Bridging the Multiscale Gap: Identifying Cellular Parameters from Multicellular Data (original) (raw)
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2017
Background The configuration of necrotic areas within the retinal pigmented epithelium is an important element in the progression of age-related macular degeneration (AMD). In the exudative (wet) and non-exudative (dry) forms of the disease, retinal pigment epithelial (RPE) cells respond to adjacent atrophied regions by secreting vascular endothelial growth factor (VEGF) that in turn recruits new blood vessels which lead to a further reduction in retinal function and vision. In vitro models exist for studying VEGF expression in wet AMD (Vargis et al., Biomaterials 35(13):3999–4004, 2014), but are limited in the patterns of necrotic and intact RPE epithelium they can produce and in their ability to finely resolve VEGF expression dynamics. Results In this work, an in silico hybrid agent-based model was developed and validated using the results of this cell culture model of VEGF expression in AMD. The computational model was used to extend the cell culture investigation to explore the dynamics of VEGF expression in different sized patches of RPE cells and the role of negative feedback in VEGF expression. Results of the simulation and the cell culture studies were in excellent qualitative agreement, and close quantitative agreement. Conclusions The model indicated that the configuration of necrotic and RPE cell-containing regions have a major impact on VEGF expression dynamics and made precise predictions of VEGF expression dynamics by groups of RPE cells of various sizes and configurations. Coupled with biological studies, this model may give insights into key molecular mechanisms of AMD progression and open routes to more effective treatments.
2015
There have been several techniques developed in recent years to develop computer models of a variety of disease behaviors. Agent-based modeling is a discrete-based modeling approach used agents to represent individual cells that mechanically interact and secrete, consume or react to soluble products. It has become a powerful modeling approach, widely used by computational researchers. In this research, we utilized agent-based modeling to study and explore disease development, particularly in two applications, breast cancer and bioengineering experiments. We further proposed an error-minimization search approach and used it to estimate cellular parameters from multicellular in vitro data. In this dissertation, in the first study, we developed a 2D agent-based model that attempted to emulate the in vivo structure of breast cancer. The model was applied to describe the progression from DCIS into DCI. This model confirms that the interaction between tumor cells and the surrounding stroma in the duct plays a critical role in tumor growth and metastasis. This interaction depends on many mechanical and chemical factors that work with each other to produce tumor invasion of the surrounding tissue. In the second study, an in silico model was developed and applied to understanding the underlying mechanism of vascular-endothelial growth factor (VEGF) auto-regulation in REP and emulate the in vitro experiments as part of bioengineering research. This model may provide a system with robust predictive modeling and visualization that could enable discovery of the molecular mechanisms involved in age-related macular degeneration (AMD) progression and provide routers to the development of effective treatmen
A model microfluidics-based system for the human and mouse retina
Biomedical Microdevices, 2015
The application of microfluidics technologies to the study of retinal function and response holds great promise for development of new and improved treatments for patients with degenerative retinal diseases. Restoration of vision via retinal transplantation therapy has been severely limited by the low numbers of motile cells observed post transplantation. Using modern soft lithographic techniques, we have developed the μRetina, a novel and convenient biomimetic microfluidics device capable of examing the migratory behavior of retinal lineage cells within biomimetic geometries of the human and mouse retina. Coupled computer simulations and experimental validations were used to characterize and confirm the formation of chemical concentration gradients within the μRetina, while real-time images within the device captured radial and theta cell migration in response to concentration gradients of stromal derived factor (SDF-1), a known chemoattractant. Our data underscore how the μRetina can be used to examine the concentrationdependent migration of retinal progenitors in order to enhance current therapies, as well as develop novel migration-targeted treatments.
An integrated approach to quantitative modelling in angiogenesis research
Journal of the Royal Society, Interface / the Royal Society, 2015
Angiogenesis, the process by which new vessels form from existing ones, plays an important role in many developmental processes and pathological conditions. We study angiogenesis in the context of a highly controllable experimental environment: the cornea micropocket assay. Using a multidisciplinary approach that combines experiments, image processing and analysis, and mathematical modelling, we aim to provide mechanistic insight into the action of two angiogenic factors, vascular endothelial growth factor A (VEGF-A) and basic fibroblast growth factor (bFGF). We use image analysis techniques to extract quantitative data, which are both spatially and temporally resolved, from experimental images, and we develop a mathematical model, in which the corneal vasculature evolves in response to both VEGF-A and bFGF. The experimental data are used for model parametrization, while the mathematical model is used to assess the utility of the cornea micropocket assay and to characterize proposed...
Quantitative inference of cellular parameters from microfluidic cell culture systems
Biotechnology and Bioengineering, 2009
Microfluidic cell culture systems offer a convenient way to measure cell biophysical parameters in conditions close to the physiological environment. We demonstrate the application of a mathematical model describing the spatial distribution of nutrient and growth factor concentrations in inferring cellular oxygen uptake rates from experimental measurements. We use experimental measurements of oxygen concentrations in a poly(dimethylsiloxane) (PDMS) microreactor culturing human hepatocellular liver carcinoma cells (HepG2) to infer quantitative information on cellular oxygen uptake rates. We use a novel microchannel design to avoid the parameter correlation problem associated with simultaneous cellular uptake and diffusion of oxygen through the PDMS surface. We find that the cellular uptake of oxygen is dependent on the cell density and can be modeled using a logistic term in the Michaelis-Menten equation. Our results are significant not only for the development of novel assays to quantitatively infer cell response to stimuli, but also for the development, design, and optimization of novel in vitro systems for drug discovery and tissue engineering.
Multiscale Angiogenesis Modeling
Lecture Notes in Computer Science, 2005
We propose a deterministic two-scale tissue-cellular approach for modeling growth factor-induced angiogenesis. The bioreaction and diffusion of capillary growth factors (CGF) are modeled at a tissue scale, whereas capillary extension, branching and anastomosis are modeled at a cellular scale. The capillary indicator function is used to bridge these two scales. To solve the equation system numerically, we construct a twogrid algorithm that involves applying a mixed finite element method to approximate concentrations of CGF on a coarse mesh and a point-topoint tracking method to simulate sprout branching and anastomosis on a fine grid. An analysis of the algorithm establishes optimal error bounds for each of the processes-CGF reaction-diffusion, capillary extension, sprout branching and anastomosis-and overall error bounds for their coupled nonlinear interactions.
Uncovering the behaviors of individual cells within a multicellular microvascular community
Proceedings of the National Academy of Sciences, 2011
Although individual cells vary in behavior during the formation of tissues, the nature of such variations are largely uncharacterized. Here, we tracked the morphologies and motilities of ∼300 human endothelial cells from an initial dispersed state to the formation of capillary-like structures, distilling the dynamics of tissue morphogenesis into an array of ∼36,000 numerical phenotypes. Quantitative analysis of population averages revealed two previously unidentified phases in which the cells spread before forming connections with neighboring cells and where the microvascular plexus stabilized before spatially reorganizing. Analysis at the single-cell level showed that in contrast to the population-averaged behavior, most cells followed distinct temporal patterns that were not reflected in the bulk average. Interestingly, some of these behavioral patterns correlated to the cells' final structural role within the plexus. Knowledge of how individual cells or groups of cells behave enhances our understanding of how native tissues self-organize and could ultimately enable more precise approaches for engineering tissues and synthesizing multicellular communities.
Microfluidics-based systems biology
Molecular BioSystems, 2006
Systems biology seeks to develop a complete understanding of cellular mechanisms by studying the functions of intra-and inter-cellular molecular interactions that trigger and coordinate cellular events. However, the complexity of biological systems causes accurate and precise systems biology experimentation to be a difficult task. Most biological experimentation focuses on highly detailed investigation of a single signaling mechanism, which lacks the throughput necessary to reconstruct the entirety of the biological system, while high-throughput testing often lacks the fidelity and detail necessary to fully comprehend the mechanisms of signal propagation. Systems biology experimentation, however, can benefit greatly from the progress in the development of microfluidic devices. Microfluidics provides the opportunity to study cells effectively on both a single-and multi-cellular level with high-resolution and localized application of experimental conditions with biomimetic physiological conditions. Additionally, the ability to massively array devices on a chip opens the door for high-throughput, high fidelity experimentation to aid in accurate and precise unraveling of the intertwined signaling systems that compose the inner workings of the cell.
AJP: Heart and Circulatory Physiology, 2013
Developing therapies aimed at manipulating microvascular remodeling requires a better understanding of angiogenesis and how angiogenesis relates to other network remodeling processes, such as lymphangiogenesis and neurogenesis. The objective of this study was to develop an angiogenesis model that enables probing of multicellular and multisystem interactions at the molecular level across an intact adult microvascular network. Adult male Wistar rat mesenteric windows were aseptically harvested and cultured in serum-free minimum essential media. Viability/cytotoxicity analysis revealed that cells remain alive for at least 7 days. Immunohistochemical labeling at 3 days for platelet endothelial cell adhesion molecule (PECAM), neuron-glial antigen 2 (NG2), lymphatic vessel endothelial hyaluronan receptor-1 (LYVE-1), and class III β-tubulin identified endothelial cells, pericytes, lymphatics, and nerves, respectively. Media supplemented with bFGF or VEGF induced an increase in endothelial ...
PLoS ONE, 2013
A growing number of studies are evaluating retinal progenitor cell (RPC) transplantation as an approach to repair retinal degeneration and restore visual function. To advance cell-replacement strategies for a practical retinal therapy, it is important to define the molecular and biochemical mechanisms guiding RPC motility. We have analyzed RPC expression of the epidermal growth factor receptor (EGFR) and evaluated whether exposure to epidermal growth factor (EGF) can coordinate motogenic activity in vitro. Using Boyden chamber analysis as an initial highthroughput screen, we determined that RPC motility was optimally stimulated by EGF concentrations in the range of 20-400ng/ml, with decreased stimulation at higher concentrations, suggesting concentration-dependence of EGFinduced motility. Using bioinformatics analysis of the EGF ligand in a retina-specific gene network pathway, we predicted a chemotactic function for EGF involving the MAPK and JAK-STAT intracellular signaling pathways. Based on targeted inhibition studies, we show that ligand binding, phosphorylation of EGFR and activation of the intracellular STAT3 and PI3kinase signaling pathways are necessary to drive RPC motility. Using engineered microfluidic devices to generate quantifiable steady-state gradients of EGF coupled with live-cell tracking, we analyzed the dynamics of individual RPC motility. Microfluidic analysis, including center of mass and maximum accumulated distance, revealed that EGF induced motility is chemokinetic with optimal activity observed in response to low concentration gradients. Our combined results show that EGFR expressing RPCs exhibit enhanced chemokinetic motility in the presence of low nanomole levels of EGF. These findings may serve to inform further studies evaluating the extent to which EGFR activity, in response to endogenous ligand, drives motility and migration of RPCs in retinal transplantation paradigms. Citation: Unachukwu UJ, Sauane M, Vazquez M, Redenti S (2013) Microfluidic Generated EGF-Gradients Induce Chemokinesis of Transplantable Retinal Progenitor Cells via the JAK/STAT and PI3Kinase Signaling Pathways. PLoS ONE 8(12): e83906.