An insulin-producing cell line, HE, was derived from immortalized human pancreatic p-cells transformed by SV40 DNA, To make serum free medium specific for the cells, it was necessary to select basal medium, hormones, growth factors and coating materials. First of all, human insulin-producing cell line was adapted serially to the culture medium containing minimum required (1%) fetal calf serum (HB-1) from 10% fetal calf serum, The cells had a tendency to attach to the culture dishes coated with collagen or gelatin in the serum-free medium and showed a stronger attachment to collagen than to gelatin, For the basal medium selection, medium containing an equal mixture of RPMI 1640, Dulbecco's modified Eagle's medium, and Ham's F-12 (RDF) was shown to be the most suitable for the cells among 7 differently mixed basal media, The cells exhibited highly positive response to 7 factors including insulin-transferrin-selenium (ITS)complex in the basal medium. In addition, other 16 factors also influenced the cell growth positively, In contrast, 4 factors including aldosterone inhibited the cell growth, Cell growth was enhanced remarkably when the cells were cultured in the serum-free medium supplemented with all the seven positive factors, compared with in RDF serum-free medium. ITS and linoleic acid-BSA were found to be essential for the cell growth. The growth was much higher in the medium containing 6 strong positive factors without either dexamethasone, glucagon, or thyrocalcitonin than in the medium containing all the 7 strong positive factors, It is suggested that the interaction between the individual positive factors must be considered in choosing the factors with which the serum-free medium is supplemented.
Aldosterone ; Calcitonin ; Cell Line* ; Collagen ; Dexamethasone ; DNA ; Gelatin ; Glucagon ; Humans* ; Insulin ; Intercellular Signaling Peptides and Proteins*