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ABSTRACT: The Cre-loxP system combined with gene targeting strategies has proven to be very useful for gene inactivation in specific tissues and/or cell types. To achieve adrenal cortex specific recombination in vivo, we used a 0.5-kb fragment of the 5'-flanking region of the akr1b7 gene to drive Cre expression in adrenocortical cells. The resulting 0.5 akr1b7-Cre mice express Cre in all steroidogenic zones of the adrenal cortex but not in the gonads. Although recombination of the ROSA26R reporter locus was not observed in all cortical cells, we provide evidence that Cre is expressed in all the cells of the cortex in adult mice. In addition, Cre activity was found in collecting ducts and maturing glomeruli of the kidney. This line is the first to show specific Cre expression in the adrenal cortex in the absence of Cre expression in the gonads. This transgene thus provides a valuable tool for specific gene recombination in the adrenal cortex and kidney.
Molecular and Cellular Endocrinology 12/2008; 300(1-2):197-204. · 4.19 Impact Factor
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ABSTRACT: The Wnt/beta-catenin and cAMP signaling pathways play an important role in adrenal cortex tumorigenesis. Somatic activating mutations of the beta-catenin gene (CTNNB1) are the most frequent genetic defects identified both in adrenocortical adenomas (ACAs) and adrenocortical cancers (ACCs). PRKAR1A mutations leading to cAMP pathway dysregulation are observed in primary pigmented nodular adrenocortical diseases (PPNADs) and some sporadic ACAs.
The objective of the investigation was to study Wnt/beta-catenin dysregulation in adrenocortical tumors (ACTs) with cAMP pathway genetic alteration and search for secondary CTNNB1 somatic mutations in heterogeneous tumors.
Nine PPNADs, including five with macronodules, three ACAs with PRKAR1A somatic mutations, and one heterogeneous tumor with ACC developed within an ACA, were studied by immunohistochemistry and DNA sequencing.
beta-Catenin accumulation was observed in all PPNADs, ACAs with PRKAR1A mutations, and the ACC component of the heterogeneous tumor. CTNNB1 somatic activating mutations were found in the macronodule of two of the five macronodular PPNADs, in one ACA with a PRKAR1A somatic mutation, and in the malignant part of the heterogeneous ACT.
The Wnt/beta-catenin pathway is activated in PPNADs and ACAs with PRKAR1A mutations, suggesting a cross talk between the cAMP and Wnt/beta-catenin pathways in ACT development. In addition, the occurrence as an additional hit of a CTNNB1 somatic mutation is associated with larger or more aggressive ACTs. This underlines the importance of the Wnt/beta-catenin pathway in adrenal cortex tumorigenesis and the importance of genetic accumulation in the progression of ACTs.
Journal of Clinical Endocrinology & Metabolism 10/2008; 93(10):4135-40. · 6.50 Impact Factor
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Rossella Libé,
Amato Fratticci,
Joel Coste,
Frédérique Tissier,
Anelia Horvath, Bruno Ragazzon,
Fernande Rene-Corail,
Lionel Groussin,
Xavier Bertagna,
Marie Laure Raffin-Sanson,
Constantine A Stratakis,
Jérome Bertherat
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ABSTRACT: We have reported previously nonsense inactivating mutations of the phosphodiesterase 11A (PDE11A) gene in patients with micronodular adrenocortical hyperplasia and Cushing syndrome. The aim of this study is to investigate the presence of somatic or germ-line PDE11A mutations in various types of adrenocortical tumors: ACTH-independent macronodular adrenocortical hyperplasia (AIMAH), adrenocortical adenoma (ACA), and adrenocortical cancer (ACC).
PDE11A was sequenced in 117 adrenocortical tumors and 192 controls subjects; immunohistochemistry for PDE11A and tumor cyclic AMP levels were studied in a subgroup of adrenocortical tumors.
One PDE11A inactivating mutation (R307X) was found in one ACA, 22 germ-line missense variants (18.8%) were found in adrenocortical tumors, and only 11 missense variants (5.7%) were found in controls. By comparing the common mutations, a higher frequency of mutations in adrenocortical tumors than in age/sex-matched controls were observed [16% versus 10% in ACC, 19% versus 10% in ACA, and 24% versus 9% in AIMAH; odds ratio (OR), 3.53; P = 0.05]. Somatic DNA from adrenocortical tumors with missense variants showed a wild-type allelic loss. A significant difference between ACC and controls was observed for a polymorphism in exon 6 (E421E; OR, 2.1; P = 0.03) and three associated polymorphisms located in intron 10-exon 11-intron 11 (OR, 0.5; P = 0.01). In AIMAH/ACA, cyclic AMP levels were higher than in normal adrenals and decreased PDE11A immunostaining was present in adrenocortical tumors with PDE11A variants.
The present investigation of a large cohort of adrenocortical tumors suggests that PDE11A sequence defects predispose to a variety of lesions (beyond micronodular adrenocortical hyperplasia) and may contribute to the development of these tumors in the general population.
Clinical Cancer Research 07/2008; 14(12):4016-24. · 7.74 Impact Factor
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ABSTRACT: We established the first adrenocortical tumor cell lines with complete zona fasciculata (ZF) cell phenotype from tumors induced in transgenic mice by large T-antigen of simian virus 40 under the control of the aldose reductase-like akr1b7 gene promoter. Adrenocortical tumor cell lines produced high amounts of corticosterone and were responsive to ACTH. All genes that are supportive for glucocorticoid synthesis including cyp21a1 and cyp11b1 were expressed, and most of them were transiently up-regulated by ACTH at transcriptional level: stimulation culminated after 3-6 h and returned to basal levels after 24 h. Taking advantage of these cells, we have examined the effect of ACTH on DAX-1 (dosage-sensitive sex reversal-adrenal hypoplasia congenita critical region on X-chromosome, gene 1) and SF-1 (steroidogenic factor 1), two transcription factors known to respectively repress and activate adrenocortical steroidogenesis by acting on common target genes. According to their antagonistic activities, DAX-1 mRNA and protein levels were transiently down-regulated by ACTH, whereas those of SF-1 were stimulated, with kinetics paralleling those of steroidogenic genes expression, notably of two known SF-1 target genes, star and akr1b7. This suggests an essential role of SF-1/DAX-1 proteins ratio to achieve proper ACTH control of steroidogenic gene expression in cells derived from ZF. This was confirmed in mice adrenals, where repression of dax-1 gene and concomitant up-regulation of sf-1, star, and akr1b7 genes were observed in response to ACTH stimulation. In conclusion, using both unique differentiated cell lines and in vivo approaches, we provide the first evidence that hormonally induced changes in SF-1/DAX-1 ratio are part of the molecular arsenal of ZF cells to fine tune ACTH responsiveness.
Endocrinology 05/2006; 147(4):1805-18. · 4.46 Impact Factor
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ABSTRACT: Primary Pigmented Nodular Adrenocortical Disease (PPNAD) is a rare primary bilateral adrenal defect causing corticotropin-independent Cushing's syndrome. It occurs mainly in children and young adults. Macroscopic appearance of the adrenals is characteristic with small pigmented micronodules observed in the cortex. PPNAD is most often diagnosed in patients with Carney complex (CNC), but it can also be observed in patients without other manifestations or familial history (isolated PPNAD). The CNC is an autosomal dominant multiple neoplasia syndrome characterized by the association of myxoma, spotty skin pigmentation and endocrine overactivity. One of the putative CNC genes has been identified as the gene of the regulatory R1A subunit of protein kinase A (PRKAR1A), located at 17q22-24. Germline heterozygous inactivating mutations of PRKAR1A have been reported in about 45% of patients with CNC, and up to 80% of CNC patients with Cushing's syndrome due to PPNAD. Interestingly, such inactivating germline PRKAR1A mutations have also been found in patients with isolated PPNAD. The hot spot PRKAR1A mutation termed c.709[-7-2]del6 predisposes mostly to isolated PPNAD, and is the first clear genotype/phenotype correlation described for this gene. Somatic inactivating mutations of PRKAR1A have been observed in macronodules of PPNAD and in sporadic cortisol secreting adrenal adenomas. Isolated PPNAD is a genetic heterogenous disease, and recently inactivating mutations of the gene of the phosphodiesterase 11A4 (PDE11A4) located at 2q31-2q35 have been identified in patients without PRKAR1A mutations. Interestingly, both PRKAR1A and PDE11A gene products control the cAMP signaling pathway, which can be altered at various levels in endocrine tumors.
Pituitary 02/2006; 9(3):211-9. · 1.83 Impact Factor
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ABSTRACT: The akr1-b7 gene encodes a scavenger enzyme expressed in steroidogenic glands under pituitary control. In the zona fasciculata of the adrenal cortex where its expression is controlled by ACTH, AKR1-B7 detoxifies isocaproaldehyde produced during the first step of steroidogenesis. Three steroidogenic factor-1 (SF-1)-responsive elements (SFREs) are contained within the -510/+41 promoter region, which was previously demonstrated to drive gene expression in transgenic mice adrenal cortex. All these sequences bind at least SF-1 in Y1 adrenocortical cell nuclear extracts and can be activated by overexpression of this factor in HeLa cells. However, the three SFREs show distinct properties regarding akr1-b7 promoter activity in Y1 cells. Whereas the proximal -102 SFRE supports basal promoter activity, the -458 bona fide SFRE is essential for both basal promoter activity and cAMP responsiveness, although it is unresponsive to cAMP when isolated from its promoter context. This suggests that SF-1 is not a cAMP-responsive factor per se. The neighboring SFRE at -503 is a palindromic sequence that binds monomeric and heteromeric SF-1 as well as an adrenal-specific complex. Using MA-10 Leydig cells and Y1-10r9 mutant cells, we provide evidence that its activity in adrenocortical cells depends on the binding of the adrenal-specific factor, which is required for basal and cAMP-induced promoter activity. Furthermore, the -503 site has intrinsic cAMP-sensing ability in Y1 cells, which is correlated with increased adrenal-specific complex binding. Collectively, our results suggest that cAMP responsiveness of the akr1-b7 promoter is achieved through cooperation between the adrenal-specific factor bound to the -503 site and SF-1 bound to the -458 site.
Endocrinology 03/2004; 145(2):508-18. · 4.46 Impact Factor
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