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Biomedical subjects

B Rapoport

Publications and source records attributed to B Rapoport.

At least 235 records · Page 13Linked to original sources
PubMed1978

Bioassay of TSH using dog thyroid cells in monolayer culture.

The cAMP response to TSH stimulation in dog thyroid cells in monolayer culture was adapted as a means to assay TSH bioactivity. Of a variety of polypeptide hormones examined, only TSH and LH stimulated thyroid cell cAMP generation, but stimulation by LH probably represented contamination with TSH. Serum was found to be a potent, noncompetitive inhibitor of the thyroid cell cAMP response to TSH stimulation. The inhibitor(s) present in serum was nondialyzable and, on Sephadex G-200 gel filtration, eluted over a wide range between the void volume and the albumin peak. Because of this inhibitory effect of serum, partial purification of TSH from serum was necessary, and was achieved using Sephacryl S-200 gel filtration. Columns were calibrated with human TSH as measured by radioimmunoassay and bovine TSH as measured by bioassay. Intraassay variation of the thyroid cell cAMP response to a 100 microU/ml TSH standard was 9.2%. TSH standards were added to human serum following which the TSH was extracted by gel filtration and assayed for bioactivity. With TSH standards of 100 microU/ml intra-assay variation was 19.9%; and with TSH standards of 25 microU/ml, interassay variation was 34.5%. TSH bioactivity was demonstrable in serum from 21 of 25 patients with primary hypothyroidism, 4 of 14 normal subjects and none of 3 patients on exogenous thyroxine replacement. Although a positive correlation was observed between serum TSH bioactivity and immunoactivity, numerous individual samples displayed a dichotomy between the two measurements. This study provides new evidence that immunoassayable TSH in the serum of patients with primary hypothyroidism is not necessarily synonymous with TSH bioactivity.

Animals↗
PubMed1978

Acute effects of thyroid-stimulating hormone on cultured thyroid cell morphology.

The acute effect of TSH on the ultrastructure of dog thyroid cells in monolayer tissue culture was examined by scanning electron microscopy. Although thyroid cells in monolayer culture remained responsive to TSH stimulation, the cell surface changes induced by TSH were different from those seen in intact thyroid tissue. Morphological changes were observed within 15 min after the addition of TSH, the earliest time point examined. These changes were fully developed within 4 h. Spontaneous morphological transformation was not observed in the absence of TSH. Morphological alterations induced by TSH were 1) the thickening and retraction of the cell border, 2) the formation of a network of cytoplasmic projections resulting from both cell retraction and active cytoplasmic extension, and 3) an increase in the number of surface microvilli. The relationship between this acute TSH action and thyroid hormone synthesis and secretion remains to be clarified.

Animals↗
PubMed1977

Cultured thyroid cell adenosine 3',5'-cyclic monophosphate response to thyrotropin: loss and restoration of sensitivity to iodide inhibition.

Unlike in all other thyroid preparations, exposure of dog thyroid cells in long-term monolayer culture to iodide (10(-7) to 10(-3) M for up to 19 h did not blunt the subsequent adenosine 3', 5'-cyclic monophosphate (cAMP) response to thyrotropin (TSH) stimulation. This lack of effect of iodide was observed even when confluent thyroid cells were "follicularized" by the action of TSH in the culture medium. Preincubation of these cells in thyroxine (T4) and triiodothyronine (T3) was similarly without effect on the subsequent cAMP response to TSH. Study of thyroid cells during the early phase of primary culture demonstrated that inhibition by iodide (10(-4) M) of the cAMP response to TSH occurred after 7 h but was lost after 48 h of cell culture. This inhibitory effect of iodide was prevented by the inclusion of methimazole in the preincubation medium. As with iodide-insensitive cells, T4 and T3 were without effect on the cAMP response to TSH in iodide-sensitive thyroid cells. Exposure of iodide-insensitive thyroid cells to iodide-containing medium obtained after 2 h of incubation with dog thyroid slices, as well as to medium enriched with the 100,000 g supernatant fraction of homogenates prepared from these thyroid slices, did not restore the inhibitory action of iodide. However, iodide-sensitivity of the cAMP response to TSH was restored by preincubation of iodide-insensitive cells in 10(-4) M iodide plus an H2O2-generating system (glucose-glucose oxidase). These data suggest that T4 and T3 are not organic iodine inhibitors of the thyroid cAMP response to TSH. In addition, they provide evidence against the existence of a soluble, freely diffusible, organic iodine inhibitor of thyroid adenylate cyclase. The loss of sensitivity to iodide inhibition of adenylate cyclase that occurs in thyroid cells shortly after initiation of primary culture appears to be related to a defect in the cellular organification mechanism, possibly the H2O2-generating system.

Adenylyl Cyclases↗
PubMed1976

Induction of refractoriness to thyrotropin stimulation in cultured thyroid cells. Dependence on new protein synthesis.

Cultured dog thyroid cells were used to investigate the mechanism by which previous exposure to thyrotropin (TSH) induces refractoriness to further TSH stimulation of cellular adenosine 3'-5'-monophosphate (cAMP). Refractoriness of the cAMP response to TSH could not be overcome by exposure of the cells to supramaximal stimulatory concentrations of TSH. Although an unknown factor present in human and fetal calf serum was found to inhibit the thyroid cell cAMP response to TSH, this factor could not account for refractoriness because refractoriness could be induced in the absence of serum. Induction of thyroid refractoriness did not appear to be related to cellular concentrations of cyclic AMP, because equal refractoriness was produced by TSH alone or TSH plus the phosphodiesterase inhibitor, 3-isobutyl-1-methyl xanthine. In addition, preincubation of thyroid cells in 10(-4) M cAMP did not result in subsequent refractoriness. Recovery from the refractory process required almost 24 h. Short term (15 min) stimulation with TSH did not produce thyroid cell refractoriness, and reversal of the stimulation was obtained by thorough washing of the cells. Long term TSH stimulation (16 h), however, resulted in both supramaximal cAMP response to TSH, and inclusion of TSH together with cycloheximide did not produce refractoriness. Cyclic AMP phosphodiesterase activity in thyroid cell homogenate was unaltered by TSH or dibutyryl cyclic AMP pretreatment of the cells for up to 24 h, or cycloheximide for up to 4 h. In contrast, TSH-stimulated, but not F--stimulated, adenylate cyclase activity was reduced in thyroid cell homogenates after preincubation of the cells in TSH. Refractoriness to TSH stimulation was not associated with an alteration in the binding of 125I-TSH to cultured thyroid cells. These studies suggest that the thyroid cAMP response to TSH is modulated by an inhibitory mechanism dependent upon new protein synthesis. TSH stimulation itself increases the degree of this inhibition through a mechanism not involving cAMP.

Animals↗
PubMed1976

Dog thyroid cells in monolayer tissue culture: adenosine 3', 5'-cyclic monophosphate response to thyrotropic hormone.

A simple, rapid, and efficient method is described for establishing dog thyroid cells in tissue culture. Thyroid cell yield from a small amount of tissue (1 g) is high and viability is excellent. The adenosine 3',5-cyclic monophosphate (cAMP) response to thyrotropin (TSH) was investigated in these thyroid cells. Peak cAMP values were achieved after 10-15 min of TSH stimulation (in the presence of 0.5 mM 3-isobutyl-1-methyl-xanthine, MIX), with a subsequent decline to about half the maximal value after approximately 4 hours. This decline in cAMP concentration was associated with the development of refractoriness to TSH stimulation. Half-maximal stimulation of thyroid cell cAMP content was observed at a TSH concentration of between 1 and 2 mU/ml. Maximal cAMP values achieved were approximately 30-fold greater than basal values in the presence of MIX. The threshold of sensitivity of the cAMP response to TSH was very low, with significant stimulation being observed at a TSH concentration of 5-10 muU/ml. As determined by double reciprocal plots, the net cAMP response to TSH appeared to represent a single function over the entire TSH concentration range tested.

Animals↗
PubMed1976

On the mechanism of inhibition by iodine of the thyroid adenylate cyclase response to thyrotropic hormone.

Rats maintained on a low-iodine diet were hypophysectomized, and their diet was than enriched with iodide. Cyclic AMP (cAMP) concentrations achieved in their thyroids following in vitro TSH stimulation were significantly lower than those in the thyroids of control animals that did not receive dietary iodide enrichment. The addition of 0.1% methimazole (MMI) or 1% KC1O4 to the diet abolished this inhibitory effect of iodide. The administration of triiodothyronine in the died did not reproduce the inhibitory effect of iodide. The effect of iodide in vitro on the thyroid cAMP response to TSH was then investigated using paired thyroid lobes obtained from intact rats fed a low-iodine diet. During a 15-min incubation period, concentrations of iodide up to 10(-3)M, together with TSH (125 mU/ml), did not affect the thyroid cAMP response to TSH. In contrast, the preincubation of the lobes in 5 X 10(-5)M Nal for 2 h preceding a final 15-min incubation in medium containing TSH alone resulted in final cAMP concentrations significantly lower than those in paired lobes not exposed to iodide. Basal cAMP concentrations in thyroids not subjected to TSH stimulation were unaffected by preincubation in iodide. The inclusion of TSH during the preincubation period augmented the inhibitory effect of iodide on the final thyroid cAMP concentration achieved. The inclusion of MMI together with iodide during the preincubation period abolished the inhibitory effect of iodide on the final cAMP concentration achieved by TSH stimulation. Direct measurement of newly formed organic iodine in vitro demonstrated it to be inversely proportional to the final cAMP concentration achieved by TSH stimulation. The preincubation of thyroid lobes in iodide was without effect on the subsequent stimulation of cAMP by PGE1, or on the stimulation by F- of adenylate cyclase activity in the thyroid homogenate. The data support the concept of an as yet unknown organic form of iodine that limits thyroid adenylate cyclase responsiveness to TSH stimulation. This may, in part, explain the diverse, and generally inhibitory, actions of iodide on thyroid function.

Adenylyl Cyclases↗
PubMed1976

Compensatory thyroid hypertrophy after hemithyroidectomy in rats.

Thyroid enlargement occurs in association with a variety of circumstances characterized by an impaired capacity of the gland to secrete adequate amounts of hormone. To elucidate the factors responsible for such compensatory thyroid growth, particularly the role of TSH, we have observed the response of the serum TSH, T3 and T4 concentrations following hemithyroidectomy in the rat, and have attempted to correlate changes in these functions with changes in the weight and histology of the thyroid remnant. Hemithyroidectomy was performed in male Sprague-Dawley rats weighing 150 to 370 g, sham-operated animals serving as controls. As compared to findings in sham-operated animals, serum T4 concentrations declined promptly after hemithyroidectomy. In Experiment I serum T4 concentrations remained low for about 10 days and then returned to initial values. In Experiment II serum T4 concentrations remained lower than initial T4 values or values found in sham-operated animals until 34 days after hemithyroidectomy. Serum T3 concentrations were not significantly altered after hemithyroidectomy in either group but tended to be lower in the hemithyroidectomized animals. Serum TSH concentrations increased within 3 days after hemithyroidectomy and, for as long as 21 weeks, remained at values higher than those present preoperatively or those seen in sham-operated animals. Thyroid lobe weight increased following removal of the contralateral lobe and this increase was also sustained throughout the duration of the experiments. Biochemical and histological observations indicated that enlargement of the residual lobe was due to hypertrophy rather than hyperplasia.

Animals↗
PubMed1975

Pituitary-thyroid responsiveness to intramuscular thyrotropin-releasing hormone based on analyses of serum thyroxine, tri-iodothyronine and thyrotropin concentrations.

To develop a test of pituitary-thyroid responsiveness to thyrotropin-releasing hormone that would obviate the need for measuring serum thyrotropin, we determined serum thyrotropin, thyroxine, and tri-iodothyronine concentrations before and at frequent intervals after the intramuscular administration of 2 mg of thyrotropin-releasing hormone in normal subjects and in patients with a variety of thyroid disorders. In specimens obtained four and five hours after administration of the hormone to normal subjects, serum thyroxine concentration increased 2.4 plus or minus 0.7 mug per 100 ml (mean plus or minus S.D.) over base-line values, the magnitude of increase being greater than 1.5 mug per 100 ml in 32 of 34 subjects. Serum thyroxine concentrations after administration of thyrotropin-releasing hormone did not increase in 11 hyperthyroid patients. Of 13 with hypothyroidism, increases in 12 were 0 to 0.7 mug per 100 ml; in one the increment was 1.2 mug per 100 ml. Measurement of the serum thyroxine response to intramuscular thyrotropin-releasing hormone will usually suffice to determine the integrity of the hypothalamic-pituitary-thyroid complex.

Adult↗
PubMed1975

Inhibitory effect of dietary iodine on the thyroid adenylate cyclase response to thyrotropin in the hypophysectomized rat.

In hypophysectomized rats given dietary regimens either rich or deficient in iodine, the increase in thyroid cyclic AMP concentration induced acutely by a single dose of TSH was significantly less in iodine-enriched than in iodine-deficient animals. Direct assays revealed that this difference was because the thyroid adenylate cyclase response to TSH was less in the iodine-enriched animals, phosphodiesterase activity being no different in the two groups. This effect may explain the inhibitory action of dietary iodine enrichment on diverse functional and anatomical responses of the thyroid to TSH.

3',5'-Cyclic-AMP Phosphodiesterases↗
PubMed1974

Hyperresponse to thyrotropin-releasing hormone accompanying small decreases in serum thyroid hormone concentrations.

To determine whether pituitary thyrotropin (TSH) responsiveness to thyrotropin-releasing hormone (TRH) is enhanced by small decreases in serum thyroxine (T4) and triiodothyronine (T3), 12 euthyroid volunteers were given 190 mg iodide po daily for 10 days to inhibit T4 and T3 release from the thyroid. Basal serum T4, T3, and TSH concentrations and the serum T4 and TSH responses to 400 mug TRH i.v. were assessed before and at the end of iodide administration. Iodide induced small but highly significant decreases in basal serum T4 (8.0+/-1.6 vs. 6.6+/-1.7 mug/100 ml; mean +/- SD) and T3 (128+/-15 vs. 110+/-22 ng/100 ml) and increases in basal serum TSH (1.3+/-0.9 vs. 2.1+/-1.0 muU/ml). During iodide administration, the TSH response to TRH was significantly increased at each of seven time points up to 120 min. The maximum increment in serum TSH after TRH increased from a control mean of 8.8+/-4.1 to a mean of 13.0+/-2.8 muU/ml during iodide administration. As evidence of the inhibitory effect of iodide on hormonal release, the increment in serum T3 at 120 min after TRH was significantly lessened during iodide administration (61+/-42 vs. 33+/-24 ng/100 ml). These findings demonstrate that small acute decreases in serum T4 and T3 concentrations, resulting in values well within the normal range, are associated both with slight increases in basal TSH concentrations and pronounced increases in the TSH response to TRH. These results demonstrate that a marked sensitivity of TSH secretion and responsiveness to TRH is applicable to decreasing, as well as increasing, concentrations of thyroid hormones.

Adult↗