Tumor necrosis factor alpha (TNFalpha) is a multifunctional cytokine present in oocytes and macrophages in the neonatal rat ovary. The presence of both TNFalpha and its receptors in the neonatal rat ovary suggests a potential role for it in follicle assembly or oocyte atresia. Previous studies have provided support for effects of TNFalpha on isolated granulosa and theca cells and intact follicles; however, to our knowledge, this is the first study to investigate the effects of TNFalpha on the earliest stages of follicular development. Effects of TNFalpha on oocyte/follicle number and apoptosis were investigated using an ovarian organ-culture system that supported assembly of primordial follicles in vitro. Ovaries were collected on the day of birth and treated with TNFalpha (0, 0.1, 1.0, 10, or 50 ng/ml), a function-blocking TNFalpha antibody (5 microg/ml), or control immunoglobulin (Ig) G. At 1 ng/ml, TNFalpha decreased follicle and oocyte numbers during 3 days of culture, whereas higher (10 and 50 ng/ml) or lower (0.1 ng/ml) doses had no effect. Treatment with TNFalpha antibodies increased the number of oocytes and follicles compared to nonspecific IgG control. To determine whether the decreased oocyte/follicle numbers were due to an apoptotic effect of TNFalpha, apoptosis was examined by DNA laddering. At 1 ng/ml, TNFalpha increased apoptotic DNA laddering twofold, with no significant effect from lower or higher doses. The cells undergoing apoptosis, as determined by in situ end-labeling, were oocytes, interstitial cells, and granulosa cells. These findings suggest that TNFalpha may be involved in oocyte atresia that normally occurs during the perinatal period.

Tumor necrosis factor-alpha (TNF) is a cytokine produced not only by various cells of the immune system, but also by various cells in the reproductive system. We have demonstrated that oocytes are an important source of TNF and that the onset of oocytic TNF expression occurs around birth. TNF receptors are localized on oocytes, granulosa cells and interstitial cells allowing for the possibility of autocrine or paracrine actions of TNF. The late fetal/early neonatal period represents a time during which several key events occur, including formation of the primordial follicle and extensive oocyte apoptosis. We have utilized an ovary culture system to examine the involvement of TNF in early ovarian function. This culture system allows both primordial follicle function and apoptosis to occur in vitro. Our results show that TNF can decrease oocyte and primordial follicle number through stimulation of oocyte apoptosis in vitro. TNF thus may serve as an important intraovarian factor involved in the determination of the size of the primordial follicle pool.