Rone J D, Goodman A L
Department of Gynecology and Obstetrics, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205.
Endocrinology. 1990 Dec;127(6):2821-8. doi: 10.1210/endo-127-6-2821.
Angiogenic activity was detected in media conditioned by ovarian cells from superovulated, pseudopregnant (PMSG/human CG treated) immature Holtzman rats. Media conditioned by cells from luteinized rat ovaries stimulated the directed migration of rabbit endothelial cells or mouse Balb/c3T3 cells, but was not mitogenic to either cell type. That endotheliotropic activity was not associated with a steroid was indicated by the finding that chemoattractant activity was detected in fractions after reversed-phase C18 chromatography, which removes more than 95% of steroids present in the media, and that chemoattractant activity was precipitated by ammonium sulfate and by ethanol. Full chemoattractant activity was recovered after boiling (95 C for 30 min), lyophilization, dialysis, Sephadex G-25 desalting columns, and pH changes from 3-10. After Sephadex G-200 chromatography, chemoattractant activity emerged at elution volumes corresponding to 20,000-30,000 mol wt. Chemoattractant activity was not retained by Concanavalin A-Sepharose or gelatin-Sepharose, and was only partially retained by heparin-agarose. Chemoattractant activity was also partially retained on both cation and anion exchange columns. Our collective findings indicate the presence of a nonsteroidal, heat-stable, pronase-sensitive factor, nominal mol wt of 20,000-30,000, in media conditioned by cells from luteinized rat ovaries; this factor is chemoattractive but not mitogenic to endothelial cells. Ovarian-derived chemoattractant activity appears to be distinct from fibroblast growth factor because it lacked detectable mitogenic activity, and because fibroblast growth factor was not active in our cell migration bioassay. Because stimulation of endothelial cell migration is a key event during angiogenesis, demonstration of an ovarian endotheliotropic chemoattractant is consistent with our hypothesis that angiogenesis factors play a role in the paracrine regulation of ovarian function.
在经超排卵、假孕（孕马血清促性腺激素/人绒毛膜促性腺激素处理）的未成熟霍尔茨曼大鼠卵巢细胞所分泌的培养基中检测到了血管生成活性。来自黄体化大鼠卵巢细胞的培养基能刺激兔内皮细胞或小鼠Balb/c3T3细胞的定向迁移，但对这两种细胞类型均无促有丝分裂作用。反相C18色谱分离后的组分中检测到趋化活性，这表明这种内皮细胞趋化活性与类固醇无关，因为该色谱法可去除培养基中95%以上的类固醇，且趋化活性可被硫酸铵和乙醇沉淀。经煮沸（95℃ 30分钟）、冻干、透析、Sephadex G - 25脱盐柱处理以及pH值从3 - 10变化后，可恢复全部趋化活性。经Sephadex G - 200色谱分离后，趋化活性在对应于20,000 - 30,000分子量的洗脱体积处出现。伴刀豆球蛋白A - Sepharose或明胶 - Sepharose不能保留趋化活性，肝素 - 琼脂糖仅能部分保留趋化活性。阳离子和阴离子交换柱也能部分保留趋化活性。我们的总体研究结果表明，在黄体化大鼠卵巢细胞所分泌的培养基中存在一种非甾体、热稳定、对链霉蛋白酶敏感的因子，其标称分子量为20,000 - 30,000；该因子对内皮细胞具有趋化作用但无促有丝分裂作用。卵巢来源的趋化活性似乎与成纤维细胞生长因子不同，因为它缺乏可检测到的促有丝分裂活性，且成纤维细胞生长因子在我们的细胞迁移生物测定中无活性。由于内皮细胞迁移的刺激是血管生成过程中的关键事件，卵巢内皮细胞趋化因子的证明与我们的假设一致，即血管生成因子在卵巢功能的旁分泌调节中起作用。
