Poly(A)-rich RNA has been isolated from calf thymus and translated in vitro in a rabbit reticulocyte translation system.Three peptides with M, = 58,000, 33,000, and 13,000 were specifically immunoprecipitated from the translation products with calf terminal deoxynucleotidyltransferase antiserum.An oligo(dT)-purified preparation of calf terminal transferase competed with only the M, = 58,000 peptide in the immunoprecipitation reaction.The anti-terminal transferase serum did not precipitate a M, = 58,000 peptide from translation products of spleen or liver mRNA, but it did precipitate the M, = 33,000 and 13,000 peptides from products of spleen mRNA and a M, = 13,000 peptide from products of liver mRNA.In addition, when an affinity-purified antibody to calf terminal transferase was used, only a M, = 68,000 peptide was immunoprecipitated from the translation products of calf thymus mRNA, and none was immunoprecipitated from spleen or liver mRNA products.This antibody also precipitated a M, = 58,000 peptide from the cell lysates of calf thymocytes labeled in vitro with [36S]methionine.These results demonstrate that calf terminal transferase is biosynthesized as a M, = 58,000 peptide.Terminal deoxynucleotidyltransferase (nucleoside triphosphate:DNA nucleotidylexotransferase; EC 2.7.7.31) is a unique DNA polymerase which catalyzes the addition of deoxyribonucleotides to the 3'-OH end of an oligodeoxynucleotide primer in the absence of a template.The enzyme is normally restricted to immature lymphocytes of the thymus and bone marrow (1-3).Terminal transferase' activity has also been found in circulating lymphocytes in certain leukemias (4, 5 ) .Calf terminal transferase was originally characterized by Chang and Bollum (6) as a M, = 32,000 peptide comprised of two subunits, a M, = 8,000 a subunit and a M , = 24,000 p subunit.However, a higher molecular weight for calf terminal transferase has recently been reported.Using immunoadsorbent chromatography in the presence of protease inhibitors, Nakamura et al. (7) have purified several immunospecific polypeptides with molecular weights ranging from 42,000 to
Utilizing sensitive and specific radioimmunoassays, serum concentrations of human thyrotropin (hTSH), the immunologically common alpha subunit of the glycoprotein hormones, and the specific beta subunit of hTSH (hTSH-0) have been measured in normal individuals, in patients with primary hypothyroidism, and in patients with other disorders of thyroid function before and after intravenous administration of thyrotropin releasing hormone (TRH). In 29 normal individuals hTSH-β was not detectable in serum (<0.5 ng/ml) before or after TRH; alpha was <0.5–2.0 ng/ml in men and premenopausal women and 1.0–5.0 ng/ml in post-menopausal women and did not increase after TRH. In 20 patients with primary hypothyroidism mean serum hTSH-β was 1.3 ng/ml and increased to a peak value of 3.7 ng/ml after TRH; mean alpha was 4.3 ng/ml and increased to 6.3 ng/ml after TRH. None of the patients with Graves' disease, a hyper-functioning thyroid nodule, or hypothyrotropic hypothyroidism had detectable serum hTSH-β concentrations or alpha concentrations higher than the normals before or after TRH. In 3 patients with primary hypothyroidism given an intravenous bolus of labeled hTSH, no dissociation of hTSH into subunits was detectable for at least 3 h, indicating that the increment in serum alpha and hTSH-β after TRH represented secretion of free subunits from the pituitary. In addition, L-thyroxine (L–T)4()administered to 2 hypothyroid patients deceased the serum concentrations of alpha and hTSH-β before and after TRH. Serum hTSH-β was fully suppressed with 100–300 fig (L–T)4 daily, but there was a residual serum alpha component, which could not be suppressed with thyroid hormone and probably represented alpha subunits arising from gonadotropin-secreting pituitary cells. Normal pituitary glands also contained a predominance of free alpha subunit relative to hTSH-β, in addition to hTSH. The secretion of free subunits in hypo-thyroidism may represent only a quantitative difference from the normal state, and subunits of hTSH appear to respond to the same control mechanisms as complete hTSH. (J Clin Endocrinol Metab40: 872, 1975)
Six hyperthyroid patients with inappropriately elevated serum thyrotropin (TSH) had serum concentrations of the common alpha subunit of the glycoprotein hormones (alpha) and the beta subunit of TSH (TSH-β) measured by radioimmunoassay. The three patients with a pituitary tumor had markedly elevated serum alpha levels of 105, 16.6, and 19.5 ng/ml and TSH of 34, 1.7, and 5.6 εU/ml, yielding molar alpha to TSH ratios of 31 to 98; TSH-β was not detected (<;0.5 ng/ml). The three patients without a pituitary tumor had serum alpha concentrations of 1.2, 0.5, and 1.0 ng/ml and serum TSH levels of 160, 9.3, and 90 εU/ml, yielding molar alpha to TSH ratios <1; TSH-β was 1.3, 0.5, and 0.5 ng/ml. Subunit and TSH secretion in the patients with a pituitary tumor demonstrated little or no change after thyrotropin releasing hormone (TRH) or thyroid hormone administration. In contrast, serum subunit and TSH levels increased after TRH in the patients withou a pituitary tumor; both basal and peak subuni and TSH levels after TRH decreased after th* administration of thyroid hormone although the] were still inappropriately high in relation to th* elevated serum thyroid hormone levels. In botl the tumor and non-tumor patients, alpha subuni and TSH concentrations decreased after the administration of dexamethasone. Inthe patients with; pituitary tumor, hypophysectomy followed by radiotherapy caused a decrease in serum alpha and TSf concentrations, as well as remission of hyperthyroidism. Thepresence of elevated serum alph; concentrations and undetectable TSH-β in patientswith TSH-induced hyperthyroidism may serve to identify those patients with a pituitary tumor and in certain patients the reduction of serum alph; may better indicate the adequacyof therapy,
The effect of carlorie intake on the development of spontaneous mammary tumors in virgin C3H mice was studied. Only about 10% of the mice fed a low-calorie diet [10 kcal/day (1 kcal = 4.184 kJ)] since weaning developed mammary tumors, compared to about 60% of those mice that were reared on high-calorie diets (16 kcal/day or lab chow ad lib). In order to understand the mechanism by which a low-calorie diet decreases the occurrence of mammary tumors in mice, we compared the sex cycle, the amounts of circulating thyroid-stimulating hormone (thyrotropin), growth hormone, and prolactin, the production of type A and B virus particles in the mammary glands, and the morphology of the mammary glands of mice fed low- and high-calorie diets. The amount of serum prolactin and the synthesis of type A and B particles in mammary tissues of mice fed a low-calorie diet was markedly decreased compared to those of age-matched mice fed high-calorie diets. In addition, in young mice fed a low-calorie diet, there were fewer mammary alveolar lesions than in mice fed a high-calorie diet, although the size of the lesions was similar. However, in older mice fed the high-calorie diet, the number and size of these lesions were greater than in the mice raised on the low-calorie diet. The other factors that we studied were not affected by calorie restriction. Our findings suggest that the reduction in serum prolactin level, mammary tumor virus production, and proliferation of mammary alveolar lesions associated with dietary calorie restriction is responsible for lowering the incidence of mammary tumors in mice.
Highly purified human TSH (hTSH) was dissociated into its alpha and beta subunits with propionic acid; the subunits were separated from undissociated hTSH and;-from each other by gel and ion exchange chromatography. During purification hTSH-a was identified by its cross reaction in a radioimmunoassay for the alpha subunit of human chorionic gonadotropin (hCG-α), and hTSH-β was identified by its cross reaction in an hTSH immunoassay. Purified hTSH-α (> 90% pure) and hTSH-β (>95% pure) were used toinduce antibodies in rabbits by a primary injection of 10 or 15 (µg, respectively, followed by 1 to 3 booster injections of 5 μg. Using hTSH-α tracer and anti-hTSH-α from each of 3 rabbits, radioimmunoassays were developed capable of detecting about 1 ng'ml hTSH-α. In these hTSH-α assays, the cross reactivity of hCG-α and hLH-α was virtually complete, while the cross reactivity of hLH was 15% and the reactivity of hTSH, hTSH-β,and hCG was less than 5%. Using hTSH-β tracer and anti-hTSH-β from each of 3 rabbits, radioimmunoassays were developed capable of detecting 0.025'0.25 ng⁄ml hTSH-β. Two of these immunoassays displayed conventional decreased bindingof tracer with increments of unlabeled hTSH-β, while one showed paradoxical enhanced binding. In these hTSH-β assays, hTSH, hTSH-α, hCG, hCG-α,hCG-β, hLH, and hLH-β displayed cross reactivity of 1% or less. The development of hTSH subunit radioimmunoassays provides additional information about the immunologic relationships of glycoprotein hormones and permits measurement of free subunit concentrations in certain unfractionated human sera. (Endocrinology94: 1411, 1974)
Concentrations of prolactin and growth hormone in the serum of rats were significantly increased by morphine. Dose response studies demonstrated that maximum prolactin release required lower doses of morphine than those needed for the maximum growth hormone response. Selective blockade of mu 1 (high affinity) opiate receptor with the irreversible antagonist naloxazone reduced morphine-induced peak concentrations of prolactin by 80 percent while increasing peak growth hormone levels by 250 percent. These results suggest different receptor mechanisms for the opiate modulation of the two hormones. The mu 1 (high affinity) receptor sites appear to mediate the morphine-induced release of prolactin but not growth hormone.
TSH was measured in human amniotic fluid after 3- to 8-fold concentration of the fluids. Amniotic fluid TSH was identical to standard pituitary TSH by immunological and gel chromatographic criteria. In 201 samples of normal second trimester amniotic fluid (16-19 weeks of pregnancy), amniotic fluid TSH concentrations had a mean value of 0.4 microunits/ml (range, less than 0.15 to 1.7 microunits/ml). In 21 samples of third trimester amniotic fluid (obtained to check fetal lung maturity), amniotic fluid TSH concentrations had a mean value of 0.25 microunits/ml (range, less than 0.15 to 0.55 microunits/ml). The capability of measuring TSH in amniotic fluid and the relative constancy of these values between the second and third trimesters of pregnancy suggest that the determination of TSH levels in amniotic fluid may be useful in the diagnosis of fetal hypothyroidism in utero.
