Division of Reproductive Sciences, Oregon National Primate Research Center, OHSU West Campus, 505 NW 185th Ave, Beaverton, OR 97006, USA.
Mol Hum Reprod. 2011 Mar;17(3):152-65. doi: 10.1093/molehr/gaq089. Epub 2010 Oct 29.
Experiments were designed to evaluate changes in the transcriptome (mRNA levels) in the ovulatory, luteinizing follicle of rhesus monkeys, using a controlled ovulation model that permits analysis of the naturally selected, dominant follicle at specific intervals (0, 12, 24 and 36 h) after exposure to an ovulatory (exogenous hCG) stimulus during the menstrual cycle. Total RNA was prepared from individual follicles (n= 4-8/timepoint), with an aliquot used for microarray analysis (Affymetrix Rhesus Macaque Genome Array) and the remainder applied to quantitative real-time PCR (q-PCR) assays. The microarray data from individual samples distinctly clustered according to timepoints, and ovulated follicles displayed markedly different expression patterns from unruptured follicles at 36 h. Between timepoint comparisons revealed profound changes in mRNA expression profiles. The dynamic pattern of mRNA expression for steroidogenic enzymes (CYP17A, CYP19A, HSD3B2, HSD11B1 and HSD11B2), steroidogenic acute regulatory protein (StAR) and gonadotrophin receptors [LH/choriogonadotrophin receptor (LHCGR), FSH receptor (FSHR)] as determined by microarray analysis correlated precisely with those from blinded q-PCR assays. Patterns of mRNA expression for epidermal-growth-factor-like factors (amphiregulin, epiregulin) and processes [hyaluronan synthase 2 (HAS2), tumor necrosis factor alpha-induced protein 6 (TNFAIP6)] implicated in cumulus-oocyte maturation/expansion were also comparable between assays. Thus, several mRNAs displayed the expected expression pattern for purported theca (e.g. CYP17A), granulosa (CYP19A, FSHR), cumulus (HAS2, TNFAIP6) cell and surface epithelium (HSD11B)-related genes in the rodent/primate pre-ovulatory follicle. This database will be of great value in analyzing molecular and cellular pathways associated with periovulatory events in the primate follicle (e.g. follicle rupture, luteinization, inflammatory response and angiogenesis), and for identifying novel gene products controlling mammalian fertility.
实验旨在评估恒河猴排卵、黄体化卵泡中转录组（mRNA 水平）的变化，采用控制性排卵模型，允许在月经周期中接触排卵（外源性 hCG）刺激后特定时间间隔（0、12、24 和 36 小时）分析自然选择的优势卵泡。从单个卵泡（n=4-8/时间点）中提取总 RNA，其中一部分用于微阵列分析（Affymetrix 恒河猴基因组微阵列），其余部分用于定量实时 PCR（q-PCR）检测。单个样本的微阵列数据根据时间点明显聚类，排卵卵泡在 36 小时与未破裂卵泡显示出明显不同的表达模式。时间点比较显示 mRNA 表达谱发生深刻变化。类固醇生成酶（CYP17A、CYP19A、HSD3B2、HSD11B1 和 HSD11B2）、类固醇急性调节蛋白（StAR）和促性腺激素受体[黄体生成素/绒毛膜促性腺激素受体（LHCGR）、卵泡刺激素受体（FSHR）]的 mRNA 表达动态模式通过微阵列分析确定与盲法 q-PCR 检测结果完全吻合。与卵丘卵母细胞成熟/扩张相关的表皮生长因子样因子（amphiregulin、epiregulin）和过程[透明质酸合酶 2（HAS2）、肿瘤坏死因子-α诱导蛋白 6（TNFAIP6）]的 mRNA 表达模式在两种检测方法之间也具有可比性。因此，几种 mRNA 显示了预期的表达模式，推测为老鼠/灵长类动物排卵前卵泡中的膜细胞（例如 CYP17A）、颗粒细胞（CYP19A、FSHR）、卵丘细胞（HAS2、TNFAIP6）和表面上皮（HSD11B）相关基因。该数据库将在分析与灵长类动物卵泡排卵相关的分子和细胞途径（例如卵泡破裂、黄体化、炎症反应和血管生成）以及鉴定控制哺乳动物生育的新基因产物方面具有重要价值。
