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아미노산의 첨가가 돼지 체외수정란의 후기배의 발달에 미치는 영향
김연수,송상현,조성근,곽대오,김철욱,방충생,정기화,Kim Y. S.,Song S. H.,Cho S. K.,Kwack D. O.,Kim C. W.,Park C. S.,Chung K. H. 한국동물번식학회 2005 Reproductive & developmental biology Vol.29 No.3
본 연구는 아미노산의 첨가가 돼지 수정란의 체외 발달율에 미치는 영향을 구명하고자 PEF가 함유된 NCSU-23을 기본배지로 체외성숙 및 체외배양액을 조성한 후 EA(Essential amino acid), NA(Non-essential amino acid) 및 EANA(EA+ NA)를 첨가하여 체외성숙, 체외수정 및 체외발달에 미치는 영향을 조사하였다. 체외성숙 배지에 아미노산을 첨가한 결과 MH 단계까지의 체외성숙율은 NA 첨가군이 $83.3\%$로 대조구 $70.0\%$에 비하여 유의적으로(p<0.05) 높았다. 그러나 체외 수정 이후의 배 발달율과 수정율에서는 아미노산 첨가군과 무첨가군 사이에 유의적인 차이는 없었다. 체외배양액에 아미노산을 첨가한 후 배반포의 내부세포괴(ICM) 세포와 영양배엽(TE) 세포의 발달에 미치는 영향을 조사한 결과, ICM에서는 유의차를 발견할 수 없었으나 TE 세포는 EANA 처리구가 $18.0{\pm}0.5$개로 대조구 $16.09{\pm}0.56$개에 비해 유의적(p<0.05)으로 많았다. 총세포수에서도 EANA 처리구가 $50.0{\pm}1.0$개로 대조구 $44.2{\pm}1.1$개보다 유의적(p<0.05)으로 많았다. 이상의 결과를 종합할 때, 돼지의 체외수정란 생상에 있어서 아미노산의 첨가는 배반포로의 발달율에는 영향을 미치지 못하였으나 체외성숙율을 높이고 배반포의 세포수 향상에 도움을 주는 것으로 판단된다. 특히, 영양배엽(W) 세포의 발달율이 높은 것으로 보아 아미노산의 첨가는 돼지수정란의 착상에 도움을 줄 것이라 기대된다. The objective of this study was to investigative the effects of amino acids supplementation on maturation, fertilization and embryo development of pig oocytes. Essential amino acids (EA), non-essential amino acids (NA) or both amino acids (EA + NA) were supple-mented to North Carolina State University (NCSU) 23 medium containing porcine follicular fluid (pFF). When the amino acids were supplemented to the maturation medium, the maturation rates were higher (p<0.05) in the NA group than control ($83.3{\pm}0.04\%\;versus\;70.0{\pm}0.05\%$, but the subsequent cleavage rates and development to morula and blstocyst stage between aminoacid supplement groups and control were not different. The developmental rates to morula and blastocysts stage were not significantly different regardless of amino acid supplementation to culture medium. In addition, supplementation of amino acids did not significantly affect the rate of fertilization and polyspermy. When the amino acids were supplement to culture medium, the number of trophectodermal (TE) cells was significantly (p<0.05) higher in amino acid supplement group than that of control ($18.6{\pm}0.5\;versus\;16.1{\pm}0.6$), whereas the numbers of inner cell mass (ICM) cells were not different among the treaonent groups and control ($29.0{\pm}0.9\~31.5{\pm}1.2$). Total cell number was also significantly (p<0.05) higher in EANA group ($50.0{\pm}1.0$) than that of control group ($44.2{\pm}1.1$). These results indicate that the amino acid supplementation to maturation and culture medium may not significantly stimulate early embryo development, but may improve the TE cell number of blastocyst stage in the pig.
수핵란의 전활성화가 토끼 핵이식 수정란의 체외발달에 미치는 효과
전병균,송상현,정기화,곽대오,이효종,최상용,박충생 경상대학교 유전공학연구소 1997 遺傳工學硏究所報 Vol.16 No.-
To examine the efficiency of nuclear transplantation the influence of electrical preactivation of recipient cytoplasm on the in vitro developmental potentyl in the nuclear transplant rabbit embryos were evaluated. The embryos of 16-cell stage were collected and synchronized to G₁phase of 32-cell stage. The recipient cytoplasms were obtained by removing the first polar body and chromosome mass by non-disruptive microsurgery procedure. The separated G₁phase blastomeres of 32-cell stage were out into the non-preactivated and/or the preactivated recipient cytoplasm by electrical stimulation. After culture until 20h post-hCG injection, the nuclear transplant oocytes were electrofused. The fused nuclear transplant embryos were co-cultured with rabbit oviduct for 120h, the nuclear transplant embryos developed to blastocyst stage were stained with Hoechst 33342 and their blastomere were counted. The electrofision rate was similar to the non-preactivated and preactivated recipient cytoplasm(81.8 and 85.7%, respectively). However, the in vitro developmental rate to blastocyst stage with the non-preactivated recipient cytoplasm(163.7cells), as compared with the preactivated recipient cytoplasm(85.4 cells). These results considered better that non-preactivated oocytes, MⅡ phase oocytes, were used for recipient cytoplasms in the rabbit nuclear transplant procedure.