|Kimberly D. Tremblay
Assistant Professor of Veterinary and Animal Science, University of Massachusetts
Ph.D.: University of Pennsylvania
Mouse Development and Organogenesis
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.
Identification and Manipulation of Liver and Pancreas Precursors
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. 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.
Clonal Analysis of Endoderm and Endodermal Organ Growth and Development
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.
Calmont, A. Wandzioch, E., Tremblay, K. D . Minowada, G., Martin, G. R., and Zaret, K. (2006). An FGF-response pathway that mediates hepatic gene induction of embryonic endoderm cells. Developmental Cell. 11 : 1-10.
Bort, R, Signore, M., Tremblay, K. D., Martinez Barbera , J.-P., Zaret, K. (2006). Hex homeobox gene controls the transition of the endoderm to a pseudostratified, cell emergent epithelium for liver bud development. Developmental Biology, 290:44-56.
Tremblay, K. D. and Zaret, K. (2005). Distinct populations of endoderm cells converge to generate the embryonic liver bud and ventral foregut tissues. Developmental Biology 280 :87-99.
Tremblay, K.D., Dunn, N. R. and Robertson, E. J. (2001). Mouse embryos lacking Smad1 signals display defects in extra-embryonic tissues and germ cell formation. Development 128: 3609-3621.
Tremblay, K.D., Hoodless, P.A., Bikoff, E, and Robertson, E.J. (2000). Formation of the definitive endoderm is a Smad2-dependent process. Development 127:3079-3090.
Doherty, A. S., Mann, M. R. W., Tremblay, K. D., Bartolomei, M. S., Schultz, R. M. (2000). Differential effects of culture on imprinted H19 expression in the preimplantation mouse embryo. Biology of Reproduction62: 1526-1535.
Schultz, R.M., Tremblay, K.D., Doherty, A.S. and Bartolomei, M.S. (2000). Effect of embryo culture on imprinted gene expression in the preimplantation mouse embryo. In "The Testis: From Stem Cell to Sperm Function". Goldberg, E., ed., Springer-Verlag, New York.
Davis, T. L.*, Tremblay, K.D.* and Bartolomei, M.S. (1998). Imprinted expression and paternal methylation of the mouse H19 gene are conserved in extraembryonic lineages. Developmental Genetics 23:111-118.
* denotes equal contribution
Tremblay, K. D. (1998). Bisulfite methylation analysis of single DNA strands. Trends in Genetics, Technical Tips Online. 01242
Tremblay, K.D., Duran, K.L. and Bartolomei, M.S. (1997). A 5’ 2 kilobase-pair region of the imprinted mouse H19 gene exhibits exclusive paternal methylation throughout development. Molecular and Cellular Biology 17: 4322-4329.
Tremblay, K.D., Saam, J.R., Ingram, R.S., Tilghman, S.M. and Bartolomei, M.S. (1995). A paternal-specific methylation imprint marks the alleles of the mouse H19 gene. Nature Genetics9: 407-413.