No abstract

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.

Osteoblasts are cells responsible for matrix deposition during bone development and although temporal expression of many genes has been related to osteoblast differentiation, a complete description of osteoblastspecific gene regulation will lead to a better understanding of osteoblast function. In this study