Latham Keith E, Wigglesworth Karen, McMenamin Malgorzata, Eppig John J
The Fels Institute for Cancer Research and Molecular Biology and Department of Biochemistry, Temple University School of Medicine, Philadelphia, Pennsylvania 19140, USA.
Biol Reprod. 2004 May;70(5):1253-62. doi: 10.1095/biolreprod.103.023937. Epub 2003 Dec 26.
The development of an ovarian follicle requires a complex set of reciprocal interactions between the oocyte and granulosa cells in order for both types of cells to develop properly. These interactions are largely orchestrated by the oocyte via paracrine factors such as growth differentiation factor 9 (GDF9). To examine these interactions further, a study was conducted of the effects of oocytes at different stages of development on proteins synthesized by mouse granulosa cells during the transition of granulosa cells (GCs) from preantral, secondary (2 degrees ) follicles (2 degrees GCs) to mural granulosa cells (3 degrees GCs) of antral tertiary (3 degrees ) follicles. The ability of recombinant GDF9 to mimic the effects of oocytes was also determined. Effects were evaluated by high- resolution, two-dimensional protein gel electrophoresis coupled to computer-assisted, quantitative gel image analysis. Coculture of the 2 degrees GCs with growing oocytes (GOs) from 2 degrees follicles brought about many of the changes in granulosa cell phenotype associated with the 2 degrees to 3 degrees follicle transition. GDF9 likewise brought about many of these changes, but only a subset of GDF9-affected protein spots were also affected by coculture with GOs. Coculture of 2 degrees GCs with the nearly fully grown oocytes (FGOs) from 3 degrees follicles had a reduced effect on 2 degrees GC phenotype, in comparison with coculture with GOs. For some proteins, oocyte coculture or GDF9 treatment appeared to have opposite effects on 2 degrees GCs and 3 degrees GCs. Additional effects of GDF9 and oocytes were seen in cultures of 2 degrees GCs for proteins other than those that differed between untreated control 2 degrees and 3 degrees GCs. These results indicate that GOs and GDF9 can each induce 2 degrees GCs to shift their phenotype toward that of 3 degrees GCs. The ability of the oocyte to produce this effect is diminished with oocyte development. The transition in the GC phenotype promoted by oocytes appears stable because differences in 2 degrees GCs promoted by oocytes and GDF9 were observed in untreated 3 degrees GCs. We conclude that the influence of the oocyte on GCs changes with the progression of their development, and so too does the response of the GCs to the oocyte. Moreover, by acting on the 2 degrees GCs, GOs are able to influence stably the phenotype of 3 degrees GCs. Thus, at or near the 2 degrees to 3 degrees follicle transition, signals from the growing oocyte contribute to the development of the mural GC phenotype.
卵巢卵泡的发育需要卵母细胞和颗粒细胞之间一系列复杂的相互作用，以使这两种细胞都能正常发育。这些相互作用在很大程度上由卵母细胞通过旁分泌因子如生长分化因子9（GDF9）来协调。为了进一步研究这些相互作用，开展了一项研究，观察处于不同发育阶段的卵母细胞对小鼠颗粒细胞在从初级卵泡、次级（2°）卵泡（2°颗粒细胞）向窦状三级（3°）卵泡的壁颗粒细胞（3°颗粒细胞）转变过程中合成蛋白质的影响。还测定了重组GDF9模拟卵母细胞作用的能力。通过高分辨率二维蛋白质凝胶电泳结合计算机辅助定量凝胶图像分析来评估作用效果。将2°颗粒细胞与来自2°卵泡的生长中卵母细胞（GOs）共培养，引发了许多与2°到3°卵泡转变相关的颗粒细胞表型变化。GDF9同样引发了许多这些变化，但与GOs共培养也受影响的GDF9作用的蛋白质斑点只是其中一部分。与GOs共培养相比，将2°颗粒细胞与来自3°卵泡的近完全成熟卵母细胞（FGOs）共培养对2°颗粒细胞表型的影响较小。对于某些蛋白质，卵母细胞共培养或GDF9处理对2°颗粒细胞和3°颗粒细胞似乎有相反的作用。在2°颗粒细胞培养中，除了未处理的2°和3°颗粒细胞之间存在差异的蛋白质外，还观察到了GDF9和卵母细胞的其他作用效果。这些结果表明，GOs和GDF9均可诱导2°颗粒细胞将其表型转变为3°颗粒细胞的表型。随着卵母细胞发育，其产生这种作用的能力会减弱。卵母细胞促进的颗粒细胞表型转变似乎是稳定的，因为在未处理的3°颗粒细胞中观察到了卵母细胞和GDF9促进的2°颗粒细胞之间的差异。我们得出结论，卵母细胞对颗粒细胞的影响随着其发育进程而变化，颗粒细胞对卵母细胞的反应也是如此。此外，通过作用于2°颗粒细胞，GOs能够稳定地影响3°颗粒细胞的表型。因此，在2°到3°卵泡转变时或接近该转变时，生长中卵母细胞发出的信号有助于壁颗粒细胞表型的发育。
