Effects of Dexamethasone, Calcium and 1,25-Dihydroxycholecalciferol on Calcitonin and Calcitonin Gene-Related Peptide mRNA Levels from the CA-77 C Cell Line

Abstract

We used the CA-77 cell, a murine C-cell line derived from a medullary thyroid carcinoma, to study the effects of glucocorticoids, calcium, and vitamin D metabolites on calcitonin (CT) gene expression. Total RNA was isolated, and CT and CT gene-related peptide (CGRP) mRNAs were measured by Northern hybridizations using specific probes. A control mRNA probe (cyclophilin) was used to quantitate the specificity of the changes in CT and CGRP mRNAs. The CA-77 C cell line cultured in basal conditions with a medium deprived of fetal calf serum, but was supplemented by insulin, expressed mainly the CGRP mRNA. Dexamethasone was found to increase both CT and CGRP mRNAs in a time- and dose-dependent way without changing the alternative splicing. A slight but significant increase in the steady-state CT mRNA level was found 3 days after addition of 10(-10) M dexamethasone; the same dose slightly decreased the CGRP mRNA level; concentrations of dexamethasone > or = 10(-9) M elevated both mRNAs. A twelve-fold increase for CT mRNA, and an eightfold increase in CGRP mRNA occurred 3 days after administration of 10(-6) M dexamethasone. Kinetic data revealed inductions of both mRNAs 24 hours after exposure to 10(-7) M dexamethasone, and highest CT and CGRP mRNAs levels were observed after 72 hours of treatment. Calcium from 1-4 mM in short-term (1 hour and 4 hour) or long-term stimulations (1 day and 4 days), with or without dexamethasone cotreatment was ineffective. CT and CGRP mRNAs levels were both half-reduced 48 hours after addition of 10(-7) M 1,25-dihydroxycholecalciferol; this effect was transient, as it disappeared 2 days later.

Effects of 17β-estradiol on calcitonin and calcitonin-gene-related peptide secretions and contents in a murine medullary thyroid carcinoma C-cell line (CA-77)

“…The dose of 17(3-estradiol used in the present in vitro studies (100 nmol/I) was shown to achieve a maximal increase of CT secretion by thyroparathyroid gland complexes of 8-d-old rat pups (Greenberg et al, 1986) (Naveh-Many et al, 1992), and also with the fact that estradiol given to ovariectomized animals prevents the decrease in CT mRNA levels (Naveh-Many et al, 1992;Lamari et al, 1994 (Russo et al, 1988;Collignon et al, 1992) and enhances the biosynthesis and release of both CT and CGRP (Lamari et al, 1994) in CA-77 C cells has already been reported. All-trans-retinoic acid also was shown to decrease both CT and CGRP mRNAs in CA-77 C cells and the secretion of both peptides (Lamari et al, 1994).…”

Decrease in CGRP and CT levels either contained in or released by CA-77 C cells after combined treatments with 1,25-dihydroxyvitamin D 3 analogues and 9-cis retinoic acid

“…Recent evidence suggests that isomerization of all-trans retinoic acid to 9-cis retinoic acid occurs in the cells (Urbach and Rando, 1994). 9-cis retinoic acid, a stereoisomer of all-trans, is a specific ligand for retinoic X receptors (RXR) (Heyman et al, 1992 (Collignon et al, 1992;Lamari et al, 1994). One of the most stricking results was enhancement of the inhibitory effect due to the combined treatment with the calcitriol analogues and 9-cis retinoic acid on the amounts of CGRP and CT, both contained in the C cells and that which was released.…”

Effect of Sympathetic Innervation on Isoproterenol-Induced Cystatin S Gene Expression in Rat Submandibular Glands during Early Development

“…Cyclophilin has been used as an internal standard to quantitate the specificity of changes in calcitonin and calcitonin gene-related peptide mRNA levels in the CA-77 cell line [9], as an internal standard in RT-PCR studies measuring the mRNAs encoding 5-HT1A and 5-HT2A proteins in human hippocampus [3] and as a standard for normalization in experiments examining the effects of steroid hormones and ß-adrenoreceptor agonists on the regulation of the rat Cys S gene [6].…”