Journal of Embryo Transfer (한국수정란이식학회지)

The Korean Society of Embryo Transfer (한국수정란이식학회)
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Apoptosis-related Genes Altered in Bovine Cystic Ovary

난소낭종 시 변화되는 세포사멸 관련 유전자

  • Tak, Hyun-Min (Department of Physiology, School of Medicine and Institute of Health Sciences, Gyeongsang National University) ;
  • Kim, Gyu-Tae (Department of Physiology, School of Medicine and Institute of Health Sciences, Gyeongsang National University) ;
  • Kim, Eun-Jin (Department of Physiology, School of Medicine and Institute of Health Sciences, Gyeongsang National University) ;
  • Mun, Yun-Ja (Department of Physiology, School of Medicine and Institute of Health Sciences, Gyeongsang National University) ;
  • Choe, Chang-Yong (Animal Genetic Resources Station, National Institute of Animal Science RDA) ;
  • Son, Dong-Soo (Animal Genetic Resources Station, National Institute of Animal Science, RDA) ;
  • Han, Jae-Hee (Department of Physiology, School of Medicine and Institute of Health Sciences, Gyeongsang National University) ;
  • Kang, Da-Won (Department of Physiology, School of Medicine and Institute of Health Sciences, Gyeongsang National University)
  • 탁현민 (경상대학교 의학전문대학원 생리학교실) ;
  • 김규태 (경상대학교 의학전문대학원 생리학교실) ;
  • 김은진 (경상대학교 의학전문대학원 생리학교실) ;
  • 문윤자 (경상대학교 의학전문대학원 생리학교실) ;
  • 최창용 (농촌진흥청 국립축산과학원 가축유전자원시험장) ;
  • 손동수 (농촌진흥청 국립축산과학원 가축유전자원시험장) ;
  • 한재희 (경상대학교 의학전문대학원 생리학교실) ;
  • 강다원 (경상대학교 의학전문대학원 생리학교실)
  • Published : 2009.03.31
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Abstract

This study was carried out to investigate expression of apoptosis-related differentially expressed gene (DEG) in ovaries of Korean cattle with follicular and luteal cysts and to identify the relationship between cyst and apoptosis using microarray, real-time PCR, TUNEL staining, and Western blot analysis. Microarray data showed that PIK3R2 and AKT1 were significantly up-regulated in follicular cyst, and TNF-RAF2, PRLR, FOXL2, STK4, and COL4A3 were up-regulated whereas INHA, CIDEB, BCL10, and FASLG were down-regulated in luteal cyst. Real-time PCR was performed to validate DEGs altered in luteal cyst. Of nine DEGs, four DEGs down-regulated in luteal cyst showed a positive corelation between microarray data and real-time PCR data. In this study, we focused on INHA, among many DEGs, which was highly down-regulated in both follicular and luteal cysts. Real-time PCR and micro array data showed that INHA was down-regulated by 12.3-fold and by 1.4-fold, respectively, in the bovine follicular cyst. TUNEL assay and Western blot analysis for ERK, JNK, p38, PI3K, and Akt, which were used to detect whether apoptosis is occurred, showed no significant changes in cystic ovaries (p>0.05). In the expression and activity of caspase-3, Bax, Bel-2, and Bel-xL, there was no significant changes between follicular cystic ovary and normal ovary. Rather, the expression levels of PI3K and p-Akt were decreased in follicular cystic ovary. These results suggest that deficiency of apoptosis in cystic ovary is associated with decreased expression of apoptotic effectors.

본 연구는 한우의 난포낭종 및 황체낭종에서 세포자멸사 관련 유전자의 발현 변화를 조사함으로써 난소낭종과 세포자 멸사간의 관계를 정리하고자 한다. 마이크로어레이 분석 결과, 난포낭종에서 PIK3R2와 AKT1가 유의하게 증가하였고, 황체낭종에서는 증가되는 세포자멸사 관련 DEGs, TNF-RAF2, PRLR, FOXL2, STK4 및 COL4A3와 감소하는 DEGs, INHA, CIDEB, BCL10 및 FASLG가 포함되었다. 정량적 역전사 중합 효소 반응을 이용하여 mRNA 발현 변화를 검증한 결과 INHA, CIDEB, BCL10 및 FASLG만 마이크로어레이와 동일한 결과를 보였다. INHA의 경우 마이크로어레이 결과에서는 1.43배 감소하고 정량적 역전사 중합효소 반응 결과에서는 12.3배 감소하였다. 세포사멸 확인을 위하여 TUNEL 형광염색과 ERK, JNK, p38, PI3K 및 Akt/PKB의 웨스턴 블랏 분석을 실시한 결과 정상난소와의 유의한 차이를 확인할 수 없었다. Caspase의 활성 및 Bax, Bcl-2, Bcl-xL의 단백질 분석 역시 정상난소와 유의한 차이를 보이지 않았다. 오히려 PI3K와 p-Akt의 발현이 정상에 비하여 감소하는 경향을 보였다. 이러한 결과들로 미루어 보아 난소낭종 시 볼 수 있는 세포자멸사의 낮은 발현율은 세포 사멸인자의 발현 감소로 나타날 수 있을 것이라 생각된다.

Keywords

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