GENE EXPRESSION AND FUNCTION OF CYTOKINE DURING MATURATION AND AFTER FERTILIZATION IN PORCINE OVA

本研究采用逆转录-聚合酶链式反应(RT-PCR)检测了猪卵母细胞体外成熟和受精过程中细胞因子[表皮生长因子(EGF)、碱性成纤维细胞生长因子(BFGF)、胰岛素样生长因子-I(IGF-I)、血小板衍生生长因子-A(PDGF-A)和转化生长因子-α(TGF-α)]及其受体(R)mRNAs(EGF-R、bFGF-R、IGF-I-R、PDGF-Ra)的表达。2)受体酪氨酸激酶抑制剂(染料木素和除草霉素A)对猪卵丘-卵母细胞复合体(COCs)体外卵丘质量扩张(CME)和核成熟的影响；3)体外受精延迟受精对猪体外成熟受精胚胎囊胚发育能力的影响。结果表明，EGF、bFGF、IGF-I和PDGF-A的mRNAs在成熟和受精的各个阶段均可检测到，但在8-16细胞期检测不到。在发育2-细胞期后，才能检测到转化生长因子-α的表达。IGF-I-R和PDGF-Ra mRNAs在成熟、受精和发育的各个阶段均可检测到。EGF-R和bFGF-RmRNAs在胚胎成熟和发育的各个阶段均可检测到，但在受精早期未检测到。在成熟培养液中添加金雀异黄素或去甲氧西林A可显著降低CME和中期II卵母细胞的比例，且呈剂量依赖关系。体外受精延迟是提高胚胎发育率至囊胚期的有效方法。这些结果表明，猪卵在体外成熟和受精后表现出细胞因子及其受体mRNAs的时间和阶段特异性表达模式，诱导CME和卵母细胞向中期II发展的信号转导途径涉及细胞因子敏感过程。

This study was carried out examine, 1) by the reverse transcription-polymerase chain reaction (RT-PCR), whether cytokine mRNAs [epidermal growth factor (EGF), basic fibroblast growth factor (bFGF), insulin-like growth factor-I (IGF-I), platelet derived growth factor-A (PDGF-A) and transforming growth factor-a (TGF-a)] and cytokine-receptor (R) mRNAs (EGF-R,bFGF-R,IGF-I-R,PDGF-Ra) express in porcine ova during maturation and after fertilization in vitro, 2) the effect of inhibitors for receptor-tyrosine kinase (genistein and herbimycin A) on cumulus mass expansion (CME) and nuclear maturation of porcine cumulus-oocytes complexes (COCs) in vitro, and 3) the effect of delayd insemination in vitro on the developmental capacity to blastocyst in porcine embryos derived from in vitro maturation and fertilization. The results are as follows ; EGF,bFGF,IGF-I and PDGF-A mRNAs were detectable at all stages of maturation and fertilization, but were not detected at 8-to 16-cell stages. TGF-a mRNA was only dctectable after 2-cell stage of development. IGF-I-R and PDGF-Ra mRNAs were detectable at all stages of maturation, fertilization and development. EGF-R and bFGF-R mRNAs were detectable at all stages of maturation and development, but were not detected at early stage of fertilization. The addition of genistein or herbimycin A to maturation medium significantly decreased the rates of CME and oocytes with metaphase II in dose-dependent fashion. The delayd insemination in vitro was effective for the increase of rate of embryos developed to the blastocyst stage. These results indicate that porcine ova show the temporal and stage-specific expression patterns of cytokine and its receptor mRNAs during in vitro maturation and after fertilization, and that the signal transduction pathway for induction of CME and the progression of oocytes to metaphase II involve cytokine-sensitive process.