Physiological significance of IGF-binding protein protease in follicular growth of human ovary

来自优势卵泡而非闭锁卵泡的卵泡液含有胰岛素样生长因子（IGF）结合蛋白-4（IGFBP-4）蛋白水解活性。使用颗粒细胞培养研究FSH和IGFs对IGFBP-4蛋白水解活性的影响。蛋白水解活性通过[^<125>I]IGFBP-4与培养基孵育来评估，裂解产物通过放射自显影来分析。当细胞与FSH或IGF孵育时，碘化IGFBP-4被切割成18 kDa的片段，而IGFBP-4在对照培养物中保持完整。几种蛋白酶抑制剂对IGFBP-4降解的抑制表明IGFBP-4降解是由金属丝氨酸蛋白酶诱导的。当细胞与IGF-I类似物LR 3-IGF-I和胰岛素一起孵育时，IGFBP-4的降解没有被刺激，所述胰岛素与IGF-I受体结合但对IGFBP具有较低的亲和力或没有亲和力。向来自未处理的GC培养物的培养基中添加IGFs而不是FSH刺激[^<125>I]IGFBP-4的蛋白水解。类似地，外源性添加的共价交联的[^<125>I]IGF-II-IGFBP-4复合物被蛋白水解，然而，IGFs没有增强复合物的降解。这些结果表明，人颗粒细胞产生IGFBP-4蛋白酶，IGFBP-4对蛋白水解的敏感性增加是由IGFs与IGFBP-4的结合诱导的。这些新的机制可能是重要的调节IGF介导的卵泡在卵巢中。

Follicular fluid from dominant follicles but not atretic follicles contains insulin-like growth factor (IGF) binding protein-4 (IGFBP-4) proteolytic activity. The effect of FSH and IGFs on IGFBP-4 proteolytic activity was studied using granulosa cell culture. Proteolytic activity was assessed by the incubation of [^<125>I]IGFBP-4 with medium and cleaved products were analyzed by autoradiography. The iodinated IGFBP-4 was cleaved into 18 kDa fragment when cells were incubated with FSH or IGFs while IGFBP-4 was remained intact in control culture. Inhibition of IGFBP-4 degradation by several protease inhibitors suggests that IGFBP-4 degradation was induced by a metalloserine protease. The degradation of IGFBP-4 was not stimulated when cells were incubated with the IGF-I analogue, LR3-IGF-I, and insulin that bind to IGF-I receptor but have less or no affinity for IGFBPs. Addition of IGFs but not FSH to medium from untreated GC culture stimulated proteolysis of [^<125>I]IGFBP-4. Similarly, exogenously added covalently cross-linked [^<125>I]IGF-II-IGFBP-4 complexes were proteolyzed, however, IGFs did not enhance the degradation of the complexes. These results suggest that human granulosa cells produce IGFBP-4 protease and that the increased susceptibility of IGFBP-4 to proteolysis is induced by the binding of IGFs to IGFBP-4. These novel mechanisms may be important to modulate IGF-mediated folliculogenesis in the ovary.