In our lab we study the development of the definitive endoderm, one of the 3 primary
germ layers that arises during gastrulation. The definitive endoderm produces the entire
gastrointestinal tract as well as accessory digestive and respiratory organs. These
organs include lung, liver, pancreas, thyroid, parathyroid and thymus. Although much is
known about the genes involved in the function of these organs in the adult, relatively
little is known about how these tissues are initially patterned and organized. An overall
goal of the lab is to understand the morphological and molecular mechanisms that give
rise to endodermal organs, focusing on liver and pancreas.
One promising research area is the creation of differentiated cell types from embryonic
stem (ES) cells. In particular, many labs are attempting to create hepatocytes (early
liver cells) and pancreatic beta cells, the insulin producing cells that are lacking in
type I diabetes patients. This task has proven extremely difficult. Recent breakthroughs
towards these goals were made possible through an understanding of the molecules and
interactions required to generate endoderm and endodermal organs during early mouse
development. The work in our lab is focused on furthering our understanding of normal
liver and pancreas formation with the goal that advancements made here will translate
into advances in ES differentiation and curing the diseased organ. Towards this end, we
are taking two broad approaches to study the early stages of endoderm organogenesis in
the mouse. They can be defined as embryological approaches, utilizing whole embryo
culture, and genetic approaches, using transgenic and homologous recombination to create
favorable environments to study this fascinating tissue layer.
The endoderm is a uniform epithelial layer that covers the ventral surface of the
pre-somitic mouse embryo. From 8-15 somites (S), the accessory organs appear, in a
temporally and spatially characteristic manner, as thickenings in this epithelium
After thickening, a bud appears that produces the endodermal component of each the mature
organ. Organ-specific gene expression follows or is coincident with the onset of these
morphological processes and as a result, little is known about the location of these
organ progenitors in the endodermal sheet, the morphological processes leading to
organogenesis, or the molecular mechanisms that initiate organogenesis. Because of the
lack of genes or promoters expressed specifically in the endoderm, more traditional
approaches, such as knock-out and transgenic experiments, have had limited success in
tackling these issues. We have decided to use a whole embryo culture system in which we
can successfully culture whole embryos, and their accompanying extraembryonic tissues,
from early somite stages (~day 8.25) until day 10 (Fig. 2). Culturing allows us to
manipulate the pre-specified (d8.25) endoderm and follow these changes through d10, when
the liver and pancreas buds have formed and begun to differentiate. We have used viable
dyes to identify precursor populations in the early somite embryo and are now using
electroporation to identify genes that disrupt early organogenesis.
Little is known about the growth or developmental potential of early individual
endodermal cells. Is an individual endodermal cell committed to a specific organ or is it
capable of giving rise to cells contributing to multiple organs? Similarly, it is unknown
whether individual cells expressing organ-specific markers are capable of giving rise to
all differentiated cell-types within the organ or are excluded from certain lineages. To
answer these questions we perform experiments with transgenic or knock-in mice that
produce embryos that have had individual endoderm cells marked with either the
fluorescent markers EGFP or the histological marker LacZ. By retrospectively analyzing
the clonal descendants of these cells, we will elucidate the normal processes that give
rise to endodermal organ, further define organ precursors and understand the
morphological processes that produce the mature organs.
