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Serum thyroxine levels in patients receiving L-thyroxine suppression or replacement therapy.

Serum thyroxine (T(4)) and protein-bound iodine (PBI) levels were measured in 140 adult patients receiving L-thyroxine, 0.2 or 0.3 mg. daily for at least six months, as suppression or replacement therapy. All were judged to be clinically euthyroid at the time of the measurements. The mean PBI was 8.3 mug./100 ml. and the mean T(4) was 10.8 mug./100 ml. The range of values for the PBI was 3.5 to 13.2 and for the T(4) was 3.1 to 17.0. Because of the wide range of values, it is considered that the judgement of whether a patient is euthyroid on L-thyroxine therapy should be based primarily on clinical evaluation and only secondarily on serum T(4) or PBI levels.

Goiter↗

Free thyroxin index and direct measurements of free thyroxin compared for evaluating postpartum autoimmune thyroid dysfunction.

Measurement of free thyroxin (FT4) by a recently introduced commercial assay (Amerlex Free T4 RIA) was compared with the calculated free thyroxin index (FT4I) for serum from 104 postpartum women. Of these, 63 had transient thyroid dysfunction due to autoimmune thyroiditis, six had transient Graves' thyrotoxicosis, and 35 were euthyroid with no signs of autoimmune thyroid disease. The correlation between results for FT4 and the calculated FTI for 95 serum samples from women with no signs of autoimmune thyroiditis (r = 0.941; p = 0.0001) was almost identical to that for 270 serum samples from women with thyroid microsomal autoantibodies characteristic of autoimmune thyroiditis (r = 0.937; p = 0.0001). Furthermore, we observed no difference when the autoimmune group was subdivided according to low or high titers of thyroid microsomal antibodies. In no case did autoantibodies to thyroxin interfere with the FT4 assay. However, one woman had a spuriously low value for FT4I owing to interference by autoantibodies to triiodothyronine with the triiodothyronine resin uptake test. We conclude that the FT4 RIA assay provided diagnostic information in this group of postpartum women equivalent to that of the more elaborate procedure of determining FT4I.

Autoantibodies↗

Serum triiodothyronine, total thyroxine, and free thyroxine concentrations in horses.

The objectives of this experiment were to determine serum concentrations of triiodothyronine (T3), thyroxine (T4), and free thyroxine (fT4) at rest, following thyroid-stimulating hormone (TSH) administration, and following phenylbutazone administration in healthy horses. This was done to determine which available laboratory test can best be used for diagnosis of hypothyroid conditions in horses. Serum T3, T4, and fT4 concentrations in serum samples obtained before and after TSH stimulation and following phenylbutazone administration for 7 days were determined. Baseline values ranged from 0.21 to 0.80 ng of T3/ml, 6.2 to 25.1 ng of T4/ml, and 0.07 to 0.47 ng of fT3/dl. After 5 IU of TSH was administered IV, serum T3 values increased to 6 times baseline values in 2 hours. Thyroxine values increased to 3 times baseline values at 4 hours and remained high at 6 hours. Free T4 values increased to 4 times baseline values at 4 hours and remained high at 6 hours. Administration of 4.4 mg of phenylbutazone/kg, every 12 hours for 7 days significantly decreased T4 and fT4 values, but did not significantly affect serum T3 concentrations. It was concluded that a TSH stimulation test should be performed when hypothyroidism is suspected. Measurement of serum fT4 concentrations, by the single-stage radioimmunoassay, does not provide any additional information about thyroid gland function over that gained by measuring T4 concentrations. Phenylbutazone given at a dosage of 4.4 mg/kg every 24 hours, for 7 days did significantly decrease resting T4 and fT4 concentrations, but did not significantly affect T3 concentrations in horses.

Analysis of Variance↗

Effect of storage conditions on cortisol, total thyroxine, and free thyroxine concentrations in serum and plasma of dogs.

OBJECTIVE: To determine for dogs stability of cortisol, thyroxine (T4), and free thyroxine (fT4) in plasma and serum stored in glass or plastic tubes at -20, 4, 25, and 37 C. DESIGN: Prospective study. ANIMALS: Phase I, 7 Greyhounds; Phase II, 6 mixed-breed dogs. PROCEDURE: Phase I: blood was obtained after administration of thyroid-stimulating hormone and adrenocorticotropin. Serum and plasma samples from each dog were divided into 8 aliquots, 4 in glass and 4 in plastic tubes. A pair of aliquots, 1 in plastic and 1 in glass, were stored at -20, 4, 25, or 37 C for 5 days and then assayed for hormones. Phase II: blood was obtained without prior stimulation. For fT4 determination, serum from each dog was placed in plastic or glass tubes, assayed immediately, stored at -20 C for 5 days, and reassayed. Aliquots from each dog were also stored for 1 day at 4 or 25 C and then assayed. Samples for cortisol determination were handled as in phase I. RESULTS: Phase I: there was no effect of tube type (glass vs plastic) on cortisol. Cortisol concentrations decreased after storage in serum at 4, 25, and 37 C, and in plasma at 37 C, compared with storage at -20 C. There was no effect of sample type (serum or plasma) on T4. Thyroxine concentrations increased after storage at 37 C in glass, compared with storage at -20 C. The fT4 concentrations were lower in serum than plasma after storage at -20 C. Concentrations of fT4 increased after storage at 37 C in glass, compared with storage at -20 C. Phase II: the fT4 concentrations did not change after storage in any condition. There was no effect of tube type on cortisol concentrations. Serum cortisol concentrations decreased after storage at 37 C, compared with storage at -20 C. CLINICAL IMPLICATIONS: For cortisol, cooling of plasma is not necessary, but serum should be shipped cold. For T4 and fT4, serum is sufficient; contained within plastic tubes, samples can be shipped without cooling if assayed within 5 days.

Analysis of Variance↗

Clinical evaluation of new indicators of free thyroxine concentration and thyroxine-binding globulin capacity.

A rapid and simple method is described for the indirect measurement of free thyroxine (T4) concentration and maximal thyroxine-binding globulin (TBG) capacity applying a single kit of Thyopac-4. Initially, a value for total T4 is determined, after which the patient's serum is added to permit estimation of T4 corrected for abnormal TBG concentration. The ratio of increased radioactivity in the supernatant after the addition of patient's serum to that obtained using pooled normal serum is designated the TBG index, being directly related to the amount of TBG in the added serum. The index is significantly increased in hypothyroidism and pregnancy, and decreased in hyperthyroidism, correlating well with the TBG capacity measured by reverse-flow paper electrophoresis. A free T4 index (FTI) can be calculated from the values for T4 and TBG index, because the TBG index is reciprocally related to the serum uptake test (T3-resin). Values for FTI are significantly increased and decreased in hyper- and hypothyroidism, respectively, whereas they remain within the normal range in pregnancy and TBG deficiency. The FTI is shown to bear a nearly straight-line relationship to that calculated from separate estimations of T4 and the serum uptake test (T3-resin).

Evaluation Studies as Topic↗

Comparison of serum free thyroxine indices and "corrected" thyroxine tests.

A dual competitive protein-binding assay for serum thyroxine (T4) is described. The assay is called the "corrected" T4 index and can be performed simultaneously with the determination of total T4. This method correlated significantly with the free T4 index and the effective T4 ratio. The "corrected" T4 index was compared for precision and diagnostic accuracy with the effective T4 ratio and two different free T4 indices. Improved modifications of the thyroxine and triiodothyronine Sephadex uptake tests, used for calculation of the free T4 indices, are also reported. In spite of better precision, the "corrected" T4 index and effective T4 ratio gave twice as many incorect classifications of both hyper- and hypothyroidism as the free T4 indices. Previous reports on the effective T4 ratio and other basically similar tests might give an overly optimistic view of what can be expected in routine clinical practice. The free T4 index, which is a good estimate of the free T4 concentration, may still be the thyroid function test of choice when serum T4 and clinical impression are not in conformity.

Adult↗

Serum free thyroxine concentrations in normal euthyroid subjects and ones with high serum thyroxine binding globulin concentration.

Serum free thyroxine (fT4) was assayed by a commercial fT4 method in 30 normal euthyroid subjects, 19 pregnant females, 13 euthyroid subjects with high thyroxine binding globulin (TBG) and three with low or undetectable serum TBG concentration. In a number of these fT4 was also calculated on the basis of the application of the law of mass action to the binding situation. In states in which TBG was altered for congenital reasons both the experimentally determined and calculated fT4 were not significantly different from their respective means in the normal euthyroid population. Pregnant females had both lower experimental and theoretical free T4 concentrations. It is inferred from these data that TBG concentration per se is without effect on serum fT4 concentration.

Blood Proteins↗

Serum thyroxine binding capacity-dependent bias in five free thyroxine immunoassays: assessment with serum dilution experiments and impact on diagnostic performance.

OBJECTIVES: To assess the presence and magnitude of a serum thyroxine binding capacity (sBC)-dependent bias in five free thyroxine (FT(4)) immunoassays, compared with equilibrium dialysis (ED). The exhibited bias is confronted with clinical results from previous studies to evaluate its impact on FT(4) determination in sera with various sBC. DESIGN AND METHODS: The sera of three pregnant women and three non thyroidal ill (NTI) patients were serially diluted in an inert buffer to progressively decrease the sBC. FT(4) values were measured in diluted and undiluted samples with the six assays. RESULTS: As a function of increasing dilution performed on pregnancy sera, except for ED and Vitros ECi FT(4,) the other four FT(4) assay results decreased to different degrees, in the following order: two-step GammaCoat RIA< Elecsys< ADVIA:Centaur< Amerlex-MAB RIA. In sera from NTI patients, the decrease was more marked and found at high dilution with the Vitros ECi assay. Data from previous studies showed that FT(4) measured with biased assays were decreased only in samples from very severely NTI patients with low sBC and that FT(4) results in pregnancy sera with high sBC were not significantly biased. CONCLUSIONS: The dilution test is a sensitive alarm to assess sBC-dependent bias in FT(4) assays. For all FT4 assays and particularly when a bias is observed, documentation should be sought on the diagnostic performance of the assay and supported by a detailed clinical study including samples with low sBC. Physicians should still be educated about the limitations of FT(4) assays.

Blood Proteins↗

Effects of thyroxine binding on the stability, conformation, and fluorescence properties of thyroxine-binding globulin.

The effects of thyroxine (T4) on several molecular properties of human thyroxine-binding globulin (TBG) have been evaluated. Changes in the sedimentation constant and relaxation time indicate that TBG becomes more symmetric and compact when T4 is bound. This modification in structure is associated with an increase in the stability of TBG to denaturation by either acid or guanidinium chloride. T4 binding also produces changes in the emission and excitation spectra of TBG, reflecting different environments of the four tryptophanyl residues. T4 preferentially quenches residues in a less polar environment. In addition, it alters the effect of the collisional quencher, acrylamide, so as to indicate a shift in the environment of some of the exposed tryptophanyl residues.

Acrylamides↗

Thyroxine-binding globulin, triiodothyronine, thyroxine and thyrotropin in newborn infants and children.

Thyroxine-binding globulin (TBG), triiodothyronine (T3), thyroxine (T4) and thyrotropin (TSH) have been determined by radioimmunoassay in plasma of newborn infants and throughout childhood until puberty. Mean maternal TBG concentration was 1.65 +/- 0.09 mg/100 ml (SEM) and significantly higher (p less than 0.01) than cord blood levels of TBG (1.16 +/- 0.08 mg/100 ml (SEM). Throughout infancy and childhood TBG remained significantly elevated (p less than 0.01) compared to a middle age control group of healthy blood donors. T3, T4 and TSH concentrations behaved postnatally as known from previous studies. The T3 and T4 increase observed immediately after birth was not a secondary phenomenon due to changes in TBG concentration since this globulin did not change significantly during this period.

Aging↗

[Concentration levels of thyroxine binding globulin and thyroxine in the blood of euthyroid children. Reliability and diagnostic value of the T4/TBG quotiens--establishing reference values].

The most important transportprotein for thyroxine (T4) is the thyroxine-binding globulin (TBG). This protein and T4 were determined using the radioimmunoassay method in bloodserum of 443 euthyreotic children. The age-specific analysis showed high TBG and T4 concentrations in the postnatal phase, nearly constancy between 1. and 9. years with concentrations in the upper physiological range of adults, elevation of the values in the age of 10/11. years and definite attainment of the normal range for adults at 12/13. years. The importance of the T4/TBG quotient for thyroid gland diagnostics was discussed.

Adolescent↗

Salsalate and salicylate binding to and their displacement of thyroxine from thyroxine-binding globulin, transthyrin, and albumin.

Salsalate and its metabolite salicylate are inhibitors of thyroid hormone binding to serum transport proteins and reduce serum total thyroxine (T4) and triiodothyronine (T3) in vivo. The present study is the first to examine and compare salsalate and salicylate binding to human thyroxine-binding globulin (TBG), transthyretin (TTR), albumin (ALB), and whole human serum, and displacement by salsalate and salicylate of T4 from isolated TBG, TTR, ALB. Salsalate and salicylate binding were studied by ultrafiltration at salsalate concentrations of 0.24-1.16 mM (62-300 microg/mL) and salicylate concentrations of 0.0375-2.25 mM (6.0-360 microg/mL). T4 displacement by salsalate and salicylate from TBG, TTR, and ALB was studied by equilibrium dialysis at salsalate concentrations of 0 to 1.52 mM (0-393 microg/mL) and salicylate concentrations of 0 to 60 mM (0-9600 microg/mL). In normal human serum, 95% of salsalate and 76% of salicylate were protein-bound. In the presence of isolated ALB, 89% of salsalate and 64% of salicylate were ALB-bound. Both salsalate and salicylate inhibited T4 binding to all 3 major T4 binding proteins. Both displaced proportionately more T4 from TBG compared with TTR and ALB. The principal site to which both salsalate and salicylate in serum is bound (ALB) is different from the principal site from which they displace T4 (TBG). Salsalate potency in displacing T4 from TBG and ALB was approximately 100-fold greater than salicylate potency.

Binding, Competitive↗

Underestimates and overestimates of total thyroxine concentrations caused by unwanted thyroxine-binding protein effects.

The first evidence of unwanted serum protein effects on analogue-based total thyroxine (T4) determinations came from a study that varied serum protein concentrations while total T4 concentrations were constant. The present study approached this issue by varying total T4 concentrations while protein concentrations were constant. Four analogue-based total T4 immunoassays were applied to solutions that contained either free T4 without binding protein, a T4-binding protein without T4, or protein-bound T4. When total T4 concentrations were 3-12 microg/dL, the assays reported total T4 determinations that ranged from none detected to 23 microg/dL. These T4 determinations reflected the protein to which T4 was bound, in addition to the level of T4. Total T4 was underrepresented when T4 was unbound, or thyroxine-binding globulin (TBG) bound. Total T4 was overrepresented when T4 was albumin-bound, or transthyretin-bound. There were substantial disparities among assays applied to the same total T4 solutions. These assays reported no detectable T4 when applied to T4-binding protein solutions without protein-bound T4. Nonetheless, T4-binding proteins contributed to the underestimates and overestimates of protein bound T4. Different forms of protein bound T4 were quantified differently, evidence that protein-T4 complexes persist during quantification. We attribute the unexpected total T4 values to a combination of incomplete protein-bound T4 release from T4-binding proteins during quantification, and variably inaccurate quantifications of the protein-bound T4 that remained.

Albumins↗

A variant of porcine thyroxine-binding globulin has reduced affinity for thyroxine and is associated with testis size.

The field of genomics applies the dissection of genetic differences toward an understanding of the biology of complex traits. Quantitative trait loci (QTL) for testis size, plasma FSH in boars, and body composition (backfat) have been identified near the centromere on the X chromosome in a Meishan-White Composite resource population. Since thyroid function affects Sertoli cell development and adult testis size in rodents, and thyroxine-binding globulin (TBG) maps to this region on the porcine X chromosome, TBG was a positional candidate gene for testis size. We discovered a polymorphism in exon 2 of the porcine TBG gene that results in an amino acid change of the consensus histidine to an asparagine. This single nucleotide polymorphism (SNP) resides in the ligand-binding domain of the mature polypeptide, and the Meishan allele is the conserved allele found in human, bovine, sheep, and rodent TBG. Binding studies indicate altered binding characteristics of the allelic variants of TBG with the asparagine (White Composite) isoform having significantly greater affinity for thyroxine than the histidine (Meishan) isoform. Alternate alleles in boars from the resource population are also significantly associated with testis weight. Therefore, this polymorphism in TBG is a candidate for the causative variation affecting testis size in boars.

Amino Acid Sequence↗

Comparisons between serum concentrations of thyroxine and thyroxine-binding proteins in samples simultaneously obtained from capillary, peripheral vein, central vein and aorta in newborn infants.

A total number of 40 newborn infants with various maturity were studied: 13 babies without perinatal events, 19 infants recovered from transient diseases, 6 infants with idiopathic respiratory distress syndrome and 2 infants with asphyxia indicating artificial ventilation. Comparisons were performed between serum concentrations of thyroxine (T4), thyroxine-binding globulin (TBG), prealbumin (TBPA) and albumin (Alb) in capillary versus peripheral vein, aorta versus central vein and, finally, in peripheral versus central veins. In healthy infants serum T4 concentrations in capillary blood and peripheral vein did not differ significantly. Although serum concentrations of thyroid hormone-binding proteins tended to be increased in aortic compared to central venous specimens no statistically significant differences appeared. In infants in good clinical conditions serum T4, TBG, TBPA, and Alb levels were 6--8% higher in peripheral than in central veins, possibly primarily due to a hemo-concentrating effect of venous stasis. Therefore, in evaluation of the thyroid variables in newborn infants the technique of blood sampling must be considered. In most infants with idiopathic respiratory distress syndrome and in one asphyxiated baby a remarkable tendency to a low serum TBG and T4 concentration in peripheral compared to central vein samples were observed.

Aorta↗

Maternal free thyroxine and thyroxine-binding globulin during pregnancy ending in congenital malformations in the offspring.

Thyroxine-binding globulin (TBG) and free thyroxine (FT4) were studied in 172 apparently healthy women giving birth to malformed children. The malformations were divided into two groups: minor malformations (group A) and major malformations (group B). The blood samples were usually drawn in the first trimester. Group A showed no significant difference in TBG values compared to women with healthy children. Group B showed lower, but not significantly lower values in earlier pregnancy. However, from the 15th-16th week of pregnancy the TBG values were significantly lower. The FT4 values were in the hyperthyroid range in as much as 20% of the women, compared to the reported rate of 0.2% hyperthyroidism in pregnant women.

Congenital Abnormalities↗

The contribution of thyroxine-binding prealbumin to the binding of thyroxine in human serum, as assessed by immunoadsorption.

An immunoadsorption technique employing a rabbit antiserum specific for human serum prealbumin has been devised to remove thyroxine (T(4))-binding prealbumin (TBPA) from serum completely without affecting the T(4)-binding activity of thyroxine-binding globulin (TBG) or the concentration of the other major proteins in serum. As judged from the proportion of T(4) associated with the antigen-antibody precipitate, only about 15% of the endogenous T(4) is bound by TBPA, a value considerably less than that indicated by electrophoretic methods. As judged from the increase in the proportion of free T(4) that followed immunoadsorption of TBPA, TBPA does act as one determinant of the proportion of free T(4) but is far less important than TBG in this respect. A decrease in the T(4)-binding capacity of TBPA cannot solely account for the increase in the proportion of free T(4) in the sera of ill patients, since a comparable increase does not occur in normal sera after complete removal of TBPA. From data obtained in normal and abnormal sera before and after immunoadsorption of TBPA, estimates of the equilibrium constants for the interactions between T(4) and its binding proteins, as they exist in serum, have been derived. The values obtained were: K(ALB), 6.2 x 10(5); K(TBPA), 2.3 x 10(8); and K(TBG), 1.7 x 10(10).

Adsorption↗

Metabolism of thyroxine-binding globulin in man. Abnormal rate of synthesis in inherited thyroxine-binding globulin deficiency and excess.

It has been previously suggested that inherited thyroxine-binding globulin (TBG) abnormalities in man may be due to mutations at a single X-chromosome-linked locus controlling TBG synthesis. However, abnormalities in TBG degradation have not been excluded. The availability of purified human TBG and its successful labeling with radioiodide allowed us to examine such possibility. Human TBG was purified by affinity chromatography, labeled under sterile conditions with 131I or 125I,, and mixed with [125I]thyroxine (T4) or [131I]T4, respectively, before their intravenous injection. Blood and urine samples were collected over a 10-day period, and the turnover parameters were calculated. In eight normal volunteers mean values +/-SD for TBG and T4 respectively, were as follows: Half time (t1/2) 5.3 +/- 0.4 and 7.0 +/- 0.6 days; distribution space (DS) 7.2 +/- 1.0 and 10.8 +/- 1.2 liters; and total daily degradation (D) 0.211 +/- 0.053 and 0.088 +/- 0.011 mumol/day. In all subjects, t1/2 of TBG was shorter than that of T4; and the DS was smaller. 2.4 mol of TBG was degraded for each mole of T4. In five of six subjects from four families, comprising hemizygous and heterozygous carriers of TBG absence, decrease, and excess, the t1/2 and DS for TBG were within the normal range. The D of TBG was proportional to the serum concentration of the protein. Changes in the T4 kinetics in these patients were compatible with euthyroidism and with the known alterations in the extrathyroidal T4 pool associated with the changes in serum TBG concentration. A striking decrease in the t1/2 of TBG was found only in a patient with acquired diminution in TBG concentration and in patients with thyrotoxicosis or other conditions apparently unrelated to thyroid dysfunction. TBG t1/2 was 2.5 days in a patient with multiple myeloma and 3.6 days in two patients with thyrotoxicosis. Decreased TBG t1/2 was also observed in three of six patients with nonthyroidal pathology and was associated with an increase in TBG D disproportionate to their level of serum TBG. These studies indicate that changes in TBG concentration in patients with X-chromosome-linked TBG abnormalities are due to alterations in its rate of synthesis. In other conditions, abnormalities of TBG degradation and/or rate of synthesis may be found.

Adult↗