Barboni B, Turriani M, Galeati G, Spinaci M, Bacci M L, Forni M, Mattioli M
Dipartimento di Scienze Veterinarie ed Agroalimentari, Fisiologia Veterinaria, Università di Teramo, 64020 Italy.
Biol Reprod. 2000 Sep;63(3):858-64. doi: 10.1095/biolreprod63.3.858.
Angiogenesis is the process that drives blood vessel development in growing tissues in response to the local production of angiogenic factors. With the present research the authors have studied vascular endothelial growth factor (VEGF) production in ovarian follicles as a potential mechanism of ovarian activity regulation. Prepubertal gilts were treated with 1250 IU equine chorionic gonadotropin (eCG) followed 60 h later by 750 IU of human chorionic gonadotropin (hCG) in order to induce follicle growth and ovulation. Ovaries were collected at different times of the treatment and single follicles were isolated and classified according to their diameter as small (<4 mm), medium (4-5 mm), or large (>5 mm). VEGF levels were measured in follicular fluid by enzyme immunoassay, and VEGF mRNA content was evaluated in isolated theca and granulosa compartments. Equine chorionic gonadotropin stimulated a prompt follicular growth and induced a parallel evident rise in VEGF levels in follicular fluid of medium and large follicles. Analysis of VEGF mRNA levels confirmed the stimulatory effect of eCG, showing that it is confined to granulosa cells, whereas theca cells maintained their VEGF steady state mRNA. Administration of hCG 60 h after eCG caused a dramatic drop in follicular fluid VEGF that reached undetectable levels in 36 h. A parallel reduction in VEGF mRNA expression was recorded in granulosa cells. The stimulating effect of eCG was also confirmed by in vitro experiments, provided that follicles in toto were used, whereas isolated follicle cells did not respond to this hormonal stimulation. Consistent with the observation in vivo, granulosa cells in culture reacted to hCG with a clear block of VEGF production. These results demonstrate that while follicles of untreated animals produce stable and low levels of the angiogenic factor, VEGF markedly rose in medium and large follicles after eCG administration. The increasing levels, essentially attributable to granulosa cells, are likely to be involved in blood vessel development in the wall of growing follicles, and may play a local key role in gonadotropin-induced follicle development. When ovulation approaches, under the effect of hCG, the production of VEGF is switched off, probably creating the safest conditions for the rupture of the follicle wall while theca cells maintained unaltered angiogenic activity, which is probably required for corpus luteum development.
血管生成是一个过程，它驱动生长组织中的血管发育，以响应血管生成因子的局部产生。在本研究中，作者研究了卵巢卵泡中血管内皮生长因子（VEGF）的产生，作为卵巢活动调节的一种潜在机制。对青春期前的后备母猪用1250国际单位的马绒毛膜促性腺激素（eCG）进行处理，60小时后再给予750国际单位的人绒毛膜促性腺激素（hCG），以诱导卵泡生长和排卵。在处理的不同时间收集卵巢，分离单个卵泡，并根据其直径分为小卵泡（<4毫米）、中卵泡（4 - 5毫米）或大卵泡（>5毫米）。通过酶免疫测定法测量卵泡液中的VEGF水平，并在分离的卵泡膜和颗粒细胞区室中评估VEGF mRNA含量。马绒毛膜促性腺激素刺激卵泡迅速生长，并导致中卵泡和大卵泡的卵泡液中VEGF水平明显平行升高。对VEGF mRNA水平的分析证实了eCG的刺激作用，表明这种作用局限于颗粒细胞，而卵泡膜细胞维持其VEGF稳态mRNA。在eCG后60小时给予hCG导致卵泡液VEGF急剧下降，在36小时内降至检测不到的水平。在颗粒细胞中记录到VEGF mRNA表达的平行降低。eCG的刺激作用也通过体外实验得到证实，前提是使用整个卵泡，而分离的卵泡细胞对这种激素刺激没有反应。与体内观察结果一致，培养中的颗粒细胞对hCG反应，VEGF产生明显受阻。这些结果表明，未处理动物的卵泡产生稳定且低水平的血管生成因子，而在给予eCG后，中卵泡和大卵泡中的VEGF显著升高。增加的水平主要归因于颗粒细胞，可能参与生长卵泡壁的血管发育，并可能在促性腺激素诱导的卵泡发育中起局部关键作用。当排卵临近时，在hCG的作用下，VEGF的产生被关闭，这可能为卵泡壁破裂创造最安全的条件，而卵泡膜细胞维持未改变的血管生成活性，这可能是黄体发育所必需的。
