Bilateral Idiopathic Adrenal Hyperplasia: Genetics and Beyond.

Bilateral adrenal hyperplasia currently accounts for up to 2 thirds of cases of primary aldosteronism. As such, it represents a major opportunity for targeted medical management as opposed to unilateral surgically correctable forms of the disease. Although the majority of cases of primary aldosteronism are sporadic, bilateral adrenal hyperplasia may occur in the context of familial hyperaldosteronism where it is associated with specific germline mutations. Over the past 5 years, impressive progress has been made in our understanding of the genetic basis underlying primary aldosteronism, allowing us to identify and characterize new familial forms of the disease and to understand the mechanisms involved in the formation of aldosterone producing adenoma. In contrast, our knowledge of the genetic contribution to the development of bilateral adrenal hyperplasia, and in a larger context, to renin and aldosterone levels in the general population, is still poor. This review summarizes our current knowledge on the genetics of bilateral adrenal hyperplasia and addresses some open questions to be addressed by future research. In particular, genome-wide association studies in large populations may provide clues to understanding the genetic susceptibility underlying the development of primary aldosteronism.

Mineralocorticoid receptor p.I180V polymorphism: association with body mass index and LDL-cholesterol levels.

BACKGROUND/AIMS: Aldosterone and the mineralocorticoid receptor (MR) play a major role in sodium balance and blood pressure control. They are also involved in adipocyte metabolism. The aim of this study was to analyze the association between the MR p.I180V polymorphism with hypertension and markers of cardiovascular risk. DESIGN AND METHODS: Case-control study nested within a cohort of 2063 subjects followed since birth to date. All subjects (age 23-25 yr old) from the entire cohort with systolic and diastolic hypertension (no.=126) were paired with 398 normotensive controls. MR p.I180V genotype association with anthropometric and biochemical markers of cardiometabolic risk was tested. RESULTS: There was a significant association of the MR p.I180V genotype with body mass index (BMI) and LDL-cholesterol level (p<0.01). Hypertensive subjects carrying the polymorphic G allele (AG or GG genotypes) presented significantly higher BMI (30.0+/-6.0 vs 28.7+/-5.6 kg/m(2); p<0.01) and higher LDL-cholesterol (139.9+/-60.3 vs 109.9+/-35.5 mg/dl; p<0.01). The frequency of the polymorphism MR p.I180V was similar between hypertensive subjects and controls (p=0.15). CONCLUSIONS: The MR p.I180V polymorphism seems to be associated with cardiovascular risk factors including BMI and LDL-cholesterol levels. This original in vivo finding reinforces the role of MR in adipocyte biology and in cardiovascular disease.

Potential role of progestogens in the control of adipose tissue and salt sensitivity via interaction with the mineralocorticoid receptor.

Beside their role in the control of water and electrolyte homeostasis, recent data clearly indicate that aldosterone and the mineralocorticoid receptor (MR) are involved in adipocyte biology. It has been recently shown that aldosterone promotes white and brown adipocyte differentiation in vitro through specific activation of the MR. In addition, a non-epithelial pro-inflammatory role for MR activation has been recently inferred from studies on mineralocorticoid/salt administration in experimental animal models and from clinical studies. The mineralocorticoid system could hence represent a potential target for new therapeutic strategies in obesity and the metabolic syndrome. Progesterone has high affinity for the MR and is a natural antagonist of aldosterone. Differently from classic synthetic progestins, which are devoid of antimineralocorticoid properties, progesterone and new progestogens show remarkable antimineralocorticoid effects. Here, we discuss the potential role of the antimineralocorticoid properties of progestogens in the control of body weight, adipose tissue proliferation and salt sensitivity; their therapeutic use in postmenopausal women, as well as in women affected by polycystic ovary syndrome, may open new and unexpected possibilities in the treatment of related metabolic disorders.

Composting of green waste: observations from windrow trials and bench-scale experiments.

The composting of green waste is currently of great importance to the waste industry. For this reason, large-scale compost windrows were investigated and the results compared with those from small, bench-scale experiments. In the two windrows sampled, temperature profiles differed, and greater organic matter loss was observed in the higher temperature windrow. However, enhanced organic matter degradation through elevated temperatures was not supported in the bench-scale experiment, where, under fixed temperatures (30 and 50 degrees C, respectively), no difference in both the total dry and organic matter content was observed. Analysis of particle size distributions in both sets of experiments strongly suggested that moisture-dependent effects were a confounding factor in determining the total LOI of the windrows. Analysis of beta-glucosidase activity in the bench-scale samples demonstrated that it was the assay temperature, rather than the in situ composting temperature, which was more important in determining the level of activity. No evidence for adaptation to local maxima due to variation in the thermal environment was observed.

Hibernoma development in transgenic mice identifies brown adipose tissue as a novel target of aldosterone action.

Aldosterone is a major regulator of salt balance and blood pressure, exerting its effects via the mineralocorticoid receptor (MR). To analyze the regulatory mechanisms controlling tissue-specific expression of the human MR (hMR) in vivo, we have developed transgenic mouse models expressing the SV40 large T antigen (TAg) under the control of each of the two promoters of the hMR gene (P1 or P2). Unexpectedly, all five P1-TAg founder animals died prematurely from voluminous malignant liposarcomas originating from brown adipose tissue, as evidenced by the expression of the mitochondrial uncoupling protein ucp1, indicating that the proximal P1 promoter was transcriptionally active in brown adipocytes. No such hibernoma occurred in P2-TAg transgenic mice. Appropriate tissue-specific usage of P1 promoter sequences was confirmed by demonstrating the presence of endogenous MR in both neoplastic and normal brown adipose tissue. Several cell lines were derived from hibernomas; among them, the T37i cells can undergo terminal differentiation into brown adipocytes, which remain capable of expressing ucp1 upon adrenergic or retinoic acid stimulation. These cells possess endogenous functional MR, thus providing a new model to explore molecular mechanisms of mineralocorticoid action. Our data broaden the known functions of aldosterone and suggest a potential role for MR in adipocyte differentiation and regulation of thermogenesis.

Characterization of the human mineralocorticoid receptor gene 5'-regulatory region: evidence for differential hormonal regulation of two alternative promoters via nonclassical mechanisms.

The mineralocorticoid receptor (MR) is a ligand-dependent transcription factor involved in the regulation of sodium homeostasis. Two distinct mRNA isoforms of the human MR (hMR) differing in their untranslated 5'-ends have recently been identified, suggesting the existence of alternative promoters. To eludicate the regulatory mechanisms controlling hMR gene expression, we have isolated and characterized approximately 15 kb of hMR 5'-flanking region. Various deletion mutants of regions located immediately upstream of the untranslated exons 1 alpha and 1 beta (P1: 1 kb and P2: 1.7 kb, respectively) were inserted into a luciferase reporter gene and used in transient transfection experiments in CV-1 and human differentiated renal H5 cells. Both regions were shown to possess significant functional promoter activity, more pronounced in renal cells, although P1 directed higher levels of basal transcription. Cotransfection experiments with hMR or human glucocorticoid receptor (hGR) revealed that, while both promoters were glucocorticoid inducible, only the distal P2 promoter was stimulated by aldosterone in a dose- and hMR-dependent manner. Furthermore, we demonstrate that hMR and hGR are able to synergistically activate the P2 promoter, consistent with cooperativity between the two transduction pathways. Mineralocorticoid induction was localized to a region between -318 and +123 bp of P2. This region does not contain any consensus hormone responsive element, and direct binding of hMR to this DNA sequence was not observed, indicating that mineralocorticoid-induced transcriptional enhancement is mediated by nonclassical mechanisms. On the other hand, Sp1 and AP-2 bind to definite sequences on both promoters, suggesting that they represent important regulators of hMR promoter activity. Our results indicate that hMR gene expression is under the control of complex regulatory mechanisms involving alternative promoters and differential hormonal control, which might allow tissue-specific modulation of aldosterone action.

A new human MR splice variant is a ligand-independent transactivator modulating corticosteroid action.

Aldosterone effects are mediated by the MR, which possesses the same affinity for mineralocorticoids and glucocorticoids. In addition to the existence of mechanisms regulating intracellular hormone availability, we searched for human MR splice variants involved in tissue-specific corticosteroid function. We have identified a new human MR isoform, hMRDelta5,6, resulting from an alternative splicing event skipping exons 5 and 6 of the human MR gene. hMRDelta5,6 mRNAs are expressed in several human tissues at different levels compared with wild-type human MR, as shown by real time PCR. Introduction of a premature stop codon results in a 75-kDa protein lacking the entire hinge region and ligand binding domain. Interestingly, hMRDelta5,6 is still capable of binding to DNA and acts as a ligand-independent transactivator, with maximal transcriptional induction corresponding to approximately 30-40% of aldosterone-activated wild-type human MR. Coexpression of hMRDelta5,6 with human MR or human GR increases their transactivation potential at high doses of hormone. Finally, hMRDelta5,6 is able to recruit the coactivators, steroid receptor coactivator 1, receptor interacting protein 140, and transcription intermediary factor 1alpha, which enhance its transcriptional activity. Ligand-independent transactivation and enhancement of both wild-type MR and GR activities by hMRDelta5,6 suggests that this new variant might play a role in modulating corticosteroid effects in target tissues.

Paracrine control of steroidogenesis by serotonin in adrenocortical neoplasms.

Serotonin (5-hydroxytryptamine; 5-HT) is able to activate the hypothalamo-pituitary-adrenal axis via multiple actions at different levels. In the human adrenal gland, 5-HT, released by subcapsular mast cells, stimulates corticosteroid production through a paracrine mode of communication which involves 5-HT receptor type 4 (5-HT4) primarily located in zona glomerulosa. As a result, 5-HT is much more efficient to stimulate aldosterone secretion than cortisol release in vitro and administration of 5-HT4 receptor agonists to healthy individuals is followed by an increase in plasma aldosterone levels without any change in plasma cortisol concentrations. Interestingly, adrenocortical hyperplasias and tumors responsible for corticosteroid hypersecretion exhibit various cellular and molecular defects which tend to reinforce the intraadrenal serotonergic tone. These pathophysiological mechanisms, which are summarized in the present review, include an increase in adrenal 5-HT production and overexpression of 5-HT receptors in adrenal neoplastic tissues. Altogether, these data support the concept of adrenal serotonergic paracrinopathy and suggest that 5-HT and its receptors may constitute valuable targets for pharmacological treatments of primary adrenal diseases.

Tissue-specific expression of alpha and beta messenger ribonucleic acid isoforms of the human mineralocorticoid receptor in normal and pathological states.

Expression of the mineralocorticoid receptor (MR) is restricted to some sodium-transporting epithelia and a few nonepithelial target tissues. Determination of the genomic structure of the human MR (hMR) revealed two different untranslated exons (1alpha and 1beta), which splice alternatively into the common exon 2, giving rise to two hMR mRNA isoforms (hMR alpha and hMR beta). We have investigated expression of hMR transcripts in renal, cardiac, skin, and colonic tissue samples by in situ hybridization with exon 1alpha and 1beta specific riboprobes, using an exon 2 probe as internal control. Specific signals for either exon 1alpha- and 1beta-containing mRNAs were detected in typically hMR-expressing cells in all tissues analyzed. hMR alpha and hMR beta were present in distal tubules of the kidney, in cardiomyocytes, in enterocytes of the colonic mucosa, and in keratinocytes and sweat glands. Interestingly, although both isoforms appear to be expressed at approximately the same level, the relative abundance of each message compared with that of exon 2-containing mRNA strikingly differs among aldosterone target tissues, suggesting the possibility of other tissue-specific transcripts originating from alternative splicing. Finally, functional hypermineralocorticism was associated with reduced expression of hMR beta in sweat glands of two patients affected by Conn's and Liddle's syndrome, whereas normal levels of hMR isoforms were found in one case of pseudohypoaldosteronism. Altogether, our results indicate a differential, tissue-specific expression of hMR mRNA isoforms, hMR beta being down-regulated in situations of positive sodium balance, independently of aldosterone levels.

No alteration in the primary structure of the mineralocorticoid receptor in a family with pseudohypoaldosteronism.

We have studied the molecular structure of the mineralocorticoid receptor (MR) complementary DNA (cDNA) in a kindred affected by pseudohypoaldosteronism (PHA). In this family, the clinical symptoms included salt wasting and failure to thrive, accompanied by high urinary levels of sodium despite hyponatremia, hyperkalemia and metabolic acidosis, elevation of PRA, and high plasma aldosterone levels. The patients were resistant to mineralocorticoid administration, but their symptoms ameliorated after a period of sodium supplementation, which was discontinued in older patients. Binding studies performed on mononuclear leukocytes of the members of the family have shown the absence of MR in two siblings and a marked reduction in another sibling of the father, suggesting either the absence of MR or a defect of the ligand-binding domain of the MR in these patients. Southern analysis of patient's DNA did not show any major rearrangement of the MR gene. To search for point mutations in the cDNA of the MR, we performed amplification of the MR cDNA by the polymerase chain reaction and direct sequencing of amplified products. No mutation was found in the entire coding sequence of the MR in patients affected by PHA. Although these results do not exclude a molecular abnormality present on the MR gene, they indicate that PHA in this family is not related to a modification of the MR primary structure.

Prolactin potentiates insulin-stimulated leptin expression and release from differentiated brown adipocytes.

The pituitary hormone prolactin (PRL) exerts pleiotropic effects, which are mediated by a membrane receptor (PRLR) present in numerous cell types including adipocytes. Brown adipose tissue (BAT) expresses uncoupling proteins (UCPs), involved in thermogenesis, but also secretes leptin, a key hormone involved in the control of body weight. To investigate PRL effects on BAT, we used the T37i brown adipose cell line, and demonstrated that PRLRs are expressed as a function of cell differentiation. Addition of PRL leads to activation of the JAK/STAT and MAP kinase signaling pathways, demonstrating that PRLRs are functional in these cells. Basal and catecholamine-induced UCP1 expression were not affected by PRL. However, PRL combined with insulin significantly increases leptin expression and release, indicating that PRL potentiates the stimulatory effect of insulin as revealed by the recruitment of insulin receptor substrates and the activation of phosphatidylinositol 3-kinase. To explore the in vivo physiological relevance of PRL action in BAT, we showed that leptin content was significantly increased in BAT of PRLR-null mice compared with wild-type mice, highlighting the involvement of PRL in the leptin secretion process. This study provides the first evidence for a functional link between PRL and energy balance via a cross-talk between insulin and PRL signaling pathways in brown adipocytes.

Inactivating mutations of the mineralocorticoid receptor in Type I pseudohypoaldosteronism.

Type I pseudohypoaldosteronism (PHA1) is a rare form of mineralocorticoid resistance characterized by neonatal renal salt wasting and failure to thrive. Typical biochemical features include high levels of plasma aldosterone and renin, hyponatremia and hyperkalemia. Different mutations of the human mineralocorticoid receptor (hMR) gene have been identified in subjects affected by the autosomal dominant or sporadic form of the disease. Our laboratory has investigated a large number of subjects with familial and sporadic PHA1. Several different mutations have been detected, which are localized in different coding exons of the hMR gene. These mutations either create truncated proteins, either affect specific amino acids involved in receptor function. In this paper, we review hMR mutations described to date in PHA1 and their functional characterization. We discuss the absence of mutations in some kindreds and the role of precise phenotypic and biological examination of patients to allow for identification of other genes potentially involved in the disease.

Pseudohypoaldosteronism and mineralocorticoid receptor abnormalities.

Pseudohypoaldosteronism is a rare inherited disease characterized by renal salt loss, hyperkalemia and metabolic acidosis despite highly elevated aldosterone values. We previously reported absent or reduced numbers of mineralocorticoid receptors in mononuclear leukocytes and defective effector mechanism as shown by no response in vitro to the incubation of aldosterone in terms of intracellular electrolyte content. We have studied the inheritance of this disorder in ten families and found two different kinds of inheritance: autosomal recessive--often in interrelated families--and autosomal dominant in unrelated families. Parallel studies in the families with the autosomal dominant form of inheritance demonstrated in addition that the effector mechanism of aldosterone is impaired in vitro both in the affected patients and in the carrier relatives characterized by a low number of mineralocorticoid receptors.

Mineralocorticoid resistance.

Pseudohypoaldosteronism was first described in 1958 by Cheek and Perry, who reported an infant with severe salt wasting in the absence of any renal or adrenal defect. Since then several reports have described patients affected by symptoms consistent with resistance to mineralocorticoid action. The clinical picture is characterized by salt wasting and failure to thrive and is resistant to the administration of exogenous mineralocorticoids. Biological features are invariably high plasma and urinary aldosterone levels and elevated plasma renin activity associated with hyponatremia, hyperkalemia, and metabolic acidosis. The discovery of abnormal binding of aldosterone to the mineralocorticoid receptor (MR) in lymphocytes from affected patients, by analogy to findings in other syndromes of steroid hormone resistance, led to the hypothesis that the disease reflected a molecular defect in MR, which has prompted a series of molecular studies to characterize the defect. In this paper we review mechanisms of mineralocorticoid action, discuss the clinical features of mineralocorticoid resistance, overview the molecular characterization of the MR, and close with some pathophysiological hypotheses and questions.

The enigma of pseudohypoaldosteronism.

The syndrome of primary pseudohypoaldosteronism (PHA) is a hereditary disease characterized by increased aldosterone secretion associated with clinical signs of hypoaldosteronism. These include salt wasting and failure to thrive in the newborn, high urinary sodium, hyponatremia, hyperkalemia, and metabolic acidosis. Plasma renin activity is usually elevated in association with aldosterone. The clinical manifestation of the disease is variable, including severely affected patients who may die in infancy and patients who are asymptomatic. The disease seems to result from a cellular resistance to mineralocorticoid action, which could be either generalized, or restricted to the kidney. The condition is inherited as an autosomal dominant or an autosomal recessive trait; however, sporadic cases have been described. In this paper we report a 20-year follow-up study of a French family affected by PHA and we discuss the pathogenesis of the disease.

Molecular characterization of the mineralocorticoid receptor in pseudohypoaldosteronism.

Pseudohypoaldosteronism (PHA) is characterized by salt-wasting and failure to thrive in the newborn, accompanied by high urinary levels of sodium despite hyponatremia, hyperkalemia and metabolic acidosis, elevation of plasma renin activity, and high plasma aldosterone levels. PHA patients are resistant to mineralocorticoid administration, but their symptoms ameliorate after a period of sodium supplementation, which can be discontinued in older subjects. Binding studies performed on mononuclear leukocytes of the family members affected by the disease have shown the absence of binding of [3H]aldosterone to the mineralocorticoid receptor (MR) in mononuclear leukocytes in two siblings and a marked reduction in another sibling and the father, suggesting either the absence of MR or a defect in the ligand binding domain of the MR in these patients. Molecular analysis of the MR in the members of this family did not reveal any major rearrangement or deletion of the MR gene. In addition, no mutation was found in the entire MR coding sequence by RT-PCR and direct sequencing of MR mRNA, and the semiquantitative RT-PCR analysis of the MR mRNA of one affected patient failed to show any quantitative abnormality in MR expression. These results do not exclude a molecular abnormality present in the MR gene being responsible for PHA. However, they indicate that in this family PHA is not related to a modification of the MR primary structure or to a major abnormality in MR expression.

Regression of left ventricular hypertrophy and improvement of diastolic function in hypertensive patients treated with telmisartan.

OBJECTIVES: The study was designed to test whether or not the angiotensin II receptor blocker telmisartan brings about regression of left ventricular (LV) concentric hypertrophy and whether or not these changes are associated with improved diastolic filling. METHODS: An echocardiographic follow-up study was performed in 85 hypertensive patients (systolic blood pressure [SBP] >140 mmHg, diastolic blood pressure [DBP] >90 mmHg) and mild-to-moderate LV hypertrophy (LV mass index related to body surface area [LVMI] 117-150 g/m2 for men and 105-150 g/m2 for women) treated with telmisartan monotherapy 40-80 mg once daily for 1 year. Blood pressure, LVMI, left atrial (LA) volumes, and diastolic function were determined at baseline and after 3, 6, 9, and 12 months of treatment. Blood pressure was also monitored at all visits. Diastolic function was assessed by examination of transmitral inflow and pulmonary vein flow patterns. RESULTS: Telmisartan reduced blood pressure; after 12 months, the mean+/-S.D. SBP and DBP were reduced from 144+/-10 to 126+/-8 mmHg (p<0.001) and from 98+/-8 to 86+/-7 mmHg (p<0.001), respectively. The LVMI was decreased from 119+/-7 to 109+/-3 g/m2 (p<0.001) after 12 months' telmisartan treatment. All patients had diastolic dysfunction at baseline. After 12 months' telmisartan treatment, a normal pattern of transmitral inflow was present in 21% of patients. The regression of LV hypertrophy observed after 12 months was associated with increased peak early diastolic velocity/peak late diastolic velocity ratio from 0.60+/-0.18 to 0.83+/-0.20 (p<0.001), shortened isovolumic relaxation time (IVRT) from 110+/-13 to 105+/-13 ms (p<0.001), and decreased deceleration time from 229+/-30 to 215+/-28 ms (p=0.002). Univariate analysis showed that shortened IVRT was related to a reduction in the LVMI and LA maximal and minimal volumes. In the multivariate analysis, the reduction in LVMI and the reduction in LA maximal and minimal volumes were independently associated with IVRT reduction. CONCLUSIONS: Telmisartan 40-80 mg is effective in LV hypertrophy regression in hypertensive patients. The reduction in LVMI due to telmisartan monotherapy was associated with a significant improvement of diastolic filling parameters and with a significant reduction of LA volumes.

Use of Amplatzer septal occluder in a case of residual atrial septal defect causing bidirectional shunting after percutaneous Inoue mitral balloon valvuloplasty.

We describe a 70-year-old woman who underwent successful percutaneous Inoue antegrade-technique mitral valvuloplasty. Three months later, the patient developed right-sided heart failure. Color Doppler echocardiography and cardiac catheterization demonstrated an atrial septal defect with bidirectional shunting and no restenosis of the mitral valve (iatrogenic Lutembacher's syndrome). Percutaneous treatment was successfully accomplished using a 13 mm Amplatzer Septal Occluder (AGA Medical Corporation, Golden Valley, Minnesota). The causes of right-to-left shunting and the effects of body position on gas exchange in such patients are also discussed.

A comparative study of the antitussive activity of levodropropizine and dropropizine in the citric acid-induced cough model in normal subjects.

Levodropropizine is the levo-rotatory (S)-enantiomer of dropropizine, a racemic non-opiate antitussive agent which has been used clinically for many years. Compared with the racemic drug, levodropropizine exhibits in animal models similar antitussive activity but considerably lower central nervous system (CNS) depressant effects. It is also less likely to cause sedation in treated patients. Since the comparative antitussive potency of the two drugs in clinical experimental models has not been evaluated, the authors performed a randomized, double blind, cross over investigation in which the effects of single oral doses (60 and 90 mg) of levodropropizine and dropropizine were assessed by using the citric acid-induced cough model in eight normal volunteers. Stimulation tests involved inhalation of individual cumulative doses of citric acid (6.3 to 53.3 mg) which at pre-study assessment had been found to induce reproducibly at least ten coughs over a 30 sec period. Each subject was studied by repeating the citric acid stimulation test four times (0 h, 1 h, 2 h and 6 h) on each of five different days separated by intervals of at least three days. In the absence of drug administration (control session), cough response to citric inhalation was remarkably reproducible throughout the 6 h period of observation. A marked and statistically significant reduction in cough response (to about one third--one sixth of the pre-drug values) was observed 1 h after intake for both compounds. At subsequent testing 2 h and 6 h after dosing, cough response was still depressed and did not differ significantly from that observed at 1 h.(ABSTRACT TRUNCATED AT 250 WORDS)