ERCAN KURAR, Mehmet Osman ATLI, Aydın GÜZELOĞLU, Ahmet SEMACAN

POMC, iNOS, PGES, IL-4, IL-5 and IL-10 gene expression in peripheral blood mononuclear cells of cyclic and pregnant mares

Kısrakların erken gebelik tanısında farklı klinik yöntemler ve laboratuvar analizleri kullanılmaktadır. Ancak pratik, güvenilir ve ekonomik bir gebelik tanı yöntemine halen ihtiyaç bulunmaktadır. Bu çalışmada, gebelikle birlikte immun sistemde meydana gelen degişimlerin moleküler düzeyde belirlenmesi, gebelik immunolojisinin anlaşılması ve sonuçta pratik bir gebelik testi geliştirilmesi için bilimsel alt yapının hazırlanması amaçlanmıştır. Kan örnekleri, 10 baş gebe ve 4 baş gebe olmayan (siklik) kısraktan ovulasyon gününden başlayarak (0. gün), 4. ve 8. günlerde alındı. Periferal kan mononükleer hücrelerinden (PKMH) total RNA izolasyonu ve cDNA sentezi gerçekleştirildi. Proopiomelanocortin (POMC), nitrik oksit sentaz (iNOS), prostaglandin E sentaz (PGES), interlökin-4 (IL-4), IL-5 ve IL-10 gen ekspresyonlarında gebeliğe bağlı muhtemel değişiklikler mRNA seviyesinde Reverz Transkriptaz-Polimeraz Zincir Reaksiyonu (PZR) ile semi-kantitatif olarak araştırıldı. İstatistiksel analizlerde General Linear Model ve modeldeki bütün etkenler için ortalama değerler (means) arasındaki farklılık olasılıkları Tukey analizi kullanılarak belirlendi. PKMH’de PGES, IL-5 ve iNOS gen ekspresyonları tespit edilemedi. İstatistiksel analizler sonucunda POMC, IL-4 ve IL-10 gen ekspresyonlarının 0., 4. ve 8. günler arasında mRNA düzeyinde herhangi bir farklılık göstermediği belirlendi. Gebeliğin veya siklik olmanın bu genlerin mRNA ekspresyonu üzerine uyarma veya baskılama şeklinde bir etkisi olmadığı tespit edildi. Yalnızca, IL-10 mRNA konsantrasyonunun gebe hayvanlarda siklik hayvanlara göre düşük olduğu belirlendi (P

Anahtar Kelimeler:

dişi hayvanlar, gen ifadesi, at, kısrak, polimeraz zincir reaksiyonu, gebelik

Siklik ve gebe kısrak periferal kan mononükleer hücrelerinde POMC, iNOS, PGES, IL-4, IL-5 ve IL-10 gen ekspresyonu

Although a number of diff erent clinical and laboratory methods are used, there is still need for a practical, reliable and economical diagnosis method for early pregnancy in mares. Objective was to provide a preliminary background for understanding immunological modifications at molecular level during early pregnancy in mare and thereby to allow development of a practical pregnancy test. Blood samples were collected from 10 pregnant and 4 cyclic mares on days of ovulation (d0), 4 and 8. Total RNA samples were isolated from peripheral blood mononuclear cells (PBMC) and cDNA synthesis was performed. Pregnancy dependent gene expression profiles of proopiomelanocortin (POMC), nitric oxide synthase (iNOS), prostaglandin E synthase (PGES), interleukin-4 (IL-4), IL-5 and IL-10 were evaluated at mRNA level using semiquantitative Reverse Transcriptase-Polymerase Chain Reaction. Data were analyzed by General Linear Model and possible diff erences between all mean factors were determined by Tukey’s analysis. No gene expressions were observed for PGES, IL-5 and iNOS in PMBCs. Expressions of POMC, IL-4 and IL-10 were not significant on d0, 4 and 8, which suggested that pregnancy or cyclic status had no eff ect on expression of these genes. Only, IL-10 mRNA expression was lower in pregnant mares than cyclic mares (P<0.05).

Keywords:

female animals, gene expression, horses, mares, polymerase chain reaction, pregnancy,

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