Enzyme kinetic and substrate-binding studies of the thyroxine to 3,5,3'-triiodothyronine converting enzyme in the rat liver microsomal fraction.

The conversion of T4 to T3 was studied in the rat liver microsomal fraction. A mean Vmax of 0.11 pmol T3 produced per mg microsomal protein/min and a mean apparent Km of 2.1 muM T4 were found. An approximation to the real Km for the experimental conditions used was obtained by applying free instead of total T4 as the substrate concentration. Thus, the Km was found to be 9.7 nM free T4, and changes of Km with different amounts of microsomal protein added were not detected. rT3 was found to be a competitive inhibitor of the T4 to T3 conversion, with a mean apparent Ki of 9.4 nM rT3. Binding studies showed that T4 is bound not only nonspecifically but also to two different classes of specific binding sites. The dissociation constants were 7.5 and 1700 nM t4, and the maximal binding capacities were 58 and 4300 pmol T4/mg microsomal protein, respectively, rT3 and T3 both had one specific binding site besides their unspecific binding to the microsomal fraction. The dissociation constants were found to be 45 nM rT3 and 850 nM T3, respectively; the maximal binding capacities amounted to 75 pmol rT3 and 4600 pmol T3 per mg microsomal protein, respectively. rT3 competes with T4 for its first (apparent Ki, 55 nM rT3) and T3 competes with T4 for its second specific binding site (apparent Ki, 960 nM T3). It is suggested, that the first specific binding site for T4 and the specific binding site for rT3 are identical, and that they represent the 5'-deiodinase. The rT3-induced inhibition of the T4 to T3 conversion is caused by a competition for the binding site of the enzyme. A competition for cofactors may play an additional role. T3 competes with T4 for a different specific binding site, which may contain the 5-deiodinase.

Estrone and estradiol in patients with cirrhosis of the liver: effects of ACTH and dexamethasone.

To study the influence of the adrenal gland on plasma estrogen levels in male patients with hepatic cirrhosis, estrone and estradiol were measured under a variety of experimental conditions. Compared to controls, estradiol levels were moderately elevated by 26% (P is less than 0.05) in patients with hepatic cirrhosis (28.5 +/- 5.4 vs. 36.0 +/- 4.7 pg/ml plasma; n: 12), whereas estrone levels exhibited a two- to threefold increase under basal conditions (32.5 +/- 5.6 vs. 67.8 +/- 20.8 pg/ml; P is less than 0.01). ACTH application resulted in a striking increase in plasma estrone levels in both patients with hepatic cirrhosis and in normal subjects (61.8 +/- 27.5 vs. 27.3 +/- 7.8 pg/ml). During stimulation with ACTH, estradiol levels showed no significant changes. After suppression of the adrenal gland by dexamethasone administered for 5 days, plasma concentrations of estrone and estradiol were found to be reduced. The absolute decrease of estrone was significantly greater in patients with hepatic cirrhosis than in healthy male subjects (35.5 +/- 12.6 vs. 21.3 +/- 6.0 pg/ml; P is less than 0.05; n: 8). Estrogen values, however, were still high in patients with hepatic cirrhosis after 5 days of dexamethasone administration (37.1 +/- 17.6 pg estrone/ml and 23.9 +/- 3.6 pg estradiol/ml plasma). It is suggested that elevated plasma values of estrogens in this disorder may be derived predominantly from adrenal production. Peripheral conversion of androgens to estrone rather than to estradiol appears to be more effective in sustaining plasma levels of estrogens in patients with hepatic cirrhosis.

Enhanced conversion of androstenedione to estrogens in obese males.

In normal and obese young males [90--120% and > 160% of ideal body weight (IBW); IBW = 100%], plasma concentrations of testosterone, androstenedione, estrone, and estradiol were measured. Metabolic clearance and production rates of androstenedione and the conversion ratios of androstenedione to testosterone, estrone, and estradiol were determined using the constant infusion technique. In the obese subjects, IBW was inversely correlated (P < 0.001) with plasma concentrations of androstenedione (r = 0.81) and testosterone (r = 0.87), while the levels of estrone (r = 0.92) and estradiol (r = 0.95) increased with IBW (P < 0.001). Thus, when normal and obese subjects were compared as groups, plasma androstenedione decreased form 1.24 +/- 0.13 to 0.93 +/- 0.15 ng/ml (mean +/- SD) and plasma testosterone decreased from 5.89 +/- 0.82 to 3.29 +/- 0.92 ng/ml (P < 0.001), while estrone increased from 28.2 +/- 3.4 to 60.0 +/- 9.4 pg/ml, and estradiol increased from 21.7 +/- 3.5 to 43.9 +/- 5.3 pg/ml. The testosterone to androstenedione and the estradiol to estrone ratios were not different in obesity, but changes in IBW were positively correlated (P < 0.001) with differences in the estrone to androstenedione (r = 0.93) and estradiol to testosterone ratios (r = 0.93), indicating that fat tissue may aromatize androgens, whereas reduction of 17-oxo-steroid appears to be of minor importance. As the MCR of androstenedione increased with IBW (from 2156 to 2636 liters/day P < 0.05) while plasma levels decreased, the apparent production rate of androstenedione was not influenced by the degree of obesity. The conversion of androstenedione to estrone (r = 0.89) and of androstenedione to estradiol (r = 0.82) was enhanced in obese subjects (P < 0.001). We suggest that enhanced aromatization of androstenedione due to an increased adipose tissue mass may account for the high plasma estrogen levels observed in obese men.

The defect of intestinal calcium transport in hyperthyroidism and its response to therapy.

Skeletal demineralization occurs in thyrotoxicosis. Fecal calcium excretion may be enhanced, and calcium balance tends to be negative. We investigated intestinal calcium transport in 12 hyperthyroid patients. Absorption was measured by segmental perfusion in the proximal jejunum at calcium concentrations commensurate with those in the fasting and postprandial states. At low luminal concentrations, under conditions where calcium is transported predominantly by active processes, the calcium absorption rate was reduced though not abolished in hyperthyroid patients (16 +/- 4 (SE) mumol/h . 30 cm segment) as compared to normal subjects (71 +/- 8 mumol/h; P less than 0.001). When perfusate calcium was raised to 5 mmol/liter there was little increment of the net absorption rate in the hyperthyroid group (45 +/- 11 mumol/h), whereas that in the normal subjects rose to 183 +/- 17 mumol/h. Likewise, the unidirectional calcium flux out of the lumen was low in hyperthyroidism (43 +/- 7 mumol/h), suggesting that low net absorption rates were not due to transmucosal calcium loss. D-Xylose permeation was similar in all study groups. Treatment of the thyroid disease led to a marked increase in calcium absorption rates from 33 +/- 10 to 124 +/- 20 mumol/h (at 2 mmol/liter P less than 0.001; n = 5) into the range of values in normal subjects (124 +/- 9 mumol/h). Circulating levels of 1,25-dihydroxyvitamin D were low in hyperthyroid patients (38 +/- 11 pg/ml) and increased during treatment to 63 +/- 11 pg/ml (P less than 0.025; n = 9), whereas 25-hydroxyvitamin D was normal and remained unchanged. We conclude that intestinal calcium transport, particularly its active component, is reversibly decreased in hyperthyroidism. The association with low plasma levels of the active vitamin D metabolite suggests that the decrease in calcium absorption may be related to calcium-regulating mechanisms as a consequence of the net calcium efflux from bone in this disease.

Conversion of androgens to estrogens in idiopathic hemochromatosis: comparison with alcoholic liver cirrhosis.

Hypogonadism is common in patients with some liver diseases, such as idiopathic hemochromatosis (IHC) and alcoholic cirrhosis (AC). However, gynecomastia, a typical feature in AC, does not occur in IHC. To determine the hormonal basis for this difference, the following parameters were determined in patients with IHC and AC as well as in normal men: plasma concentrations of androgens and estrogens, metabolic clearance and production rates of androstenedione and testosterone, and the contribution of peripheral conversion of androstenedione and testosterone to the circulating estrogens. Severe impotence in both patients with IHC and those with AC was associated with more than 50% reduction in plasma testosterone. The reduction was due to 63% and 70% decreases in testosterone production in IHC and AC, respectively. The MCRs were less affected in IHC and AC (19% and 37% reductions, respectively). In IHC, the fall in testosterone concentrations was accompanied by decreased production and plasma concentrations of androstenedione, a precursor for estrogen synthesis. In contrast, production and plasma concentrations of androstenedione were significantly increased in AC. Patients with IHC had estradiol und estrone levels similar to those in normal men (mean +/- SD, 16.2 +/- 4.6 vs. 20.3 +/- 3.7 pg/ml; P = NS), whereas in AC, estradiol and estrone were significantly elevated (38.0 +/- 5.3 and 68.5 +/- 17.2 pg/ml, respectively). In IHC, sex hormone-binding globulin levels were in the same range as in the normal men, whereas sex hormone-binding globulin was increased in AC. In IHC, the instantaneous contribution of plasma androstenedione to estrone and estradiol was normal, whereas that of plasma testosterone to plasma estrogens was decreased by about 50%. In contrast, in AC, the instantaneous contribution of plasma androstenedione to estrogens was greatly enhanced, and that of testosterone was in the normal range. Since the MCRs of androgens and the conversion ratios of androgens to estrogens indicate normal peripheral metabolism of sex hormones in IHC, decreased androgen formation implies decreased testicular synthesis. This was confirmed by a significantly decreased LH level in IHC (5.5 +/- 1.9 vs. 10.5 +/- 3.1 mU/ml in normal men), indicating pituitary failure. In AC, however, increased LH (20.0 +/- 2.7 mU/ml) may be indicative of primary testicular failure. These results confirm clinical features of hypogonadism and normal estrogenic activity in patients with IHC.(ABSTRACT TRUNCATED AT 250 WORDS)

Digoxin does not alter plasma steroid levels in health men.

Digoxin has been reported to induce feminizing effects in man. It does not compete for estradiol cytosol receptors in human breast carcinoma cells, however, and has no uterotrophic effect. We therefore investigated whether feminization might be due to digoxin action on plasma concentrations of sex steroids. Six healthy men (31.5 +/- 4 yr old) received therapeutic doses of digoxin for 43 days. We measured plasma concentrations of testosterone, androstenedione, dehydroepiandrosterone, estrone, estradiol, progesterone, 17-hydroxyprogesterone, cortisol, and aldosterone. During 35 days on digoxin levels of these steroids remained in the normal range and there was no change from before-drug values. Digoxin was in the therapeutic range of 1.9 +/- 3 nmol/l throughout. After stimulation by adrenocorticotropic hormone or human choriongonadotropin, the rise in plasma steroids was in the same range as when digoxin was given, as well as 16 wk after it had been discontinued. A normal rise in luteinizing hormone after luteinizing hormone-releasing hormone showed that the hypothalamogonadal feedback was not altered by digoxin. Free testosterone, estradiol, and cortisol concentrations under basal conditions and after stimulation were also the same before and after drug. It is concluded that the estrogen-like activity of digoxin cannot be explained by altered steroid availability from plasma. Feminizing effects attributed to digoxin may be caused by other conditions known to influence sex steroid hormones that are common in patients with heart disease. Our data suggest that digoxin may be the preferred digitalis therapy to avoid feminizing effects.

Abnormalities in estrogen, androgen, and insulin metabolism in idiopathic hemochromatosis.

Of 44 male patients with idiopathic hemochromatosis who were diagnosed at an early stage without morphological or biochemical evidence of liver disease, 25% suffered from impotence and 34% manifested glucose intolerance. Impotence was correlated with a 50% reduction in plasma testosterone, resulting from a 63% decrease in testosterone production. Testicular atrophy was caused by insufficient secretion of gonadotropins due to the selective accumulation of iron in gonadotropic cells of the pituitary gland. However, peripheral sexual hormone metabolism, in particular the conversion of androgens to estrogens, remained unaltered. It was therefore possible to employ substitution therapy successfully with testosterone in these men, and hyperestrogenism was not observed as a side effect. The pathogenetic factors in the development of diabetes mellitus in patients with idiopathic hemochromatosis include impaired insulin secretion caused by the selective deposition of iron in B-cells of the pancreas and insulin resistance due to iron accumulation in the liver. In particular, the insulin resistance is markedly improved after depletion of body iron stores by phlebotomy treatment, resulting in lower insulin requirements in patients with insulin-dependent diabetes as well as improvement of carbohydrate metabolisms in about half of the patients with non-insulin-dependent diabetes. We have concluded that hypogonadism and carbohydrate intolerance are caused by the specific distribution pattern of excess iron in the organism, accompanied by functional impairment of affected parenchymal cells.

Relationship of plasma sex hormones to different parameters of obesity in male subjects.

Relationships between plasma sex hormones and different parameters of obesity (weight, ideal body weight [IBW], overweight, fat mass, and body surface) were investigated in 70 healthy nonobese and obese males, 20-40 yr of age and with a body weight of 85%-245% of IBW. Plasma sex hormones remained unaffected by weight up to approximately 160% of the IBW. Only in the massively obese subjects was plasma testosterone decreased to 40% of controls (from 6.2 to 2.5 ng/ml), whereas free testosterone remained almost constant. On the other hand, plasma estrone and estradiol exhibited significant increases in obese subjects, ranging from 31.5 +/- 52.3 +/- 5.8 pg/ml for estrone, and 25.4 +/- 5.4 increasing to 44.7 +/0 5.0 pg/ml for estradiol. Similarly, free estradiol was shown to significantly increase with obesity in men from 505 +/- 118 to 991 +/- 123 fg/ml (p < 0.001). The ratios of testosterone/androstenedione, as well as of estradiol/estrone, were not affected by obesity, suggesting that reduction of the 17-oxo-group of the steroids is not influenced by the amount of fat tissue. A significant (p < 0.001) correlation was found between IBW and estrone (r = 0.80) and estradiol (r = 0.75), as well as the ratios of estrone/androstenedione (r = 0.62) and estradiol/testosterone (r = 0.86). This is consistent in its evidence indicating that fat tissue may be able to aromatize androgens. In the obese subjects, there were significant correlations between plasma sex hormones (testosterone, estrone, estradiol, and free estradiol) and the parameters of obesity used. Among these, correlations were best with IBW, overweight, and fat mass (r = 0.74-0.89; p < 0.001); body weight and body surface were less favorable.

The measurement of 4-androstene-3, 11, 17-trione (11-oxo-androstenedione) by radioimmunoassay in human plasma.

A radioimmunoassay (RIA) method is described for the determination of 4-androstene-3, 11, 17-trione (11-oxo-androstenedione) in human plasma. 4-androstene-3, 11, 17-trione 3-(0-carboxymethyl) oxime-bovine serum albumin conjugate was used to generate highly specific antiserum in rabbits. Cross reactivities of several other steroids with the antiserum were less than 4%. [1,2-3H] 4-androstene-3, 11, 17-trione was synthesized from [1,2-3H] 17 alpha, 21-dihydroxy-4-pregnene-3, 11, 20-trione. The intra- and interassay variation was 7.3% and 9.8%, respectively. The mean serum 4-androstene-3, 11, 17-trione level for healthy young subjects was 2.37 +/- 0.56 nM (X +/- SD) in males and 3.16 +/- 0.43 nM in females at 8 a.m. During the night, there was a marked decrease in serum level, giving at 11 p.m. 0.87 +/- 0.33 and 1.15 +/- 0.52 nM, respectively. During ACTH stimulation tests, 4-androstene-3, 11, 17-trione increased from 1.81 +/- 0.58 to 2.32 +/- 0.69 nM, while in dexamethasone suppression tests a decrease from 3.20 +/- 0.03 nM was seen. In contrast, HCG administration on 3 consecutive days did not influence plasma concentrations of 4-androstene-3, 11, 17-trione.

Testicular testosterone concentration and in vitro response to HCG in normal and in testosterone immunized rabbits.

The testosterone concentration, the in vitro response to HCG and the percentage Leydig cells in testes of normal and of testosterone-3-BSA immunized rabbits were determined. Following immunization all three parameters increased in the same order of magnitude (1.8-2.6fold). The results indicate that active immunization with testosterone has no deleterious effects on the endocrine capacity of the Leydig cells. The observed functional and morpholigical alterations of the testes are due solely to increased trophic hormone secretion from the pituitary caused by antibody binding of circulating androgens. The basic testosterone concentration in the testes of the control rabbits were in the range of values reported for other species.

Attempts towards a serological diagnosis of ankylosing spondylitis.

Polytene chromosomes of salivary glands as well as nuclear proteins from Kc-cells of Drosophila melanogaster have been used as substrate to identify and evaluate the diagnostic value of crossreacting antibodies present in sera of AS patients. The diagnostic significance of the recently described anti-93D antibody (Lakomek et al., 1984) was confirmed by screening sera of patients with definite or suspected AS using cytoimmunofluorescence on the polytene chromosomes. In addition, four new antibodies could be identified in AS sera by immunoblotting. Simultaneous detection of these antibodies supports the diagnosis of AS and is most useful in diagnosis of early stages of this disease.

[Subtotal thyroidectomy as a superior therapeutic principle in the treatment of endocrine ophthalmopathy: a comparison with retrobulbar irradiation and thyrostatic therapy alone]. / Subtotale Thyreoidektomie als überlegenes Therapieprinzip bei der Behandlung der endokrinen Ophthalmopathie: Ein Vergleich mit Retrobulbärbestrahlung bzw. alleiniger thyreostatischer Therapie.

In order to investigate the influence of near total thyroidectomy on the course of endocrine ophthalmopathy (E.O.) in patients with Graves' disease, 29 patients with goitre and E.O. were classified before and after (up to 18 months) operation by use of a special ophthalmopathy index. 14 patients without goitre served as controls; they get only antithyroid drug treatment (ADT) (E.O. I and II, n = 7) or additional retoorbital irradiation (E.O. III and IV, n = 7, linear accelerator, 20 Gray). 20 out of 29 operated patients showed an improvement in the E.O., 4 a deterioration, 5 were unchanged. 3 out of 7 not operated patients with mild E.O. showed an amelioration during ADT, 4 no change. Additional radiotherapy in 7 patients with severe E.O. caused an improvement in the clinical condition of 3 patients, 3 patients deteriorated and 1 patient showed no change. It is concluded that adequate near total thyroidectomy has a positive effect on the clinical course of E.O. in patients with Graves' disease and E.O.