A comprehensive knowledge of the genetic programs and embryonic inductive events regulating human pancreas development would advance our understanding of congenital pancreatic malformations and aid in developing rational treatment strategies for diabetes mellitus. Significant insights to the mechanisms of pancreatic organogenesis have been gained through recent experiments in mice and chickens. However, critical differences in early developmental pathways between these species and humans places the direct applicability of these data to human development in question. Furthermore, functional studies in human development are restricted by the ethical implications of directly manipulating human embryos. Consequently, a more """"""""human"""""""" model based on nonhuman primates would have broad application for studying human development. Recently, embryonic stem (ES) cell lines from nonhuman primates (rhesus macaque and common marmoset) have been derived and their potential for differentiation into derivatives of all three embryonic germ layers (ectoderm, mesoderm, and endoderm) in vivo has been characterized. Their capacity for lineage-restricted differentiation in vitro, however, has not been thoroughly tested to date. in contrast, neural, hematopoietic and muscle lineage differentiation of mouse ES cells in vitro has been extensively studied. Recently, the ability of mouse ES cells to undergo elements of endoderm and pancreatic differentiation in culture has also been demonstrated. Therefore, we will test the hypothesis that pancreatic endocrine and/or exocrine lineage specification can be recapitulated in rhesus ES cells induced to differentiate.
The specific aims of this proposal are: l) To determine the expression pattern of endoderm related genes as well as pancreatic exocrine/endocrine-specific genes in rhesus ES cells differentiating in vitro, and 2) To determine the potential of rhesus ES cells for pancreatic lineage differentiation in ES cell derived tumors in immunocompromised mice. A combination of RT-PCR analysis, Northern hybridization, ribonuclease protection assay, in situ hybridization, and immunofluorescence studies will be used to characterize the pattern of pancreatic gene expression. These studies will provide a basis for the refinement of an in vitro primate model of human pancreatic development. Such a model system would have important implications for investigating the mechanisms controlling pancreatic differentiation and for developing new cell-based strategies for treating diabetes.
