordered. The as obtained patterns were used as substrate for culturing HDFs and 11 HUVECs aiming to develop viable monolayers and bilayers tissue constructs in vitro. The results of this study demonstrated that μ-scaffolds patterning directed cells colonization and biosynthesis and guided the morphology and distribution of newly formed vasculature. All of the findings reported in this work demonstrated the vital role of μ-scaffolds architectural features and assembly on in vitro tissue growth and, pay the way about the possibility to create in silico-designed vasculatures inside modularly engineered biohybrids tissues

Development of three-dimensional, ex vivo optical imaging

The ability to analyse tissue in 3-D at the mesoscopic scale (resolution: 2-50 µm) has proven essential in the study of whole specimens and individual organs. Techniques such as ex vivo magnetic resonance imaging (MRI) and X-ray computed tomography (CT) have been successful in a number of applications. Although MRI has been used to image embryo development and gene expression in 3-D, its resolution is not sufficient to discriminate between the small structures in embryos and individual organs. Furthermore, since neither MRI nor X-ray CT are optical imaging techniques, none of them is able to make use of common staining techniques. 3-D images can be generated with confocal microscopy by focusing a laser beam to a point within the sample and collecting the fluorescent light coming from that specific plane, eliminating therefore out-of-focus light. However, the main drawback of this microscopy technique is the limited depth penetration of light (~1 mm). Tomographic techniques such as optical projection tomography (OPT) and light sheet fluorescence microscopy (also known as single plane illumination microscopy, SPIM) are novel methods that fulfil a requirement for imaging of specimens which are too large for confocal imaging and too small for conventional MRI. To allow sufficient depth penetration, these approaches require specimens to be rendered transparent via a process known as optical clearing, which can be achieved using a number of techniques. The aim of the work presented in this thesis was to develop methods for threedimensional, ex vivo optical imaging. This required, in first instance, sample preparation to clear (render transparent) biological tissue. In this project several optical clearing techniques have been tested in order to find the optimal method per each kind of tissue, focusing on tumour tissue. Indeed, depending on its structure and composition (e.g. amount of lipids or pigments within the tissue) every tissue clears at a different degree. Though there is currently no literature reporting quantification of the degree of optical clearing. Hence a novel, spectroscopic technique for measuring the light attenuation in optically cleared samples is described in this thesis and evaluated on mouse brain. 5 Optical clearing was applied to the study of cancer. The main cancer model investigated in this report is colorectal carcinoma. Fluorescently labelled proteins were used to analyse the vascular network of colorectal xenograft tumours and to prove the effect of vascular disrupting agents on the vascular tumour network. Furthermore, optical clearing and fluorescent compounds were used for ex vivo analysis of perfusion of a human colorectal liver metastasis model

In vitro Tissue Engineering of Liver and Primary Lymphoid Tissues with Inverted Colloidal Crystal Scaffolds for Drug Testing Application.

Effective early stage drug toxicity testing is imperative to minimize failures in the late clinical stages of the drug development process. 2D cell cultures have been dominantly used, but they cannot adequately estimate actual toxic effects of drug molecules due to the limited capability in restoring original cellular behaviors in 3D tissues. As a potential solution to improve the predictive power of in vitro screening procedures, this dissertation explored a new opportunity of in vitro tissue engineering as a part of the drug development process. Besides the biological significance in functional tissue formation, scaffolds should be transparent and support standardized tissue growth. Inverted colloidal crystal (ICC) hydrogel scaffolds having standardized 3D structure and materials as well as retaining a high analytical capability were developed for this purpose. Uniform size spherical pore arrays prepared with cell repulsive polyacrylamide promoted homogenous HepG2 liver tissue spheroid formation, while the transparent hydrogel matrix allowed convenient characterization of cellular processes. The standardized spheroid culture model was successfully applied to the in vitro toxicity testing of CdTe and Au nanoparticles. Significantly reduced toxic effects were observed compared to the conventional 2D culture attributed by tissue-like morphology and cell phenotypic change in the spheroid culture. In addition, ICC scaffolds combined with a LBL surface modification technique served as a platform for engineering primary lymphoid tissue, i.e. bone marrow and thymus. Under dynamic culture condition, hematopoietic stem cells (HSCs) could travel deep into the scaffold via interconnecting channels, while they were temporarily entrapped due to limited channel size and number. As a result, HSCs extensively interacted with stromal cells growing along the LBL coated pore surface. Such intimate cell-cell and cell-matrix interaction is the key process in HSCs survival and differentiation that was substantiated by ex vivo expansion and B-/T-cell differentiation of HSCs. Overall this thesis introduces a promising application of in vitro tissue engineering as a practical and valuable early stage toxicity testing tool. ICC scaffolds exhibited unique advantage in preparation of spheroid culture model and lymphoid tissue engineering. Standardized in vitro tissue models substantiate the capability to extend current cellular level cytotoxicity to the tissue level.Ph.D.Biomedical EngineeringUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/64654/1/jungwoo_1.pd

Application of Nanomaterials in Biomedical Imaging and Cancer Therapy

To mark the recent advances in nanomaterials and nanotechnology in biomedical imaging and cancer therapy, this book, entitled Application of Nanomaterials in Biomedical Imaging and Cancer Therapy includes a collection of important nanomaterial studies on medical imaging and therapy. The book covers recent works on hyperthermia, external beam radiotherapy, MRI-guided radiotherapy, immunotherapy, photothermal therapy, and photodynamic therapy, as well as medical imaging, including high-contrast and deep-tissue imaging, quantum sensing, super-resolution microscopy, and three-dimensional correlative light and electron microscopy. The significant research results and findings explored in this work are expected to help students, researchers and teachers working in the field of nanomaterials and nanotechnology in biomedical physics, to keep pace with the rapid development and the applications of nanomaterials in precise imaging and targeted therapy

Topical Issues of Theoretical and Clinical Medicine

Abstract book of International scientific and practical conference of students, postgraduates and young scientists, Sumy, October 17-19

Novel cell models to study breast tumour microenvironment and disease progression

Breast cancer is the most prevalent and deadly in woman. ER+ breast cancer represents around two-thirds of all cases and has a favourable prognosis due to good response to endocrine therapy. However, these tumours present 25% of disease relapse due to drug resistance and metastatic behaviour. Tumour progression and acquired drug resistance are modulated by the interactions between tumour cells and the surrounding microenvironment. Most models employed to address these mechanisms fail to reflect the complex tumour microenvironment and do not allow long-term monitoring of tumour progression. (...