본 연구에서는 MAPK 저해제인 U0126이 난자성숙과정에서 특히 감수분열, 미세소관 형성 그리고 액틴 필 라먼트 형성에 미치는 영향을 조사하였다. 그 결과 MAPK 단백질은 12시간째에 인산화되기 시작하여, 24시간째에 대부분 인산화 되었고 metaphase II에 이르기 까지 유지되었다. 배포단계(GV)에 있는 난자를 U0126의 $20{\mu}M$ 농도로 처리하였을 때 MAPK의 인산화가 완전히 억제되었으나 배포의 파열 단계(GVBD)로의 성숙에는 진행하였으나, metaphase I까지는 발달하지 못하였다. 또한 MAPK 저해제로 인해 비정상적인 방추사의 형성을 초래하였다. 난자를 배포의 파열단계(GVBD) 이후에 U0126을 처리하였을 때 극체의 방출은 정상 이였으나 중기 판의 배열과 염색체의 분열은 비정상적 이였다. 결론적으로, 유사분열 활성화 효소단백질인 MAPK의 활성은 돼지 난자의 체외성숙과정에서 배포단계(GV)의 염색체의 배열과 감수분열의 완성에 중요한 조절 인자임을 이번 연구를 통해 알 수 있었다.
본 연구에서는 MAPK 저해제인 U0126이 난자성숙과정에서 특히 감수분열, 미세소관 형성 그리고 액틴 필 라먼트 형성에 미치는 영향을 조사하였다. 그 결과 MAPK 단백질은 12시간째에 인산화되기 시작하여, 24시간째에 대부분 인산화 되었고 metaphase II에 이르기 까지 유지되었다. 배포단계(GV)에 있는 난자를 U0126의 $20{\mu}M$ 농도로 처리하였을 때 MAPK의 인산화가 완전히 억제되었으나 배포의 파열 단계(GVBD)로의 성숙에는 진행하였으나, metaphase I까지는 발달하지 못하였다. 또한 MAPK 저해제로 인해 비정상적인 방추사의 형성을 초래하였다. 난자를 배포의 파열단계(GVBD) 이후에 U0126을 처리하였을 때 극체의 방출은 정상 이였으나 중기 판의 배열과 염색체의 분열은 비정상적 이였다. 결론적으로, 유사분열 활성화 효소단백질인 MAPK의 활성은 돼지 난자의 체외성숙과정에서 배포단계(GV)의 염색체의 배열과 감수분열의 완성에 중요한 조절 인자임을 이번 연구를 통해 알 수 있었다.
In this study, we determined effects of the mitogen-activated protein kinase (MAPK) inhibitor, U0126 on meiotic maturation, microtubule organization and actin filament assembly in the porcine oocyte. The phosphorylated MAPK was first detected at 12 h after the initiation of maturation cultures, full...
In this study, we determined effects of the mitogen-activated protein kinase (MAPK) inhibitor, U0126 on meiotic maturation, microtubule organization and actin filament assembly in the porcine oocyte. The phosphorylated MAPK was first detected at 12 h after the initiation of maturation cultures, fully activated at 24h, and remained until metaphase II. Treatment of germinal vesicle (GV) stage oocytes with $20{\mu}M$ U0126 completely blocked MAPK phosphorylation, but germinal vesicle breakdown (GVBD) was normally proceeded. However, the oocytes didn‘t progress to the metaphase I. The inhibition of MAPK resulted in abnormal spindles. In oocytes treated with U0126 after GVBD, polar body extrusion was normal, but the organization of the metaphase plate and chromosome segregation were abnormal. In conclusion, MAPK activity plays an important regulatory role in GV chromatin configuration and meiotic progress in porcine oocyte maturation.
In this study, we determined effects of the mitogen-activated protein kinase (MAPK) inhibitor, U0126 on meiotic maturation, microtubule organization and actin filament assembly in the porcine oocyte. The phosphorylated MAPK was first detected at 12 h after the initiation of maturation cultures, fully activated at 24h, and remained until metaphase II. Treatment of germinal vesicle (GV) stage oocytes with $20{\mu}M$ U0126 completely blocked MAPK phosphorylation, but germinal vesicle breakdown (GVBD) was normally proceeded. However, the oocytes didn‘t progress to the metaphase I. The inhibition of MAPK resulted in abnormal spindles. In oocytes treated with U0126 after GVBD, polar body extrusion was normal, but the organization of the metaphase plate and chromosome segregation were abnormal. In conclusion, MAPK activity plays an important regulatory role in GV chromatin configuration and meiotic progress in porcine oocyte maturation.
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대상 데이터
COCs with intact unexpended cumulus cells were isolated from cellular debris and rinsed three times in HEPES. These experiments use a basic culture medium consisting of TCM199 medium supplemented with 0.4% (w/v) BSA, 10 IU EGF, 10 IU hCG, and 10 IU PMSG. The effects of U0126 (Cell Signaling Technology, Beverly, MA) on IVM of pig oocytes were examined as follows.
성능/효과
Our results, which are consistent with previous observations [9]. showed that MAPK activation occurred after GVBD, indicating that MAPK is not required for GVBD but is involved in regulating post-GVBD events .
In conclusion, the meiotic abnormalities caused by U0126, a specific inhibitor of MEK, indicate that the MEK/MAPK pathway is an important regulator of microtubule and centrosome assembly, but not actin filament assembly.
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