CAJ | 학술논문

[目的]研究5,7,4'-三羟基异黄酮(genistein,GEN)对日本比目鱼(牙鲆)生殖腺性分化的作用,并在基因表达上探索其分子机制.[方法]运用日本比目鱼随饲养水温决定其性别的生物学特性,在鱼卵孵化后第30天到第100天性分化敏感期内,在27 ℃水温下饲养,会使幼鱼分化成雄性.同时,分别给予幼鱼投食GEN 10和100 μg/g饲料进行喂养染毒,直到第100天(青年期)和第300天(成年期),分别对各组比目鱼样本做病理组织切片,判断其性别和雌雄比例,并通过原位杂交和RT-PCR分析,探索它们的作用机制.[结果]GEN对27 ℃水温饲养下的比目鱼幼鱼有雌性化诱导作用,两组成年比目鱼雌性分别有不同的百分比:10μg/g组为46.7%,100μg/g组为96.7%,其诱导效应呈现一定剂量依赖性;比目鱼幼鱼出生后第100天时原位杂交实验显示,P450芳香化酶的mRNA在各组的雌鱼卵巢中有明显表达,但在雄鱼精巢中未见表达;而MIS(苗勒氏管抑制物质)的mRNA在各组的雌鱼卵巢中不表达,而在雄鱼精巢中能见到非常明显的表达.RT-PCR实验显示,在雌鱼卵巢中P450芳香化酶的mRNA表达被诱导,而MIS基因表达被抑制,进一步证实了原位杂交的结果.[结论]GEN对日本比目鱼的生殖腺性分化有雌性化诱导作用,其分子机制很可能是通过诱导P450芳香化酶基因表达和抑制MIS的基因表达从而发挥雌激素样干扰作用的.
[목적]연구5,7,4'-삼간기이황동(genistein,GEN)대일본비목어(아평)생식선성분화적작용,병재기인표체상탐색기분자궤제.[방법]운용일본비목어수사양수온결정기성별적생물학특성,재어란부화후제30천도제100천성분화민감기내,재27 ℃수온하사양,회사유어분화성웅성.동시,분별급여유어투식GEN 10화100 μg/g사료진행위양염독,직도제100천(청년기)화제300천(성년기),분별대각조비목어양본주병리조직절편,판단기성별화자웅비례,병통과원위잡교화RT-PCR분석,탐색타문적작용궤제.[결과]GEN대27 ℃수온사양하적비목어유어유자성화유도작용,량조성년비목어자성분별유불동적백분비:10μg/g조위46.7%,100μg/g조위96.7%,기유도효응정현일정제량의뢰성;비목어유어출생후제100천시원위잡교실험현시,P450방향화매적mRNA재각조적자어란소중유명현표체,단재웅어정소중미견표체;이MIS(묘륵씨관억제물질)적mRNA재각조적자어란소중불표체,이재웅어정소중능견도비상명현적표체.RT-PCR실험현시,재자어란소중P450방향화매적mRNA표체피유도,이MIS기인표체피억제,진일보증실료원위잡교적결과.[결론]GEN대일본비목어적생식선성분화유자성화유도작용,기분자궤제흔가능시통과유도P450방향화매기인표체화억제MIS적기인표체종이발휘자격소양간우작용적.
[ Objective ] To investigate the estrogenic effects of genistein(GEN) on the gonadal sex differentiation in Japanese flounder, a teleost that exhibits temperature-dependent sex determination. [ Methods ] Treatment with GEN was carried out by providing the flounder larvae with an artificial diet mixed with 10 μg/g and 100μg/g diet of GEN at a masculinizing temperature (27 ℃ ) from 30th to 100th day after hatching(dah). Phenotypic sex of the fish was determined at 100th (juvenile stage)inhibiting substance) mRNAs expression in the genetically female flounder treated with GEN were examined by in situ hybridization and RT-PCR analyses. [ Results ] The results showed that treatment with 10 μg/g and 100 tg/g diet of GEN dose- dependently induced feminization of the flounder larvae reared at 27 ℃ with female percentages of 46.7% and 96.7%, respectively. The result of in situ hybridization and RT-PCR analyses showed that feminization of the larvae by GEN was accompanied by up-regulation of P450arom mRNA expression and down-regulation of MIS mRNA expression in female gonad at 100th day after hatching.[ Conclusion ] GEN has estrogenic action on gonadal sex differentiation in Japanese flounder and its mechanism may be due to the induction of P450arom mRNA expression and the suppression of MIS mRNA expression.