SİNEM ÖZLEM ENGİNLER, Özen Banu ÖZDAŞ, ASİYE İZEM SANDAL, RAMAZAN ARICI, Ezgi ERTÜRK, ALPER BARAN, T. F. Seval TOYDEMİR, ÇAĞATAY TEK, MEHMET RAGIP KILIÇARSLAN, KEMAL AK

The Effect of Cysteamine and Oviductal Cells in Different Culture Media on the Development of Sheep Embryos

Koyun yün, et ve süt kaynağı olarak dünya çapında önemli bir çiftlik hayvanıdır. İn vitro kültür esnasında oksidatif stres embriyo ve gametlerin gelişiminde defektlere neden olur. Sisteamin, L-askorbik asit, beta merkaptoetanol, sistein, glutatyon, proteinler, vitaminler gibi birçok antioksidanlar oksidatif stresi engellemek için kültür medyumunda kullanılır. Bu çalışmada maturasyon medyumuna sisteamin ve kültür medyumuna ovidukt hücreleri eklenen oositlerin kontrol grubu oluşturularak kültürleri sonucundaki gelişim aşamalarının değerlendirilmesi hedeflendi. Oositler mezbahada kesilen koyunların ovaryumlarından elde edildi. Seçilen oositler 100 µM sisteamin ilaveli ve ilavesiz TCM-199 medyumunda 38.5-38.8ºC'de %5 CO2 altında 23 saat boyunca inkübe edildi. IVF aşamasında koçlardan elektroejakülasyon yöntemiyle elde edilen taze sperma H-SOF medyumunda yıkandıktan sonra fertilizasyon amacıyla B-SOF medyumunda bulunan oositler ile 18 saat 38.5-38.8ºC'de %5 CO2, %5 O2 ve %90 N2'de inkübe edildi. Sisteaminli/sisteaminsiz (S +, -) maturasyon medyumundan çıkan oositler, SOF veya CR1aa medyumlarında ovidukt hücreli/ovidukt hücresiz (Ov +, -) şu şekilde gruplara bölündü; Grup Ia: SOF+(S+ Ov-), Grup Ib: SOF+(S+Ov+), Grup Ic: SOF+(S- Ov-) Grup Id: SOF+(S- Ov+); Grup IIa: CR1aa+(S+ Ov-), Grup IIb: CR1aa+(S+ Ov+), Grup IIc: CR1aa+(S- Ov-), Grup IId: CR1aa+(S- Ov+). Embriyolar 38.5-38.8ºC'de %5 CO2, %5 O2, %90 N2 içeren inkubatörde 7 gün boyunca kültüre edildi. Kültür sonucunda embriyoların blastosit aşamasına kadar olan gelişimleri günlük olarak izlenerek kaydedildi. Bu çalışmada istatistik analiz için SPSS paket programında bulunan GLM prosedürü kullanıldı. Sonuç olarak, SOF ve CR1aa medyumlarında kültüre edilen koyun embriyolarının blastosiste ulaşma kapasitesi açısından in vitro üretim medyumlarında sisteamin veya ovidukt hücresi ilavesinin gelişimleri üzerine bir etkisinin olmadığı ve gruplar arasında istatistiksel bir önemin olmadığı tespit edildi

Kültür Medyumlarında Sisteamin ve Ovidukt Hücrelerinin Koyun Embriyo Gelişimi Üzerine Etkisi

Sheep is a very important source of wool, meat and milk all over the world. Oxidative stress during in vitro culture leads to defects in development of gametes and embryos. Several antioxidants such as cysteamine, L-ascorbic acid, beta mercaptoethanol, cysteine, glutathione, proteins, vitamins are used to supplement culture media to counter the oxidative stress. This study was aimed to detect the effect of cysteamine supplementation to the maturation medium and oviductal cell supplementation to culture medium on the subsequent development rates of sheep embryos with the control group. Oocytes were obtained from slaughtered sheep ovaries. Selected oocytes were incubated with or without 100 µM cysteamine in TCM199 medium under 38.5-38.8ºC 5% CO2 for 23 h. During IVF fresh semen was collected from ram by electroejaculation, they were washed in H-SOF medium and were fertilized in B-SOF medium with oocytes incubated for 18 hours under 38.5-38.8ºC 5% CO2, 5% O2 and 90% N2. The oocytes were obtained from maturation medium with/without cysteamine (C+,-) and were cultured in SOF or CR1aa media with/without oviductal cells (Ov+,-) and were grouped as; Group Ia: SOF+(C+ Ov-), Group Ib: SOF+(C+Ov+), Group Ic: SOF+(C- Ov-) Group Id: SOF+(C- Ov+); Group IIa: CR1aa+(C+ Ov-), Group IIb: CR1aa+(C+ Ov+), Group IIc: CR1aa+(C- Ov-), Group IId: CR1aa+(C- Ov+). Embryos were incubated under 38.5-38.8ºC 5% CO2, 5% O2, 90% N2 in culture medium for 7 days. Embryo developments were observed and recorded daily. GLM procedure found in SPSS packet program was used for statistical analysis in this study. In conclusion, the addition of cysteamine or oviductal cells in vitro culture media found to have any effect in terms of the capacity of reaching to blastocyst stage in SOF or CR1aa media and no statistical difference is detected between groups

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