Vascular Endothelial Growth Factor and Its Receptor, Flk-1/KDR, Are Cytoprotective in the Extravascular Compartment of the Ovarian Follicle

Abstract

Vascular endothelial growth factor (VEGF) is a potent mitogen and cytoprotective factor for vascular endothelial cells. Although VEGF is ubiquitously expressed, its role in nonvascular tissues is poorly understood. VEGF interacts with various cell surface receptors to mediate its cellular effects. It previously has been thought that the VEGF receptor Flk-1/KDR, its main signaling receptor, was expressed exclusively by endothelial cells. However, in the present study using bovine and rodent models, we demonstrate that VEGF and Flk-1/KDR are coexpressed in ovarian granulosa cells. VEGF and Flk-1/KDR mRNA and protein were both detectable in follicle tissue sections and in vitro cultured granulosa cells. Expression of both ligand and receptor increased in healthy follicles throughout follicular development. VEGF treatment of serum-starved and cytokine-exposed granulosa cells resulted in enhanced survival, and this cytoprotection was ameliorated when Flk-1/KDR signaling was inhibited. Reduced expression of Flk-1/KDR was also associated with the onset and progression of follicle atresia, suggesting involvement in follicular health in vivo. The results of this study demonstrate for the first time expression of Flk-1/KDR in ovarian granulosa cells and identify a novel extravascular role for VEGF and its receptor in ovarian function.

Vascular endothelial growth factor regulates germ cell survival during establishment of spermatogenesis in the bovine testis

“…An increasing number of reports suggest that VEGFA has direct effects on nonvascular cells including neuronal cells (Marti & Risau 1998, Wada et al 2006, Nishijima et al 2007), muscle cells (Germani et al 2003, Bryan et al 2008, and granulosa cells (Greenaway et al 2004). Data also suggest potential functional roles for VEGFA in the postnatal testis, as KDR and FLT1 are expressed in testicular germ cells in humans (Ergun et al 1997), rats (Rudolfsson et al 2004), and mice (Nalbandian et al 2003), while somatic Sertoli and Leydig cells produce VEGFA (Liu & Yang 2004).…”

Inhibition of vascular endothelial growth factor A expression in mouse granulosa cells by lentivector-mediated RNAi

“…This method provides a novel tool to study the function of VEGFA and its receptors and apoptosis genes of MGC in vitro. Previous reports have indicated that VEGF and its receptors, VEGFR-1 and VEGFR-2, are expressed in pig follicle cells (Barboni et al, 2000), bovine granulosa cells (Einspanier et al, 2002;Greenaway et al, 2004) and MGC (Wulff et al, 2002) in a coordinated fashion in which they protect these cells from apoptosis, which demonstrates that VEGF functions as a survival factor for ovarian follicle and granulosa cells (Kosaka et al, 2007). Therefore, the inhibition of the VEGF gene by RNAi strategy to study its function in MGC requires an understanding of the related effects of this strategy downstream of the VEGF-related genes.…”

“…Unlike VEGF, which is produced by most cell types, research has previously demonstrated that the VEGF receptors Flt-1 and Flk-1 are coexpressed in vascular cells (Ferrara, 2002) including granulosa cells in which they interact to protect these cells against apoptotic cell death, and inhibition of VEGF causes a decrease in Flt and Flk expression (Greenaway et al, 2004;Kosaka et al, 2007). Flt-1 is believed to be expressed in ovarian follicles that consist of oocyte, granulosa cells and the cells of the internal and external theca layer (Okamura et al 2001).…”

“…Its expression in ovarian follicles depends on follicular size. Previous research with bovine and porcine follicles has shown that VEGFA is weakly expressed during early ovarian follicular development and becomes more pronounced in granulosa and theca cells, along with dominant follicle development (Barboni et al, 2000;Greenaway et al, 2004). The rat ovary also shows similar characteristics, with some secondary follicles displaying extremely strong VEGFA immunoreactivity in the zona pellucida (Celik-Ozenci et al, 2003).…”

“…VEGF receptor genes (Flt-1 and Flk-1/KDR) have also been shown to be expressed in granulosa cells (Einspanier et al, 2002;Greenaway et al, 2004). Hence, we investigated the effects of the shRNAs on both Flt-1 and Flk-1 mRNA levels, against housekeeping GAPDH mRNA, and found that there was significant knockdown of all shRNA constructs compared to the shNC ( Figure 5A and B).…”