title: Evidence that FOXO3a is involved in oocyte apoptosis in the neonatal rat ovary.
abstract: Previous studies have proposed that the forkhead transcription factor FOXO3a is involved in cell cycle arrest and apoptosis and that it may also repress follicular development by inducing cell cycle arrest in ovaries. We have recently demonstrated that FOXO3a induces oocyte apoptosis of neonatal rat ovaries under in vitro conditions. In the present study, we evaluated the role of FOXO3a in oocyte apoptosis under in vivo conditions. Ovaries from rats were obtained from newborns on postnatal day (PD) 1, 2, 3, and 4. TUNEL assay results showed that oocyte apoptosis occurred mainly on PD 1 and 2. Immunohistochemical staining of FOXO3a, Bim, Fas ligand (FasL), p27KIP1, caspase-8, and caspase-3 showed that they were all expressed mainly in naked oocytes on PD 1 and 2. The percentage of positive FOXO3a staining of oocytes reached peak levels in the ovaries of 2-day-old rats, which was consistent with the rate of the apoptotic profiles determined by TUNEL. The percentage between TUNEL-positive and FOXO3a-positive oocytes in the nucleus showed no statistical differences within the 4-day-old rat ovaries. Furthermore, the positive oocyte percentage of the target factors of FOXO3a (Bim, p27KIP1, and FasL) and pro-apoptotic proteins (caspase-3 and caspase-8) also reached peak levels in the ovaries of 2-day-old rats, which was similar to the rate of FOXO3a-positive oocytes. These results suggest that FOXO3a in the oocyte nucleus is involved in oocyte apoptosis; that is, FOXO3a-positive oocytes may be the apoptotic cells. To verify this, rat oocytes were subjected to TUNEL and immunofluorescent double-labeling assays. We found that TUNEL-positive cells were also FOXO3a-, Bim-, or FasL-positive. To identify the downstream target of FOXO3a, double immunofluorescent staining with antibodies to Bim and FasL was performed. We found that FOXO3a-positive cells were also Bim- and FasL-positive. We conclude that the overexpression of FOXO3a in the oocyte nucleus of neonatal rat ovaries may play an important role in the apoptosis of naked oocytes, and that Bim, FasL, and p27KIP1 are the key downstream factors of FOXO3a.
authors.affiliation: Laboratory of Cell Senescence, Shantou University Medical College, Shantou, China.
authors.forename: Xu-Xia
authors.initials: XX
authors.lastname: Sui
authors.forename: Li-Li
authors.initials: LL
authors.lastname: Luo
authors.forename: Jin-Jie
authors.initials: JJ
authors.lastname: Xu
authors.forename: Yu-Cai
authors.initials: YC
authors.lastname: Fu
mesh_terms: D000818:Animals; D000831:Animals, Newborn; D017209:Apoptosis / Q000502:physiology; D051017:Apoptosis Regulatory Proteins / Q000378:metabolism; D000072224:Bcl-2-Like Protein 11; D020169:Caspases / Q000378:metabolism; D050760:Cyclin-Dependent Kinase Inhibitor p27 / Q000378:metabolism; D053222:Fas Ligand Protein / Q000378:metabolism; D005260:Female; D000071316:Forkhead Box Protein O3; D051858:Forkhead Transcription Factors / Q000378:metabolism / Q000502:physiology*; D048788:Growth and Development / Q000502:physiology; D008297:Male; D008565:Membrane Proteins / Q000378:metabolism; D009865:Oocytes / Q000378:metabolism / Q000502:physiology*; D006080:Ovarian Follicle / Q000378:metabolism / Q000502:physiology; D010053:Ovary / Q000254:growth & development / Q000378:metabolism / Q000502:physiology*; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: D051017:Apoptosis Regulatory Proteins; D000072224:Bcl-2-Like Protein 11; C000606778:Bcl2l11 protein, rat; C493962:Cdkn1b protein, rat; C499515:FOXO3 protein, rat; D053222:Fas Ligand Protein; C505662:Faslg protein, rat; D000071316:Forkhead Box Protein O3; D051858:Forkhead Transcription Factors; D008565:Membrane Proteins; D011518:Proto-Oncogene Proteins; D050760:Cyclin-Dependent Kinase Inhibitor p27; D020169:Caspases