Programmed Cell Death in Human Ovary Is a Function of Follicle and Corpus Luteum Status*

Abstract

Although extensive investigation on follicular apoptosis (programmed cell death) has been conducted in the infraprimate ovary, there is little information regarding apoptosis and its relationship to follicular status in the human. In this study, apoptosis was investigated in 116 human ovarian follicles (primordial to dominant) and 5 corpora lutea from a total of 27 premenopausal women. Follicles and corpora lutea were evaluated for the presence of DNA fragmentation, characteristic of apoptosis, by two methods: in situ hybridization using 3' end-labeling of DNA with digoxigenin-labeled nucleotides and subsequent digoxigenin antibody and peroxidase staining, and/or biochemical analysis of low molecular weight DNA laddering. Follicle functional status was evaluated by determining follicle sizes and follicular fluid androgen/estrogen (A/E) ratios. No apoptosis was observed in 67 primordial, primary, or secondary follicles. Positive staining for DNA fragmentation was found in a few granulosa cells in 0.1- to 2-mm follicles, whereas abundant staining in granulosa was detected in 2.1- to 9.9-mm follicles. In contrast, no DNA fragmentation was detected in dominant follicles (10-16 mm). The frequency of apoptosis in follicles was calculated to be 37% in 0.1- to 2-mm follicles, 50% in 2.1- to 5-mm follicles, and 27% in 5.1- to 9.9-mm follicles. Abundant low molecular weight DNA laddering was only found in androgen-dominant follicles and not in estrogen-dominant follicles. Positive staining for DNA fragmentation and low molecular weight DNA laddering were observed in degenerating but not healthy-appearing corpora lutea. In the former, DNA fragmentation was found primarily in large luteal cells. These data suggest that follicular atresia in human ovary results from normal programmed cell death and primarily occurs in the granulosa cell layers of the early antral and < 10-mm antral follicles primarily. Furthermore, because apoptosis occurs as early as the 200-mm stage, follicle selection may begin as early as the initial formation of the antrum. The results also suggest that degeneration of the corpus luteum occurs by apoptotic mechanisms.

Positive and negative effects of cellular senescence during female reproductive aging and pregnancy

“…During folliculogenesis, one or a few ovarian follicles become dominant and go through ovulation, while others fail to reach preovulatory stage, trigger apoptosis and succumb to atresia (Block 1951, Yuan & Giudice 1997. Because support cells are important for selecting follicles, which will either become dominant or undergo atresia (Matsuda et al 2012), lack of proliferation and cellular expansion in support cells due to replicative senescence may suppress the ability of growing follicles to advance to the preovulatory stage, making all of these immature follicles destined for apoptosis instead (Fig.…”

The effect of ovarian reserve and receptor signalling on granulosa cell apoptosis during human follicle development

“…Apoptosis is a normal regulatory process that contributes to the maintenance of a healthy complement of follicles and their constituent oocytes (Yuan and Giudice, 1997). The granulosa cells are more susceptible to apoptosis in the follicle than the theca or cumulus cells (Bencomo, Pérez, Arteaga et al, 2006).…”

“…Earlier studies on apoptosis of follicular cells have employed a range of analyses based on morphological assessment of pyknotic cell counts, TUNEL, and propidium iodide assessment, all with pooled follicle samples of unknown size (Yuan and Giudice, 1997,Nakahara et al, 1997,Oosterhuis et al, 1998,Seifer, 1996,Giampietro, Sancilio, Tiboni et al, 2006,Austin, Mihm, Evans et al, 2001,Bomsel-Helmreich, Gougeon, Thebault et al Many of these studies suffer from technical limitations because they have relied on pooling follicles of different sizes, counting a small portion of the granulosa cells (~100-1000), and have excluded follicles because of blood contamination, or failed to exclude white blood cells (Nakahara et al, 1997,Oosterhuis et al, 1998,Seifer, 1996. In addition, when propidium iodide and Annexin V-FITC are combined, spectral overlap was not compensated for, and made the incorrect interpretation of the quadrants as being apoptosis-induced necrotic cells (Jančar et al, 2007,Seifer, 1996,Giampietro et al, 2006.…”

Follicular Atresia and Luteolysis Evidence of a Role for N‐Cadherin

“…Although several hundred thousands of primordial and primary follicles are present in the mammalian ovary before puberty, only very few will fully mature and ovulate. The others will be eliminated by atresia, a process that exhibits both the biochemical and morphological features of programmed cell death 1–4. Moreover, in each reproductive cycle, a corpus luteum will form and eventually degenerate by luteolysis, which is also an apoptotic process 5–8.…”

“…The primary follicular constituents that undergo apoptosis are the granulosa cells (GCs) 9. Because follicular degeneration, or corpus luteum regression, is associated with loss of cell‐cell adhesion sites, 4 it can be hypothesized that cell adhesion molecules (CAMs) are implicated in GC survival and death. In addition to cell‐cell adhesion, recent studies suggest that an interaction with extracellular matrix proteins plays an important role in regulating cell survival 10.…”