Involvement of the KIT/KITL Signaling Pathway in 4-Vinylcyclohexene Diepoxide-Induced Ovarian Follicle Loss in Rats1

Abstract

Repeated daily dosing of rats with the occupational chemical 4-vinylcyclohexene diepoxide (VCD) depletes the ovary of primordial and primary follicles through an increase in the natural process of atresia. Additionally, in vitro exposure of Postnatal Day 4 (PND 4) rat ovaries to VCD causes similar follicular depletion. This study was designed to investigate survival signaling pathways that may be associated with VCD-induced ovotoxicity in small preantral follicles. Female Fischer 344 rats (PND 28) were dosed daily (80 mg/kg/day VCD i.p.; 12 days in vivo), and PND 4 ovaries were cultured (VCD 20 or 30 microM; 8 days in vitro). Microarray analysis identified a subset of 14 genes whose expression was increased or decreased by VCD in both experiments (i.e., via both exposure routes). Particularly, the analysis showed that relative to controls, VCD did not affect mRNA expression of growth and differentiation factor 9 (Gdf9), whereas there were decreases in mRNA encoding bone morphogenic protein receptor 1a (Bmpr1a) and Kit. To confirm findings from microarray, the genes Gdf9, Bmpr1a, and Kit were further examined. When growth factors associated with these pathways were added to ovarian cultures during VCD exposure, GDF9 and BMP4 had no effect on VCD-induced ovotoxicity; however, KITL attenuated this follicle loss. Additionally, there was a decrease in Kit and an increase in Kitl expression (mRNA and protein) following VCD exposure, relative to control. These results support that VCD compromises KIT/KITL signaling, which is critical for follicular survival in primordial and primary follicles.

Small RNAs: Their Possible Roles in Reproductive Failure

“…However, damage to oocytes by chemical exposures in utero and/or during childhood presents a concern, which would not be detected until the reproductive years. Ovotoxic chemicals can accelerate activation of primordial follicles from the ovarian reserve (Fernandez et al 2008 ;Keating et al 2009 ), leading to POF. Since this process is, at least partially, regulated by PI3K signaling, the involvement of miRNAs is plausible.…”

Mechanisms of Ovarian Atresia Induced by Xenobiotic Exposures

“…When PI3K was inhibited in PND4 ovaries exposed to VCD, primordial follicles were protected from VCD-induced ovotoxicity . It was subsequently demonstrated that VCD induces a decrease in c-KIT autophosphorylation (Mark-Kappeler et al, 2011) and AKT phosphorylation (Keating et al, 2011) after two days followed by a decrease in c-Kit mRNA expression on day 4, prior to an increase in KL mRNA and follicle loss on day 6 of exposure (Fernandez et al, 2008). Further, exogenous KL in culture partially attenuated VCD-induced follicle loss, while there was no effect of exogeneous BMP15 or GDF9 (Fernandez et al, 2008).…”

“…It was subsequently demonstrated that VCD induces a decrease in c-KIT autophosphorylation (Mark-Kappeler et al, 2011) and AKT phosphorylation (Keating et al, 2011) after two days followed by a decrease in c-Kit mRNA expression on day 4, prior to an increase in KL mRNA and follicle loss on day 6 of exposure (Fernandez et al, 2008). Further, exogenous KL in culture partially attenuated VCD-induced follicle loss, while there was no effect of exogeneous BMP15 or GDF9 (Fernandez et al, 2008). Thus, the initial ovarian target of VCD is the oocyte, and VCD-induced inhibition of the PI3K pathway is thought to be an early initiating ovotoxic event, which precedes the activation of the classical apoptotic pathways.…”

Inhibition of PIK3 Signaling Pathway Members by the Ovotoxicant 4-Vinylcyclohexene Diepoxide in Rats

“…Oligoarray analysis demonstrated that, following follicle loss, mRNA encoding Kit was reduced in rat ovaries in response to VCD exposure via in vivo dosing (d15) or in vitro culture (d8) [26].…”