School of Biomedical Sciences, the Chinese University of Hong Kong, Shatin, New Territories, Hong Kong, China.
Biol Reprod. 2011 Mar;84(3):476-86. doi: 10.1095/biolreprod.110.086363. Epub 2010 Nov 17.
The gonad-specific expression of a recently discovered Igf subtype (Igf3) in teleost has indicated the important role of this novel Igf in the reproductive functions of fish. In the present study using zebrafish as the model organism, we have further examined the gene expression patterns and the physiological role of Igf3 in the ovary. The igf3 gene in zebrafish was found to be alternatively spliced into two transcripts, with transcript variant 1 exclusively expressed in the gonads, and transcript variant 2 only expressed during early development. Using specific antibodies developed for zebrafish Igf3, both the Igf3 prepropeptide and the mature peptide forms of Igf3 were found to be predominantly expressed in the zebrafish ovary. Real-time PCR and in situ hybridization revealed that igf3 mRNA is relatively low in the early follicles, but is significantly increased after the mid-vitellogenic stage (midstage III), and is high in the full-grown follicles. In the full-grown follicles, igf3 mRNA was detected mainly in the somatic follicular cells with a low level of expression in the oocytes. Igf3 immunoreactivity was confined to the follicular cells only. The expression of igf3 was significantly up-regulated in both ovarian fragments and isolated follicles upon treatment with human chorionic gonadotropin in dose- and time-dependent manners. Treatment with 8-bromoadenosine 3',5'-cyclic monophosphate or 3-isobutyl-1-methylxanthine also up-regulated the expression of igf3 in full-grown follicles. Incubation of follicles with recombinant zebrafish Igf3 significantly enhanced oocyte maturation in time-, dose-, and stage-dependent manners. The actions of Igf3 could be blocked by cycloheximide, but not by actinomycin D. Taken together, these results support an important role of Igf3 in the ovarian functions of zebrafish, especially in oocyte maturation.
最近发现的一种 Igf 亚型（Igf3）在硬骨鱼中的性腺特异性表达表明，这种新型 Igf 在鱼类生殖功能中具有重要作用。本研究以斑马鱼为模型生物，进一步研究了 Igf3 在卵巢中的基因表达模式和生理作用。发现斑马鱼的 igf3 基因被剪接为两种转录本，转录变体 1 仅在性腺中表达，转录变体 2 仅在早期发育过程中表达。使用针对斑马鱼 Igf3 开发的特异性抗体，发现 Igf3 前肽和成熟肽形式的 Igf3 主要在斑马鱼卵巢中表达。实时 PCR 和原位杂交显示，igf3 mRNA 在早期卵泡中相对较低，但在中卵黄生成期（中期 III）后显著增加，并在完全成熟的卵泡中升高。在完全成熟的卵泡中，igf3 mRNA 主要在体细胞卵泡细胞中检测到，卵母细胞中的表达水平较低。Igf3 免疫反应性仅局限于卵泡细胞。卵巢片段和分离的卵泡在人绒毛膜促性腺激素处理下，igf3 的表达呈剂量和时间依赖性显著上调。8-溴腺苷 3'，5'-环单磷酸或 3-异丁基-1-甲基黄嘌呤处理也可上调完全成熟卵泡中 igf3 的表达。用重组斑马鱼 Igf3 孵育卵泡可显著增强卵母细胞成熟，具有时间、剂量和阶段依赖性。Igf3 的作用可被环己酰亚胺阻断，但不能被放线菌素 D 阻断。综上所述，这些结果支持 Igf3 在斑马鱼卵巢功能中的重要作用，尤其是在卵母细胞成熟中。
