School of Paediatrics and Reproductive Health, Robinson Institute, Research Centre for Reproductive Health, Discipline of Obstetrics and Gynaecology, University of Adelaide, Adelaide, South Australia 5005, Australia.
Reproduction. 2011 Nov;142(5):647-57. doi: 10.1530/REP-11-0196. Epub 2011 Sep 6.
It is widely held that mammalian cumulus cell (CC) expansion requires oocyte-paracrine signalling, however in three of the four species studied to date, CC expansion occurs in the absence of the oocyte. This study was conducted to examine the paracrine and SMAD/MAPK intracellular signalling mechanism mediating porcine CC expansion, and to compare these to the mouse. Cumulus-oocyte complexes (COCs) and oocyte-free complexes (OOXs) from pigs and eCG-primed mice were treated in vitro with FSH and a broad range of TGFB superfamily antagonists. Expansion of porcine COCs and OOXs was unaffected by neutralisation of growth differentiation factor 9, TGFB, activin A, activin B and a broad spectrum bone morphogenetic protein antagonist. A SMAD-responsive luciferase reporter assay confirmed that porcine oocytes secreted factors that activate SMAD3 and SMAD1/5/8 in granulosa cells, but murine oocytes activated SMAD3 only. Treatment of COCs with a SMAD2/3 phosphorylation inhibitor (SB431542) partially inhibited porcine CC expansion and expression of TNFAIP6, but ablated murine CC expansion. SB431542 was equally effective at attenuating porcine CC expansion in the presence or absence of the oocyte. By contrast, a SMAD1/5/8 phosphorylation inhibitor (dorsomorphin) had no effect on porcine or murine CC function. Inhibition of ERK1/2 and p38 MAPK signalling pathways prevented porcine COC expansion and expression of most matrix genes examined. The activation of CC SMAD signalling by oocytes, and the requirement of SMAD2/3 signalling for expansion, is notably contrasted in pigs and mice. Nonetheless, porcine CC SMAD2/3 signalling is likely to be needed for optimal matrix formation, possibly by facilitating essential MAPK signals.
人们普遍认为哺乳动物卵丘细胞（CC）的扩展需要卵母细胞旁分泌信号，但迄今为止在研究的四个物种中的三个中，CC 的扩展发生在没有卵母细胞的情况下。本研究旨在研究介导猪 CC 扩展的旁分泌和 SMAD/MAPK 细胞内信号机制，并将其与小鼠进行比较。来自猪和 eCG 诱导的小鼠的卵丘 - 卵母细胞复合物（COC）和卵母细胞 - 自由复合物（OOX）在体外用 FSH 和广泛的 TGFB 超家族拮抗剂处理。猪 COC 和 OOX 的扩展不受生长分化因子 9、TGFB、激活素 A、激活素 B 和广谱骨形态发生蛋白拮抗剂的中和影响。SMAD 反应性荧光素酶报告基因测定证实，猪卵母细胞分泌的因子可激活颗粒细胞中的 SMAD3 和 SMAD1/5/8，但鼠卵母细胞仅激活 SMAD3。用 SMAD2/3 磷酸化抑制剂（SB431542）处理 COC 可部分抑制猪 CC 的扩展和 TNFAIP6 的表达，但使鼠 CC 的扩展消失。SB431542 在存在或不存在卵母细胞的情况下对猪 CC 扩展同样有效。相比之下，SMAD1/5/8 磷酸化抑制剂（dorsomorphin）对猪或鼠 CC 功能没有影响。ERK1/2 和 p38 MAPK 信号通路的抑制阻止了猪 COC 的扩展和大多数基质基因的表达。卵母细胞对 CC SMAD 信号的激活以及 SMAD2/3 信号对扩展的要求，在猪和小鼠中形成鲜明对比。尽管如此，猪 CC SMAD2/3 信号可能需要最佳的基质形成，可能通过促进必要的 MAPK 信号。
