Insulin-degrading enzyme higher in subjects with metabolic syndrome.

Metabolic syndrome (MS) is comprised of a cluster of abnormalities in glucose, lipid, and vascular homeostasis, which is most commonly linked to abdominal obesity. MS heralds increased risk for development of diabetes and is linked to impairment in insulin signaling. Insulin-degrading enzyme (IDE) is one of the mechanisms through which insulin blood levels are maintained. It has been previously suggested that controlling IDE levels could provide yet another potential therapeutic approach in diabetes. Here we aim to investigate whether changes in serum IDE levels correlate with the severity of MS. Using a highly sensitive ELISA assay of active IDE in human serum, we found a strong correlation between circulating IDE levels and circulating levels of triglycerides, insulin, and c-peptide and an inverse correlation with HDL cholesterol (HDLc). Serum IDE levels were higher in MS subjects than in control subjects. Hence, circulating IDE may serve as a tool to identify subjects with abnormal insulin metabolism, possibly those with MS that are at risk to develop diabetes.

Circulating 3,3', 5'-triiodothyronine (reverse T3) in the human newborn.

Serum concentrations of 3,3',5'-triiodothyronine (reverse T3 rT3), 3,3',5-triiodothyronine (T), and thyroxine (T4) were measured in cord blood and invenous blood samples obtained between 2 h and 30 days of postnatal life from healthy full-term newborn infants. The mean serum rT3 concentration of (mean plus or minus SE) 151 plus or minus 12 ng per 100 ml in 18 cord blood samples was significantly higher than the level (41 plus or minus 2 ng per 100 ml) in 27 normal adult sera; the corresponding mean serum T4 of 12.7 plus or minus 0.8 mug per 100 ml in cord blood also was significantly higher than that (8.6 plus or minus 1.9 mug per 100 ml) in 108 normal adults. By contrast, the mean serum T3 concentration in 15 cord blood samples, 24 plus or minus 3 mg per 100 ml, was significantly lower than the value of 126 plus or minus 3.2 ng per 100 ml measured in 108 normal adults. At 4 h of age the mean serum rT3 concentration (165 plus or minus 13 ng per 100 ml) in six newborns was 4ot significantly different from that in paired cord blood samples (194 plus or minus 25 ng per 100 ml); on the other hand, whenever, studied, the mean serum T3 and T4 levels were significantly higher at 4 h than at birth. The failure of serum rT3 concentrations to rise after delivery in response to the early neonatal thyrotropin (TSH) surge and at a time when serum T3 and T4 levels increase significantly prompted a study of the rT3 response to 10 IU of intramuscular TSH in three healthy adult subjects. Just as in the newborns, serum rT3 failed to rise appreciably in these subjects, even though serum T3 and T4 showed the expected increments. Serum rT3 concentrations in 1-4 day-old newborn infants did not differ significantly from values in the cord blood but were significantly lower in older neonates. The mean serum rT3 level in 5-7-day-old infants was higher than that in normal adults, but in 9-11 day and 20-30-day-old infants, mean rT3 values were statistically similar to the adult value. The mean serum T3 concentrations in neonates between 1-30 days old were either higher than or comparable to the values of normal adults. The mean serum T4 concentrations in neonates between birth and 30 days of age were significantly higher than the mean adult level. The mean serum rT3 to T4 ratios (rT3/T4) were elevated in 1-4-day-old neonates; the values in older neonates were similar to those in adults. These results suggest that (a) factors other than TSH are important modulators of serum rT3 in man; (b) high serum rT3 concentration in the newborn becomes comparable to that in the normal adult by 9-11 days of neonatal life.

Identification of imidazo[1,2-b]pyridazine TYK2 pseudokinase ligands as potent and selective allosteric inhibitors of TYK2 signalling.

As a member of the Janus (JAK) family of non-receptor tyrosine kinases, TYK2 mediates the signaling of pro-inflammatory cytokines including IL-12, IL-23 and type 1 interferon (IFN), and therefore represents an attractive potential target for treating the various immuno-inflammatory diseases in which these cytokines have been shown to play a role. Following up on our previous report that ligands to the pseudokinase domain (JH2) of TYK2 suppress cytokine-mediated receptor activation of the catalytic (JH1) domain, the imidazo[1,2-b]pyridazine (IZP) 7 was identified as a promising hit compound. Through iterative modification of each of the substituents of the IZP scaffold, the cellular potency was improved while maintaining selectivity over the JH1 domain. These studies led to the discovery of the JH2-selective TYK2 inhibitor 29, which provided encouraging systemic exposures after oral dosing in mice. Phosphodiesterase 4 (PDE4) was identified as an off-target and potential liability of the IZP ligands, and selectivity for TYK2 JH2 over this enzyme was obtained by elaborating along selectivity vectors determined from analyses of X-ray co-crystal structures of representative ligands of the IZP class bound to both proteins.

The structure of a calmodulin mutant with a deletion in the central helix: implications for molecular recognition and protein binding.

BACKGROUND: Calmodulin (CaM) is the major calcium-dependent regulator of a large variety of important intracellular processes in eukaryotes. The structure of CaM consists of two globular calcium-binding domains joined by a central 28-residue alpha helix. This linker helix has been hypothesized to act as a flexible tether and is crucial for the binding and activation of numerous target proteins. Although the way in which alterations of the central helix modulate the molecular recognition mechanism is not known exactly, the relative orientation of the globular domains seems to be of great importance. The structural analysis of central helix mutants may contribute to a better understanding of how changes in the conformation of CaM effect its function. RESULTS: We have determined the crystal structure of a calcium-saturated mutant of chicken CaM (mut-2) that lacks two residues in the central helix, Thr79 and Asp80, at 1.8 A resolution. The mutated shorter central helix is straight, relative to that of the wild-type structure. The loss of a partial turn of the central alpha helix causes the C-terminal domain to rotate 220 degrees around the helix axis, with respect to the N-terminal domain. This rotation places the two domains on the same side of the central helix, in a cis orientation, rather than in the trans orientation found in wild-type structures. CONCLUSIONS: The deletion of two residues in the central helix of CaM does not distort or cause a bending of the linker alpha helix. The main consequence of the mutation is a change in the relative orientation of the two globular calcium-binding domains, causing the hydrophobic patches in these domains to be closer and much less accessible to interact with the target enzymes. This may explain why this mutant of CaM shows a marked decrease in its ability to activate some enzymes while the mutation has little or no effect on its ability to activate others.

A human monoclonal antibody to insulin.

To elucidate the immune aspects of insulin-dependent diabetes mellitus (IDDM), we attempted to generate human monoclonal anti-insulin antibodies by fusing peripheral blood lymphocytes obtained from 10 insulin-treated IDDM patients with cells from a human lymphoblastoid cell line. Hybridomas that secreted immunoglobulins appeared in 9 of 400 wells. One of these hybridomas secreted anti-insulin antibody of the IgM class. The lymphocytic partner of this hybridoma was obtained from an IDDM patient who had undetectable levels of antibodies to insulin in his serum. Thus, by employing the hybridoma technique, it was possible to reveal the presence of insulin-sensitized B-lymphocytes in a patient who was serologically negative for anti-insulin antibodies. The monoclonal antibody recognized intact human insulin and insulins of other species, but not isolated A- and B-chains. This indicates that the antibody was functionally an autoantibody directed to an epitope formed by the native conformation of a highly conserved portion of the insulin molecule. This is the first report of a human hybridoma antibody to insulin.

Periplasmic binding protein structure and function. Refined X-ray structures of the leucine/isoleucine/valine-binding protein and its complex with leucine.

The three-dimensional structure of the native unliganded form of the Leu/Ile/Val-binding protein (Mr = 36,700), an essential component of the high-affinity active transport system for the branched aliphatic amino acids in Escherichia coli, has been determined and further refined to a crystallographic R-factor of 0.17 at 2.4 A resolution. The entire structure consists of 2710 non-hydrogen atoms from the complete sequence of 344 residues and 121 ordered water molecules. Bond lengths and angle distances in the refined model have root-mean-square deviations from ideal values of 0.05 A and 0.10 A, respectively. The overall shape of the protein is a prolate ellipsoid with dimensions of 35 A x 40 A x 70 A. The protein consists of two distinct globular domains linked by three short peptide segments which, though widely separated in the sequence, are proximal in the tertiary structure and form the base of the deep cleft between the two domains. Although each domain is built from polypeptide segments located in both the amino (N) and the carboxy (C) terminal halves, both domains exhibit very similar supersecondary structures, consisting of a central beta-sheet of seven strands flanked on either side by two or three helices. The two domains are far apart from each other, leaving the cleft wide open by about 18 A. The cleft has a depth of about 15 A and a base of about 14 A x 16 A. Refining independently the structure of native Leu/Ile/Val-binding protein crystals soaked in a solution containing L-leucine at 2.8 A resolution (R-factor = 0.15), we have been able to locate and characterize an initial, major portion of the substrate-binding site of the Leu/Ile/Val-binding protein. The binding of the L-leucine substrate does not alter the native crystal structure, and the L-leucine is lodged in a crevice on the wall of the N-domain, which is in the inter-domain cleft. The L-leucine is held in place primarily by hydrogen-bonding of its alpha-ammonium and alpha-carboxylate groups with main-chain peptide units and hydroxyl side-chain groups; there are no salt-linkages. The charges on the leucine zwitterion are stabilized by hydrogen-bond dipoles. The side-chain of the L-leucine substrate lies in a depression lined with non-polar residues, including Leu77, which confers specificity to the site by stacking with the side-chain of the leucine substrate.(ABSTRACT TRUNCATED AT 400 WORDS)

Structure of the L-leucine-binding protein refined at 2.4 A resolution and comparison with the Leu/Ile/Val-binding protein structure.

The three-dimensional X-ray structure of the leucine-binding protein (36,900 Mr and 346 residues), an active transport component of Escherichia coli, has been determined by the method of molecular replacement, using the refined structure of the Leu/Ile/Val-binding protein (344 residues) as the model structure. The two amino acid-binding proteins have 80% sequence identity and, although both crystallize in the same space group, they have very different unit cell dimensions. The rotation function yielded one significant peak, which subsequently led to a single self-consistent translation function solution. The model was first refined by the constrained least-squares method, with each of the two domains of the molecule treated separately to allow for any small change in the relative orientation of the two domains. The model was then modified in order to reflect the 72 changes in amino acid side-chains and two insertions in going from the Leu/Ile/Val-binding protein sequence to that of the L-leucine-binding protein. Final structure refinement, using the restrained least-squares technique, resulted in an R-factor of 0.20 for 13,797 reflections to a resolution of 2.4 A. The model is comprised of 2600 protein atoms and 91 solvent molecules. The L-leucine-binding protein structure is, as expected, very similar to the Leu/Ile/Val-binding protein structure; both are in the unliganded conformation with the cleft between the two domains wide open and easily accessible. The superimposing of the structures yields a root-mean-square difference of 0.68 A in the alpha-carbon atoms of the 317 equivalent residues. The five regions of the leucine-binding protein structure that differ by more than 1.6 A from the Leu/Ile/Val-binding protein structure are far from the major portion of the ligand-binding site, which is located in one domain of the bilobate protein. Between the structures, there are three differences in the amino acid side-chains that form the major portion of the substrate-binding sites. These substitutions, by themselves, fail to clearly explain the differences in the specificities for branched aliphatic amino acids.

Structure of a retro-binding peptide inhibitor complexed with human alpha-thrombin.

The crystallographic structure of the ternary complex between human alpha-thrombin, hirugen and the peptidyl inhibitor Phe-alloThr-Phe-O-CH3, which is acylated at its N terminus with 4-guanidino butanoic acid (BMS-183507), has been determined at 2.6 A resolution. The structure reveals a unique "retro-binding" mode for this tripeptide active site inhibitor. The inhibitor binds with its alkyl-guanidine moiety in the primary specificity pocket and its two phenyl rings occupying the hydrophobic proximal and distal pockets of the thrombin active site. In this arrangement the backbone of the tripeptide forms a parallel beta-strand to the thrombin main-chain at the binding site. This is opposite to the orientation of the natural substrate, fibrinogen, and all the small active site-directed thrombin inhibitors whose bound structures have been previously reported. BMS-183507 is the first synthetic inhibitor proved to bind in a retro-binding fashion to thrombin, in a fashion similar to that of the N-terminal residues of the natural inhibitor hirudin. Furthermore, this new potent thrombin inhibitor (Ki = 17.2 nM) is selective for thrombin over other serine proteases tested and may be a template to be considered in designing hirudin-based thrombin inhibitors with interactions at the specificity pocket.

Crystallographic determination of the structures of human alpha-thrombin complexed with BMS-186282 and BMS-189090.

The crystallographic structures of the ternary complexes of human alpha-thrombin with hirugen (a sulfated hirudin fragment) and the small-molecule active site thrombin inhibitors BMS-186282 and BMS-189090 have been determined at 2.6 and 2.8 A. In both cases, the inhibitors, which adopt very similar bound conformations, bind in an antiparallel beta-strand arrangement relative to the thrombin main chain in a manner like that reported for PPACK, D-Phe-Pro-Arg-CH2Cl. They do, however, exhibit differences in the binding of the alkyl guanidine moiety in the specificity pocket. Numerous hydrophilic and hydrophobic interactions serve to stabilize the inhibitors in the binding pocket. Although PPACK forms covalent bonds to both serine and the histidine of the catalytic triad of thrombin, neither BMS-186282 nor BMS-189090 bind covalently and only BMS-186282 forms a hydrogen bond to the serine of the catalytic triad. Both inhibitors bind with high affinity (Ki = 79 nM and 3.6 nM, respectively) and are highly selective for thrombin over trypsin and other serine proteases.

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.

The response of newborn sheep to TRH with and without somatostatin.

Hormone responses to a bolus injection of thyrotropin releasing hormone (TRH) were studied in 8 newborn lambs between 6 and 19 hours of age. The effect of a bolus injection and 45 min infusion of somatostatin (SRIF) on these responses was studied in 2 other animals. Serial measurements of serum TSH, prolactin, triiodothyronine (T3) and thyroxine (T4) were conducted for 2 to 6 h in all animals. Mean baseline T4 and T3 concentrations were 12.6 microng/dl and 221 ng/dl, respectively, both significantly higher than values in fetal or adult animals. These high values were due to the events of parturition. In spite of the high baseline T4 and T3 levels, there were rapid and significant increases in both TSH and prolactin concentrations in response to TRH alone. The TSH response evoked further increments in serum T3 and T4 concentrations observed at 30 min and 60 min, respectively, both subsequently increasing progressively through 6 h. During the 45 min period of SRIF infusion, the TSH T4 and T3 responses to the zero time TRH injection were minimal. However, after discontinuing SRIF, late increases in TSH, T4 and T3 were observed. The results indicate that the hyperiodothyroninemia characteristic of the newborn period does not block the response to exogenous TRH, whereas the inhibitory effect of exogenous SRIF is observed in the newborn as in the adult. The increased endogenous TRH secretion presumably responsible for the neonatal TSH surge may be overriding the negative feedback effect of thyroid hormones.

Serum 3,5,3'-triiodothyronine and 3,3',5'-triiodothyronine concentrations during acute heat load.

To determine the changes in thyroid hormone metabolism during short periods of exposure to heat, 30 euthyroid healthy male volunteers (aged 23--40 yr) were placed in a climatic chamber for 2 h (35 C, 50% relative humidity). The subjects were at complete rest during the first hour and performed light work (40 watts) during the second hour. Blood samples for T4, T3 and rT3 were drawn at 0, 60, and 120 min. Rectal temperature and heart rate were monitored continuously. No significant changes in T4, T3, rT3, rectal temperature, or heart rate were observed after the first hour (basal levels, 8.5 +/- 0.3 microgram/dl, 160 +/- 5 ng/dl, 14.5 +/- 2.5 ng/dl, 37.2 +/- 0.1 C, and 78 +/- 8 beats/min, respectively; mean +/- SEM). During the second hour, a significant rise in body temperature was recorded (38.5 +/- 0.1 C), accompanied by a significant decrease in mean serum T3 concentration and a rise in mean serum rT3 concentration, T4 concentration remained unchanged. Our findings suggest that, parallel to the elevation in body temperature, there is a shift in the conversion of T4 to the noncalorigenic rT3 metabolite rather than to T3.

Thyroxine concentration in human milk.

Thyroxine (T4) concentrations were measured by RIA in 37 milk samples from 19 healthy eutryroid mothers, obtained between 3-165 days postpartum. The mean milk T4 concentration in the first week postpartum was 0.38 +/- 0.07 microgram/100 ml (mean +/- SEM). The mean T4 concentrations between 8 and 48 days rose to 4.27 +/- 0.50 microgram/100 ml (mean +/- SEM), and decreased to 1.11 +/- 0.25 microgram/100 ml (mean +/- SEM) after 50 days postpartum. The data suggest that human milk can provide a significant exogenous source of T4 to the infant. In the hypothyroid infant, the amount of T4 in human milk may delay clinical recognition of the disease. Although this exogenous source of T4 may alleviate the disease, it is insufficient to prevent the detrimetal effects of hypothyroidism.

Hypothalamic-pituitary function in patients with prolonged coma.

In eight patients (six males and two females) hospitalized for prolonged coma (164-1320 days; five patients were in traumatic coma, two after cardiac arrest during surgery, and one after a venomous scorpion sting), diurnal variation of cortisol secretion, human (L) GH, PRL, TSH secretion as well as the release of the pituitary hormones after TRH and L-dopa stimulation were assessed. Serum T4 cortisol, hGH, hPRL, and hTSH levels were normal. The cortisol diurnal variation was preserved in six patients. In seven of eight patients, the hPRL and hTSH responses to TRH test were normal and in five patients, there was a rise of more than 150% in serum hGH concentrations. After L-dopa administration, five patients responded with a rise in hGH. In three patients, there was no response of hGH to TRH or L-dopa. Five patients responded with significant reduction in the serum hPRL concentration after L-dopa, and in three patients, no change was observed. It was concluded that patients with prolonged coma have variable hypothalamic pituitary function. They preserve the cortisol diurnal variation and the hypothalamic-pituitary-thyroid axis as well as PRL secretion; however, hGH secretion may respond abnormally to various stimuli.

Cross genotype sex hormone treatment in two cases of hypogonadal osteoporosis.

BACKGROUND: Sex hormone deficiency is the most common cause of bone loss. Reduced bone mass and an increased risk for osteoporotic fractures have been described in hypogonadal subjects of both sexes. We present here the results of treating two patients showing abnormal sexual differentiation (an XX male and an XY female), who suffered from bone loss related to sex hormone deficiency, with cross genotype sex hormones. SUBJECTS AND METHODS: Patient 1 was an asymptomatic 39-yr-old XY female with complete androgen insensitivity. Her testes had been removed, and she later discontinued estrogen treatment. Patient 2, a 37-yr-old XX male, had congenital adrenal hyperplasia, which led to a masculine phenotype. He was ovariectomized and reared as a male. He was treated with glucocorticoids but refused androgen treatment for many years. We treated both patients with phenotypically matched sex hormones (patient 1 received conjugated estrogens 1.25 mg/day, and patient 2 received 250 mg testosterone every 4 weeks) and followed their bone mineral density (BMD) using dual-energy X-ray absorptiometry, urine calcium, and hydroxyproline excretion. RESULTS: Before treatment both patients had low sex hormones and highly elevated gonadotropins. As a result of treatment urine hydroxyproline excretion decreased from 45 and 26.7 mg/g creatinine to 15 and 15.9 mg/g creatinine in patients 1 and 2 respectively. In patient 1, lumbar BMD rose from 0.912gr/cm2 to 0.976gr/cm2 and femoral neck BMD rose from 0.716gr/cm2 to 0.836gr/cm2 after 4 years of treatment. In patient 2, lumbar BMD rose from 0.717gr/cm2 to 0.815gr/cm2 and the femoral neck BMD rose from 0.509gr/cm2 to 0.635gr/cm2 after 27 months of treatment. CONCLUSIONS: Phenotypically-matched sex hormone therapy in patients with abnormal sexual differentiation is essential not only to maintain external appearance but also for the preservation of bone mass.

Effect of intraamniotic fluid thyroxine injection on fetal serum and amniotic fluid iodothyronine concentrations.

Seven hundred micrograms of T4 were injected into the amniotic cavity 24 h before delivery of five pregnant women scheduled for elective cesarean section at term. T4, T3, and rT3 concentrations were measured by RIA in amniotic fluid obtained at the time of the injection and in amniotic fluid and cord serum samples collected at delivery. Iodothyronine concentrations also were determined on cord samples from 24 full term control infants. The geometric mean serum T4 concentration in the experimental infants was 27.2 micrograms/dl, almost 3 times that of the control population (10.3 micrograms/dl); serum rT3 concentrations were markedly elevated to a mean of 657 ng/dl, compared to 254 ng/dl in control infants. The mean serum T3 concentration was slightly but significantly increased to 61.3 ng/dl (control, 48.3 ng/dl; P less than 0.02). Amniotic fluid T4, T3, and rT3 concentrations all increased significantly. T4 injection into the amniotic fluid is an effective method of increasing fetal serum T4 concentrations. The preferential pathway of monodeiodination of the injected T4 in the human fetus is to rT3 rather than T3.

11 Beta-hydroxylase deficiency congenital adrenal hyperplasia: update of prenatal diagnosis.

Hormonal measurements in maternal urine and amniotic fluid (AF) during pregnancy and/or at delivery correctly predicted the postnatal diagnosis of 11 beta-hydroxylase deficiency congenital adrenal hyperplasia (11 beta-OH deficiency CAH) in 7 fetuses at risk. In the 4 affected ones, maternal urinary tetrahydro-11-deoxycortisol (THS) excretion was high during the first trimester [0.3-2.2 mg/day (1.1-7.7 mumol/day)] and rose further during the third trimester [0.5-3.5 mg/day (1.8-12.3 mumol/day)] compared to urinary THS excretion in 20 normal pregnancies of the same gestational age (P less than 0.01). In 1 mother, dexamethasone administration (2 mg/day for 72 h) greatly reduced urinary THS excretion (and plasma steroid levels). Urinary THS excretion was low after delivery in these mothers, in normal pregnancies, and in parents of affected individuals [less than 0.05 mg/day (less than 0.08 mumol/day); P = NS]. However, 2 of the 3 heterozygous mothers who carried nonaffected fetuses excreted moderately increased amounts of THS during pregnancy, ranging from 0.15-0.26 mg/day (0.53-0.91 mumol/day), significantly higher than normal (P less than 0.01). Although urinary THS excretion in these mothers was similar to that in 2 mothers with affected fetuses early in pregnancy, urinary THS excretion was higher in mothers with affected compared to those with nonaffected fetuses after the first trimester (P less than 0.01). AF THS and 11-deoxycortisol concentrations were markedly elevated in pregnancies with affected fetuses (P less than 0.01), but normal in nonaffected ones. AF delta 4-androstenedione levels were high in 2 pregnancies and borderline elevated in a third. Although the AF tetrahydrocortisol and tetrahydrocortisone levels were always within the normal range, the AF THS to tetrahydrocortisol plus tetrahydrocortisone ratio was significantly elevated in all pregnancies with affected fetuses (2.8-5.5; P less than 0.01) and normal in nonaffected ones (0.48-1.2; P = NS) compared to that in 160 normal pregnancies [0.64 +/- 0.34 (+/- SD)]. AF 17-hydroxyprogesterone, testosterone, and 11-deoxycorticosterone levels were normal in all pregnancies. Maternal plasma 11-deoxycortisol and delta 4-androstenedione concentrations, determined sequentially throughout gestation, were variable and did not contribute to prenatal diagnosis. All affected infants were born hyperpigmented, 2 were large for gestational age, and the female was severely virilized. In the first week of life 2 males developed severe hypertension with seizures and adrenal insufficiency, respectively.(ABSTRACT TRUNCATED AT 400 WORDS)

Amniotic fluid 3,3',5'-triiodothyronine in the detection of congenital hypothyroidism.

Amniotic fluid rT3 levels were measured during pregnancy in two women who previously gave birth to infants suffering from neonatal hypothyroidism. In the first case, hypothyroidism was strongly suspected because of repeated low levels of rT3 in the amniotic fluid (20-64 ng/dl) at 16 and 31 weeks of gestation. A normal infant was delivered. He is now 10 months old and taking no treatment; he has no clinical or laboratory signs of hypothyroidism. In the second case, amniotic rT3 levels (140-180 ng/dl) were well within the normal range for 15-19 weeks of pregnancy, but an affected hypothyroid infant was born. These data suggest that amniotic fluid rT3 levels may not be a reliable tool in diagnosing intrauterine hypothyroidism.

Killer cell activity and antibody-dependent cell-mediated cytotoxicity are normal in Hashimoto's disease.

Antibody-dependent cell-mediated cytotoxicity (ADCC) is the process by which antibodies interact with killer cells to effect cell lysis, whereas natural killing (NK) refers to the ability of peripheral blood killer cells to lyse target cells in the absence of specific antibody. The purpose of the present study was to determine if either NK cells or ADCC might play a role in the development of Hashimoto's thyroiditis (HD) by testing the ability of killer cells to cause lysis of K562 erythroleukemia tumor cells and human thyrocytes in the presence and absence of serum from normal and HD patients. Using K562 target cells, NK activity was 70 +/- 4% (mean +/- SEM) for HD effector cells and 66 +/- 5% for normal effector cells at an effector to target ratio of 100:1. Similarly, with thyrocytes as targets, effector cells from HD patients (38 +/- 3%) and normal subjects (34 +/- 5%) caused comparable lysis (at an effector to target ratio of 100:1). Using K562 target cells, ADCC was 35% when effector cells from HD or normal subjects were coincubated with either normal or HD sera. Using thyrocyte target cells, lysis was about 25-30%, but, again, no differences were found between HD and normal effector cells or serum. There was a significant correlation between lysis for K562 and thyrocyte target cells, but there was no significant correlation between the titer of serum antithyroid microsomal antibodies and specific lysis. Intrathyroidal lymphocytes and peripheral lymphocytes from one patient with HD caused comparable lysis of labeled thyrocyte targets, as did normal peripheral lymphocytes. We conclude that ADCC and NK activities in peripheral lymphocytes were normal in HD patients and, therefore, may not have a primary role in mediating thyrocyte destruction in Hashimoto's thyroiditis.