Endothelial cell transduction in primary cultures from regressing mesonephros

Abstract

Loss of renal function during normal aging is associated with vascular alterations. Consequently, new therapeutic approaches, including gene therapy, to protect renal endothelial cells are expected to be greatly beneficial. Quail mesonephros is a transitory embryonic kidney that has been used for the study of vascular development and involution. Vascular alterations in regressing mesonephros are similar to those observed in aging kidney. In the present study, we examined adenovirus-mediated gene transfer to endothelial cells in primary cultures from developing and regressing quail mesonephros. Quail embryos with developing and regressing mesonephros were examined on day 6 (30HH) and day 11 (40HH) of incubation, respectively. The senescence markers, associated beta-galactosidase activity and p16(INK4a), were examined in whole mesonephros. Quail embryos were injected intracardiacally with adenoviral vectors (rAd-CMV-LacZ) and endothelial cell transduction examined. In addition, primary cell cultures from mesonephros were exposed to adenoviral vectors. Endothelial cells in primary cultures were identified as QH1(+), LEP100(-) and acidic phosphatase(-) cells and adenovirus-transduced cells were those positive for bacterial-associated beta-galactosidase activity. We report that endothelial cells in the whole regressing mesonephros and primary cell cultures expressed senescence markers. In addition, we observed that adenoviral vectors were able to transduce endothelial cells in the whole regressing mesonephros, and that cultured endothelial and macrophagic cells from the regressing mesonephros were more efficiently transduced than those derived from the developing mesonephros. Our results suggest that quail mesonephros provides a practical model to assay gene transfer to endothelial cells in regressing/senescent vessels.