Modeling Morphogenesis in silico and in vitro: Towards Quantitative, Predictive, Cell-based Modeling

Cell-based, mathematical models help make sense of morphogenesis—i.e. cells organizing into shape and pattern—by capturing cell behavior in simple, purely descriptive models. Cell-based models then predict the tissue-level patterns the cells produce collectively. The first step in a cell-based modeling approach is to isolate sub-processes, e.g. the patterning capabilities of one or a few cell types in cell cultures. Cell-based models can then identify the mechanisms responsible for patterning in vitro. This review discusses two cell culture models of morphogenesis that have been studied using this combined experimental-mathematical approach: chondrogenesis (cartilage patterning) and vasculogenesis (de novo blood vessel growth). In both these systems, radically dif- ferent models can equally plausibly explain the in vitro patterns. Quantitative descriptions of cell behavior would help choose between alternative models. We will briefly review the experimental methodology (microfluidics technology and traction force microscopy) used to measure responses of individual cells to their micro-environment, including chemical gradients, physical forces and neighboring cells. We conclude by discussing how to include quantitative cell descriptions into a cell-based model: the Cellular Potts model

Computational modeling of angiogenesis: towards a multi-scale understanding of cell-cell and cell-matrix interactions

Combined with in vitro and in vivo experiments, mathematical and com- putational modeling are key to unraveling how mechanical and chemical signaling by endothelial cells coordinates their organization into capillary-like tubes. While in vitro and in vivo experiments can unveil the effects of for example environmental changes or gene knockouts, computational models provide a way to formalize and understand the mechanisms underlying these observations. This chapter reviews re- cent computational approaches to model angiogenesis, and discusses the insights they provide in the mechanisms of angiogenesis. We introduce a new cell-based computational model of an in vitro assay of angio- genic sprouting from endothelial monolayers in fibrin matrices. Endothelial cells are modeled by the Cellular Potts Model, combined with continuum descriptions to model haptotaxis and proteolysis of the extracellular matrix. The computational model demonstrates how a variety of cellular structural properties and behaviors determine the dynamics of tube formation. We aim to extend this model to a multi-scale model in the sense that cells, extracellular matrix and cell-regulation are de- scribed at different levels of detail and feedback on each other. Finally we discuss how computational modeling, combined with in vitro and in vivo modeling steers experiments, and how it generates new experimental hypotheses and insights on the mechanics of angiogenesis

Single and combined effects of αvβ3- and α5β1-integrins on capillary tube formation in a human fibrinous matrix

The fibrinous exudate of a wound or tumor stroma facilitates angiogenesis. We studied the involvement of RGD-binding integrins during tube formation in human plasma-derived fibrin clots and human purified fibrin matrices. Capillary-like tube formation by human microvascular endothelial cells in a 3D plasma-derived fibrinous matrix was induced by FGF-2 and TNF-α and depended largely on cell-bound u-PA and plasmin activities. While tube formation was minimally affected by the addition of either the αvβ3-integrin inhibiting mAb LM609 or the α5-integrin inhibiting mAb IIA1, the general RGD-antagonist echistatin completely inhibited this process. Remarkably, when αvβ3- and α5β1-integrins were inhibited simultaneously, tube formation was reduced by 78%. It was accompanied by a 44% reduction of u-PA antigen accumulation and 41% less production of fibrin degradation products. αvβ5-integrin-blocking antibodies further enhanced the inhibition by mAb LM609 and mAb IIA1 to 94%, but had no effect by themselves. αv-specific cRGD only inhibited angiogenesis when α5β1-integrin was simultaneously blocked. Endostatin mimicked the effect of α5β1-integrin and inhibited tube formation only in the presence of LM609 or cRGD (73 and 80%, respectively). Comparable results were obtained when purified fibrin matrices were used instead of the plasma-derived fibrinous matrices. These data show that blocking of tube formation in a fibrinous exudate requires the simultaneous inhibition of αvβ3- and α5β1-integrins. This may bear impact on attempts to influence angiogenesis in a fibrinous environment

A local uPAR-plasmin-TGFβ1 positive feedback loop in a qualitative computational model of angiogenic sprouting explains the in vitro effect of fibrinogen variants

In experimental assays of angiogenesis in three-dimensional fibrin matrices, a temporary scaffold formed during wound healing, the type and composition of fibrin impacts the level of sprouting. More sprouts form on high molecular weight (HMW) than on low molecular weight (LMW) fibrin. It is unclear what mechanisms regulate the number and the positions of the vascular-like structures in cell cultures. To address this question, we propose a mechanistic simulation model of endothelial cell migration and fibrin proteolysis by the plasmin system. The model is a hybrid, cell-based and continuum, computational model based on the cellular Potts model and sets of partial-differential equations. Based on the model results, we propose that a positive feedback mechanism between uPAR, plasmin and transforming growth factor β1 (TGFβ1) selects cells in the monolayer for matrix invasion. Invading cells releases TGFβ1 from the extracellular matrix through plasmin-mediated fibrin degradation. The activated TGFβ1 further stimulates fibrin degradation and keeps proteolysis active as the sprout invades the fibrin matrix. The binding capacity for TGFβ1 of LMW is reduced relative to that of HMW. This leads to reduced activation of proteolysis and, consequently, reduced cell ingrowth in LMW fibrin compared to HMW fibrin. Thus our model predicts that endothelial cells in LMW fibrin matrices compared to HMW matrices show reduced sprouting due to a lower bio-availability of TGFβ1

Hypoxia Impairs Initial Outgrowth of Endothelial Colony Forming Cells and Reduces Their Proliferative and Sprouting Potential

Vascular homeostasis and regeneration in ischemic tissue relies on intrinsic competence of the tissue to rapidly recruit endothelial cells for vascularization. The mononuclear cell (MNC) fraction of blood contains circulating progenitors committed to endothelial lineage. These progenitors give rise to endothelial colony-forming cells (ECFCs) that actively participate in neovascularization of ischemic tissue. To evaluate if the initial clonal outgrowth of ECFCs from cord (CB) and peripheral blood (PB) was stimulated by hypoxic conditions, MNCs obtained from CB and PB were subjected to 20 and 1% O2 cell culture conditions. Clonal outgrowth was followed during a 30 day incubation period. Hypoxia impaired the initial outgrowth of ECFC colonies from CB and also reduced their number that were developing from PB MNCs. Three days of oxygenation (20% O2) prior to hypoxia could overcome the initial CB-ECFC outgrowth. Once proliferating and subcultured the CB-ECFCs growth was only modestly affected by hypoxia; proliferation of PB-ECFCs was reduced to a similar extent (18–30% reduction). Early passages of subcultured CB- and PB-ECFCs contained only viable cells and few if any senescent cells. Tube formation by subcultured PB-ECFCs was also markedly inhibited by continuous exposure to 1% O2. Gene expression profiles point to regulation of the cell cycle and metabolism as major altered gene clusters. Finally we discuss our counterintuitive observations in the context of the important role that hypoxia has in promoting neovascularization

Consensus guidelines for the use and interpretation of angiogenesis assays

The formation of new blood vessels, or angiogenesis, is a complex process that plays important roles in growth and development, tissue and organ regeneration, as well as numerous pathological conditions. Angiogenesis undergoes multiple discrete steps that can be individually evaluated and quantified by a large number of bioassays. These independent assessments hold advantages but also have limitations. This article describes in vivo, ex vivo, and in vitro bioassays that are available for the evaluation of angiogenesis and highlights critical aspects that are relevant for their execution and proper interpretation. As such, this collaborative work is the first edition of consensus guidelines on angiogenesis bioassays to serve for current and future reference

Samenwerkende Cellen : Hoe?Zo! Radio, 14.03.2011 (NTR) [23:50]

Je lichaam bestaat uit zo’n 50.000 miljard cellen die allemaal nauwkeurig met elkaar samenwerken. Maar hoe doen ze dat? Dat is precies de vraag waar systeembioloog Roeland Merks zich mee bezig houdt. Samen met experimenteel bioloog Pieter Koolwijk onderzoekt hij hoe complexe ontwikkelingen, zoals bloedvatgroei bij tumoren, tot stand komen