Studies of the notch signaling pathway using transgenic mouse models

Description

The Notch signaling pathway is a cell communication pathway essential for formation of multiple systems during mammalian development. Aberrant Notch signaling is associated with a variety of human diseases. Functional studies of Notch in mice have been limited because both the absence and overexpression of Notch results in embryonic lethality. To investigate the effects of Notch signaling in vivo, three lines of Notch transgenic mice have been created that have a floxed beta-geo/stop signal between a strong promoter and the constitutively active intracellular domain of Nothch1 (IC-Notch1). IC-Notch1 can be activated after the introduction of Cre recombinase and its expression is detected through a co-expressed EGFP or hPLAP. Double transgenic IC-Notch1/pCX-Cre embryos in which IC-Notch1 expression was globally activated died at E9.5 with lack of neural tube closure, disrupted vasculature and irregular somites, demonstrating that expression of IC-Notch1 can be effectively activated by Cre recombinase. Endothelial/hematopoietic specific expression of IC-Notch1 in double transgenic IC-Notch1/Tie2-Cre embryos induced embryonic lethality at E9.5 with defects in vascular development, but did not affect primitive hematopoiesis. The Snail repressor, a mediator of endothelial-to-mesenchymal transition, was upregulated by IC-Notch1 expression in embryonic heart.To avoid the embryonic lethality, inducible IC-Notch1 expression in adult mice was achieved by crossing IC-Notch1 mice with a Cre transgene under the tetracycline operator controlled Cre (tet-O-Cre) and tetracycline transactivator under the control of Tie2 promoter (Tie2-tTA). Using this system, IC-Notch1/tet-O-Cre/Tie2-tTA mice survived embryonic development when maintained on tetracycline. After withdrawing tetracycline post-natally, expression of IC-Notch1 was detected in endothelial and hematopoietic cells by immunostaining of the GFP reporter. The IC-Notch1 expressing females were less fertile with lack of mature follicles. Matrigel plug assay showed that IC-Notch1 expression in adult mice inhibited bFGF-induced, but not VEGF induced neovascularization. In addition, 50% of transgenic mice with IC-Notch1 expression developed enlarged hematopoietic organs. Immunohistochemistry showed extensive T cell infiltration in various organs. Thus, constitutive active Notch signaling inhibited angiogenesis and induced T cell hyperproliferation in adults. This study provided a series of mouse models and valuable insights to design therapies for vessel related diseases and T cell lymphoma.

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