ACTH- and Cytochalasin-Related Changes in Adrenal Cell Morphology and Cytoskeleton

Following 1 hr incubation with ACTH, cytochalasin D or ACTH/cytochalasin, detergent-solubilized mouse adrenal tumor cells cytoskeletal changes were examined using scanning and transmission electron microscopy. Steroid production was also examined

G-protein-coupled receptors in aldosterone-producing adenomas: a potential cause of hyperaldosteronism

The source of aldosterone in 30–40% of patients with primary hyperaldosteronism (PA) is unilateral aldosterone-producing adenoma (APA). The mechanisms causing elevated aldosterone production in APA are unknown. Herein, we examined the expression of G-protein-coupled receptors (GPCRs) in APA and demonstrated that when compared with normal adrenals, there is a general elevation of certain GPCR in many APA and/or ectopic expression of GPCR in others. RNA samples from normal adrenals (n = 5), APAs (n = 10), and cortisol-producing adenomas (CPAs; n = 13) were used on 15 genomic expression arrays, each of which included 223 GPCR transcripts presented in at least 1 out of 15 of the independent microarrays. The array results were confirmed using real-time RT-PCR (qPCR). Four GPCR transcripts exhibited a statistically significant increase that was greater than threefold when compared with normal adrenals, suggesting a general increase in expression when compared with normal adrenal glands. Four GPCR transcripts exhibited a > 15-fold increase of expression in one or more of the APA samples when compared with normal adrenals. qPCR analysis confirmed array data and found the receptors with the highest fold increase in APA expression to be LH receptor, serotonin receptor 4, GnRH receptor, glutamate receptor metabotropic 3, endothelin receptor type B-like protein, and ACTH receptor. There are also sporadic increased expressions of these genes in the CPAs. Together, these findings suggest a potential role of altered GPCR expression in many cases of PA and provide candidate GPCR for further study

Discordance between imaging and immunohistochemistry in unilateral primary aldosteronism

Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/139975/1/cen13442.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/139975/2/cen13442_am.pd

Single-Center Prospective Cohort Study on the Histopathology, Genotype, and Postsurgical Outcomes of Patients With Primary Aldosteronism

Unilateral forms of primary aldosteronism are usually surgically treated to remove the source of aldosterone excess. After adrenalectomy, aldosteronism persists in some patients indicating abnormal aldosterone production from the unresected gland. Our objective was to investigate histopathology, genotype, and postsurgical outcomes in a 3-year prospective cohort of surgically treated patients for primary aldosteronism (from 2016 to 2018). The cohort comprised 60 consecutively operated patients categorized with classical or nonclassical histopathologic findings of unilateral primary aldosteronism. In the classical group were 45 solitary aldosterone-producing adenomas or dominant aldosterone-producing nodules; in the nonclassical group were 15 cases of multiple aldosterone-producing micronodules or nodules (12 cases) or aldosterone-producing diffuse hyperplasia (3 cases). The classical group displayed higher baseline plasma aldosterone concentrations (262 versus 155 pg/mL, P=0.008) and an increased aldosterone-to-renin ratio (81 versus 42, P=0.002). A high proportion of the classical group achieved complete biochemical success (97.6% versus 66.7% in the nonclassical group, P=0.002). The nonclassical versus classical group displayed an increased ratio of absolute aldosterone concentration in the contralateral adrenal vein to peripheral vein at adrenal venous sampling (3.8 versus 2.0, P=0.004). Variants in aldosterone-driver genes were identified in 85% of 41 aldosterone-producing adenomas and were excluded in the remaining 15% by CYP11B2 guided next-generation sequencing. There were no differences in clinical or biochemical outcomes in patients with a solitary aldosterone-producing adenoma categorized by KCNJ5 mutation status. In conclusion, adrenals with a nonclassical histopathology of unilateral primary aldosteronism are associated with a higher incidence of postsurgical disease persistence and increased aldosterone production from the unresected adrenal

PPARG dysregulation as a potential molecular target in adrenal Cushing's syndrome

BACKGROUND We performed a transcriptomic analysis of adrenal signaling pathways in various forms of endogenous Cushing's syndrome (CS) to define areas of dysregulated and druggable targets. METHODOLOGY Next-generation sequencing was performed on adrenal samples of patients with primary bilateral macronodular adrenal hyperplasia (PBMAH, n=10) and control adrenal samples (n=8). The validation groups included cortisol-producing adenoma (CPA, n=9) and samples from patients undergoing bilateral adrenalectomy for Cushing's disease (BADX-CD, n=8). In vivo findings were further characterized using three adrenocortical cell-lines (NCI-H295R, CU-ACC2, MUC1). RESULTS Pathway mapping based on significant expression patterns identified PPARG (peroxisome proliferator-activated receptor gamma) pathway as the top hit. Quantitative PCR (QPCR) confirmed that PPARG (l2fc<-1.5) and related genes - FABP4 (l2fc<-5.5), PLIN1 (l2fc<-4.1) and ADIPOQ (l2fc<-3.3) - were significantly downregulated (p<0.005) in PBMAH. Significant downregulation of PPARG was also found in BADX-CD (l2fc<-1.9, p<0.0001) and CPA (l2fc<-1.4, p<0.0001). In vitro studies demonstrated that the PPARG activator rosiglitazone resulted in decreased cell viability in MUC1 and NCI-H295R (p<0.0001). There was also a significant reduction in the production of aldosterone, cortisol, and cortisone in NCI-H295R and in Dihydrotestosterone (DHT) in MUC1 (p<0.05), respectively. OUTCOME This therapeutic effect was independent of the actions of ACTH, postulating a promising application of PPARG activation in endogenous hypercortisolism

Evaluation Research and Institutional Pressures: Challenges in Public-Nonprofit Contracting

This article examines the connection between program evaluation research and decision-making by public managers. Drawing on neo-institutional theory, a framework is presented for diagnosing the pressures and conditions that lead alternatively toward or away the rational use of evaluation research. Three cases of public-nonprofit contracting for the delivery of major programs are presented to clarify the way coercive, mimetic, and normative pressures interfere with a sound connection being made between research and implementation. The article concludes by considering how public managers can respond to the isomorphic pressures in their environment that make it hard to act on data relating to program performance.This publication is Hauser Center Working Paper No. 23. The Hauser Center Working Paper Series was launched during the summer of 2000. The Series enables the Hauser Center to share with a broad audience important works-in-progress written by Hauser Center scholars and researchers

International Histopathology Consensus for Unilateral Primary Aldosteronism

Objective: Develop a consensus for the nomenclature and definition of adrenal histopathologic features in unilateral primary aldosteronism (PA). Context: Unilateral PA is the most common surgically treated form of hypertension. Morphologic examination combined with CYP11B2 (aldosterone synthase) immunostaining reveals diverse histopathologic features of lesions in the resected adrenals. Patients and methods: Surgically removed adrenals (n = 37) from 90 patients operated from 2015 to 2018 in Munich, Germany, were selected to represent the broad histologic spectrum of unilateral PA. Five pathologists (Group 1 from Germany, Italy, and Japan) evaluated the histopathology of hematoxylin-eosin (HE) and CYP11B2 immunostained sections, and a consensus was established to define the identifiable features. The consensus was subsequently used by 6 additional pathologists (Group 2 from Australia, Brazil, Canada, Japan, United Kingdom, United States) for the assessment of all adrenals with disagreement for histopathologic diagnoses among group 1 pathologists. Results: Consensus was achieved to define histopathologic features associated with PA. Use of CYP11B2 immunostaining resulted in a change of the original HE morphology-driven diagnosis in 5 (14%) of 37 cases. Using the consensus criteria, group 2 pathologists agreed for the evaluation of 11 of the 12 cases of disagreement among group 1 pathologists. Conclusion: The HISTALDO (histopathology of primary aldosteronism) consensus is useful to standardize nomenclature and achieve consistency among pathologists for the histopathologic diagnosis of unilateral PA. CYP11B2 immunohistochemistry should be incorporated into the routine clinical diagnostic workup to localize the likely source of aldosterone production

The farnesoid X receptor regulates transcription of 3 beta-hydroxysteroid dehydrogenase type 2 in human adrenal cells

Recent studies have shown that the adrenal cortex expresses high levels of farnesoid X receptor (FXR), but its function remains not known. Herein, using microarray technology, we tried to identify candidate FXR targeting genes in the adrenal glands, and showed that FXR regulates 3β-hydroxysteroid dehydrogenase type 2 (HSD3B2) expression in human adrenocortical cells. We further demonstrated that FXR stimulated HSD3B2 promoter activity and have defined the cis-element responsible for FXR regulation of HSD3B2 transcription. Transfection of H295R adrenocortical cells with FXR expression vector effectively increased FXR expression levels and additional treatment with chenodeoxycholic acid (CDCA) caused a 25-fold increase in the mRNA for organic solute transporter alpha (OSTα), a known FXR target gene. HSD3B2 mRNA levels also increased following CDCA treatment in a concentration-dependent manner. Cells transfected with a HSD3B2 promoter construct and FXR expression vector responded to CDCA with a 20-fold increase in reporter activity compared to control. Analysis of constructs containing sequential deletions of the HSD3B2 promoter suggested a putative regulatory element between -166 and -101. Mutation of an inverted repeat between -137 and -124 completely blocked CDCA/FXR induced reporter activity. Chromatin immunoprecipitation assays further confirmed the presence of a FXR response element in the HSD3B2 promoter. In view of the emerging role of FXR agonists as therapeutic treatment of diabetes and certain liver diseases, the effects of such agonists on other FXR expressing tissues should be considered. Our findings suggest that in human adrenal cells, FXR increases transcription and expression of HSD3B2. Alterations in this enzyme would influence the capacity of the adrenal gland to produce corticosteroids

Genetic Architecture of Soybean Yield and Agronomic Traits

Soybean is the world’s leading source of vegetable protein and demand for its seed continues to grow. Breeders have successfully increased soybean yield, but the genetic architecture of yield and key agronomic traits is poorly understood. We developed a 40-mating soybean nested association mapping (NAM) population of 5,600 inbred lines that were characterized by single nucleotide polymorphism (SNP) markers and six agronomic traits in field trials in 22 environments. Analysis of the yield, agronomic, and SNP data revealed 23 significant marker-trait associations for yield, 19 for maturity, 15 for plant height, 17 for plant lodging, and 29 for seed mass. A higher frequency of estimated positive yield alleles was evident from elite founder parents than from exotic founders, although unique desirable alleles from the exotic group were identified, demonstrating the value of expanding the genetic base of US soybean breeding

PPARG dysregulation as a potential molecular target in adrenal Cushing's syndrome

BackgroundWe performed a transcriptomic analysis of adrenal signaling pathways in various forms of endogenous Cushing’s syndrome (CS) to define areas of dysregulated and druggable targets.MethodologyNext-generation sequencing was performed on adrenal samples of patients with primary bilateral macronodular adrenal hyperplasia (PBMAH, n=10) and control adrenal samples (n=8). The validation groups included cortisol-producing adenoma (CPA, n=9) and samples from patients undergoing bilateral adrenalectomy for Cushing’s disease (BADX-CD, n=8). In vivo findings were further characterized using three adrenocortical cell-lines (NCI-H295R, CU-ACC2, MUC1).ResultsPathway mapping based on significant expression patterns identified PPARG (peroxisome proliferator-activated receptor gamma) pathway as the top hit. Quantitative PCR (QPCR) confirmed that PPARG (l2fc&lt;-1.5) and related genes – FABP4 (l2fc&lt;-5.5), PLIN1 (l2fc&lt;-4.1) and ADIPOQ (l2fc&lt;-3.3) – were significantly downregulated (p&lt;0.005) in PBMAH. Significant downregulation of PPARG was also found in BADX-CD (l2fc&lt;-1.9, p&lt;0.0001) and CPA (l2fc&lt;-1.4, p&lt;0.0001). In vitro studies demonstrated that the PPARG activator rosiglitazone resulted in decreased cell viability in MUC1 and NCI-H295R (p&lt;0.0001). There was also a significant reduction in the production of aldosterone, cortisol, and cortisone in NCI-H295R and in Dihydrotestosterone (DHT) in MUC1 (p&lt;0.05), respectively.OutcomeThis therapeutic effect was independent of the actions of ACTH, postulating a promising application of PPARG activation in endogenous hypercortisolism