Ramazan Arici

Kedi Oositlerinin In Vitro Olgunlaştırılması ve Kumulus Hücrelerinin Apoptoz Oranları Üzerine, Ovaryum Taşıma ve Saklama Sıcaklığının Etkisi

The objective of the present study was to examine the effects of two different transport temperature (37°C vs 4°C) and cold storage of ovaries for 24 h on cumulus cell apoptosis and maturation rates of cat oocytes in vitro. Ovaries were collected from 15 ovariohysterectomized domestic cats and maintained and transported to the laboratory in phosphate buffer saline at 37°C and 4°C. In order to determine the effects of storing time, some ovaries transported at 4°C were stored at the same temperature for 24 h. Selected cumulus oocyte complexes (COCs) were matured for 48 h at 38°C in four-well petri dishes containing 500 μL of modified oviduct medium (mSOF) under mineral oil in a 5% CO2 incubator with nearly 100% humidified. The morphological features of apoptosis were analysed in the cumulus cells at the beginning of in vitro maturation in both transporting temperature groups and after 24 h of cold stored group. The degree of apoptosis in cumulus cells were measured by terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling (TUNEL). The IVM rates of oocytes were determined using Hoechst (33342) staining. Although the apoptotic morphological features were seen rarely and in similar rates in 37 and 4°C transporting groups (19.40 and 21.55%, P>0.001), it was seen more intensely in the 24 h cold stored group (34.80%, P<0.001). The IVM findings were similar (49.77, 44.55%) at 37°C and 4°C groups (P>0.05), and importantly lower at 4°C transporting and 24 h cold stored groups (18.90%, P<0.05). In conclusion, the results of this study suggest that (I) cumulus cells of cat oocytes are partially exposed to apoptosis during transportation at warm or cold temperature, (II) storing of ovaries for 24 h at 4°C causes apoptosis of the cumulus cells at much higher rates and (III) storing of ovaries for 24 h at 4°C affects negatively IVM rate of oocytes.

Kedilerde MI ve MII Oositleri Kullanilarak Somatik Klonlama

Animal production via SCNT provides a unique tool for protection of valuable individuals, conservation of vulnerable and endangered species and production of transgenic animals. A total of 167 MI and 219 MII stage oocytes were used as the material of the study. The oocytes were enucleated at 44 h after in vitro maturation by aspiration of the polar body and the MI or MII plates. Cycling granulosa cells were used for nuclear transfer. Cell fusion was induced with DC pulses of 2.0 kV/cm 60µs, 0.1s apart (2x) delivered by a BTX Electrocell Manipulator 200 (BTX, San Diego, CA, USA). After fusion, the embryos were activated by 1.0 kV/cm 20µs DC pulses 0.1s apart (2x) followed by 2 mM 6-DMAP (6-dimethylaminopurine) incubation in culture medium for 4 h in a humidified atmosphere of 5% CO2, 5% O2, and 90% N2 at 38°C. The somatic cell transferred embryos were cultured for 8 days in mSOF medium supplemented with 0.4% BSA in a humidified 5% CO2, 5% O2, and 90% N2 atmosphere at 38°C. After in vitro culture period, all embryos transferred to HSOF containing Hoechst 33342 (5 μg/mL) and the cell numbers were counted under ultraviolet light using a fluorescent microscope. The fusion (66.66 vs 21.55%) and cleavage rates (15.75 vs 11.11%) were significantly higher in MII stage oocytes than MI stage oocytes (P<0.02). While SCNT embryos were developed to morula stage in MII group (14; 9.58%), all the cleaved embryos were arrested at the 2-4 cell stage in MI group. None of the embryos was developed to blastocyst stage in both groups.

The effect of oviductal cells on in vitro maturation of canine oocytes in different culture media

Sinem Özlem ENGİNLER*, Asiye İzem SANDAL, Özen Banu ÖZDAŞ, Ramazan ARICI, Ezgi ERTÜRK, Elif Merve ÇINAR, Israa Faris MOHAMMED, Alper BARAN, Çağatay TEK, Mehmet Can GÜNDÜZ Department of Obstetrics and Gynecology, Faculty of Veterinary Medicine, Istanbul University, Avcılar, İstanbul, Turkey Department of Reproduction and Artificial Insemination, Faculty of Veterinary Medicine, Istanbul University, Avcılar, İstanbul, Turkey