Influence of Lysophosphatidic Acid on Nitric Oxide-Induced Luteolysis in Steroidogenic Luteal Cells in Cows1

For decades, the mechanisms responsible for germ cell depletion from the ovary, either directly during the perinatal period or indirectly via follicular atresia during postnatal life, have remained relatively obscure. The recent application of sensitive biochemical techniques for the study of cell death to the analysis of ovarian function has revealed that these two events, as well as a third instance of ovarian cell degeneration (luteolysis), are dependent upon the activation of physiological cell death mechanisms. It is therefore hypothesized that the controlled deletion of ovarian cell populations is accomplished via activation of a 'universal' pathway of cellular suicide involving altered expression of a conserved cohort of genes. The identity of the hormonal and intracellular effectors responsible for the coordination of life and death decisions made by ovarian cells during development as well as the biological and clinical implications of gene-directed cell death in the ovary are explored in this review.

The corpus luteum (CL) is a transient reproductive gland that produces progesterone (P), required for the establishment and maintenance of pregnancy. Although the regulation of bovine luteal function has been studied for several decades, many of the regulatory mechanisms involved are incompletely understood. We are far from understanding how these complex mechanisms function in unison. The purpose of this overview is to stress important steps of regulation during the lifetime of CL. In the first part, the importance and regulation of angiogenesis and blood flow during CL formation is described. The results underline the importance of growth factors especially of vascular endothelial growth factor A (VEGF A) and basic fibroblast growth factor (FGF-2) for development and completion of a dense network of capillaries. In the second part, the regulation of function by endocrine/paracrine- and autocrine-acting regulators is discussed. There is now more evidence that besides the main endocrine hormones LH and GH local regulators as growth factors, peptides, steroids and prostaglandins are important modulators of luteal function. During early CL development until mid-luteal stage oxytocin, prostaglandins and P itself stimulate luteal cell proliferation and function supported by the luteotropic action of a number of growth factors. The still high mRNA expression, protein concentration and localization of growth factors [VEGF, FGF-1, FGF-2, insulin-like growth factors (IGFs)] in the cytoplasm of luteal cells during mid-luteal stage suggest maintenance (survival) functions for growth factors. In the absence of pregnancy regression (luteolysis) of CL occurs. Progesterone itself regulates the length of the oestrous cycle by influencing the timing of the luteolytic signal prostaglandin F2alpha (PGF2alpha) from the endometrium. The cascade of mediators afterwards is very complex and still not well-elucidated. Evidence is given for participation of blood flow, inflammatory cytokines, vasoactive peptides (angiotensin II and endothelin-1), reactive oxygen species, angiogenic growth factors (VEGFs, FGFs, IGFs) and decrease of the classical luteotropic components as LH-R, GH-R, P450(scc) and 3beta-HSD. Despite of differences in methodology and interpretations, progress has been made and will continue to be made.

Tumour necrosis factor alpha (TNF-alpha) and gamma-interferon (IFN-gamma) are cytotoxic to bovine luteal cells in vitro and may contribute to cell death during luteolysis in vivo. In this study, the mechanism by which luteal cells are killed by TNF-alpha and IFN-gamma was investigated. Luteal cells were cultured for 7 days in the presence or absence of TNF-alpha and IFN-gamma. Inhibitors of arachidonate metabolism or scavengers of free radicals were included in the culture media. In addition, the effect of IFN-beta on the viability of cytokine-treated luteal cells was tested. Lastly, untreated and cytokine-treated cells were subjected to single cell gel electrophoresis for quantification of DNA fragmentation. Neither indomethacin nor nordihydroguaiaretic acid, which are inhibitors of cyclooxygenase and lipoxygenase, respectively, were able to prevent cytokine-induced cell death. Similarly, both the phospholipase A(2) inhibitor arachidonyltrifluoromethyl ketone and the nitric oxide synthase inhibitor N(G)-monomethyl-L-arginine, were largely without effect. In contrast, while vitamin C did not significantly affect viability, superoxide dismutase plus catalase increased viability of cytokine-treated cells (P < 0.05), and IFN-beta prevented cell death (P < 0.05). Finally, while control cells remained free of DNA damage, TNF-alpha plus IFN-gamma induced significant amounts of DNA damage by 48 h after initiation of treatment (P < 0.05). In conclusion, reactive oxygen species, but not arachidonate metabolism or nitric oxide, contribute to cytokine-induced luteal cell death in vitro, and the process of cell death may be via apoptosis. Furthermore, IFN-beta may confer protective effects against cytokine-induced cell death in bovine luteal cells.