ESTABLISHMENT AND CHARACTERIZATION OF A CHO CELL-LINE PRODUCING A SECRETED FORM OF HUMAN CILIARY NEUROTROPHIC FACTOR - NEUROPROTECTIVE EFFECTS OF THE RECOMBINANT PROTEIN

Human ciliary neurotrophic factor (CNTF) was inserted into a mammalian expression vector linked to the prepro sequence of human nerve growth factor. A Chinese hamster ovary cell line was established by resistan ce to neomycin and the plasmid integrated DNA was amplified using the metallothionein gene. This cell line contained several hundred copies of the human CNTF gene and produced an NH2 terminal truncated form of human CNTF (22 kDa) which was secreted into the medium. Although the c opy number of the human CNTF gene was high and its mRNA was actively t ranscribed, the recombinant protein secreted into the medium constitut ed only 35-40% of the total amount of human CNTF synthesized by these cells. Both wild-type human CNTF produced in bacterial cells and the h uman CNTF obtained by forced secretion were effective in protecting hi ppocampal pyramidal neurons from injury induced by glucose deprivation , a form of excitotoxic neurodegeneration.