Big picture: human embryonic stem cells as a model for the embryo
How an amorphous lump of identical cells is able to develop into a human being is a profound question of equal intellectual and practical importance. Its fascination springs in part from the desire to know where we come from, and how we differ from other animals. Yet the ability to direct differentiation of cultured cells into distinct cell types for therapeutic use, perhaps even coax them to form complete organs, relies on answering the same question. Ethical considerations make experimentation on human embryos unacceptable and on other mammals undesirable. Moreover, significant interspecies differences complicate inference about human development from animal models. This challenges us to find answers in alternative ways.
Remarkably, human embryonic stem cells (hESC) retain the embryo’s innate capacity to self-organize. Upon treatment with the differentiation-inducing molecule BMP4, hESCs confined by micropatterning technology to a disc shaped like the early embryo will mimic gastrulation, the first step in embryonic development when pluripotent (i.e. uncommitted) cells commit to one of three different lineages -ectoderm, mesoderm or endoderm- that organize into three germ layers. Although the resultant arrangement of the lineages differs from the embryo, reproducibly forming concentric rings rather than layers, there is substantial evidence that the same genetic circuitry is orchestrating this pattern. Different is also not necessarily worse. The behavior of cells in different conditions provides valuable information, and by systematically varying the environment we can probe mechanisms of self-organization with a level of precision unthinkable in living embryos.
Read more about stem cells as a model for the embryo in an article by Aryeh Warmflash and me.
Interpretation of morphogen dynamics
The paradigm for embryonic patterning is the French flag model, according to which a gradient of a substance called a morphogen, determines cell fates in a concentration-dependent manner. This model is believed to describe the mammalian embryo, but the evidence is limited, and the model is incomplete in that it does not address the effect of changing concentrations. Theoretical consideration shows that response to e.g. change in concentration rather than concentration itself can have a dramatic effect on the way pattern formation takes place. Nodal is the quintessential signaling factor shaping the mammalian embryo, without which gastrulation is completely disrupted, and is believed to act as a textbook morphogen. Experimentally, we have found contrasting signaling response to change in ligand concentration for Nodal and the related signaling molecule BMP4. We are now uncovering how this affects transcription and cell fate decisions.
Effect of tissue topology on patterning
Pattern formation depends on the geometry and the topology of the tissue, and varying these -something that could never be done in real embryos- provides valuable information. Theoretically, the simplest mechanisms for spatial differentiation and morphogen gradient formation depend on the existence of a colony border. However, by growing cells on spheres we found that patterning still occurs. This suggests that the underlying mechanism is the formation of a spontaneous morphogen gradient through an activator-inhibitor system, suggested as the mechanism for gastrulation long ago by the mathematician Alan Turing. Visualizing endogenous gradients will allow us to conclusively show this (see next paragraph). Simultaneously we are working to develop hESCs on curved surfaces of different topology as a more general experimental platform.
Morphogen gradient formation in vitro
Not only is it poorly understood how cells in the mammalian embryo respond to morphogens, it is also not clear if and how morphogen gradients form in space and time, and therefore what the cells are actually exposed to. Indirect evidence suggests that differentiation in micropatterned colonies depends on self-organized morphogen gradients, in particular of Nodal (see interpretation of morphogens above). We are working on visualizing endogenous Nodal in these colonies to determine if a gradient is present and if so how it forms. Recently it has been shown that Wnt does not signal long range as was previously thought, and other signaling molecules may have similar surprises in store.
Role of tissue mechanics in germ layer specification
In recent years there has been a surge of work on cellular and tissue mechanics, and an important role for mechanics in growth regulation and differentiation has been demonstrated in a number of systems. Little is known about the role of mechanics in gastrulation, but there is good reason to believe that it is significant. The primitive streak is the embryonic region where mesoderm and endoderm form and ingress, and is likely to have distinct mechanical properties. By combining our stem cell model for gastrulation with systematic manipulation of substrate and intracellular forces, we can understand the effect of mechanics on germ layer specification and spatial organization of hESCs.