Investigation of the Distribution and Expression Level of Pro and Anti-Apoptotic Bax and Bcl-2 in Ovarian Follicles at Different Developmental Stages

Öz

Background: In this study we aimed to investigate the distribution and expression level of pro and anti-apoptotic proteins, Bax and Bcl-2, in different developmental stages of ovarian follicles and any relations between these proteins and follicle atresia. Materials and Methods: For that purpose, bilateral 16 ovaries of adult 8 mice were received and the tissues were fixed in 10% neutral buffered formalin. Routine tissue processing protocol was performed and the samples were embedded into paraffin blocks. Five µm thick sections were received and the tissue sections were stained with Bax and Bcl-2 immunohistochemistry. The ovarian follicles were classified as primordial, primary, secondary and antral. Distribution and expression levels of Bax and Bcl-2 were evaluated among and within the developmental stages. The expression levels of Bax and Bcl-2 were also compared with atretic follicle ratio. Results: Immunopositivity of Bax and Bcl-2 were observed in ovarian stromal cells, granulosa, oocytes, and lutheal cells in a varying range. Despite of some immunpositivity, most of the primordial and primary follicle granulosa cells and oocytes were negative for these apoptosis regulator proteins. The intensity of immunopositivity increased at the farther developmental process in follicles. In addition, the immunoexpression level significantly increased just with the beginning of the secondary follicular stage and the expression levels were the most intense in antral follicles. Furthermore, some of the antral follicles were intense Bax positive which were observed with atretic follicle morphology. Conclusions: Bax and Bcl-2 are crucial regulators of ovarian follicle development. Although Bcl-2 contributes on development, correlation analyses indicated that Bax decides stronger than Bcl-2 on the atresia or development fate of follicle.

Anahtar Kelimeler

• Bax
• Bcl-2
• Ovary
• Follicle
• Atresia

Farklı Gelişim Dönemlerindeki Ovaryum Foliküllerinde Pro ve Anti-Apoptotik Bax ve Bcl-2'nin Dağılım ve Ekspresyon Düzeyinin İncelenmesi

Öz

Amaç: Bu çalışmada, farklı gelişimsel evredeki ovaryum foliküllerinde Bax ve Bcl-2 ekspresyon düzeylerini ve bu proteinlerle folikül atrezisi arasındaki ilişkiyi araştırmayı amaçladı. Materyal ve Metod: Bu amaçla 8 fareye ait bilateral toplam 16 ovaryum toplandı ve dokular 10%’luk tamponlanmış formalin içerisinde fikse edildi. Dokulara rutin doku takibi protokolü uygulandı ve örnekler erimiş parafine gömülerek bloklandı. Parafin bloklara gömülü doku örneklerinden alınan 5 µm kalınlığındaki kesitler Bax ve Bcl-2 immunohistokimya boyandı. Ovaryum folikülleri primordiyal, primer, seconder ve antral şeklinde sınıflandırılıd. Gelişim aşamalarının içinde ve aşamalar arasındaki Bax ve Bcl-2 dağılımı ve ekspresyon düzeyi değerlendirildi. Ekspresyon düzeyleri aynı zamanda folikül atrezisi ile kıyaslandı. Bulgular: Ovaryum stroma hücrelerinde, granuloza, oosit ve luteal hücrelerde Bax ve Bcl-2 immunopozitivitesi değişen oranlarda izlendi. Yer yer gözlenen immunopozitiviteye karşın, primordiyal ve primer folikül granuloza hücreleri ve oositleri büyük bir oranda apoptozis düzenleyici proteinler yönünden negatifti. İmmunopozitivite yoğunluğu ileri gelişim dönemlerinde artış eğilimindeydi. Ayrıca, immunoekspresyon düzeyi sekonder foliküler aşamadan sonra önemli bir artış gösteriyordu ve antral foliküllerde en yüksek düzeyde ekspresyon yoğunluğu izlendi. Dahası bazı antral foliküllerdeki yoğun Bax pozitifliğine atretik folikül morfolojisi eşlik etmekteydi. Sonuç: Bax ve Bcl-2 ovaryum folikül gelişiminin önemli düzenleyicileridir. Her ne kadar Bcl-2 gelişim sürecinde etkili olsa da, korelasyon analizleri Bax’ın folikül atrezi veya gelişim kararını vermede Bcl-2’den daha güçlü olduğunu göstermiştir.

Anahtar Kelimeler

• Bax
• Bcl-2
• Ovaryum
• Folikül
• Atrezi

Kaynakça

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