On Multiscale Approaches to 3-Dimensional Modeling of Morphogenesis

In this paper we present the foundation of a unified, object-oriented, three-dimensional (3D) biomodeling environment, which allows us to integrate multiple submodels at scales from subcellular to tissues and organs. Our current implementation combines a modified discrete model from statistical mechanics, the Cellular Potts Model (CPM), with a continuum reaction-diffusion (RD) model and a state automaton with well-defined conditions for cell differentiation transitions to model genetic regulation. This environment allows us to rapidly and compactly create computational models of a class of complex developmental phenomena. To illustrate model development, we simulate a simplified version of the formation of the skeletal pattern in a growing embryonic vertebrate limb

Conserved but flexible modularity in the zebrafish skull: implications for craniofacial evolvability

Morphological variation is the outward manifestation of development and provides fodder for adaptive evolution. Because of this contingency, evolution is often thought to be biased by developmental processes and functional interactions among structures, which are statistically detectable through forms of covariance among traits. This can take the form of substructures of integrated traits, termed modules, which together comprise patterns of variational modularity. While modularity is essential to an understanding of evolutionary potential, biologists currently have little understanding of its genetic basis and its temporal dynamics over generations. To address these open questions, we compared patterns of craniofacial modularity among laboratory strains, defined mutant lines and a wild population of zebrafish ( ). Our findings suggest that relatively simple genetic changes can have profound effects on covariance, without greatly affecting craniofacial shape. Moreover, we show that instead of completely deconstructing the covariance structure among sets of traits, mutations cause shifts among seemingly latent patterns of modularity suggesting that the skull may be predisposed towards a limited number of phenotypes. This new insight may serve to greatly increase the evolvability of a population by providing a range of 'preset' patterns of modularity that can appear readily and allow for rapid evolution

Factors Regulating Chondrogenic Differentiation

Chondrogenesis is a co-ordinated differentiation process in which mesenchymal cells condensate, differentiate into chondrocytes and begin to secrete molecules that form the extracellular matrix. It is regulated in a spatio-temporal manner by cellular interactions and growth and differentiation factors that modulate cellular signalling pathways and transcription of specific genes. Moreover, post-transcriptional regulation by microRNAs (miRNAs) has appeared to play a central role in diverse biological processes, but their role in skeletal development is not fully understood. Mesenchymal stromal cells (MSCs) are multipotent cells present in a variety of adult tissues, including bone marrow and adipose tissue. They can be isolated, expanded and, under defined conditions, induced to differentiate into multiple cell lineages including chondrocytes, osteoblasts and adipocytes in vitro and in vivo. Owing to their intrinsic capability to self-renew and differentiate into functional cell types, MSCs provide a promising source for cell-based therapeutic strategies for various degenerative diseases, such as osteoarthritis (OA). Due to the potential therapeutic applications, it is of importance to better understand the MSC biology and the regulatory mechanisms of their differentiation. In this study, an in vitro assay for chondrogenic differentiation of mouse MSCs (mMSCs) was developed for the screening of various factors for their chondrogenic potential. Conditions were optimized for pellet cultures by inducing mMSC with different bone morphogenetic proteins (BMPs) that were selected based on their known chondrogenic relevance. Characterization of the surface epitope profile, differentiation capacity and molecular signature of mMSCs illustrated the importance of cell population composition and the interaction between different populations in the cell fate determination and differentiation of MSCs. Regulation of Wnt signalling activity by Wnt antagonist sFRP-1 was elucidated as a potential modulator of lineage commitment. Delta-like 1 (dlk1), a factor regulating adipogenesis and osteogenesis, was shown to exhibit stage-specific expression during embryonic chondrogenesis and identified as a novel regulator of chondrogenesis, possibly through mediating the effect of TGF-beta1. Moreover, miRNA profiling demonstrated that MSCs differentiating into a certain lineage exhibit a specific miRNA expression profile. The complex regulatory network between miRNAs and transcription factors is suggested to play a crucial role in fine-tuning the differentiation of MSCs. These results demonstrate that commitment of mesenchymal stromal cells and further differentiation into specific lineages is regulated by interactions between MSCs, various growth and transcription factors, and miRNA-mediated translational repression of lineage-specific genes.Siirretty Doriast

Simulation of the spatial structure and cellular organization evolution of cell aggregates arranged in various simple geometries, using a kinetic monte carlo method applied to a lattice model

ilustraciones, graficasEsta tesis trata los modelos de morfogénesis, en particular los modelos de evolución guiada por contacto que son coherentes con la hipótesis de la adhesión diferencial. Se presenta una revisión de algunos modelos, sus principios biológicos subyacentes, la relevancia y aplicaciones en el marco de la bioimpresión, la ingeniería de tejidos y la bioconvergencia. Luego, se presentan los detalles de los modelos basados en métodos de Monte Carlo para profundizar más adelante en el modelo basados en algoritmos Kinetic Monte Carlo (KMC) , más específicamente, se describe en detalle un modelo KMC de autoaprendizaje (SL-KMC). Se presenta y explica la estructura algorítmica del código implementado, se evalúa el rendimiento del modelo y se compara con un modelo KMC tradicional. Finalmente, se realizan los procesos de calibración y validación, se observó que el modelo es capaz de replicar la evolución del sistema multicelular cuando las condiciones de energía interfacial del sistema simulado son similares a las del sistema de calibraciones. (Texto tomado de la fuente)This thesis treats the models for morphogenesis, in particular the contact-guided evolution models that are coherent with the differential adhesion hypothesis. A review of some models, their biological underpinning principles, the relevance and applications in the framework of bioprinting, tissue engineering and bioconvergence are presented. Then the details for the Monte Carlo methods-based models are presented to later deep dive into the Kinetic Monte Carlo (KMC) based model, and more specifically a Self-Learning KMC (SL-KMC) model is described to detail. The algorithmic structure of the implemented code is presented and explained, the model performance is assessed and compared with a traditional KMC model. Finally, the calibration and validation processes have been carried out, it was observed that the model is able to replicate the multicellular system evolution when the interfacial energy conditions of the simulated system are similar to those of the calibrations system.MaestríaMagíster en Ingeniería - Ingeniería Químic

Multiscale developments of cellular Potts models

Multiscale problems are ubiquitous and fundamental in all biological phenomena that emerge naturally from the complex interaction of processes which occur at various levels. A number of both discrete and continuous mathematical models and methods have been developed to address such an intricate network of organization. One of the most suitable individual cell-based model for this purpose is the well-known cellular Potts model (CPM). The CPM is a discrete, lattice-based, flexible technique that is able to accurately identify and describe the phenomenological mechanisms which are responsible for innumerable biological (and nonbiological) phenomena. In this work, we first give a brief overview of its biophysical basis and discuss its main limitations. We then propose some innovative extensions, focusing on ways of integrating the basic mesoscopic CPM with accurate continuous models of microscopic dynamics of individuals. The aim is to create a multiscale hybrid framework that is able to deal with the typical multilevel organization of biological development, where the behavior of the simulated individuals is realistically driven by their internal state. Our CPM extensions are then tested with sample applications that show a qualitative and quantitative agreement with experimental data. Finally, we conclude by discussing further possible developments of the metho

A Cellular Potts Model of single cell migration in presence of durotaxis

Cell migration is a fundamental biological phenomenon during which cells sense their surroundings and respond to different types of signals. In presence of durotaxis, cells preferentially crawl from soft to stiff substrates by reorganizing their cytoskeleton from an isotropic to an anisotropic distribution of actin filaments. In the present paper, we propose a Cellular Potts Model to simulate single cell migration over flat substrates with variable stiffness. We have tested five configurations: (i) a substrate including a soft and a stiff region, (ii) a soft substrate including two parallel stiff stripes, (iii) a substrate made of successive stripes with increasing stiffness to create a gradient and (iv) a stiff substrate with four embedded soft squares. For each simulation, we have evaluated the morphology of the cell, the distance covered, the spreading area and the migration speed. We have then compared the numerical results to specific experimental observations showing a consistent agreement

Biofidelic simulations of embryonic joint growth and morphogenesis

During skeletal development, the opposing surfaces in the joint mould into interlocking and reciprocal shapes in a process called morphogenesis. Morphogenesis is critical to the health and function of the joint, and yet, little is known about the process of joint morphogenesis. For example, how do different joints acquire their specific shapes? Which cellular processes underlie joint shaping and how are they regulated? However, it is known that fetal movements are critical to joint development, with alterations or absences of movement being implicated in multiple pre- and post-natal musculoskeletal conditions. This doctorate explored the cell-level dynamics governing joint growth and the implication of movements in regulating them, using novel biofidelic and mechanobiological models of joint growth. Cell-level data from wild type zebrafish larvae were tracked and synthesised in a biofidelic simulation of zebrafish jaw joint growth. Growth characteristics were quantified revealing a strong anisotropy (Chapter 3). Next, zebrafish larvae were immobilised using drug treatment. The material properties of the zebrafish jaw cartilage were measured using nano-indentation in the presence or absence of movement showing a delay in cartilage stiffening in immobilised larvae (Chapter 4). Then, I developed a novel mechanobiological model of zebrafish jaw joint growth, which identified a correlation between growth characteristics and the dynamic patterns of mechanical stimuli experienced by joint elements over jaw motion (Chapter 5). Finally, local growth rates were characterised in the mouse elbow in the presence or absence of skeletal muscles. Spatial heterogeneity in the growth rates correlated with the emergence of specific shape features at the level of the condyles. Immobilisation led to disruption of the local growth rates correlated with failed shape differentiation of the condyles. The relative contribution of key cell activities to growth such as cell volume expansion, cell number increases and extracellular matrix expansion, were shown to vary over time in both wild types and muscleless-limbs and to be altered in the absence of skeletal muscles (Chapter 6). This research offers avenues for improvement in simulations of joint development and potentially other organs. It provides fundamental advance in our understanding of mechanoregulation in the developing joint and increases our understanding of the origins of musculoskeletal abnormalities.Open Acces

Transcriptional regulation of the Ccn2 gene

Cellular Communication Network Factor 2 (CCN2) is a matricellular protein which functions in many tissues, and is most notably expressed by chondrocytes, with knockout of the Ccn2 gene expression causing severe chondrodysplasia in mice. Regulation at the prerequisite stage of transcription by non-coding genomic elements is fundamental in the expression of every gene. Until now, the capacity for cis-acting regulatory modules to control the expression of Ccn2 has been obscure. The current project sought to delineate the capacity for cis-acting enhancer regions to regulate Ccn2 transcription within highly specific temporospatial contexts. A 300 kilobase intergenic genomic region upstream of Ccn2 was examined in silico in order to identify putative enhancer regions based on chromatin characteristics of enhancers; histone modification, DNase I hypersensitivity and interspecies evolutionary conservation of DNA sequence. Transgenic mice were created using constructs consisting of each putative enhancer region driving the expression of a LacZ reporter gene in conjunction with a silent Hsp68 minimal promoter. Expression of the protein product of LacZ, β-galactosidase was assayed at several developmental time-points in order to determine whether candidate enhancers were able to control gene transcription. Five enhancer regions of murine Ccn2 expression, located -230kb, -198kb, -148kb, -137kb and -102kb upstream of the gene each drove LacZ expression in a tissue-specific manner at embryonic day E15.5. -230kb drove transgene activity within osteoblasts, whereas -198kb, -148kb and -137kb all exhibited function within chondrocytes. The -102kb enhancer was active within the superficial vasculature. More comprehensive examination of the - 148kb enhancer revealed function in adulthood, and that a truncated region of this sequence is also capable of enhancing gene transcriptional output. Enhancer sequences were also examined in vitro using Electrophoretic Mobility Shift Assay in order to test the capacity of transcription factors of interest to bind to enhancers and modulate function. Master regulator of chondrocyte physiology, the transcription factor SOX9 was found to bind to sequences within the -137kb, -148kb and -230kb enhancers. The findings described herein are the first to characterise the role of cis-acting enhancer regions in the transcription of Ccn2 within the murine genome. Understanding the mechanisms that underpin temporospatial control of Ccn2 expression will inform both the characterisation and amelioration of profound pathological conditions that result from loss of CCN2 transcriptional regulation such as osteoarthritis, fibrosis and cancer in humans

A Cellular Potts Model of single cell migration in presence of durotaxis

Cell migration is a fundamental biological phenomenon during which cells sense their surroundings and respond to different types of signals. In presence of durotaxis, cells preferentially crawl from soft to stiffsubstrates by reorganizing their cytoskeleton from an isotropic to an anisotropic distribution of actin fil- aments. In the present paper, we propose a Cellular Potts Model to simulate single cell migration over flat substrates with variable stiffness. We have tested five configurations: (i) a substrate including a soft and a stiffregion, (ii) a soft substrate including two parallel stiffstripes, (iii) a substrate made of succes- sive stripes with increasing stiffness to create a gradient and (iv) a stiffsubstrate with four embedded soft squares. For each simulation, we have evaluated the morphology of the cell, the distance covered, the spreading area and the migration speed. We have then compared the numerical results to specific experimental observations showing a consistent agreement

A node-based version of the cellular Potts model

The cellular Potts model (CPM) is a lattice-based Monte Carlo method that uses an energetic formalism to describe the phenomenological mechanisms underlying the biophysical problem of interest. We here propose a CPM-derived framework that relies on a node-based representation of cell-scale elements. This feature has relevant consequences on the overall simulation environment. First, our model can be implemented on any given domain, provided a proper discretization (which can be regular or irregular, fixed or time evolving). Then, it allowed an explicit representation of cell membranes, whose displacements realistically result in cell movement. Finally, our node-based approach can be easily interfaced with continuous mechanics or fluid dynamics models. The proposed computational environment is here applied to some simple biological phenomena, such as cell sorting and chemotactic migration, also in order to achieve an analysis of the performance of the underlying algorithm. This work is finally equipped with a critical comparison between the advantages and disadvantages of our model with respect to the traditional CPM and to some similar vertex-based approaches