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1.
Hypertension ; 74(5): 1152-1159, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31564164

RESUMO

Microarray comparison of the transcriptomes of human adrenal zona glomerulosa (ZG) and zona fasciculata found several ZG-specific genes that negatively regulate aldosterone secretion. The third and most significantly upregulated ZG-gene (19.9-fold compared with zona fasciculata, P=6.58×10-24) was ANO4, a putative Ca2+-activated chloride channel. We have investigated the role of ANO4 in human adrenal, and whether it functions like the prototype anoctamin, ANO1. We evaluated ANO4 mRNA and protein expression in human adrenal by qPCR and immunohistochemistry, compared the effects of ANO4 and ANO1 overexpression on baseline and stimulated aldosterone secretion and cell proliferation in H295R cells, and analyzed ANO4 activity as a Ca2+-activated chloride channel in comparison with other anoctamins by a fluorescence-based functional assay. The expression of ANO4 in ZG was confirmed by qPCR as 23.21-fold upregulated compared with zona fasciculata (n=18; P=4.93×10-7). Immunohistochemistry found cytoplasmic, ZG-selective expression of ANO4 (anoctamin 4) protein. ANO4 overexpression in H295R cells attenuated calcium-mediated aldosterone secretion and cell proliferation in comparison to controls. The latter effects were in a different direction to those of ANO1. The functional assay showed that, in contrast to ANO1, ANO4 expression results in low levels of calcium-dependent anion transport. In conclusion, ANO4 is one of the most highly expressed genes in ZG. It attenuates stimulated aldosterone secretion and cell proliferation. Although belonging to a family of Ca2+-activated chloride channels, it does not generate significant plasma membrane chloride channel activity.


Assuntos
Aldosterona/biossíntese , Anoctaminas/genética , Regulação da Expressão Gênica , Hiperaldosteronismo/genética , Hipertensão/fisiopatologia , Transdução de Sinais/genética , Zona Glomerulosa/metabolismo , Córtex Suprarrenal/citologia , Córtex Suprarrenal/metabolismo , Córtex Suprarrenal/patologia , Análise de Variância , Comunicação Celular/genética , Proliferação de Células , Células Cultivadas , Imunofluorescência , Humanos , Hiperaldosteronismo/patologia , Hipertensão/etiologia , Reação em Cadeia da Polimerase em Tempo Real/métodos , Análise Serial de Tecidos , Técnicas de Cultura de Tecidos , Transcriptoma/genética , Regulação para Cima , Zona Fasciculada/metabolismo , Zona Fasciculada/patologia , Zona Glomerulosa/patologia
2.
Medicina (Kaunas) ; 54(5)2018 Nov 19.
Artigo em Inglês | MEDLINE | ID: mdl-30463213

RESUMO

Background and objectives: Energy drinks are popular non-alcoholic beverages. They are consumed in large amounts, mainly by active, young people. Although they are easily accessible and marketed as safe, numerous cases of adverse effects have been published, including cardiac arrest, arrythmias, acute hepatitis, and renal failure. The aim of the current study is the assessment of energy drink influence on the histological structure of adrenal cortex in rats. Material and Methods: 15 male young Wistar rats were equally divided into three groups: control (C), experimental (E) and reversibility control (RC). C group received water and standard rodent food ad libitum while both E and RC groups had additionally unlimited access to energy drinks. C and E groups were decapitated after 8 weeks and RC was given another 8 weeks without energy drinks. Adrenal glands were embedded in paraffin blocks and 5 µm slides were prepared and stained according to standard H&E and Masson's trichrome protocols. Additionally, immunohistochemical stainings against Ki-67, p53, CTGF and caspase-3 were prepared. Results: Decreased vacuolization and numerous pyknotic nuclei were noted in E and RC groups. Overexpression of caspase-3 was noted both subcapsular in zona glomerulosa and along sinusoids in zona fasciculata. Increased collagen deposition in zona glomerulosa and zona fasciculata of E and RC was observed. Insular and irregular overexpression of CTGF was noted. The overall picture of CTGF expression matched the Masson's trichrome. No significant difference was observed in Ki-67 expression. Conclusions: The results of the current study suggest that the stimulation is so intense that it causes significant damage to adrenal cortical cells, resulting in their apoptosis. It seems, however, that the observed effects are at least partially reversible.


Assuntos
Cafeína/efeitos adversos , Bebidas Energéticas/efeitos adversos , Gotículas Lipídicas , Taurina/efeitos adversos , Zona Fasciculada/metabolismo , Zona Fasciculada/patologia , Zona Glomerulosa/metabolismo , Zona Glomerulosa/patologia , Animais , Apoptose , Caspase 3/biossíntese , Colágeno/biossíntese , Fator de Crescimento do Tecido Conjuntivo/biossíntese , Antígeno Ki-67/biossíntese , Masculino , Ratos , Ratos Wistar , Zona Fasciculada/citologia , Zona Glomerulosa/citologia
3.
Hypertension ; 72(4): 874-880, 2018 10.
Artigo em Inglês | MEDLINE | ID: mdl-30354720

RESUMO

Primary aldosteronism affects ≈5% to 10% of hypertensive patients and has unilateral and bilateral forms. Most unilateral primary aldosteronism is caused by computed tomography-detectable aldosterone-producing adenomas, which express CYP11B2 (aldosterone synthase) and frequently harbor somatic mutations in aldosterone-regulating genes. The cause of the most common bilateral form of primary aldosteronism, idiopathic hyperaldosteronism (IHA), is believed to be diffuse hyperplasia of aldosterone-producing cells within the adrenal cortex. Herein, a multi-institution cohort of 15 IHA adrenals was examined with CYP11B2 immunohistochemistry and next-generation sequencing. CYP11B2 immunoreactivity in adrenal glomerulosa harboring non-nodular hyperplasia was only observed in 4/15 IHA adrenals suggesting that hyperplasia of CYP11B2-expressing cells may not be the major cause of IHA. However, the adrenal cortex of all IHA adrenals harbored at least 1 CYP11B2-positive aldosterone-producing cell cluster (APCC) or micro-aldosterone-producing adenomas. The number of APCCs per case (and individual APCC area) in IHA adrenals was significantly larger than in normotensive controls. Next-generation sequencing of DNA from 99 IHA APCCs demonstrated somatic mutations in genes encoding the L-type calcium voltage-gated channel subunit α 1-D ( CACNA1D, n=57; 58%) and potassium voltage-gated channel subfamily J-5 ( KCNJ5, n=1; 1%). These data suggest that IHA may result from not only hyperplasia but also the accumulation or enlargement of computed tomography-undetectable APCC harboring somatic aldosterone-driver gene mutations. The high prevalence of mutations in the CACNA1D L-type calcium channel provides a potential actionable therapeutic target that could complement mineralocorticoid blockade and inhibit aldosterone overproduction in some IHA patients.


Assuntos
Aldosterona , Canais de Cálcio Tipo L/genética , Canais de Potássio Corretores do Fluxo de Internalização Acoplados a Proteínas G/genética , Hiperaldosteronismo , Hipertensão , Zona Glomerulosa , Aldosterona/biossíntese , Aldosterona/genética , Citocromo P-450 CYP11B2/metabolismo , Feminino , Regulação da Expressão Gênica , Humanos , Hiperaldosteronismo/complicações , Hiperaldosteronismo/genética , Hiperaldosteronismo/metabolismo , Hiperplasia , Hipertensão/etiologia , Hipertensão/metabolismo , Imuno-Histoquímica , Masculino , Mutação , Zona Glomerulosa/metabolismo , Zona Glomerulosa/patologia
4.
Endocr Rev ; 39(6): 1029-1056, 2018 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-30007283

RESUMO

The identification of several germline and somatic ion channel mutations in aldosterone-producing adenomas (APAs) and detection of cell clusters that can be responsible for excess aldosterone production, as well as the isolation of autoantibodies activating the angiotensin II type 1 receptor, have rapidly advanced the understanding of the biology of primary aldosteronism (PA), particularly that of APA. Hence, the main purpose of this review is to discuss how discoveries of the last decade could affect histopathology analysis and clinical practice. The structural remodeling through development and aging of the human adrenal cortex, particularly of the zona glomerulosa, and the complex regulation of aldosterone, with emphasis on the concepts of zonation and channelopathies, will be addressed. Finally, the diagnostic workup for PA and its subtyping to optimize treatment are reviewed.


Assuntos
Neoplasias do Córtex Suprarrenal , Adenoma Adrenocortical , Envelhecimento , Aldosterona/metabolismo , Hiperaldosteronismo , Zona Glomerulosa , Neoplasias do Córtex Suprarrenal/diagnóstico , Neoplasias do Córtex Suprarrenal/metabolismo , Neoplasias do Córtex Suprarrenal/patologia , Adenoma Adrenocortical/diagnóstico , Adenoma Adrenocortical/metabolismo , Adenoma Adrenocortical/patologia , Envelhecimento/metabolismo , Envelhecimento/patologia , Humanos , Hiperaldosteronismo/diagnóstico , Hiperaldosteronismo/metabolismo , Hiperaldosteronismo/patologia , Zona Glomerulosa/metabolismo , Zona Glomerulosa/patologia
5.
Folia Morphol (Warsz) ; 77(4): 629-641, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29611160

RESUMO

BACKGROUND: Stress exposure exerts direct effects on the morphology and functionality of the adrenal cortex. In addition, ageing effects growth, differentiation, apoptosis and cellularity of the cortex. The missing data is the combined effect of stress and ageing on the adrenal cortex. The aim of this study was to demonstrate the structural changes in the adrenal cortex following the exposure to stress in the adult and aged albino rats. MATERIALS AND METHODS: Forty rats were divided into groups I and II (adult and senile). Each group was further subdivided into subgroups a and b (control and stressed). Light and electron microscopic studies were done. Area per cent of collagen fibres (Masson's trichrome-stained sections), number of proliferating cells (optical density immunoreactivity in the Ki67 stained sections) and thickness of the three adrenal zones were also measured. RESULTS: Lamellar separation of the capsule with subcapsular spindle cell hyperplasia and areas of ghost cells were observed in zona glomerulosa (ZG) and zona fasciculata (ZF) in group I-b. Separation and indentation of the capsule with its lamellar separation were observed in group II-a with the existence of multiple scattered degenerative foci in ZF and zona reticularis (ZR). Similar and aggressive was the architectural pattern of ZF in group II-b with the presence of areas of homogenous degeneration. The nuclei of ZG had marginated chromatin in group I-b and were pyknotic with deformed irregular outlines in group II-b. Multiple lysosomes and vacuolar degeneration mitochondria were also seen in group I-b. The nuclei of ZF were irregular with condensed marginated heterochromatin in group I-b, irregular with scattered chromatin in group II-a and indented with areas of chromatin destruction in group II-b. Mitochondria with disrupted cristae and cristolysis were also detected in group I-b. Numerous lipofuscin granules and dilated smooth endoplasmic reticulum were revealed in group II-b. The mean collagen fibre area per cent and the mean number of the proliferating cells in group II-b were significantly higher by 39% and 23%. The thickness of ZG decreased significantly by 20% in group I-b. Contrary, the thickness of both ZF and ZR increased significantly by 10% in group I-b. CONCLUSIONS: Histological alterations occurred in the adrenal cortex in response to stress, especially when coupled with the advance of age. This was accompanied by increase in the area per cent of collagen fibres and increase in the mean number of the proliferating cells in the adrenal cortex.


Assuntos
Envelhecimento/patologia , Estresse Psicológico , Zona Fasciculada , Zona Glomerulosa , Zona Reticular , Animais , Masculino , Ratos , Ratos Wistar , Estresse Psicológico/metabolismo , Estresse Psicológico/patologia , Zona Fasciculada/metabolismo , Zona Fasciculada/patologia , Zona Glomerulosa/metabolismo , Zona Glomerulosa/patologia , Zona Reticular/metabolismo , Zona Reticular/patologia
6.
Bull Exp Biol Med ; 164(4): 493-496, 2018 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-29504097

RESUMO

Prenatal and postnatal exposure to low doses of the endocrine disruptor dichlorodiphenyltrichloroethane (DDT) leads to delayed activation of the canonical ß-catenin/Wnt signaling in zona glomerulosa and zona reticularis of the adrenal cortex in rats, which changed the rate of their postnatal development. Suppression of the Wnt pathway in zona fasciculata promotes its regeneration after DDT-induced blood circulation disorders and cell death.


Assuntos
Animais Recém-Nascidos/genética , DDT/farmacologia , Disruptores Endócrinos/farmacologia , Zona Glomerulosa/efeitos dos fármacos , Zona Reticular/efeitos dos fármacos , beta Catenina/genética , Animais , Animais Recém-Nascidos/metabolismo , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Masculino , Gravidez , Efeitos Tardios da Exposição Pré-Natal , Ratos , Ratos Wistar , Via de Sinalização Wnt , Zona Glomerulosa/crescimento & desenvolvimento , Zona Glomerulosa/metabolismo , Zona Glomerulosa/patologia , Zona Reticular/crescimento & desenvolvimento , Zona Reticular/metabolismo , Zona Reticular/patologia , beta Catenina/metabolismo
7.
Clin Sci (Lond) ; 132(6): 627-640, 2018 03 30.
Artigo em Inglês | MEDLINE | ID: mdl-29436482

RESUMO

The angiotensin II (Ang II) type 2 receptor (AT2R) and the angiotensin-(1-7) (Ang-(1-7)) receptor (MasR) play a cardiovascular protective role by counter-regulating Ang II type 1 receptor (AT1R)-mediated effects, but whether this involves blunting of adrenocortical hormone secretion is unknown. We investigated the presence of AT1R, AT2R, and MasR in aldosterone-producing adenoma (APA), a condition featuring hyperaldosteronism, and in APA-adjacent tissue. The effect of Compound 21 (C21), an AT2R agonist, on CYP11B1 (cortisol synthase) and CYP11B2 (aldosterone synthase) gene expression in NCI-H295R and HAC15 cell lines, and in APA and APA-adjacent tissue, was also assessed using the AT1R antagonist irbesartan to ascertain the specificity of C21 effect. We found that the AT1R, AT2R, and MasR were expressed in APA and APA-adjacent tissue, albeit heterogeneously. The gene expression of AT1R and AT2R was lower, and that of the MasR higher in APAs than in APA-adjacent tissue. In steroid-producing NCI-H295R and HAC15 cell lines, and in APA and APA-adjacent tissue, C21 was ineffective at nanomolar concentrations, but increased CYP11B1 and CYP11B2 gene expression at micromolar concentrations through AT1R, as this effect was blunted by irbesartan. The scant expression of the AT2R, along with the lack of any effect of C21 at low concentrations on CYP11B2, do not support the contention that the protective arm of renin-angiotensin system (RAS) blunts aldosterone synthase in the normal adrenal cortex and primary aldosteronism.


Assuntos
Neoplasias do Córtex Suprarrenal/metabolismo , Adenoma Adrenocortical/metabolismo , Aldosterona/metabolismo , Hiperaldosteronismo/metabolismo , Receptor Tipo 2 de Angiotensina/metabolismo , Sistema Renina-Angiotensina , Zona Glomerulosa/metabolismo , Neoplasias do Córtex Suprarrenal/genética , Neoplasias do Córtex Suprarrenal/patologia , Adenoma Adrenocortical/genética , Adenoma Adrenocortical/patologia , Bloqueadores do Receptor Tipo 1 de Angiotensina II/farmacologia , Estudos de Casos e Controles , Linhagem Celular , Citocromo P-450 CYP11B2/genética , Citocromo P-450 CYP11B2/metabolismo , Humanos , Hiperaldosteronismo/genética , Hiperaldosteronismo/patologia , Irbesartana/farmacologia , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Receptor Tipo 1 de Angiotensina/genética , Receptor Tipo 1 de Angiotensina/metabolismo , Receptor Tipo 2 de Angiotensina/agonistas , Receptor Tipo 2 de Angiotensina/genética , Receptores Acoplados a Proteínas-G/genética , Receptores Acoplados a Proteínas-G/metabolismo , Sistema Renina-Angiotensina/efeitos dos fármacos , Sistema Renina-Angiotensina/genética , Esteroide 11-beta-Hidroxilase/genética , Esteroide 11-beta-Hidroxilase/metabolismo , Sulfonamidas/farmacologia , Tiofenos/farmacologia , Zona Glomerulosa/efeitos dos fármacos , Zona Glomerulosa/patologia
8.
Nat Genet ; 50(3): 355-361, 2018 03.
Artigo em Inglês | MEDLINE | ID: mdl-29403012

RESUMO

Primary aldosteronism is the most common and curable form of secondary arterial hypertension. We performed whole-exome sequencing in patients with early-onset primary aldosteronism and identified a de novo heterozygous c.71G>A/p.Gly24Asp mutation in the CLCN2 gene, encoding the voltage-gated ClC-2 chloride channel 1 , in a patient diagnosed at 9 years of age. Patch-clamp analysis of glomerulosa cells of mouse adrenal gland slices showed hyperpolarization-activated Cl- currents that were abolished in Clcn2-/- mice. The p.Gly24Asp variant, located in a well-conserved 'inactivation domain'2,3, abolished the voltage- and time-dependent gating of ClC-2 and strongly increased Cl- conductance at resting potentials. Expression of ClC-2Asp24 in adrenocortical cells increased expression of aldosterone synthase and aldosterone production. Our data indicate that CLCN2 mutations cause primary aldosteronism. They highlight the important role of chloride in aldosterone biosynthesis and identify ClC-2 as the foremost chloride conductor of resting glomerulosa cells.


Assuntos
Canais de Cloreto/genética , Mutação com Ganho de Função , Hiperaldosteronismo/genética , Adulto , Animais , Criança , Canais de Cloreto/metabolismo , Estudos de Coortes , Análise Mutacional de DNA , Feminino , Mutação em Linhagem Germinativa , Humanos , Hiperaldosteronismo/patologia , Camundongos , Camundongos Knockout , Pessoa de Meia-Idade , Linhagem , Sequenciamento Completo do Exoma , Adulto Jovem , Zona Glomerulosa/metabolismo , Zona Glomerulosa/patologia
9.
JCI Insight ; 2(23)2017 12 07.
Artigo em Inglês | MEDLINE | ID: mdl-29212953

RESUMO

Primary and secondary hypertension are major risk factors for cardiovascular disease, the leading cause of death worldwide. Elevated secretion of aldosterone resulting from primary aldosteronism (PA) is a key driver of secondary hypertension. Here, we report an unexpected role for the ubiquitin ligase Siah1 in adrenal gland development and PA. Siah1a-/- mice exhibit altered adrenal gland morphology, as reflected by a diminished X-zone, enlarged medulla, and dysregulated zonation of the glomerulosa as well as increased aldosterone levels and aldosterone target gene expression and reduced plasma potassium levels. Genes involved in catecholamine biosynthesis and cAMP signaling are upregulated in the adrenal glands of Siah1a-/- mice, while genes related to retinoic acid signaling and cholesterol biosynthesis are downregulated. Loss of Siah1 leads to increased expression of the Siah1 substrate PIAS1, an E3 SUMO protein ligase implicated in the suppression of LXR, a key regulator of cholesterol levels in the adrenal gland. In addition, SIAH1 sequence variants were identified in patients with PA; such variants impaired SIAH1 ubiquitin ligase activity, resulting in elevated PIAS1 expression. These data identify a role for the Siah1-PIAS1 axis in adrenal gland organization and function and point to possible therapeutic targets for hyperaldosteronism.


Assuntos
Glândulas Suprarrenais/metabolismo , Aldosterona/metabolismo , Proteínas/fisiologia , Glândulas Suprarrenais/patologia , Medula Suprarrenal/patologia , Adulto , Animais , Colesterol/biossíntese , Feminino , Regulação da Expressão Gênica/fisiologia , Humanos , Hiperaldosteronismo/genética , Hiperaldosteronismo/metabolismo , Rim/metabolismo , Masculino , Camundongos Knockout , Pessoa de Meia-Idade , Mutação , Proteínas Nucleares/genética , Potássio/sangue , Proteínas Inibidoras de STAT Ativados/biossíntese , Proteínas Inibidoras de STAT Ativados/genética , Proteínas Inibidoras de STAT Ativados/metabolismo , Proteínas/genética , Transdução de Sinais/genética , Proteínas Modificadoras Pequenas Relacionadas à Ubiquitina/metabolismo , Tretinoína/fisiologia , Ubiquitina-Proteína Ligases/genética , Zona Glomerulosa/metabolismo , Zona Glomerulosa/patologia
11.
J Clin Endocrinol Metab ; 102(6): 1792-1796, 2017 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-28368480

RESUMO

Context: Whether primary aldosteronism (PA) is the consequence of a monoclonal or multiclonal process is unclear. Case Description: A 48-year-old man with severe bilateral PA refractory to medical therapy underwent unilateral adrenalectomy of the dominant adrenal. Although computed tomography showed three left-sided cortical nodules, postsurgical histopathology and genetic analysis revealed five different adrenocortical adenomas. Two zona fasciculata (ZF)-like aldosterone-producing adenomas (APAs) each harbored distinct known somatic KCNJ5 mutations (L168R and T158A). A zona glomerulosa-like APA harbored a known CACNA1D G403R somatic mutation, whereas a zona reticularis-like adenoma, which was grossly black in pigmentation with histologic characteristics more associated with cortisol-producing adenomas, expressed CYP11B2, CYP17, and DHEA-ST by immunohistochemistry (IHC) and harbored no known somatic mutations. The fifth adenoma was ZF-type, negative for CYP11B2 and CYP17 IHC, and harbored no known somatic mutations. Conclusions: This case highlights complex intertumor heterogeneity in histology, steroidogenesis, and somatic mutations in multiple adrenocortical adenomas arising in a single patient with PA. These findings suggest that the syndrome of PA can involve heterogeneous and multiclonal functional adrenal adenomas.


Assuntos
Neoplasias do Córtex Suprarrenal/genética , Adenoma Adrenocortical/genética , Hiperaldosteronismo/genética , Neoplasias Primárias Múltiplas/genética , Neoplasias do Córtex Suprarrenal/metabolismo , Neoplasias do Córtex Suprarrenal/patologia , Neoplasias do Córtex Suprarrenal/cirurgia , Adenoma Adrenocortical/metabolismo , Adenoma Adrenocortical/patologia , Adenoma Adrenocortical/cirurgia , Canais de Cálcio Tipo L/genética , Citocromo P-450 CYP11B2/metabolismo , Canais de Potássio Corretores do Fluxo de Internalização Acoplados a Proteínas G/genética , Humanos , Hiperaldosteronismo/metabolismo , Hiperaldosteronismo/patologia , Hiperaldosteronismo/cirurgia , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Neoplasias Primárias Múltiplas/metabolismo , Neoplasias Primárias Múltiplas/patologia , Neoplasias Primárias Múltiplas/cirurgia , Esteroide 17-alfa-Hidroxilase/metabolismo , Sulfotransferases/metabolismo , Zona Fasciculada/patologia , Zona Glomerulosa/patologia , Zona Reticular/patologia
12.
Endocr J ; 64(1): 39-47, 2017 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-27681703

RESUMO

Somatic mutations in KCNJ5 gene have been identified in patients with adrenal aldosterone-producing adenomas (APAs). We previously reported that Japanese patients with APAs had distinct characteristics from patients in Western countries; i.e. they had a high frequency of KCNJ5 mutations and exhibited a frequent association with cortisol co-secretion. Therefore, APAs among Japanese patients may have different features from those in Western countries. We added recent cases, examined 47 cases (43% male) of APAs, including clinicopathological features, KCNJ5 mutations, and the mRNA levels of several steroidogenic enzymes, and compared the results obtained to those reported in other countries. While the prevalence of KCNJ5 mutations is approximately 40% in Western countries, 37 APA cases (78.7%) showed mutations: 26 with p.G151R and 11 with p.L168R. Although a significant gender difference has been reported in the frequency of KCNJ5 mutations in Europe, we did not find any gender difference. However, the phenotypes of Japanese patients with mutations were similar to those of patients in Western countries; patients were younger and had higher plasma aldosterone levels, lower potassium levels, and higher diastolic blood pressure. Reflecting these phenotypes, APAs with mutations had higher CYP11B2 mRNA levels. However, in contrast to APAs in Western countries, Japanese APAs with mutations showed lower CYP11B1, CYP17A1, and CYP11A1 mRNA levels. These findings demonstrated that Japanese APA patients may have distinct features including a higher prevalence of KCNJ5 mutations, no gender difference in the frequency of these mutations, and characteristics similar to the zona glomerulosa.


Assuntos
Neoplasias do Córtex Suprarrenal/genética , Adenoma Adrenocortical/genética , Aldosterona/metabolismo , Canais de Potássio Corretores do Fluxo de Internalização Acoplados a Proteínas G/genética , Hiperaldosteronismo/genética , Mutação , Neoplasias do Córtex Suprarrenal/metabolismo , Neoplasias do Córtex Suprarrenal/patologia , Adenoma Adrenocortical/metabolismo , Adenoma Adrenocortical/patologia , Adulto , Idoso , Feminino , Humanos , Hiperaldosteronismo/metabolismo , Hiperaldosteronismo/patologia , Japão , Masculino , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único , Fatores Sexuais , Zona Glomerulosa/patologia
13.
Elife ; 4: e06315, 2015 Apr 24.
Artigo em Inglês | MEDLINE | ID: mdl-25907736

RESUMO

Many Mendelian traits are likely unrecognized owing to absence of traditional segregation patterns in families due to causation by de novo mutations, incomplete penetrance, and/or variable expressivity. Genome-level sequencing can overcome these complications. Extreme childhood phenotypes are promising candidates for new Mendelian traits. One example is early onset hypertension, a rare form of a global cause of morbidity and mortality. We performed exome sequencing of 40 unrelated subjects with hypertension due to primary aldosteronism by age 10. Five subjects (12.5%) shared the identical, previously unidentified, heterozygous CACNA1H(M1549V) mutation. Two mutations were demonstrated to be de novo events, and all mutations occurred independently. CACNA1H encodes a voltage-gated calcium channel (CaV3.2) expressed in adrenal glomerulosa. CACNA1H(M1549V) showed drastically impaired channel inactivation and activation at more hyperpolarized potentials, producing increased intracellular Ca(2+), the signal for aldosterone production. This mutation explains disease pathogenesis and provides new insight into mechanisms mediating aldosterone production and hypertension.


Assuntos
Canais de Cálcio Tipo T/genética , Cálcio/metabolismo , Hiperaldosteronismo/genética , Hipertensão/genética , Mutação , Adolescente , Adulto , Idade de Início , Aldosterona/biossíntese , Aldosterona/metabolismo , Sequência de Aminoácidos , Canais de Cálcio Tipo T/metabolismo , Sinalização do Cálcio , Criança , Pré-Escolar , Feminino , Expressão Gênica , Genótipo , Heterozigoto , Humanos , Hiperaldosteronismo/complicações , Hiperaldosteronismo/metabolismo , Hiperaldosteronismo/patologia , Hipertensão/complicações , Hipertensão/metabolismo , Hipertensão/patologia , Lactente , Masculino , Potenciais da Membrana , Pessoa de Meia-Idade , Dados de Sequência Molecular , Fenótipo , Recidiva , Alinhamento de Sequência , Zona Glomerulosa/metabolismo , Zona Glomerulosa/patologia
14.
Hypertension ; 65(5): 1103-10, 2015 May.
Artigo em Inglês | MEDLINE | ID: mdl-25776071

RESUMO

Common somatic mutations in CACNAID and ATP1A1 may define a subgroup of smaller, zona glomerulosa (ZG)-like aldosterone-producing adenomas. We have therefore sought signature ZG genes, which may provide insight into the frequency and pathogenesis of ZG-like aldosterone-producing adenomas. Twenty-one pairs of zona fasciculata and ZG and 14 paired aldosterone-producing adenomas from 14 patients with Conn's syndrome and 7 patients with pheochromocytoma were assayed by the Affymetrix Human Genome U133 Plus 2.0 Array. Validation by quantitative real-time polymerase chain reaction was performed on genes >10-fold upregulated in ZG (compared with zona fasciculata) and >10-fold upregulated in aldosterone-producing adenomas (compared with ZG). DACH1, a gene associated with tumor progression, was further analyzed. The role of DACH1 on steroidogenesis, transforming growth factor-ß, and Wnt signaling activity was assessed in the human adrenocortical cell line, H295R. Immunohistochemistry confirmed selective expression of DACH1 in human ZG. Silencing of DACH1 in H295R cells increased CYP11B2 mRNA levels and aldosterone production, whereas overexpression of DACH1 decreased aldosterone production. Overexpression of DACH1 in H295R cells activated the transforming growth factor-ß and canonical Wnt signaling pathways but inhibited the noncanonical Wnt signaling pathway. Stimulation of primary human adrenal cells with angiotensin II decreased DACH1 mRNA expression. Interestingly, there was little overlap between our top ZG genes and those in rodent ZG. In conclusion, (1) the transcriptome profile of human ZG differs from rodent ZG, (2) DACH1 inhibits aldosterone secretion in human adrenals, and (3) transforming growth factor-ß signaling pathway is activated in DACH1 overexpressed cells and may mediate inhibition of aldosterone secretion in human adrenals.


Assuntos
Carcinoma Adrenocortical/genética , Aldosterona/metabolismo , Proteínas do Olho/genética , Regulação Neoplásica da Expressão Gênica , RNA Neoplásico/genética , Fatores de Transcrição/genética , Fator de Crescimento Transformador beta/genética , Zona Glomerulosa/metabolismo , Carcinoma Adrenocortical/metabolismo , Carcinoma Adrenocortical/patologia , Progressão da Doença , Proteínas do Olho/biossíntese , Feminino , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase em Tempo Real , Transdução de Sinais , Fatores de Transcrição/biossíntese , Fator de Crescimento Transformador beta/biossíntese , Células Tumorais Cultivadas , Zona Glomerulosa/patologia
15.
Mol Cell Endocrinol ; 408: 220-6, 2015 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-25617716

RESUMO

The current pathological diagnosis of Aldosterone Producing Adenoma (APA) is limited to the description of nodules and/or hyperplasia in the resected adrenal gland, independent of their functional characteristics. The aim of our study was to characterize histopathological markers to confirm the presence and identify the sites of aldosterone production and to discriminate KCNJ5-related APA. We investigated 18 adrenals with APA and 15 with non-functioning adrenal incidentaloma (NFAI) for expression of Disabled-2 and GIRK4, two markers of zona glomerulosa (ZG), and 77 adrenals with APA with known mutational status for GIRK4 expression. Two-thirds of APA and only one NFAI exhibited both GIRK4 and Disabled-2 membrane staining, allowing to correctly classify 79% of adenomas. Remarkably, 28/32 APA with KCNJ5 mutations exhibited lower GIRK4 expression in APA relative to peritumoral ZG. This was highly specific for KCNJ5 mutations, indicating that GIRK4 immunohistochemistry might be used for initial screening of the somatic mutation status.


Assuntos
Adenoma/genética , Adenoma/patologia , Aldosterona/biossíntese , Biomarcadores Tumorais/metabolismo , Canais de Potássio Corretores do Fluxo de Internalização Acoplados a Proteínas G/genética , Mutação/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Neoplasias das Glândulas Suprarrenais/patologia , Adulto , Análise Mutacional de DNA , Diagnóstico Diferencial , Feminino , Canais de Potássio Corretores do Fluxo de Internalização Acoplados a Proteínas G/metabolismo , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Proteínas Supressoras de Tumor/metabolismo , Zona Glomerulosa/patologia
16.
Endocr J ; 61(4): 393-401, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24531914

RESUMO

The majority of the cases diagnosed as primary aldosteronism (PA) are caused by aldosterone-producing adenoma (APA) or idiopathic hyperaldosteronism (IHA). Histopathologically, both IHA and adjacent adrenal glands of APA demonstrate remodeled subcapsular zone (RSZ) but these zones in two disorders are markedly different in terms of steroidogenesis. 3ß-Hydroxysteroid dehydrogenase/Δ5-Δ4 isomerase (3ß-HSD) expression has been known to be activated synergistically by GATA6 and SF1, and repressed by DAX1 through abolishing the activation. Nerve growth factor-induced clone B (NGFIB) is also known as one of the transcription factors to bind to and activate 3ß-HSD promoter. The results of our immunohistochemical analysis demonstrated the expression levels of 3ß-HSD in RSZ of IHA were higher than in RSZ of adjacent adrenals of APA, while those in the zona glomerulosa (ZG) of normal adrenal gland (NA) were in between these two RSZs. The expression levels of GATA6, SF1 and DAX1 did not prominently differ among these three types of adrenals, especially between in RSZs of IHA and APA cases, indicating the marked difference of 3ß-HSD expression was unlikely to be explained by the levels of these three factors. However, the levels of NGFIB expression were significantly higher in RSZ of IHA than in RSZ of adjacent adrenals of APA and the ZG of NA (P<0.05), which may partly account for the expression levels of 3ß-HSD among the three groups of adrenals. These results may imply NGFIB plays important roles in the marked differences in steroidogenic functions in the two distinct types of RSZ of PA cases.


Assuntos
Receptor Nuclear Órfão DAX-1/metabolismo , Fator de Transcrição GATA6/metabolismo , Hiperaldosteronismo/metabolismo , Membro 1 do Grupo A da Subfamília 4 de Receptores Nucleares/metabolismo , Fator Esteroidogênico 1/metabolismo , Regulação para Cima , Zona Glomerulosa/metabolismo , 3-Hidroxiesteroide Desidrogenases/metabolismo , Neoplasias do Córtex Suprarrenal/metabolismo , Neoplasias do Córtex Suprarrenal/patologia , Neoplasias do Córtex Suprarrenal/fisiopatologia , Neoplasias do Córtex Suprarrenal/cirurgia , Adenoma Adrenocortical/metabolismo , Adenoma Adrenocortical/patologia , Adenoma Adrenocortical/fisiopatologia , Adenoma Adrenocortical/cirurgia , Biomarcadores/metabolismo , Regulação para Baixo , Humanos , Hiperaldosteronismo/etiologia , Hiperaldosteronismo/patologia , Hiperaldosteronismo/cirurgia , Imuno-Histoquímica , Zona Glomerulosa/patologia , Zona Glomerulosa/cirurgia
17.
J Clin Endocrinol Metab ; 99(2): E257-62, 2014 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-24423300

RESUMO

CONTEXT: Therapeutic management of primary aldosteronism requires accurate differentiation between aldosterone-producing adenoma (APA) and idiopathic hyperaldosteronism (IHA). However, little is known about the molecular features that delineate the difference between APA and IHA. Two different isoforms of 3ß-hydroxysteroid dehydrogenase (HSD3B1 and HSD3B2) are thought to be expressed in the human adrenal gland, but the lack of isoform-specific antibody has so far hampered mapping of these isoforms in APA and IHA. OBJECTIVES: The aim of our study is to develop and characterize isoform-specific monoclonal antibodies against HSD3B1 and HSD3B2. Using these antibodies, we determined for the first time the immunolocalization of HSD3B1 and HSD3B2 in normal human adrenal cortex as well as in adrenal specimens from APA and IHA. RESULTS: Immunohistochemical analysis with isoform-specific antibodies revealed zone-specific expression of HSD3B1 and HSD3B2 in the adrenal cortex. HSD3B1 immunoreactivities were essentially confined to the zona glomerulosa (ZG), in which aldosterone is produced. In contrast, HSD3B2 was not confined to the ZG but was found across the zona fasciculata, which is where cortisol is produced. Moreover, immunohistopathological analysis of primary aldosteronism revealed a previously uncharacterized difference between APA and IHA. Notably, hyperplasia of ZG seen for IHA was accompanied by a robust expression of ZG isoform HSD3B1. In contrast, tumor cells in APA were not immunopositive to HSD3B1. Rather, a strong and dominant expression of HSD3B2 characterized APA. Moreover, perhaps due to compensatory responses to excess aldosterone, APA had an adjacent ZG whose immunoreactivities to HSD3B1 and HSD3B2 were profoundly reduced. CONCLUSIONS: Isoform-specific monoclonal antibodies against HSD3B1 and HSD3B2 may be of great value for immunohistochemical differentiation between APA and IHA.


Assuntos
3-Hidroxiesteroide Desidrogenases/imunologia , Córtex Suprarrenal/metabolismo , Hiperaldosteronismo/imunologia , Adenoma/metabolismo , Adenoma/patologia , Córtex Suprarrenal/patologia , Neoplasias do Córtex Suprarrenal/metabolismo , Neoplasias do Córtex Suprarrenal/patologia , Anticorpos Monoclonais/metabolismo , Humanos , Hiperaldosteronismo/classificação , Hiperaldosteronismo/metabolismo , Zona Glomerulosa/metabolismo , Zona Glomerulosa/patologia
18.
Endocr Pathol ; 25(3): 229-35, 2014 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-24242700

RESUMO

Metallothionein-3 (MT-3) is an intracellular, low molecular weight protein mainly distributed in the central nervous system but also in various peripheral organs and several types of human neoplasms. However, details of MT-3 expression have not been examined in human adrenal cortex and its disorders. The mRNA levels of MT-3 were first evaluated by quantitative RT-PCR (qPCR) in adrenocortical aldosterone-producing adenoma (APA: 11) and cortisol-producing adenoma (CPA: 14). In addition, MT-3 immunohistochemistry was performed in non-pathological adrenal glands (NA: 19), idiopathic hyperaldosteronism (IHA: 10), APA (20), CPA (24), adjacent non-neoplastic adrenal glands of adenoma (AAG: 20), and adrenocortical carcinoma (ACC: 8). H295R cells were also treated with angiotensin-II or forskolin in a time-dependent manner, and the changes of MT-3 mRNA levels were evaluated by qPCR. Results of qPCR analysis demonstrated that MT-3 mRNA levels were significantly higher in APA than CPA (P = 0.0004). MT-3 immunoreactivity was detected in the zona glomerulosa of NA, IHA, and AAG, as well as in APA, CPA, and ACC. When treated with angiotensin-II and forskolin, MT-3 mRNA levels reached a peak by 12 h in H295R cells, with significantly higher levels compared to control non-treated cells (P < 0.01). The presence of MT-3 in the ZG of NA, IHA, and AAG, as well as APA may imply a role in the pathophysiology of aldosterone-producing tissues.


Assuntos
Adenoma/metabolismo , Neoplasias do Córtex Suprarrenal/metabolismo , Córtex Suprarrenal/metabolismo , Hiperaldosteronismo/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Adenoma/genética , Adenoma/patologia , Córtex Suprarrenal/patologia , Neoplasias do Córtex Suprarrenal/genética , Neoplasias do Córtex Suprarrenal/patologia , Linhagem Celular Tumoral , Humanos , Hiperaldosteronismo/genética , Hiperaldosteronismo/patologia , Proteínas do Tecido Nervoso/genética , Zona Glomerulosa/metabolismo , Zona Glomerulosa/patologia
19.
Endocr J ; 61(1): 35-40, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24077222

RESUMO

The Research Committee of Disorders of Adrenal Hormones, Japan, undertook a nationwide epidemiological study of primary aldosteronism (PA). The present study was undertaken as a part of this study to reveal the relationship between type of treatment and the prognosis of PA. In the primary survey, 4161 patients with PA during the period January 1, 2003-December 31, 2007 were reported from 3252 departments of internal medicine, pediatrics and urology. In the secondary survey, a questionnaire that requested detailed clinical information on individual patients was sent to those departments reporting patients in the primary survey. In total, data on 1706 patients with PA were available in the present study. Among patients with bilateral or unilateral aldosterone-producing adenoma, after adjustment for age at which prognosis was examined, sex, surgical treatment and medical treatment, surgical treatment was significantly associated with amelioration of hypertension (adjusted odds ratio [OR]: 0.47 [95% confidence interval (CI): 0.29-0.77]) and hypokalemia (adjusted OR: 0.17 [95% CI: 0.11-0.29]). No significant relationship was observed between medical treatment and such prognosis in this group of patients. Among patients with bilateral or unilateral adrenal hyperplasia, surgical, but not medical, treatment was significantly associated with amelioration of hypokalemia (adjusted OR: 0.23 [95% CI: 0.06-0.74]), while there was no relationship between surgical or medical treatment and the prognosis of hypertension. In conclusion, surgery offered a better prognosis of PA than medication with regards to hypertension and hypokalemia, with the limitation that a new anti-aldosterone drug, eplerenone, was not available during the study period.


Assuntos
Hiperaldosteronismo/epidemiologia , Adenoma/metabolismo , Aldosterona/biossíntese , Estudos Epidemiológicos , Feminino , Inquéritos Epidemiológicos , Humanos , Hiperaldosteronismo/tratamento farmacológico , Hiperaldosteronismo/cirurgia , Hiperplasia , Hipertensão/complicações , Hipertensão/terapia , Hipopotassemia/complicações , Hipopotassemia/terapia , Japão/epidemiologia , Masculino , Prognóstico , Inquéritos e Questionários , Zona Glomerulosa/patologia
20.
J Clin Invest ; 122(6): 2046-53, 2012 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-22546854

RESUMO

Aldosterone, which plays a central role in the regulation of blood pressure, is produced by zona glomerulosa (ZG) cells of the adrenal gland. When dysregulated, aldosterone is pathogenic and contributes to the development and progression of cardiovascular and renal disease. Although sustained production of aldosterone requires persistent Ca2+ entry through low-voltage activated Ca2+ channels, isolated ZG cells are considered nonexcitable, with recorded membrane voltages that are too hyperpolarized to permit Ca2+ entry. Here, we show that mouse ZG cells within adrenal slices spontaneously generate membrane potential oscillations of low periodicity. This innate electrical excitability of ZG cells provides a platform for the production of a recurrent Ca2+ signal that can be controlled by Ang II and extracellular potassium, the 2 major regulators of aldosterone production. We conclude that native ZG cells are electrical oscillators, and that this behavior provides what we believe to be a new molecular explanation for the control of Ca2+ entry in these steroidogenic cells.


Assuntos
Relógios Biológicos , Canais de Cálcio/metabolismo , Cálcio/metabolismo , Potenciais da Membrana , Zona Glomerulosa/metabolismo , Angiotensina II/metabolismo , Animais , Doenças Cardiovasculares/metabolismo , Doenças Cardiovasculares/patologia , Doenças Cardiovasculares/fisiopatologia , Nefropatias/metabolismo , Nefropatias/patologia , Nefropatias/fisiopatologia , Masculino , Camundongos , Potássio/metabolismo , Zona Glomerulosa/patologia
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