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Human promyelocytic leukaemia cells (HL-60) can be induced to differentiate into mature granulocytes in vitro by 1 alpha,25-dihydroxycholecalciferol [1 alpha,25(OH)2D3], the active form of cholecalciferol. The differentiation-associated properties, such as phagocytosis and C3 rosette formation, were induced by as little as 0.12 nM-1 alpha,25(OH)2D3, and, at 12 nM, about half of the cells exhibited differentiation on day 3 of incubation. Concomitantly the viable cell number was decreased to less than half of the control. Among various derivatives of cholecalciferol examined, 1 alpha,25(OH)2D3 and 1 alpha,24R-dihydroxycholecalciferol were the most potent in inducing differentiation, followed successively by 1 alpha,24S-dihydroxycholecalciferol, 1 alpha-hydroxycholecalciferol, 25-hydroxycholecalciferol and 24R,25-dihydroxycholecalciferol. A cytosol protein specifically bound to 1 alpha,25 (OH)2D3 was found in HL-60 cells. Its physical properties closely resembled those found in such target tissues as intestine and parathyroid glands. 1 alpha,25(OH)2D3 bound to the cytosol receptor was transferred quantitatively to the chromatin fraction. The specificity of various derivatives of cholecalciferol in inducing differentiation was well correlated with that of their association with the cytosol receptor. These results are compatible with the hypothesis that the active form of cholecalciferol induces differentiation of human myeloid leukaemia cells by a mechanism similar to that proposed for the classical concept of steroid hormone action.

Extensive fusion was induced in mouse alveolar macrophages by treatment with conditioned media obtained from spleen cell cultures treated with 15 ,ug of phytohemagglutinin or concanavalin A per ml or with 12 nM la,25-dihydroxyvitamin D3 [la,25(OH)2D31. The fusion rate was 80-90% on day 3. In addition, 1a,25(OH)2D3 added directly to alveolar macrophages induced fusion of about 35% of the cells on day 3, whereas direct addition of phytohemagglutinin and concanavalin A did not enhance fusion at all. When conditioned media from spleen cell or T cell cultures treated with 12 nM Ia,25(OH)2D3 were applied to a Sephadex G-100 column, a fusion factor (Mr 37,000-70,000) could be separated from Ia,25(OH)2D3. la,25(OH)2D3 induced fusion at 0.012-120 nM in a dose-dependent manner both by direct action and by spleen cell-mediated indirect action, but the fusion rate was always much greater in the latter than in the former at each concentration of the vitamin. Of the vitamin D3 derivatives tested, Ia,25(OH)2D3 was the most potent, followed successively by la,24R,25-trihydroxyvitamin D3, la-hydroxyvitamin D3, 25-hydroxyvitamin D3, and 24R,25-dihydroxyvitamin D3. These results clearly indicate that 1a,25(OH)2D3 induces fusion of mouse alveolar macrophages by both a direct and an indirect mechanism, the latter mediated by spleen cells, probably by T cells.Vitamin D3 is well known as an antirachitic agent that stimulates bone mineralization. In 1952, Carlsson demonstrated for the first time that vitamin D stimulates not only bone mineralization but also bone mineral mobilization (1). Vitamin D3 is metabolized first in the liver to 25-hydroxyvitamin D3 [25-(OH)D3] and then in the kidney mainly to 24R,25-dihydroxyvitamin D3 [24R,25(OH)2D3] and la,25-dihydroxyvitamin D3

Objective: One of the thyroid-speci®c transcription factors, thyroid transcription factor-2 (TTF-2), performs a crucial role in the development of the thyroid gland. We performed genetic analysis of the TITF2 gene (encoding TTF-2) in patients with thyroid dysgenesis. Methods: By direct sequencing of the PCR products of TITF2, we screened the genomic DNA from 46 patients with thyroid dysgenesis (®ve had agenesis, six had hypoplasia, 15 had ectopy, and 20 were undetermined). We also studied the transcriptional activities of TITF2 by co-expressing the luciferase gene directed by the human thyroglobulin gene promoter. Results: Human TITF2 consists of a forkhead domain, a polyalanine tract, and unique C-terminal residues. In one of the patients with an ectopic sublingual thyroid, we found a polyalanine tract of 11 alanine residues on one chromosome instead of the 14 alanine residues found in normal controls. In one patient with hypoplasia, the polyalanine tract consisted of 12 heterozygous alanine residues. The reduced polyalanine tracts were not detected in 101 normal individuals. However, the expression study showed that the transcriptional activities of TITF2 with reduced polyalanine-tract lengths were equal to that of TITF2 with an unreduced polyalanine tract. Conclusion: These results suggest that the polymorphism of the polyalanine tract of TITF2 is not a frequent cause of developmental defects of the human thyroid gland.

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