Poster Sessions

DEV-15

Jeff Hsu

J.C. Hsu, G. Di Pasquale, T. Onodera, J.S. Harunaga, M.P. Hoffman, J.A. Chiorini, K.M. Yamada

Viral Gene Transfer to Developing Mouse Salivary Glands

Branching morphogenesis is essential for the formation of salivary glands, kidneys, lung, pancreas, and many other organs during development, but the mechanisms of this crucial process are not adequately understood. Microarray and serial analysis of gene expression (SAGE) have been powerful approaches to identify candidate genes that could potentially regulate branching morphogenesis. However, functional validation of these genes has been severely hampered by the difficulty to genetically manipulate cells within organs. Using ex-vivo cultured embryonic mouse salivary glands as a model organ system to study branching morphogenesis, we have identified new vectors for viral gene transfer with high efficiency and cell-type specificity to developing salivary glands. We screened Adenovirus and 20 different types of Adeno-Associated Viruses (AAV) for their ability to transduce early stage (embryonic day 12 or 13) salivary glands. The panel of AAVs tested included not only different serotypes but also viral capsid mutants of AAVs. Enhanced green fluorescent protein (EGFP) was engineered into each of these viruses to facilitate calculations of transduction efficiency. We identified two AAV types, self-complementary AAV2 (scAAV2) and bovine AAV (BAAV), that are highly selective at targeting expression efficiently and differentially to salivary epithelial and mesenchymal cell populations, respectively. Transduction of salivary epithelia with scAAV2 that expresses fibroblast growth factor 7 (FGF7) resulted in an enhanced rate of salivary branching morphogenesis. Our findings represent the first successful selective targeting to epithelial versus mesenchymal cells in an organ undergoing branching morphogenesis, and the first use of gene transfer to analyze roles of growth factors in this process of mammalian organ morphogenesis. The vectors that we have identified can also be used for gene knockdown approaches. They may also be useful for experimental manipulation of other branching organ systems. The information derived from these studies may also facilitate gene therapeutic strategies involving viral gene transfer to salivary glands.