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 β-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 β-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.

Dynamic Spatiotemporal Expression Pattern of the Senescence-Associated Factor p16Ink4a in Development and Aging

“…Interestingly, absence of p21 was compensated by apoptosis, but still slight developmental abnormalities were detectable [43]. Although these studies focused mainly on p21, p16 loss has also been shown to result in developmental defects in the eye [79]; inactivation of p16 and p19 induced cardiomyocyte proliferation [80]; p16 has been detected in the ventricular and subventricular zones at embryonic and early postnatal stages in the rat brain; SA-β-galactosidase activity and p16 expression has been detected in regressing mesonephros of quails [81]; p16 expression in mouse embryos has been detected in motoneurons and the senolytic ABT-263 decreased the number of these cells [82]. Nevertheless, not all highly p16-positive cells are necessarily senescent [83,84].…”

Expression and Localization of Angiogenic Growth Factors in Developing Porcine Mesonephric Glomeruli

“…In the early fetal stage, the mesonephros regresses and the metanephros, i.e., the definitive kidney, starts to develop. The normal physiological regression of the mesonephros can serve as a model for the study of pathologically degenerating renal glomeruli because the morphological characteristics of developing mesonephric and metanephric glomeruli are highly similar (Gersh 1937;Leeson and Baxter 1957;Tiedemann and Egerer 1984;Smith and Mackay 1991;Nacher et al 2006Nacher et al ,2010. However, the molecular mechanisms of mesonephric glomerulogenesis are not well-known.…”