Adrenal incidentaloma

Incidentally discovered adrenal masses, or adrenal incidentalomas, have become a common clinical problem owing to wide application of radiologic imaging techniques. This definition encompasses a heterogeneous spectrum of pathologic entities, including primary adrenocortical and medullary tumors, benign or malignant lesions, hormonally active or inactive lesions, metastases, and infections. Once an adrenal mass is detected, the clinician needs to address two crucial questions: is the mass malignant, and is it hormonally active? This article provides an overview of the diagnostic clinical approach and management of the adrenal incidentaloma. Mass size is the most reliable variable to distinguish benign and malignant adrenal masses. Adrenalectomy should be recommended for masses greater than 4.0 cm because of the increased risk of malignancy. Adrenal scintigraphy has proved useful in discriminating between benign and malignant lesions. Finally, fine-needle aspiration biopsy is an important tool in the evaluation of oncological patients and it may be useful in establishing the presence of metastatic disease. The majority of adrenal incidentalomas are non-hypersecretory cortical adenomas but an endocrine evaluation can lead to the identification of a significant number of cases with subclinical Cushing's syndrome (5-15%), pheochromocytoma (1.5-13%) and aldosteronoma (0-7%). The first step of hormonal screening should include an overnight low dose dexamethasone suppression test, the measure of urinary catecholamines or metanephrines, serum potassium and, in hypertensive patients, upright plasma aldosterone/plasma renin activity ratio. Dehydroepiandrosterone sulfate measurement may show evidence of adrenal androgen excess.

The diversity of abnormal hormone receptors in adrenal Cushing's syndrome allows novel pharmacological therapies

Recent studies from several groups have indicated that abnormal or ectopic expression and function of adrenal receptors for various hormones may regulate cortisol production in ACTH-independent hypercortisolism. Gastric inhibitory polypeptide (GIP)-dependent Cushing's syndrome has been described in patients with either unilateral adenoma or bilateral macronodular adrenal hyperplasia; this syndrome results from the large adrenal overexpression of the GIP receptor without any activating mutation. We have conducted a systematic in vivo evaluation of patients with adrenal Cushing's syndrome in order to identify the presence of abnormal hormone receptors. In macronodular adrenal hyperplasia, we have identified, in addition to GIP-dependent Cushing's syndrome, other patients in whom cortisol production was regulated abnormally by vasopressin, ß-adrenergic receptor agonists, hCG/LH, or serotonin 5HT-4 receptor agonists. In patients with unilateral adrenal adenoma, the abnormal expression or function of GIP or vasopressin receptor has been found, but the presence of ectopic or abnormal hormone receptors appears to be less prevalent than in macronodular adrenal hyperplasia. The identification of the presence of an abnormal adrenal receptor offers the possibility of a new pharmacological approach to control hypercortisolism by suppressing the endogenous ligands or by using specific antagonists for the abnormal receptors.

Biosynthesis and metabolism of steroid hormones by human adrenal carcinomas

Over a 15-year period, our university-based laboratory obtained 125 adrenal tumors, of which 15 (12%) were adrenal cortical carcinomas. Of these, 6 (40% of the carcinomas) occurred in patients with clear clinical manifestations of steroid hormone excess. Adrenal cortical carcinoma cells derived from the surgically resected tumors in 4 of these patients were isolated and established in primary culture. Radiotracer steroid interconversion studies were carried out with these cultures and also on mitochondria isolated from homogenized tissues. Large tumors had the lowest steroidogenic activities per weight, whereas small tumors had more moderately depressed enzyme activities relative to cells from normal glands. In incubations with pregnenolone as substrate, 1 mM metyrapone blocked the synthesis of corticosterone and cortisol and also the formation of aldosterone. Metyrapone inhibition was associated with a concomitant increase in the formation of androgens (androstenedione and testosterone) from pregnenolone. Administration of metyrapone in vivo before surgery in one patient resulted in a similar increase in plasma androstenedione, though plasma testosterone levels were not significantly affected. In cultures of two of four tumors examined, dibutyryl cAMP stimulated 11ß-hydroxylase activity modestly; ACTH also had a significant stimulatory effect in one of these tumors. Unlike results obtained with normal or adenomatous adrenal cortical tissues, mitochondria from carcinomatous cells showed a lack of support of either cholesterol side-chain cleavage enzyme complex or steroid 11ß-hydroxylase activity by Krebs cycle intermediates (10 mM isocitrate, succinate or malate). This finding is consistent with the concept that these carcinomas may tend to function predominantly in an anaerobic manner, rather than through the oxidation of Krebs cycle intermediates.

Phenotype and genotype correlation of the microconversion from the CYP21A1P to the CYP21A2 gene in congenital adrenal hyperplasia

Deficiency of 21-hydroxylase is the most common form of congenital adrenal hyperplasia (CAH-21OH). We determined by allele-specific PCR the frequency of microconversion in the CYP21A2 gene in 50 Brazilian patients with the classical (salt wasting: SW and simple virilizing: SV) forms and nonclassical (NC) form of CAH-21OH and correlated genotype with phenotype. Genotypes were classified into three mutation groups (A, B, and C) based on the amount of enzymatic activity in in vitro studies using adrenal cells. In 94 unrelated alleles, we diagnosed 76% of the affected alleles after screening for 7 microconversions. The most frequent point mutations observed in this series were I172N (19%), V281L (18%), and IVS2,A/C>G,-12 (15%). In the SW form, the most frequent mutation was IVS2,A/C>G,-12 (38%), in the SV form it was I172N (53%), and in the NC form it was V281L (57.7%). We observed a good correlation between genotype and phenotype. Discordance between genotype and phenotype was found in one SV patient with a mild mutation in one of the alleles (R356W/V281L). However, we cannot rule out the presence of an additional mutation in these alleles. We also observed a good correlation of genotype with 17alpha-hydroxyprogesterone, testosterone, and androstenedione levels. The severity of external genitalia virilization correlated with the severity of mutation. In conclusion, the frequencies described in the present study did not differ from worldwide studies, including the Brazilian population. The few differences observed may reflect individual sample variations. This new Brazilian cohort study suggests the presence of new mutations in Brazilian patients with different forms of CAH-21OH.

Prolactin induces adrenal hypertrophy

Although adrenocorticotropic hormone is generally considered to play a major role in the regulation of adrenal glucocorticoid secretion, several reports have suggested that other pituitary hormones (e.g., prolactin) also play a significant role in the regulation of adrenal function. The aim of the present study was to measure the adrenocortical cell area and to determine the effects of the transition from the prepubertal to the postpubertal period on the hyperprolactinemic state induced by domperidone (4.0 mg kg-1 day-1, sc). In hyperprolactinemic adult and young rats, the adrenals were heavier, as determined at necropsy, than in the respective controls: adults (30 days: 0.16 ± 0.008 and 0.11 ± 0.007; 46 days: 0.17 ± 0.006 and 0.12 ± 0.008, and 61 days: 0.17 ± 0.008 and 0.10 ± 0.004 mg for treated and control animals, respectively; P < 0.05), and young rats (30 days: 0.19 ± 0.003 and 0.16 ± 0.007, and 60 days: 0.16 ± 0.006 and 0.13 ± 0.009 mg; P < 0.05). We selected randomly a circular area in which we counted the nuclei of adrenocortical cells. The area of zona fasciculata cells was increased in hyperprolactinemic adult and young rats compared to controls: adults: (61 days: 524.90 ± 47.85 and 244.84 ± 9.03 µm² for treated and control animals, respectively; P < 0.05), and young rats: (15 days: 462.30 ± 16.24 and 414.28 ± 18.19; 60 days: 640.51 ± 12.91 and 480.24 ± 22.79 µm²; P < 0.05). Based on these data we conclude that the increase in adrenal weight observed in the hyperprolactinemic animals may be due to prolactin-induced adrenocortical cell hypertrophy.

A connection between extracellular matrix and hormonal signals during the development of the human fetal adrenal gland

The human adrenal cortex, involved in adaptive responses to stress, body homeostasis and secondary sexual characters, emerges from a tightly regulated development of a zone-specific secretion pattern during fetal life. Its development during fetal life is critical for the well being of pregnancy, the initiation of delivery, and even for an adequate adaptation to extra-uterine life. As early as from the sixth week of pregnancy, the fetal adrenal gland is characterized by a highly proliferative zone at the periphery, a concentric migration accompanied by cell differentiation (cortisol secretion) and apoptosis in the central androgen-secreting fetal zone. After birth, a strong reorganization occurs in the adrenal gland so that it better fulfills the newborn's needs, with aldosterone production in the external zona glomerulosa, cortisol secretion in the zona fasciculata and androgens in the central zona reticularis. In addition to the major hormonal stimuli provided by angiotensin II and adrenocorticotropin, we have tested for some years the hypotheses that such plasticity may be under the control of the extracellular matrix. A growing number of data have been harvested during the last years, in particular about extracellular matrix expression and its putative role in the development of the human adrenal cortex. Laminin, collagen and fibronectin have been shown to play important roles not only in the plasticity of the adrenal cortex, but also in cell responsiveness to hormones, thus clarifying some of the unexplained observations that used to feed controversies.

MMP-1/PAR-1 signal transduction axis and its prognostic impact in esophageal squamous cell carcinoma

The matrix metalloprotease-1 (MMP-1)/protease-activated receptor-1 (PAR-1) signal transduction axis plays an important role in tumorigenesis. To explore the expression and prognostic value of MMP-1 and PAR-1 in esophageal squamous cell carcinoma (ESCC), we evaluated the expression of two proteins in resected specimens from 85 patients with ESCC by immunohistochemistry. Sixty-two (72.9%) and 58 (68.2%) tumors were MMP-1- and PAR-1-positive, respectively, while no significant staining was observed in normal esophageal squamous epithelium. MMP-1 and PAR-1 overexpression was significantly associated with tumor node metastasis (TNM) stage and regional lymph node involvement. Patients with MMP-1- and PAR-1-positive tumors, respectively, had poorer disease-free survival (DFS) than those with negative ESCC (P = 0.002 and 0.003, respectively). Univariate analysis showed a significant relationship between TNM stage [hazard ratio (HR) = 2.836, 95% confidence interval (CI) = 1.866-4.308], regional lymph node involvement (HR = 2.955, 95%CI = 1.713-5.068), MMP-1 expression (HR = 2.669, 95%CI = 1.229-6.127), and PAR-1 expression (HR = 1.762, 95%CI = 1.156-2.883) and DFS. Multivariate analysis including the above four parameters identified TNM stage (HR = 2.035, 95%CI = 1.167-3.681), MMP-1 expression (HR = 2.109, 95%CI = 1.293-3.279), and PAR-1 expression (HR = 1.967, 95%CI = 1.256-2.881) as independent and significant prognostic factors for DFS. Our data suggest for the first time that MMP-1 and PAR-1 were both overexpressed in ESCC and are novel predictors of poor patient prognosis after curative resection. The MMP-1/PAR-1 signal transduction axis might be a new therapeutic target for future therapies tailored against ESCC.

Neural regulation of the stress response: glucocorticoid feedback mechanisms

The mammalian stress response is an integrated physiological and psychological reaction to real or perceived adversity. Glucocorticoids are an important component of this response, acting to redistribute energy resources to both optimize survival in the face of challenge and to restore homeostasis after the immediate challenge has subsided. Release of glucocorticoids is mediated by the hypothalamo-pituitary-adrenal (HPA) axis, driven by a neural signal originating in the paraventricular nucleus (PVN). Stress levels of glucocorticoids bind to glucocorticoid receptors in multiple body compartments, including the brain, and consequently have wide-reaching actions. For this reason, glucocorticoids serve a vital function in negative feedback inhibition of their own secretion. Negative feedback inhibition is mediated by a diverse collection of mechanisms, including fast, non-genomic feedback at the level of the PVN, stress-shut-off at the level of the limbic system, and attenuation of ascending excitatory input through destabilization of mRNAs encoding neuropeptide drivers of the HPA axis. In addition, there is evidence that glucocorticoids participate in stress activation via feed-forward mechanisms at the level of the amygdala. Feedback deficits are associated with numerous disease states, underscoring the necessity for adequate control of glucocorticoid homeostasis. Thus, rather than having a single, defined feedback ‘switch’, control of the stress response requires a wide-reaching feedback ‘network’ that coordinates HPA activity to suit the overall needs of multiple body systems.

POD-1/Tcf21 overexpression reduces endogenous SF-1 and StAR expression in rat adrenal cells

During gonad and adrenal development, the POD-1/capsulin/TCF21transcription factor negatively regulates SF-1/NR5A1expression, with higher SF-1 levels being associated with increased adrenal cell proliferation and tumorigenesis. In adrenocortical tumor cells, POD-1 binds to the SF-1 E-box promoter region, decreasing SF-1 expression. However, the modulation of SF-1 expression by POD-1 has not previously been described in normal adrenal cells. Here, we analyzed the basal expression of Pod-1 and Sf-1 in primary cultures of glomerulosa (G) and fasciculata/reticularis (F/R) cells isolated from male Sprague-Dawley rats, and investigated whether POD-1 overexpression modulates the expression of endogenous Sf-1 and its target genes in these cells. POD-1 overexpression, following the transfection of pCMVMycPod-1, significantly decreased the endogenous levels of Sf-1 mRNA and protein in F/R cells, but not in G cells, and also decreased the expression of the SF-1 target StAR in F/R cells. In G cells overexpressing POD-1, no modulation of the expression of SF-1 targets, StAR and CYP11B2, was observed. Our data showing that G and F/R cells respond differently to ectopic POD-1 expression emphasize the functional differences between the outer and inner zones of the adrenal cortex, and support the hypothesis that SF-1 is regulated by POD-1/Tcf21 in normal adrenocortical cells lacking the alterations in cellular physiology found in tumor cells.

Partial characterization of genes from the embryonic axis of Melanoxylon brauna Schott. (Leguminosae-Caesalpinioideae) seeds

The objective of this study was to partially characterize some genes involved in the desiccation tolerance of the embryonic axis of Melanoxylon brauna seeds subjected, or not, to oven fast-drying. Seeds were initially dried rapidly in an oven at 40 ºC, 50 ºC, 60 ºC, 70 ºC, and 80 °C, for 24, 48 and 72 h and then subjected to germination tests and moisture content determination. Degenerate primers were designed for 19 genes. The CDNA was used as a template for PCR amplifications using the degenerate primers, and the PCR products obtained were purified, cloned and sequenced. The seeds showed a gradual reduction in percent germination with increasing temperature and drying time. Nucleotide sequences of the cloned fragments related to genes CAT1, SPS1, Abi5, Transk and PM25 were obtained. The similarity analysis with the sequences deposited in databases revealed similarities with genes CAT1, SPS1, Transk and PM25 from other plant species. The nucleotide sequences obtained from the respective genes will be used for designing specific primers for gene expression analyses during seed germination in order to understand the causes for loss of physiological quality of Melanoxylon brauna seeds.

Far infared reflectance along the C-axis of the charge stripe superconductor La1.875Ba0.125CuO4.

Reflectance measurements along the c-axis of La1.875 Bao.125CU04 at temperatures above(6K) and below(O.5K) the bulk superconducting transition temperature(3K) were performed using a Bruker rapid scan spectrometer and a Martin-Puplett polarizing spectrometer. It was found that when polarized light reflected from a sample the Bruker rapid scan spectrometer has a low frequency cutoff of lOcm-1 while the Martin-Puplett polarizing spectrometer has a low frequency cutoff of 6cm-1 A superconducting pla ma edge was absent in all measurements taken. It was concluded that if a superconducting plasma edge exists in La1.875Bao.125CU04 it is below 6cm-1.

Negative affect mediates the relationship between the Cortisol Awakening Response and Conduct Problems in boys

Ce mémoire débute avec deux chapitres portant sur les problèmes des conduites et la régulation du stress, notamment sur l’axe hypothalamique-pituitaire-surrénal (HPS). Ensuite, la littérature est résumée et nous voyons que les études qui cherchent à établir un lien entre les problèmes des conduites et l’axe HPS ont trouvé des résultats différents et parfois contradictoires. Le chapitre suivant illustre les problèmes méthodologiques qui pourraient expliquer ces résultats différents. Vient ensuite l’étude présentée dans ce mémoire qui cherche à établir un lien entre la réponse cortisolaire à l’éveil (RCE), considérée comme un bon indice du fonctionnent de l’axe HPS, et les problèmes de conduites chez l’enfant. De plus, les émotions négatives ont été associées avec les problèmes des conduites ainsi qu’aux dysfonctions de l’axe HPS, notamment le RCE. L’étude présentée dans ce mémoire cherche aussi à établir si les émotions négatives pourrait être une variable médiatrice dans la relation potentielle entre la RCE et les problèmes des conduites. L’étude révèle que pour les garçons mais pas pour les filles, une RCE réduite est associée avec les émotions négatives, ce qui est successivement associé avec les problèmes des conduites. Le dernier chapitre du mémoire examine les implications théoriques de cette médiatisation et propose également des pistes psychobiologiques pour expliquer les différences sexuelles observées.

Génomique fonctionnelle des cellules corticotropes hypophysaires : contrôle génétique de la gestion systémique des stress

L'axe hypothalamo-hypophyso-surrénalien (HPA) permet de maintenir l'homéostasie de l'organisme face à divers stress. Qu'ils soient de nature psychologique, physique ou inflammatoire/infectieux, les stress provoquent la synthèse et la libération de CRH par l'hypothalamus. Les cellules corticotropes hypophysaires perçoivent ce signal et en réaction, produisent et sécrètent l'ACTH. Ceci induit la synthèse des glucocorticoïdes (Gc) par le cortex surrénalien; ces stéroïdes mettent le système métabolique en état d’alerte pour la réponse au stress et à l’agression. Les Gc ont le rôle essentiel de contrôler les défenses de l'organisme, en plus d'exercer une rétro-inhibition sur l'axe HPA. L'ACTH est une petite hormone peptidique produite par le clivage d'un précurseur: la pro-opiomélanocortine (POMC). À cause de sa position critique dans la normalisation de l'homéostasie, le contrôle transcriptionnel du gène Pomc a fait l'objet d'études approfondies au cours des dernières décennies. Nous savons maintenant que la région promotrice du gène Pomc permet une expression ciblée dans les cellules POMC hypophysaires. L'étude du locus Pomc par des technologies génomiques m'a permis de découvrir un nouvel élément de régulation qui est conservé à travers l'évolution des mammifères. La caractérisation de cet enhancer a démontré qu'il dirige une expression restreinte à l'hypophyse, et plus particulièrement dans les cellules corticotropes. De façon intéressante, l'activité de cet élément dépend d'un nouveau site de liaison recrutant un homodimère du facteur de transcription Tpit, dont l'expression est également limitée aux cellules POMC de l'hypophyse. La découverte de cet enhancer ajoute une toute nouvelle dimension à la régulation de l'expression de POMC. Les cytokines pro-inflammatoires IL6/LIF et les Gc sont connus pour leur antagonisme sur la réaction inflammatoire et sur le promoteur Pomc via l'action des facteurs de transcription Stat3 et GR respectivement. L'analyse génomique des sites liés ii par ces deux facteurs nous a révélé une interrelation complexe et a permis de définir un code transcriptionnel entre ces voies de signalisation. En plus de leur action par interaction directe avec l’ADN au niveau des séquences régulatrices, ces facteurs interagissent directement entre eux avec des résultats transcriptionnels différents. Ainsi, le recrutement de GR par contact protéine:protéine (tethering) sur Stat3 étant lié à l'ADN provoque un antagonisme transcriptionnel. Inversement, le tethering de Stat3 sur GR supporte une action synergique, tout comme leur co-recrutement à l'ADN sur des sites contigus ou composites. Lors d'une activation soutenue, ce synergisme entre les voies IL6/LIF et Gc induit une réponse innée de défense cellulaire. Ainsi lors d'un stress majeur, ce mécanisme de défense est mis en branle dans toutes les cellules et tissus. En somme, les travaux présentés dans cette thèse définissent les mécanismes transcriptionnels engagés dans le combat de l'organisme contre les stress. Plus particulièrement, ces mécanismes ont été décrits au niveau de la réponse globale des corticotropes et du gène Pomc. Il est essentiel pour l'organisme d'induire adéquatement ces mécanismes afin de faire face aux stress et d'éviter des dérèglements comme les maladies inflammatoires et métaboliques.

Spatiotemporal regulation of the Greatwall : PP2A axis is required for mitotic progression

Le cycle cellulaire est hautement régulé par la phosphorylation réversible de plusieurs effecteurs. La kinase dépendante des cyclines Cdk1 déclenche la mitose en induisant le bris de l’enveloppe nucléaire, la condensation des chromosomes et la formation du fuseau mitotique. Chez les animaux métazoaires, ces évènements sont contrés par la protéine phosphatase PP2A-B55, qui déphosphoryle plusieurs substrats de Cdk1. La kinase Greatwall (Gwl) est activée par le complexe cycline B-Cdk1 en début de mitose et induit ensuite l’inhibition de PP2A-B55 via Endos/Arpp19. Toutefois, les mécanismes moléculaires qui régulent Gwl sont encore peu connus. Nous avons montré que Gwl a une activité s’opposant à PP2A-B55, qui collabore avec la kinase Polo pour assurer l’attachement du centrosome au noyau et la progression du cycle cellulaire dans le syncytium de l’embryon de la drosophile. Ensuite, nous avons trouvé dans des cellules de drosophile que Gwl est localisée au noyau pendant l’interphase, mais qu’elle se relocalise au cytoplasme dès la prophase, avant le bris de l’enveloppe nucléaire. Nous avons montré que cette translocation de Gwl est cruciale pour sa fonction et qu’elle dépend de la phosphorylation de plusieurs résidus de la région centrale de Gwl par les kinases Polo et Cdk1. Cette région centrale contient également deux séquences de localisation nucléaire (respectivement NLS1 et NLS2). De plus, nos résultats suggèrent que la phosphorylation de Gwl par la kinase Polo promeut sa liaison avec la protéine 14-3-3ε, ce qui favorise la rétention cytoplasmique de Gwl. Le rôle de Cdk1 dans cette translocation reste quant à lui inconnu. De plus, nous avons montré que le complexe cycline B-Cdk1 entre dans le noyau avant que Gwl ne soit transportée dans le cytoplasme. Cdk1 pourrait donc activer Gwl et phosphoryler ses substrats nucléaires, à l’abri de PP2A-B55 qui est largement cytoplasmique. Gwl est ensuite exclue du noyau et relocalisée dans le cytoplasme afin d’induire l’inhibition de PP2A-B55. Cela permet de synchroniser les événements de phosphorylation se produisant dans le noyau et dans le cytoplasme. Fait intéressant, un mécanisme de régulation de la localisation de Gwl similaire à cela a été découvert chez l’humain et chez la levure, suggérant que ce mécanisme est conservé entre différentes espèces.