IMMORTALIZED HUMAN PITUITARY-CELLS EXPRESS GLYCOPROTEIN ALPHA-SUBUNITAND THYROTROPIN-BETA (TSH-BETA)

A major problem in the study of human pituitary cells is their lack of proliferative capacity in vitro. To address this issue, we have infec ted normal human, postmortem pituitary cells in monolayer culture with a temperature-sensitive (tsA58) mutant of SV40 large T antigen. Sever al epithelial-like colonies were isolated; and one, designated CHP2, h as been studied in detail to identify its functional characteristics. CHP2 cells have undergone more than 150 culture passages and retain an epithelial morphology. They exhibit tight temperature-dependent growt h, in the presence and absence of serum, with cell division at 33 C an d growth inhibition at 39 C. CHPB cells, at both temperatures, showed diffuse immunostaining for human alpha-subunit and focal staining for TSH beta. Gene expression was confirmed by RT-PCR and sequencing. TRH and GnRH receptors were not detectable, and their absence was confirme d by their lack of effects on intracellular calcium and inositol phosp holipids. Cytogenetic analysis showed that the cells had a modal peak in the diploid range and a smaller peak in the tetraploid range. There was also a consistent loss of chromosome 22 and a normal chromosome 2 homologue, the latter being replaced by one of two chromosome 2 marke rs, M2A or M2B. In conclusion, we have immortalized human pituitary ce lls using SV40 tsT, from which we have cloned a cell line expressing a lpha-subunit and TSH beta.