Renal handling of thyroxine, 3,5,3'- and 3,3',5'-triiodothyronine, 3,3'- and 3',5'-diiodothyronine in man

The 24-h urinary excretion and renal clearance of thyroxine (T4), 3,5,3'-triiodothyronine (T3), 3,3',5'-triiodothyronine (rT3), 3,3'-diiodothyronine (3,3'-T2), and 3',5'-diiodothyronine (3',5'-T2) were measured in 17 healthy subjects. The median urinary excretion was (pmol/24h) T4: 1242, T3: 828, rT3: 12.9, 3,3'-T2: 331, and 3',5'-T2: 5.8. The corresponding renal clearances were in median (ml/min) T4: 31, T3: 133, rT3: 15, 3,3'-T2: 683, and 3',5'-T2: 4.5. The clearances differed mutually (P less than 0.01) as well as from the creatinine clearance (P less than 0.01) which was in median 87 ml/min. Thus, all iodothyronines studied were subject to tubular transport mechanisms besides glomerular filtration. The 3 iodothyronines with 2 iodine atoms in the phenolic ring of the thyronine molecule, T4, rT3 and 3',5'-T2, were mainly tubularly reabsorbed, whereas those with only one iodine atom in the phenolic ring, T3 and 3,3'-T2, were mainly tubularly secreted. It might be hypothesized that the number of iodine atoms in the phenolic ring determines the direction of the tubular transport (presence of 2 iodine atoms is associated with tubular reabsorption, and of one iodine atom with secretion), whereas the rate of tubular transport decreases with decreasing number of iodine atoms in the tyrosylic ring.     

Plasma thyroxine, 3,3',5-triiodothyronine and 3,3',5'-triiodothyronine during beta-adrenergic blockade in hyperthyroidism

Plasma thyroxine (T4), 3,3',5-triiodothyronine (T3) and 3,3',5'-triiodothyronine (rT3) were measured in 16 patients with Graves' disease. Patients were studied under the following conditions: first without any treatment, then, during beta-adrenergic blockade with propranolol, and finally after euthyroidism had been attained by carbimazole. During propranolol T3/T4 ratio decreased, whereas T4 remained unchanged. After carbimazole T3/T4 ratio returned to its pretreatment value. rT3/T4 ratio showed opposite changes. These results suggest that peripheral conversion of T4 into T3 and rT3 in hyperthyroidism is, at least partly, dependent on the functional status of the beta-adrenergic system. Suppressed peripheral conversion of T4 into T3 during beta-adrenergic blocking agents may contribute to the beneficial effects of these drugs in thyrotoxicosis.     

A radioimmunoassay for 3',5'-diiodothyronine.

The present report describes a RIA for 3',5'-diiodothyronine (T2) that can be performed on unextracted serum and which has a lower limit of detectability of 2 ng/dl. Cross-reactivity with other iodothyronines was negligible, except for rT3 which began to demonstrate cross-reactivity when rT3 levels were elevated to 180 ng/dl. Employing this RIA for T2, we have determined that 83 healthy individuals had a mean (+/-SE) serum T2 concentration of 5.0 +/- 0.3 ng/dl, thyrotoxic subjects (n = 12) had a mean T2 level that was elevated to 10.8 +/- 0.8 ng/dl, and each of 6 hypothyroid subjects had undetectable (less than 2 ng/dl) concentrations. Athyreotic patients (n = 8), receiving 0.4 mg T4 daily, had serum T2 concentrations of 15.0 +/- 3.0 ng/dl. Fasting in obese subjects was associated with an increase in serum T2 to 6.9 +/- 0.6 ng/dl from a basal level of 4.4 +/- 0.4 ng/dl in the fed state (P less than 0.01). Despite the fact that rT3 levels may be elevated in amniotic fluid and that rT3 is expected to represent the major source from which extrathyroidal T2 arises, T2 levels were low in amniotic fluid, being undetectable (less than 2 ng/dl) in 9 of 19 samples; the mean (+/-SE) T2 concentration in the 10 detectable samples was 5.4 +/- 1 ng/dl. These data indicate T2 is a normal component of serum and that the majority of serum T2 is probably derived from peripheral conversion. Furthermore, these observations suggest that situations associated with elevated rT3 levels (e.g. thyrotoxicosis and fasting) may also have increased T2 values.     

The effects of phenytoin (diphenylhydantoin) on the extrathyroidal turnover of thyroxine, 3,5,3'-triiodothyronine, 3,3',5'-triiodothyronine, and 3',5'-diiodothyronine in man

The extrathyroidal metabolism of T4, T3, rT3, and 3',5'-diiodothyronine (3',5'-T2) was studied before and after treatment with 350 mg phenytoin (DPH) daily for 14 days in six hypothyroid patients receiving constant L-T4 replacement. The total and free serum concentrations of the four iodothyronines were reduced by approximately 30% during DPH treatment, whereas the free fractions in serum were unaltered. Concomitantly, serum TSH increased 137% (P less than 0.02). The production rate (PR) of T4 decreased 16% (P less than 0.005), indicating decreased intestinal absorption (bioavailability) of oral L-T4 during DPH treatment. The fractional rate of 5'-deiodination of T4 to T3 increased from 27% to 31% (P less than 0.05), whereas the rate of 5-deiodination of T4 to rT3 decreased from 45% to 25% (P less than 0.05). The urinary excretion of free and conjugated T4 was 2.3% of the T4 PR and was unaffected by DPH. Thus, the amount of T4 metabolized through nondeiodinative pathways apart from urinary excretion increased from 25% to 44% (P less than 0.05). The apparent distribution volume (Vd) of T4 increased (P less than 0.05), whereas the pool size was unchanged. The PR of T3 did not change during DPH treatment, nor did the mean transit time or the cellular clearance. The rT3 PR was reduced by 54% (P less than 0.02) during DPH treatment. Concomitantly, the transit time increased 10-fold (P less than 0.05), whereas Vd and pool size increased 5-fold (P less than 0.01 and P less than 0.05, respectively). The turnover of 3',5'-T2, in contrast to that of the other iodothyronines, did not change significantly during DPH treatment. T3 formation from T4 was measured in liver microsomal fractions from rats treated for 8 days with DPH and was almost identical to that in untreated animals. The data demonstrate that DPH in therapeutic concentrations did not affect serum protein binding of the iodothyronines. DPH reduced the intestinal absorption of T4 and increased the nondeiodinative metabolism of T4. The resulting decrease in total and free serum T4 and T3 was associated with an increase in serum TSH, demonstrating reduced negative feedback on the pituitary. Our data do not support the assumption that DPH induces increased hepatic deiodinating enzyme activity.(ABSTRACT TRUNCATED AT 400 WORDS)     

The metabolism of 3,3'-diiodothyronine in man

The production rate of 3,3'-Diiodothyronine (3,3'-T2) was measured in five healthy subjects after a single injection of [125I]3,3'-T2. The [125I]3,3'-T2 was measured by immunoprecipitation. To reduce the large amount of nonspecific serum radioactivity (iodides, 3,3'-T2 metabolites, and protein-bound iodine), the sear were treated before the immunoprecipitation with an anion exchanger and polyethylene glycol (final concentration, 10%). The noncompartmental analysis of the data gave the following results: MCR, 0.52 +/- 0.07 liters/min or 926 +/- 142 liters/day (mean +/- SD); and production rate, 23.7 +/- 8.2 ng/min or 34 +/- 12 micrograms/day.     

Simultaneous turnover studies of thyroxine, 3,5,3' and 3,3',5'-triiodothyronine, 3,5-, 3,3'-, and 3',5'- diiodothyronine, and 3'-monoiodothyronine in chronic renal failure

The present study evaluates the sequential extra-thyroidal monodeiodination of thyroid hormones through tri-, di-, and monoiodothyronines in chronic renal failure (CRF) in man. Simultaneous turnover studies of T4, T3, rT3, 3,5-diiodothyronine (3,5-T2), 3,3'-T2, 3',5'-T2, 3'5'-T2, and 3'-monoiodothyronine (3--T1) were conducted in six patients with CRF (creatinine clearance, 9-18 ml/min) using the single-injection, noncompartmental approach. Serum levels of T4, T3, and 3,5-T2 were reduced to two thirds of control levels (P less than 0.05), whereas serum rT3 and 3,3'-T2 levels were reduced to a minor degree. Serum 3'-5'-T1 was doubled (p less than 0.05). The MCRs of T4, rT3, and 3',5'-T2 were enhanced to 168%, 127%, and 187% of normal (P less than 0.05), respectively, whereas those of T3, 3,5-T2, 3,3'-T2, and 3'-T1 were unaffected. The mean production rates (PRs) of the iodothyronines in CRF were as follows (CRF vs. control values, expressed as nanomoles per day/70 kg): T4, 119 vs. 125; T3, 26 vs. 44 (P less than 0.01); rT3, 49 vs, 48; 3,5-T2, 3.5 vs. 7.2 (P less than 0.001); 3,3'-T2, 25 vs. 35 (P less than 0.01); 3',5'-T2, 25 vs. 14 (P less than 0.01); and 3'-T1, 39 vs. 30. Previous studies have demonstrated reduced phenolic ring (5'-) deiodination of T4 in CRF, which is supported by the present finding of unaltered PR of T4 and reduced PR of T3. In contrast the 5'-deiodination of T3 leading to the formation of 3,5-T2 was found unaffected by CRF, since the conversion rate (CR) of T3 to 3,5-T2 (PR 3,5-T2/PR T3) was unaltered (16% vs. 15% in controls). The tyrosylic ring (5-) deiodination of T4 to rT3 was unaffected in patients with CRF, the CR being 42% vs. 40% in controls, in contrast to an enhanced CR of rT3 to 3',5'-T2 (53% vs. 29%, P less than 0.01), which also is a 5-deiodination step. In conclusion, our data show that CRF profoundly changes the kinetics of all iodothyronines studied. Furthermore, our data are compatible with the existence of more than one 5'-deiodinase as well as more than one 5-deiodinase in man.     

Opposite effects of dexamethasone on serum concentrations of 3,3',5'-triiodothyronine (reverse T3) and 3,3'5-triiodothyronine (T3).

Dexamethasone, 2 mg every 6 hours for 4 doses, was given to 4 hypothyroid patients receiving treatment with synthetic thyroxine (T4) and to 8 untreated hyperthyroid patients with Graves' disease, and serum concentrations of thyroid hormones were measured by radioimmunoassays. Serum concentration of 3,3'5'-triiodothyronine (reverse T3, rT3) increased appreciably within 8 hours after the first dose of dexamethasone, was maximum at 24-32 hours after beginning dexamethasone, and remained elevated for about 24 hours after discontinuing the steroid. The mean baseline serum rT3 was 58 ng/per 100 ml in treated hypothyroid patients and 119 ng per 100 ml in patients with Graves' disease; the corresponding maximal post-dexamethasone serum rT3 values were 87 and 170 serum concentration of 3,3',5-triiodothyronine (T3) decreased. The decrease in serum T3 was significant at about 24 hours after beginning dexamethasone and was maximal at about 30 hours in both groups of cases under study. The decrease in serum T3 persisted in treated hypothyroid cases for about 24-48 hours and in Graves' disease cases as long as studied, at least 5 days after discontinuing hexamethasone. The changes in serum rT3 and T3 could not be attributed to the effect of dexamethasone on serum protein binding of the iodothyronines because the dialyzable fractions of rT3 and T3 following steroid administration were not different from those before it. Serum T4 did not change appreciably in treated hypothyroid cases, but decreased in Graves' disease cases from a mean baseline value of 23.5 mug per 100 ml to 18.4 mug per 100 ml 3 days after beginning dexamethasone. In addition, 3 hyperthyroid cases were studied before, during, and after administration of dexamethasone, 2 mg every 6 h for 5 days. Serum rT3 increased again as noted above and the increase persisted until about 24 hours after the last dose of the steroid. Serum T3 decreased considerably and remained decreased as long as studied, at least 4 days after discontinuing the steroid. Serum T4 decreased appreciably in 2 of the 3 cases studied. The data suggest that 1) conversion of T4 to T3 and to rT3 may occur via two distinct pathways in the metabolism of T4; 2) the changes in serum rT3 and T3 observed in our study may be due in part at least to a steroid-induced 'shift' in the metabolism of T4 whereby conversion of T4 to T3 is diminished and that to rT3 is enhanced; 3) in addition to the effect on peripheral metabolism of T4, steroids appear to reduce the circulating thyroid hormones in Graves' disease by another mechanism, probably by reduction in thyroid secretion.     

URINARY EXCRETION OF 3,3',5'-TRIIODOTHYRONINE (REVERSE T3)

A simple and sensitive radioimmunoassay for reverse T3 in urine using small Sephadex G25 fine columns is described. The recovery of rT3 added to urine was on average 101.0 ± 4.2% (mean ± SEM). Detection limit was 4 pg/column. Urine excretion of rT3 (mean ± SD) was 72.0 ± 32.1 ng/24 hin 61 healthy euthyroid subjects with a slight increase with age (P < 0.05), 28.8 ± 18.2 ng/24 h in 12 hypothyroid patients and 183.6 ± 79.7 ng/24 h in 25 hyperthyroid patients.     

Comparison of inhibitory effects of 3,5,3'-triiodothyronine (T3), thyroxine (T4), 3,3,',5'-triiodothyronine (rT3), and 3,3'-diiodothyronine (T2) on thyrotropin-releasing hormone-induced release of thyrotropin in the rat in vitro.

In order to compare, in vitro, the TSH suppressive effects of iodothyronines, rat pituitary quarters were first preincubated with T4, T3, rT3, or 3,3'-diiodothyronine (T2) in Gey and Gey buffer containing 1% bovine serum albumin for 2 h at 37 C and then incubated at 37 C for 1 h with the iodothyronine under study and TRH. TSH released into the medium during incubation was compared to that released by control pituitary fragments, which were not exposed to iodothyronines. All four iodothyronines (T3, T4, rT3, and T2) were able to significantly inhibit the TRH-induced release of TSH from pituitary fragments in a dose range of 0.015-2.2 microgram/ml. However, much larger doses of sodium iodide (1.25 mg/ml) and diiodotyrosine (10 and 30 microgram/ml) had no significant effect on the release of TSH. Among T3, rT3, and T4, T3 was the most potent and rT3 was the least potent. The relative potency of T3:T4:rT3 appeared to be approximately 100:12:1 when estimated from the lowest doses that caused significant inhibition of TRH-induced release of TSH, and approximately 100:6:0.5 when estimated from the doses that caused 50% inhibition of TSH release; the TSH inhibiting potency of T2 was similar to that of rT3. The activity of T4 could not be explained entirely on the basis of contamination of T4 with T3 or by in vitro conversion of T4 to T3. Similarly, the available data suggested that rT3 and T2 possess some, albeit modest, intrinsic TSH-Suppressive activity. TSH-inhibiting activities of T3, T4, and rT3 were also studied using pituitary fragments from starved and iodine-deficient rats. There was no evidence of a change in the sensitivity of the thyrotroph to either T3 or T4 in starvation. Similarly, comparison of the responses to several doses of rT3 did not indicate any significant abnormality in the sensitivity of the thyrotroph to rT3 in starvation or iodine deficiency. However, comparison of the TSH-suppressive effects of T4 in the iodine-deficient and normal rat indicated a significant increase in the sensitivity of the thyrotroph to T4 in iodine deficiency. A similar trend was also evident in the effect of T3 in iodine deficiency, but it fell short of statistical significance.     

Properties of 5'-deiodinase of 3,3',5'-triiodothyronine in rat skeletal muscle

To characterize rT3 5'-deiodinase (5'D) in rat skeletal muscle, the effects of altered thyroid status and PTU on rT3 5'D were studied. rT3 5'D activity was measured by incubating homogenates of rat skeletal muscle with [125]rT3, iodine labelled in the outer ring, in the presence of 20 mmol/l DL-dithiothreitol. This activity was observed to increase significantly 24 h after a single sc injection of T3 (75 micrograms/kg). The increase following the daily administration of this drug (15 or 75 micrograms/kg) for 3 and 14 days was dependent on the dose and number of previous days of injection. A significant decrease in activity was observed 2 weeks after thyroidectomy. The addition of 0.1 mmol/l 6-n-propyl-2-thiouracil (PTU) to the incubation medium in vitro caused a marked reduction in the activity in homogenates of skeletal muscle from hypothyroid, euthyroid and hyperthyroid rats. PTU, present at 0.05% in the drinking water for 2 weeks virtually abolished it. The properties of rT3 5'D in rat skeletal muscle thus appear to be essentially the same as those of type I enzyme with respect to response toward altered thyroid status and PTU.     

3,3',5'-Triiodothyronine (reverse T3) and 3,3',5-triiodothyronine (T3) in fetal and adult sheep: studies of metabolic clearance rates, production rates, serum binding, and thyroidal content relative to thyroxine.

To examine the mechanism(s) responsible for high serum concentration of 3,3',5'-triiodothyronine (reverse T3, rT3) and low serum concentration of 3,3',5-triiodothyronine (T3) in the fetus, we studied metabolic clearance rates (MCR) and production rates (PR) of rT3, T3, and thyroxine (T4) in adult nonpregnant sheep and sheep fetuses in utero. The mean fetal MCR-rT3 was significantly lower than that in adult sheep, and the mean fetal PR-rT3 significantly higher. The mean fetal MCR-T3 was higher than, and the mean fetal PR-T3 similar to that in adult sheep. The mean fetal MCR-T4 and PR-T4 were both significantly higher than the corresponding values in adult sheep. The ratios of mean PR-rT3 to PR-T4 (rT3/T4) were similar in fetal and adult sheep. However, the ratio of mean PR-T3 to PR-T4 (T3/T4) in the fetal sheep was much lower than that in the adult sheep. The low fetal MCR-rT3 was not attributable to high serum binding of rT3. On the basis of the thyroidal content and kinetics of iodothyronines, it was estimated that whereas thyroidal secretion may account for nearly all of serum T3 (or PR-T3) in the fetus and about 50% of serum T3 in adults, it accounts for only about 3% of the serum rT3 (or PR-rT3) in both fetal and adult sheep. The results suggest a) that elevated serum rT3 in the fetus is due to its decreased clearance and increased production by mono-deiodination of T4, and b) that low serum T3 in the fetus is due to its increased clearance and decreased production by mono-deiodination of T4. In addition, on the basis of discordant changes in the production of T3 and rT3 from T4, it appears that there may exist two separate, apparently specific, iodothyronine deiodinating activities--one cleaving the iodine atom at the 5'-position and the other acting in the iodine atom at the 5-position of the T4 molecule; 5'-iodothyronine deiodinating activity is apparently reduced in the fetus.     

Secretion of thyroxine, 3,5,3'-triiodothyronine and 3,3'5'-triiodothyronine in euthyroid man

The secretion of iodothyronines from the normal human thyroid gland was assessed by radioimmunoassay analyses of the concentrations of thyroxine (T4), 3,5,3'-triiodothyronine (T3) and 3,3',5'-triiodothyronine (reverse T3, rT3) in thyroid venous and peripheral venous blood. The subjects studied were euthyroid patients undergoing parathyroid surgery. Measurements were carried out both under apparently normal conditions, following peroral T3 pre-treatment, and before and after acute administration of TSH into a thyroid artery. In the control subjects, significant gradients between thyroid venous and peripheral venous concentrations were recorded both for T4, T3 and rT3, suggesting that all three iodothyronines are secreted by the normal human thyroid. T3 pre-treatment seemed to reduce this secretion, and acute administration of TSH promoted rapid, marked, and concomitant increments in the thyroid venous concentrations of all three iodothyronines. Hence, it appears that not only T4 but also T3 and rT3 are secreted by the normal human thyroid gland, and that TSH stimulates the secretion of all three iodothyronines. On the other hand, calculations of the relative secretion rates uielded the relation T4:T3:rT3 as 85:9:1. This indicates that, in euthyroid subjects, most of T3, and almost all of rT3, is produced by extrathyroidal conversion of T4 and not by direct thyroidal secretion.     

Influence of fasting and refeeding on 3,3',5'-triiodothyronine metabolism in man

To determine the influence of prolonged fasting and refeeding on rT3 metabolism in man, five euthyroid obese subjects underwent a 13-day fast, followed by a refeeding period. Each patient received an iv dose of 25 muCi [125I]rT3 during the fed control period, on days 7 and 13 of the fast, and on the fourth day after refeeding with a regular diet. Serial blood and urine samples were obtained to determine serum rT3 clearance and production rates and the urinary tracer rT3 deiodination fraction. Significant increases in serum rT3 values were noted by day 7 and remained elevated for the duration of the fast (P less than 0.01). Normalization of rT3 levels occurred after 4 days of refeeding. Both 7 and 13 days of fasting decreased rT3 clearance [132.6 +/- 8.3 L/day (P less than 0.001) and 132.2 +/- 9.5 L/day (P less than 0.001), respectively] without changing rT3 production (36.8 +/- 5.3 and 33.0 +/- 3.7 nmol/D, respectively) compared to control values (207.0 +/- 10.9 L/day and 31.8 +/- 3.8 nmol/day, respectively). Refeeding did not restore rT3 clearance (151.2 +/- 6.9 L/day; P less than 0.002), but significantly reduced blood rT3 production (18.4 +/- 3.8 nmol/day; P less than 0.003). The fractional deiodination of rT3 was significantly reduced on day 7 (42.5 +/- 4.6%; P less than 0.01) and day 13 (41.9 +/- 3.7%; P less than 0.01) of fasting compared to the control value (69.2 +/- 2.8%), while refeeding only partially restored deiodination to baseline (48.4 +/- 5.1%; P less than 0.04). The clearance of rT3 was highly dependent on the fractional deiodination rate (r = 0.83; P less than 0.001). Although rT3 production remained constant during fasting, reduced rT3 production was seen on the fourth day of refeeding. This unique observation explained the fall in serum rT3 to prefasting levels after 4 days of refeeding when rT3 clearance was still inhibited. This study, in context with previous investigations, indicates that T4 conversion to circulating T3 and rT3 in fasting is a highly complex and multifaceted process requiring further investigation to elucidate the mechanism responsible for these alterations.     

3,3',5'-Triiodothyronine inhibits collagen-induced human platelet aggregation.

To clarify further the activity of rT3, we examined the effect of rT3 on collagen-induced platelet activation as reflected by aggregation, serotonin release, and protein phosphorylation. rT3, T4, T3, and triiodothyroacetic acid inhibited collagen-induced platelet aggregation and serotonin release from platelets in a dose-dependent manner. However, thyronine did not inhibit collagen-induced platelet aggregation. The concentration at which rT3 inhibited by 50% collagen-induced platelet aggregation was 30 +/- 4 (mean +/- SE) mumol/L. rT3, T4, and T3 did not differ significantly in their abilities to inhibit platelet aggregation. Moreover, rT3 inhibited collagen-induced phosphorylation of the 20-kilodalton protein (myosin light chain) in platelets. In contrast, rT3 did not inhibit 12-O-tetradecanoylphorbol 13-acetate (TPA)- or thrombin-induced platelet aggregation and inhibited only minimally TPA-induced 40-kilodalton protein phosphorylation. These results suggest that rT3 inhibits collagen-induced platelet activation by inhibiting the activity of myosin light chain kinase and that it may be interesting to investigate some kinds of activity of rT3.