CULTIVATION OF RECOMBINANT, INSULIN-SECRETING ATT-20 CELLS AS FREE AND ENTRAPPED SPHEROIDS

Animal cells from endocrine glands have potential applications in biop rocessing, for the production of hormones, enzymes, possibly also reco mbinant proteins, and in tissue engineering, for the development of im munoisolated, implantable devices for long-term treatment of endocrine disorders. Immunoisolation can be achieved by surrounding cells with a biocompatible polymer which allows diffusion of nutrients and metabo lites, including hormones, but excludes higher molecular weight antibo dies and cytotoxic cells. Primary hormone-secreting cells cannot be ef fectively amplified in culture, so the large-scale application of impl antable systems based on such cells is limited by cell availability. I n this study, we conducted an initial assessment of the feasibility of using transformed, continuous cell lines in immunoisolated devices. T he model system employed consisted of mouse pituitary tumor AtT-20 cel ls which secrete recombinant proinsulin and an insulin-like peptide an d exhibit a high growth potential. Cells were cultivated as spheroids in spinner flasks and entrapped as such in alginate/polylysine/alginat e beads. Free and entrapped spheroids were propagated in fed-batch, su spension cultures. Entrapment did not significantly affect spheroid me tabolism or basal secretion. Entrapped spheroids did not increase in s ize or number and maintained roughly constant metabolic and basal secr etory activities over a 15-day period. Free spheroids in suspension in creased in size during the same period, but also maintained constant m etabolism and basal secretion, apparently because of a concomitant inc rease in hypoxic and/or necrotic cells. The potential of using continu ous cell lines in the development of bioartificial endocrine organs is discussed.