Testing a Developmental Mechanism by an Integrated Empirical-Computational Approach

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Testing a Developmental Mechanism by an Integrated Empirical-Computational ApproachStuart A. Newman, Ph.D., New York Medical CollegeMark Alber, Ph.D., University of Notre DamePROJECT SUMMARYThe objective of this project is to define and test an activator-inhibitor based mechanism for skeletal pattern formation in vertebrate limb mesenchymal cells. The strategy involves working with three distinct but interrelated "models" for this process: (i) a readily manipulable in vitro experimental model utilizing high-density ('micromass") cultures of avian limb mesenchymal cells; (ii) a conceptual developmental model that focuses on experimentally confirmed gene expression-growth factor-cell behavioral interactions common to the in vivo and in vitro patterning process; and (iii) a discrete cellular automaton (CA)-based computational model that has been shown to capture, in a semi-quantitative sense, the major features of the in vitro model, under assumptions that derive from the developmental model. The key interactions specified by thecomputational model ultimately can be integrated into a multiscale, continuous representation of limb development in vivo. Before this is possible, however, it is necessary to determine whether these interactions uniquely simulate chondrogenic pattern formation in vitro, and whether introduction of additional, experimentally-confirmed molecular interactions and geometrical considerations increases or decreases the fidelity of the model to the in vitro results.The computational model is an "agent-oriented" model that represents cells by points on a lattice which obey rules motivated by experimental findings. The "cells" follow these rules as autonomous agents, interacting with other cells and with microenvironments produced by cell activities. The rules include random cell motion, production and lateral deposition of a substrate adhesion molecule corresponding to fibronectin, production and release of a diffusible activator, corresponding to TGF-B that stimulates production of the SAM, and another diffusible factor ("inhibitor") that suppresses the activity of the activator. The cellular automaton is modeled on a 2- dimensional square lattice to emulate the quasi-2D micromass culture.The Newman laboratory will misexpress and inhibit expression of TGF-B , fibronectin, FGF2 and 8 (elicitors of lateral inhibition) and candidate inhibitory molecules in vitro and compare pattern results with corresponding manipulations of the CA model; Dr. Alber and his associates will refine the CA model so as to provide cells with more realistic shapes, allow them to move and accumulate in a third dimension, cause them to exhibit differential adhesion, and will develop a set of quantitative methods for pattern analysis, applicable to both micromass cultures and simulations, so as to facilitate detailed comparison between in vitro and in silico results.Broader impact: The project can be expected to yield several benefits that extend beyond the particular scientific problem addressed. The cellular automata approach is a general tool that can provide developmental biologists working in the area of pattern formation with a way of designing experiments, making predictions, and testing hypotheses. The cross-disciplinary nature of the project will provide a prototype for collaborative efforts by experimental biologists and mathematical, physical, and computational scientists. The work will also have educational value by providing interdisciplinary research experience to graduate students in experimental biology and applied mathematics, who will all spend time in the research groups of the alternate field.