Olijve W, de Boer W, Mulders J W, van Wezenbeek P M
NV Organon, Oss, The Netherlands.
Mol Hum Reprod. 1996 May;2(5):371-82. doi: 10.1093/molehr/2.5.371.
Follicle stimulating hormone (FSH) is a heterodimeric glycoprotein hormone produced in the anterior pituitary gland. The hormone is essential in the regulation of reproductive processes, such as follicular development and ovulation. It is clinically used for treatment of anovulation and in assisted reproduction technologies such as in-vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI). Until recently, the only source for human FSH has been the urine from post-menopausal women. Such a natural source implies limited availability and potential product variability. Thus, we have cloned the genes encoding the alpha- and beta-subunits of human FSH and transfected these into Chinese hamster ovary (CHO) cells. A CHO-clone was isolated capable of secreting intact glycosylated FSH with identical amino acid sequences to natural FSH. This cell line was grown in perfusion culture and enabled us to isolate highly pure FSH (> 99%). The complexity of the charge distribution of human recombinant FSH was demonstrated by Isoelectric focusing. The observed microheterogeneity is caused by the large number of carbohydrate chain structures which are added to the four potential glycosylation sites in the alpha beta-dimer. Furthermore, the carbohydrates show a variation in their degree of sialylation which reflects the different pl values of the individual isohormones. Despite the complexity of post-translational modification, the isoform distribution of recombinant FSH produced in a CHO-cell line and grown in perfusion culture is surprisingly similar to that observed with pituitary FSH and urinary FSH. In conclusion, we have shown that FSH-gene transfected CHO-cells are capable of stable serum-free production of recombinant FSH. A process has been developed which assures the consistent and reproducible production of highly-purified recombinant FSH.
促卵泡激素（FSH）是一种在前脑垂体中产生的异源二聚体糖蛋白激素。该激素在调节生殖过程中至关重要，例如卵泡发育和排卵。它在临床上用于治疗无排卵以及辅助生殖技术，如体外受精（IVF）和胞浆内单精子注射（ICSI）。直到最近，人促卵泡激素的唯一来源一直是绝经后妇女的尿液。这种天然来源意味着供应有限且产品可能存在变异性。因此，我们克隆了编码人促卵泡激素α亚基和β亚基的基因，并将其转染到中国仓鼠卵巢（CHO）细胞中。分离出了一种CHO克隆，它能够分泌与天然促卵泡激素氨基酸序列相同的完整糖基化促卵泡激素。该细胞系在灌注培养中生长，使我们能够分离出高纯度的促卵泡激素（>99%）。等电聚焦证明了重组人促卵泡激素电荷分布的复杂性。观察到的微异质性是由添加到αβ二聚体中四个潜在糖基化位点的大量碳水化合物链结构引起的。此外，碳水化合物的唾液酸化程度存在差异，这反映了各个同工激素不同的等电点值。尽管翻译后修饰很复杂，但在CHO细胞系中生长并在灌注培养中产生的重组促卵泡激素的同工型分布与垂体促卵泡激素和尿促卵泡激素的分布惊人地相似。总之，我们已经表明，转染了促卵泡激素基因的CHO细胞能够在无血清条件下稳定产生重组促卵泡激素。已经开发出一种方法，可确保一致且可重复地生产高度纯化的重组促卵泡激素。
