Hypogonadotropic hypogonadism in idiopathic hemochromatosis: evidence for combined hypothalamic and pituitary involvement

Hypogonadism is a common finding in idiopathic hemochromatosis. Most studies have localized the defect to either the pituitary gland or the testes. We describe a case with evidence that favors the likely concomitant involvement of the hypothalamus as a factor in the observed hypogonadism. A clinically hypogonadal male with hemochromatosis had a low testosterone concentration with inappropriately normal serum LH levels. Leydig cell function was intact, as demonstrated by a normal increase in serum testosterone following HCG administration. However, although the pituitary secretion of LH was normal in response to GnRH stimulation, clomiphene administration did not produce an increase in LH and FSH, suggesting that there was a defect in the hypothalamic GnRH response. Since the FSH and prolactin responses to stimulatory testing were inadequate, coexisting pituitary dysfunction was likely also present. We conclude that this man had hypogonadism with laboratory evidence for a combined defect in hypothalamic and pituitary function.     

Androgen and estrogen metabolism: relationship to obesity

Adipose tissue contains both aromatase and 17 beta-steroid dehydrogenase activity. Therefore, to see whether there was a relationship between obesity and certain parameters of androgen and estrogen metabolism we infused 88 women, mean age 51.1 +/- 0.3 years and mean weight 140 +/- 3 lbs, with 3H-testosterone (T)/14C-estradiol (E2) and 3H-androstenedione (A)/14C-estrone (E1) on separate occasions. Blood samples were obtained during the infusion and all urine was collected for 4 days following the start of the infusion. The blood samples were analyzed for radioactivity as A, T, E1, E2, and dihydrotestosterone (DHT) and the urines were analyzed for radioactivity as E1 and E2. From these data we calculated the percent of A converted to T ([rho]A,TBB = percent of A infused measured as T in the blood), [rho]T,ABB [rho]E1,E2BB, and [rho]E2,E1BB. We also measured the ratio of radioactivity as 3H-DHT to radioactivity as 3H-A (CRA,DHT) and 3H-T (CRT,DHT) during the respective androgen infusions. From the ratio of 3H/14C as estrone or estradiol in the urine following 3H-A or 3H-T infusions, we calculated the percent of A or T that was aromatized to E1 or E2 ([rho]A,E1BM; [rho]T,E2BM). When the data from these women were related to weight or Quetelet's Index (QI = wt/ht2) by unweighted linear regression, the only values that were significantly correlated with weight and QI were [rho]T,E2BM and [rho]A,E1BM; for all other [rho] and CR values there was no correlation with weight or QI. We conclude that peripheral aromatization is positively correlated with adiposity but androgen interconversions and estrogen interconversions are not related to adiposity.     

Augmented 17α-hydroxyprogesterone response to ACTH stimulation as evidence of decreased 21-hydroxylase activity in patients with incidentally discovered adrenal tumours ('incidentalomas')

OBJECTIVE Recent studies have indicated that the syndrome of congenital adrenal hyperplasia due to 21-hydroxylase deficiency is closely associated with the development of benign adrenocortical tumours. Tumour formation is thought to be a consequence of ACTH hypersecretion which results from the lack of glucocorticoid synthesis. The aim of this study was to evaluate 21-hydroxylase activity in patients with an incidentally discovered adrenal mass ('Incidentaloma') without a history of congenital adrenal hyperplasia. DESIGN A prospective study of 52 patients admitted to a single hospital. PATIENTS Fifty-two consecutive subjects (mean age 56·0 years, range 24–81 years) with an incidentally discovered adrenal tumour were studied. MEASUREMENTS The 21-hydroxylase reserve was assessed by intravenous bolus administration of 1-24 ACTH (tetracosactrin) with measurement of basal and stimulated serum 17α-hydroxyprogesterone (17-OHP) concentrations. Impaired 21-hydroxylase activity was defined as an exaggerated 17-OHP response, with a 17-OHP increment exceeding 7-9nmolll. Basal and stimulated cortisol concentrations, and basal ACTH were also measured. RESULTS Baseline levels of 17-OHP were normal in 44 and elevated in 8 subjects. In 37 patients (71·2%), the 17-OHP increment following ACTH administration exceeded 7·9nmol/l, demonstrating mildly decreased 21-hydroxylase activity. In these subjects, the peak serum 17-OHP correlated with the tumour diameter. In the patients with apparently normal 21-hydroxylase activity, no significant correlation was found between 17-OHP concentrations and tumour size. All patients had a stimulated serum cortisol above 550 nmol/l reflecting intact adrenal gluco-corticoid reserve. There were no other differences between the group with exaggerated and the group with normal 17-OHP Increment. The tumours were removed from two women with augmented 17-OHP responses and this was followed by normalization of 17-OHP dynamics. CONCLUSIONS Biochemical evidence for partial 21-hydroxyiase defiency is a common finding in patients with an adrenal Incidentaloma, even in the absence of a congential adrenal hyperpiasia history. Exaggerated 17-OHP increment is not accompanied by decreased adrenal glucocorticoid reserve. Normalization of the 17-OHP response after surgical treatment suggests that the phenomenon results from reduced 21-hydroxyiase activity in the tumour, which retains ACTH responsiveness.     

Immunohistochemical evidence for a lack of ferritin in duodenal absorptive epithelial cells in idiopathic hemochromatosis

Patients with idiopathic hemochromatosis exhibit an unexplained increase in intestinal iron absorption. The aim of this work was to study immunohistochemical H- and L-ferritin distribution in duodenal mucosal cells of patients with idiopathic hemochromatosis, and of subjects with various degrees of iron loading. Biopsy sections of gastrointestinal mucosa from 24 patients with idiopathic hemochromatosis, 10 patients with secondary iron overload, 6 normal subjects, and 13 iron-deficient subjects were analyzed with monoclonal antibodies for the presence of immunohistochemical H and L ferritin types, and with Perls' stain for hemosiderin. Ferritin content of duodenal homogenates was evaluated in 5 cases. The absorptive duodenal cells were found to contain ferritin, mostly of the L type, in apical granules; these ferritin granules were present in all normal, iron-deficient, and iron-over-loaded subjects, but were absent in 21 (87%) of the patients with established idiopathic hemochromatosis. In cells other than those of the duodenal epithelium, such as lamina propria or antral mucosa, ferritin and hemosiderin contents were related to iron loading and no difference was evident between primary and secondary iron overload. These findings indicate that (a) idiopathic hemochromatosis is associated with an altered ferritin expression in the duodenal absorptive epithelial cells, (b) this alteration cannot be detected by analysis of duodenal homogenates, (c) idiopathic hemochromatosis does not affect ferritin accumulation in the other cell types analyzed, and (d) ferritin in absorptive duodenal cells may have a regulatory role in iron absorption.     

Androgen insensitivity in man: evidence for genetic heterogeneity.

We have studied ten phenotypically similar patients with complete androgen insensitivity. All of the patients tested had significantly elevated serum luteinizing hormone and plasma androgens within or above the normal adult male range. On the basis of specific dihydrotestosterone binding by skin fibroblasts, we identified two subgroups. Six patients from five different families had undetectable dihydrotestosterone binding, while four patients from two families had normal binding activity. Our results indicate that within the clinical syndrome of androgen insensitivity there are at least two distinct genetic variants. These variants may result from allelic mutations of the same X-linked gene specifying the dihydrotestosterone receptor or, alternatively, from mutations of separate genes both being essential for androgen action in responsive cells.     

Dendritic growth and spine formation in response to estrogen in the developing Purkinje cell

Neurosteroids are synthesized de novo in the brain, and the cerebellar Purkinje cell is a major site for neurosteroid formation. We have demonstrated that the Purkinje cell possesses intranuclear receptor for progesterone and actively produces progesterone de novo from cholesterol only during rat neonatal life, when cerebellar cortical formation occurs dramatically. We have further demonstrated that progesterone promotes dendritic growth, spinogenesis, and synaptogenesis via its receptor in this neuron in the neonate. On the other hand, estrogen may also play an important role in the process of cerebellar cortical formation, because the neonatal rat Purkinje cell possesses estrogen receptor (ER)beta. However, estrogen formation in the neonatal cerebellum is still unclear. In this study, we therefore analyzed the biosynthesis and action of estrogen in Purkinje cells during neonatal life. RT-PCR-Southern and in situ hybridization analyses showed that Purkinje cells expressed the key enzyme of estrogen formation, cytochrome P450 aromatase, in neonatal rats. A specific enzyme immunoassay for estradiol further indicated that cerebellar estradiol concentrations in the neonate were significantly higher than those in the prepuberty and adult. Both in vitro and in vivo studies with newborn rats showed that estradiol promoted dose-dependent dendritic growth of Purkinje cells. Estradiol also increased the density of Purkinje dendritic spines. These effects were inhibited by the ER antagonist tamoxifen. These results suggest that estradiol in the developing Purkinje cell promotes dendritic growth and spinogenesis via ERbeta in this neuron. Estradiol as well as progesterone may contribute to the growth of Purkinje cells during the cerebellar cortical formation.     

Assessment of adrenal reserve in pregnancy: defining the normal response to the adrenocorticotropin stimulation test

CONTEXT: Normal pregnancy is a state of hypercortisolism, making adrenal insufficiency difficult to diagnose. Objective: We sought to identify a normative, minimum-response threshold for the ACTH stimulation test in pregnancy. We hypothesized that salivary free cortisol (SaFC) would prove a more physiological and less variable measure of adrenal reserve in pregnancy than serum cortisol (SC). DESIGN: This is a prospective study of normal controls. SETTING: The study was conducted in an obstetrical clinic in a tertiary care hospital. PATIENTS: Patients included 36 healthy ambulatory pregnant women (aged 18-37 yr) with singleton pregnancies. INTERVENTION: The 250-microg ACTH stimulation test was performed in the healthy pregnant volunteers. Based on their gestational age at the time of recruitment, women were studied in one of the trimesters and were restudied at 11-14 wk postpartum. MAIN OUTCOME MEASURES: Total SC, aldosterone, and SaFC concentrations were measured before and after ACTH. The response in pregnancy was compared with postpartum values. RESULTS: Basal SC (P = 0.01), aldosterone (P = 0.001), and SaFC (P = 0.01) values progressively increased during the trimesters of pregnancy and decreased postpartum, confirming that pregnant women have increased basal glucocorticoid and mineralocorticoid production. There was enhanced responsiveness of the maternal adrenal glands to ACTH stimulation as pregnancy progressed, as measured by peak stimulated SaFC (P = 0.009) and aldosterone (P = 0.01). In the milieu of altered binding globulins, SaFC is a more consistent, binding-globulin-independent measure of stimulated adrenal function than total SC. Minimum criteria for the normal SaFC response to ACTH stimulation in the second and third trimesters of pregnancy and postpartum have been generated based on a predominantly African-American group of subjects. CONCLUSIONS: Reliable data are available for the evaluation of the adrenal axis in pregnancy with a noninvasive, outpatient measure of SaFC. Glucocorticoid therapy in pregnancy should take into account that adrenal reserve increases as pregnancy progresses.     

New diagnostic and therapeutic possibilities in manifest idiopathic hemochromatosis

Up until recently in clinical practice suspected hemochromatosis with a pathological iron-screening test (plasma iron, percentage transferrin saturation, serum ferritin, desferrioxamine-induced urinary iron excretion) made a liver biopsy necessary. Today, as a first step, the density of the liver parenchyma can be measured by means of computed tomography. Normal findings obviate the need for laparoscopy. Since the late forties weekly or twice weekly phlebotomy has been the sole form of treatment for manifest idiopathic hemochromatosis. In the mid-sixties the hopes placed in chelating substances (desferrioxamine) were not fulfilled, because the plasma half-life (only 7-10 minutes) of this drug was too short. Even with several daily injections only a small amount of iron was removed from the body tissue (10-25 mg daily urinary iron excretion). The introduction of portable infusion pumps in the late seventies offered us a new possibility of administering desferrioxamine by subcutaneous injection (Propper et al., 1976). Until that time such treatment was successfully used only in the field of pediatrics to treat secondary transfusion hemochromatosis in thalassemia. In one case of idiopathic hemochromatosis with severe organic involvement (right heart failure, repeated esophageal hemorrhage and bronzed diabetes) we had to achieve rapid iron elimination, and for this purpose we used continuous long-term desferrioxamine administration by means of a portable infusion pump (Autosyringe) in addition to phlebotomy. Since, particularly in the critical initial phase of treatment when heart failure was always threatening, great care had to be exercised in the use of phlebotomy, iron removal was achieved largely by desferrioxamine administration (daily up to 240 mg iron elimination in urine and stools).(ABSTRACT TRUNCATED AT 250 WORDS)     

Biliary excretion of iron and ferritin in idiopathic hemochromatosis

The role of biliary excretion of iron and ferritin in iron overload was studied and evaluated. Ten patients with idiopathic hemochromatosis and two groups of controls (14 gallstone patients and 16 healthy subjects) were included. Liver tissue (obtained by percutaneous or operative biopsy) was investigated with light microscopy and transmission electron microscopy in combination with x-ray microanalysis. Fasting bile samples were obtained through duodenal aspiration or at cholecystectomy. Iron was determined in liver tissue and bile using atomic absorption spectroscopy, and ferritin was determined in serum and bile with a radioimmunoassay technique. All patients with hemochromatosis had iron-positive staining as seen in light microscopy. Electron microscopy showed iron-containing proteins in the lysosomes and cytosol of liver parenchymal cells, and this observation was supported by x-ray microanalysis. Hepatic iron concentration was increased about eightfold in the patients with hemochromatosis (p less than 0.001). Biliary iron concentration, expressed per millimole of bile acid, was increased about twofold (p less than 0.05) and biliary ferritin concentration about fivefold (p less than 0.001) in hemochromatosis. Four of the patients with hemochromatosis were reexamined after completed treatment with venesection; this resulted in normalized biliary concentrations of iron and ferritin. We conclude that biliary secretion of ferritin occurs in humans and that both iron and ferritin excretion are enhanced in hepatic iron overload. The apparently limited capacity of biliary iron excretion may be of importance for the hepatic iron accumulation in hemochromatosis.     

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)     

Myocardial involvement in idiopathic hemochromatosis: Morphologic and clinical improvement following venesection

We describe a 31 year old male patient who presented with severe cardiomyopathy caused by primary hemochromatosis. After a stormy course, complicated by heart failure and severe ventricular arrythmias, improvement in clinical status and myocardial function occurred. Depletion of myocardial iron was documented by the technique of serial endomyocardial biopsy. Myocardial iron stores were not yet depleted when hypoferremia and iron deficiency anemia occurred. This is the first reported study of myocardial morphology in a successfully treated patient with hemochromatotic cardiomyopathy.     

HFE mutations analysis in 711 hemochromatosis probands: evidence for S65C implication in mild form of hemochromatosis

Hereditary hemochromatosis (HH) is a common autosomal recessive genetic disorder of iron metabolism. The HFE candidate gene encoding an HLA class I-like protein involved in HH was identified in 1996. Two missense mutations have been described: C282Y, accounting for 80% to 90% of HH chromosomes, and H63D, which is associated with a milder form of the disease representing 40% to 70% of non-C282Y HH chromosomes. We report here on the analysis of C282Y, H63D, and the 193A-->T substitution leading to the S65C missense substitution in a large series of probands and controls. The results confirm that the C282Y substitution was the main mutation involved in hemochromatosis, accounting for 85% of carrier chromosomes, whereas the H63D substitution represented 39% of the HH chromosomes that did not carry the C282Y mutation. In addition, our screening showed that the S65C substitution was significantly enriched in probands with at least one chromosome without an assigned mutation. This substitution accounted for 7.8% of HH chromosomes that were neither C282Y nor H63D. This enrichment of S65C among HH chromosomes suggests that the S65C substitution is associated with the mild form of hemochromatosis.     

The longitudinal study of adrenal maturation during gonadal suppression: evidence that adrenarche is a gradual process

The physical changes that herald the onset of puberty result from the combination of adrenarche and gonadarche. To examine adrenal maturation and associated changes in growth without the confounding effects of changes in the gonadal steroid milieu, we performed a longitudinal study in 14 young girls with idiopathic central precocious puberty during long-term pituitary-gonadal suppression. Beginning at the mean age of 2.9 yr, dehydroepiandrosterone sulfate levels, linear growth, skeletal maturation, body mass index, and secondary sexual development were evaluated at 3- to 6-month intervals for up to 12.3 yr. In 12 of the girls, levels of dehydroepiandrosterone, androstenedione, 17-hydroxypregnenolone, and 17alpha-hydroxyprogesterone were determined before and after acute ACTH stimulation every 6 months to investigate the maturation of adrenal steroidogenic enzyme activity. Serum dehydroepiandrosterone sulfate levels rose progressively throughout the study. An exponential model fit the longitudinal datasets well and indicated that dehydroepiandrosterone sulfate levels increased approximately 22%/yr from the youngest age onward. Increasing activity of 17-20 lyase (CYP17) and decreasing activity of 3beta-hydroxysteroid dehydrogenase were also evident in preadrenarchal subjects. When controlled for chronological age, no significant associations were noted between weight, body mass index, or body surface area and dehydroepiandrosterone sulfate levels. However, similar analyses revealed modest correlations of both height and growth velocity with dehydroepiandrosterone sulfate levels. Our results suggest that adrenarche is not the result of sudden rapid changes in adrenal enzyme activities or adrenal androgen concentrations; rather, adrenarche may be a gradual maturational process that begins in early childhood.     

In vitro conversion of androgen to estrogen in amphioxus gonadal tissues

The ability to convert androgen to estrogen (aromatization) is a constant feature of gonadal and neural tissues in all major vertebrate groups. In experiments reported here, the existence of this pathway was investigated in the protochordate amphioxus (Branchiostoma lanceolatum). Following incubation with [³H]19-hydroxyandrostenedione, gonadal homogenates contained authentic estrone and estradiol-17β, as determined by derivative formation and recrystallization to constant specific activity. Cephalic (“brain”) and other segments were aromatase negative. The results indicate that a potential for estrogen biosynthesis in the gonads predates that in other tissues and arises prior to the evolution of true vertebrates.