Expression and Activity of the Fas Antigen in Bovine Ovarian Follicle Cells1

Abstract

The Fas antigen is a cell surface receptor that triggers apoptosis when bound to Fas ligand (FasL). Studies were undertaken to determine whether the cow provides a suitable model to study the role of the Fas pathway in inducing apoptosis of ovarian cells during follicular atresia. Expression of Fas antigen mRNA and responsiveness to FasL-induced killing in vitro were measured. Effects of the cytokines tumor necrosis factor (TNF)-alpha and interferon-gamma (IFN) were studied because of previous demonstrations of their role in Fas-mediated apoptosis in other cell types. Fas antigen mRNA was detectable in cultured granulosa and theca cells, and expression was increased by treatment with IFN but not TNF. Granulosa and theca cells were resistant to FasL-induced killing unless pretreated with IFN. TNF had no effect on FasL-induced killing. Granulosa and theca cell cultures in which killing occurred in response to FasL stained positively for annexin V, an early marker for cells undergoing apoptosis. These results provide a basis for further studies using the bovine ovary to examine the role of the Fas antigen in follicular atresia.

Molecular Characteristics of Porcine Fas-associated Death Domain (FADD) and Procaspase-8

“…Cell death ligand and receptor systems [such as the Fas ligand (FasL)/Fas system and TNF relating apoptosis inducing ligand (TRAIL)/TRAIL-receptor system] mediate apoptosis during follicular aresia in porcine ovaries [18][19][20][21][22][23][24][25][26][27]. In porcine granulosa cells, apoptosis is regulated by a cell death ligand and receptor system, TRAIL/TRAIL-R, as follows [22].…”

Ovarian Follicular Atresia

“…They include TNF (Basini et al, 2002;Sasson et al, 2002), Fas ligand , and TNF-related apoptosis-inducing ligand (TRAIL) (Johnson et al, 2007;Jaaskelainen et al, 2009). Additional extrinsic factors that influence apoptosis of granulosa cells include interferon-gamma Vickers et al, 2000) and several types of growth factors . Intrinsic factors of apoptosis are those that are generally provoked by aspects of stress.…”

“…In the human, bovine, and other species, Fas ligand and the Fas-mediated pathway of apoptosis are considered prominent mechanisms of granulosa cell death during follicular atresia (Quirk et al, 1995;Hakuno et al, 1996;Kondo et al, 1996;Vickers et al, 2000). The targeted, cell-specific nature of granulosa cell death without any accompanying inflammatory response or collateral damage to adjacent cells is a unique feature of the Fas ligand-Fas system, consistent with its renowned role(s) in immune response, the establishment of immune tolerance, and the activation-induced cell death of lymphocytes.…”

“…In fact for some studies, serum-withdrawal during the culture period was implemented to induce the onset of apoptosis (Quirk et al, 1995;Porter et al, 2000;Quirk et al, 2000;Vickers et al, 2000). We now know that the various growth factors contained within serum, specifically insulin-like growth factor (IGF), basic fibroblast growth factor (bFGF), and epidermal growth factor (EGF), are what provide the beneficial, anti-apoptotic effects Yang & Rajamahendran, 2000;Mani et al, 2010).…”

Apoptosis of bovine ovarian surface epithelial cells by Fas antigen/Fas ligand signaling

“…RNA was prepared (Chomczynski & Sacchi 1987) and reverse transcribed in the presence of increasing concentrations of an internal Fas or FasL RNA standard. The Fas and FasL RNA standards were prepared by in vitro transcription of mutated Fas or FasL cDNA, each containing a 50 bp deletion internal to the PCR primer-binding sites (described in Vickers et al 2000. Complementary DNA was amplified by PCR in the presence of [ 32 P]dCTP.…”