Isolation of a normal human thyroid cell line:: Hormonal requirement for thyroglobulin regulation

The long-term culture of functional follicular cells from normal adult human thyroid tissue has been obtained. They were expanded using a 1:2 split ratio until passage 28 (present status) in Click-RPMI medium enhanced with 5% fetal calf serum and diverse associations of hormones or components including porcine insulin and bovine thyrotropin. At passages 10 and 20, chromosome countings showed a normal diploid number and a normal karyotype. In calf serum containing media, cells are epithelial in the presence of thyrotropin (TSH) but present a slight elongated form in the absence of TSH. In serum-free media, 30 minutes after TSH stimulation, both epithelial and elongated cells changed in morphology to stellate-shaped, arborized forms, indicating the presence of functional TSH-receptors even in long term (18 months) TSH-free cultures. Cells produce thyroglobulin constitutively and large amounts of thyroglobulin are easily recovered in TSH-supplemented media, especially in the presence of insulin. Thyroglobulin production was increased versus days under TSH or insulin stimulation. Combination of the two hormones clearly resulted in a synergistic and not an additive effect. The other hormones present in the 6H components (transferrin, glycylhistidyl-lysine, somatostatin, and hydrocortisone) had no positive effect on thyroglobulin accumulation in media in our experimental conditions. Addition of TSH to hormone-free cultures or to insulin-, insulin plus hydrocortisone-, or 5H-containing cultures resulted in a clear increase in thyroglobulin production. Withdrawal of TSH from 6H cultures resulted in a decrease in thyroglobulin accumulation in media. Six months were required to select fibroblast-free cultures and to get passage 6. But only 17 months separated passage 6 to passage 28, indicating that the proliferative rate is increasing with in vitro cell adaptation. Such normal adult thyroid cells, thyroglobulin-producing, TSH, and insulin-sensitive, represent a new normal human thyroid cell line allowing comparative studies with cells originating from pathologic thyroid tissues.