Abstract. Bovine parathyroid glands incubated in organ culture synthesize and secrete parathyroid hormone in response to physiologic alterations in cation concentration. Hormone synthesized in the glands is immunologically different and higher in molecular weight than that released into the culture medium or circulating in bovine blood, although it is identical to purified hormone extracted from the glands. On the basis of the physiologic responses of tissue and medium hormone to cation, the transfer of radioactive label from one moiety to another, and interconversion of the two forms by a gland homogenate, it is suggested that the parathyroid glands synthesize a precursor hormone (proparathormone) that is modified to the smaller molecule that circulates in blood.Studies in vivoI and in vitro2'3 have provided direct evidence that the secretion of parathyroid hormone (PTH) is regulated by the concentration of extracellular calcium. Using a radioimmunoassay to measure the hormone4 and an in vitro test system in which physiologic function of the parathyroid glands can readily be controlled,5 we now report investigations of the synthesis of PTH. These studies provide evidence that the hormone is initially synthesized in the glands as a polypeptide with a molecular weight similar to that of purified bovine PTH,6 but that it is released as a smaller molecule which is immunologically different. Both the synthesis of hormone in the tissue and the secretion of the circulating molecule vary inversely with the extracellular calcium concentration.Materials and Methods. The external parathyroid glands were removed from cows within minutes after slaughter, placed in sterile Hanks solution and kept at 4VC until the experiment was initiated (usually 1-2 hr). The glands were washed, stripped of surrounding fat, and cut into 1-to 2-mm3 pieces for organ culture. Explants were placed in organ culture dishes as previously described5 and incubated in BGJ7 or F10 medium with 10% fetal calf serum under 5% CO2 in air. Incubations were performed with leucine-free medium containing 3 ,uCi of [U-4C ]leucine per ml, and the concentrations of calcium and magnesium were varied according to the experimental protocol. Incubations were conducted for 1-72 hr, and immediately thereafter the tissue was homogenized in a solution of 8 M urea, 0.2 M HC1, and 0.14 M j3-mercaptoethanol. Extraction was performed for 24 hr at 40C and the suspension was centrifuged at 5000 X g for 15 min with the supernatant fraction being saved. Protein content of each tissue homogenate was determined by method of Lowry et al.8Aliquots of tissue extract and supernatant medium were passed separately over columns of Sephadex G-100 (90 X 2.5 cm) and chromatographed at a rate of 20 ml/hr in 0.15 M ammonium acetate buffer (pH 5.0) containing 1 mg/ml egg white lysozyme to minimize 1631

No abstract

No abstract

No abstract