EFFECTS OF ANTIOXIDANT AND OXYGEN TENSION ON REACTIVE OXYGEN SPECIES-RESPONSIVE GENE EXPRESSION AFTER TRANSPLANTATION USING HYPOXIA INDUCED BOVINE OVARIAN TISSUE

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A large proportion of childhood and young cancer patients will lose their fertility after aggressive cancer therapy because of the gonadotoxicities of chemotherapy and radiotherapy. One of the strategies to preserve fertility for those facing premature ovarian failure is ovarian tissue banking. The efficacy of ovarian tissue banking for fertility restoration in humans has not been proved yet. However, there is ample evidence of its efficacy in animal studies. Ovarian tissue banking requires further investigations and should be considered experimental in humans at present. Probably, the most important and urgent issue for successful ovarian transplantation is how to minimize ischemic injuries before and during establishment of angiogenesis in the ovarian grafts. It could be speculated that reduced oxygen tension and antioxidant treatment would protect ovarian grafts from hypoxic injuries, as hypoxia induces the generation of reactive oxygen species (ROS). The aim of this study was to determine the effect of reduced oxygen tension and antioxidants treatment on the protection of apoptotic cell death in transplanted ovarian tissue. Bovine ovarian tissue was sliced into a size of 2 mm3 using aseptic technique, then cultured in 5% or 20% O2 for 0h, 3h, 24h and 48h in a-MEM media with/without antioxidants (ascrobic acid andS1P). mRNA expressions of ROS-responsive genes (Mn-SOD, CuZn-SOD, catalase, GPx, GCS, Bax and GAPDH) in the bovine ovary tissue were determined by RT-PCR. Apoptosis was quantified in situ by terminaldeoxynucleotidyl transferase labeling (TUNEL) assay. There was no significant association with ROS-responsive genes expression and O2 concentration after 3 hours. After 24 and 48 hours, however, culture in 5% O2 significantly increased ROS-responsive genes expression than 20% O2. In the ascrobic acid and S1P treated groups, the mRNA expressions of catalase, CuZn-SOD, Mn-SOD were increased compared with control group after 24 hours, whereas the mRNA expressionsof GPx, GCS, Bax were not different. There was no difference in ROS-responsive genes expression between control and antioxidant groups after48 hours. These results show that it is desirable to perform ovarian tissue transplantation within 3 hours post-thawing to minimize ROS production. Treatments with antioxidants play some role in protecting transplanted cells from oxidative stress-induced cell death. The strategies to reduce reactive oxygen species should be considered as helpful options in ovarian tissue transplantation. This work was supported by the Korea Research Foundation Grant funded by the Korean Government (KRF-2005-003- E00145). (poster)