Background

Our research focuses on investigating the role of Akt1 in vascular tumor development. To determine whether the activation of Akt1 in endothelial cells is sufficient to drive de novo vascular tumor formation, we have developed and published a skin graft model of vascular tumor (Phung et al., 2015). We developed the current protocol of grafting skin from transgenic mice onto the skin of wild type mice because this is a way to study the localized and skin-specific effects of the overexpression of constitutively active Akt1 in the vasculature. We have observed that overexpression of Akt1 in the systemic vasculature leads to generalized edema in the lungs and skin, resulting in premature death of transgenic animals within days due to massive pulmonary edema. A procedure to graft transgenic skin onto wild type host mouse allows one to examine the long-term effects of Akt1 overexpression in only the transgenic skin vasculature without the lethality associated with systemic Akt1 overexpression. In our studies, we are particularly interested in investigating the long-term effects of Akt1 in the skin vasculature on the development of vascular tumors based on the hypothesis that hyperactivation of Akt1 in endothelial cells is sufficient to induce vascular tumor formation, thus demonstrating the endothelial cell-autonomous effects of Akt1 in these tumors.

In the model, skin from double transgenic mice with inducible expression of activated Akt1 (myrAkt1) in endothelial cells is grafted onto the back of immunodeficient nu/nu recipient mice. Since myrAkt1 expression is inducible, we can turn off myrAkt1 expression at will by giving the animals tetracycline in their drinking water, and we can turn on myrAkt1 expression by removing tetracycline from the water. A detailed description of the myrAkt1 double transgenic mice has been published (Sun et al., 2005). A schematic of the construction of these mice is shown (Figure 1). Using the skin graft model, we were able to demonstrate that expression of myrAkt1 in endothelial cells is sufficient for the development of vascular tumor. The procedural steps of this model are described below.


Figure 1. myrAkt1 double transgenic mouse model. The VE-Cad:tTA mouse line contains the VE-cadherin promoter cloned upstream of the tetracycline-regulated transcriptional activator (tTA) gene. VE-cadherin promoter is mainly active in endothelial cells. The TET:myrAkt1 line carries a constitutively activated Akt1 with an HA tag (myrAk1) under the control of the tetracycline responsive promoter (TET). To suppress myrAkt1 expression in the double transgenic mice, 1.5 mg/ml tetracycline with 5% sucrose is given to the mice in their drinking water. To turn on myrAkt1 expression, the mice are given pure water without tetracycline. In this system, double transgenic mice express myrAkt1 in endothelial cells, and the expression of myrAkt1 can be regulated with tetracycline (Reference: Sun JF, Phung T, Shiojima I, Felske T, Upalakalin JN, Feng D, Kornaga T, Dor T, Dvorak AM, Walsh K, Benjamin LE. Microvascular patterning is controlled by fine-tuning the Akt signal. Proc Natl Acad Sci USA 2005; 02: 128-33).