Normal Adrenal Glands Research Articles

Functional Characteristic and Significance of Aldosterone-Producing Cell Clusters in Primary Aldosteronism and Age-Related Hypertension.

Primary aldosteronism (PA) is one of the most frequent curable forms of secondary hypertension. It can be caused by the overproduction of aldosterone in one or both adrenal glands. The most common subtypes of PA are unilateral aldosterone over-production due to aldosterone-producing adenomas (APA) or bilateral aldosterone over-production due to bilateral hyperaldosteronism (BHA). Utilizing the immunohistochemical (IHC) detection of aldosterone synthase (CYP11B2) has allowed the identification of aldosterone-producing cell clusters (APCCs) with unique focal localization positive for CYP11B2 expression in the subcapsular portion of the human adult adrenal cortex. The presence of CYP11B2 supports that synthesis of aldosterone can occur in these cell clusters and therefore might contribute to hyperaldosteronism. However, the significance of the steroidogenic properties of APCCs especially in regards to PA remains unclear. Herein, we review the available evidence on the presence of APCCs in normal adrenals and adrenal tissues adjacent to APAs, their aldosterone-stimulating somatic gene mutations, and their accumulation during the ageing process; raising the possibility that APCCs may play a role in the development of PA and age-related hypertension.

Sum of High-Risk Gene Mutation (SHGM): A Novel Attempt to Assist Differential Diagnosis for Adrenocortical Carcinoma with Benign Adenoma, Based on Detection of Mutations of Nine Target Genes

There has been no research on applying gene detection to differential diagnosis of adrenocortical carcinoma (ACC). We attempted to explore a novel auxiliary method for differential diagnosis between ACC with benign adrenocortical adenoma (ACA), based on mutations of target genes in tissues. Nine genes were chosen as target genes, including TP53, CTNNB1, ARMC5, PRKAR1A, ZNRF3, RB1, APC, MEN1, and RPL22. Exons sequencing of target genes were performed in 98 cases of tissue samples by FastTarget technology, including 41 ACC tissues, 32 ACA tissues, and 25 normal adrenal gland tissues. Significant mutations were detected and identified, and the clinical information was collected, for further comparative analysis and application to assist differential diagnosis of ACC. We identified 132 significant gene mutations and 227 significant mutation sites in 37 ACC tissues, much more than ACA and normal adrenal gland tissues. Mutation rates of 6 genes in ACC tissues were obviously higher than ACA tissues, including ZNRF3, ARMC5, TP53, APC, RB1, and PRKAR1A, regarded as high-risk genes. The sum of mutated high-risk genes detected in each sample was denominated sum of high-risk gene mutation (SHGM), and the rates of SHGM > 0 and SHGM > 1 in ACC tissues were 73.0% and 62.2%, respectively, both obviously higher than those in ACA tissues, with significant statistic differences. Especially for 8 cases of ACC with diameter < 5 cm, SHGM > 0 and SHGM > 1 were found in 6 samples (75%) and 4 samples (50%), respectively. However, no relevance was found between SHGM and clinical characteristics of ACC. We identified 6 high-risk genes in ACC tissues, with significantly higher mutation rates than ACA or normal adrenal gland tissues. The sum of mutated high-risk genes detected in ACC tissues was denominated SHGM, which was potential to assist the differential diagnosis of ACC with ACA, especially for the small-size ACC.

Association of high-risk neuroblastoma classification based on expression profiles with differentiation and metabolism.

Neuroblastoma, the most common extracranial solid malignancy among children, originates from undifferentiated neural crest cells (NCC). Despite recent intensified treatment, high-risk patients still have a high mortality rate. To explore a new therapeutic strategy, we performed an integrated genomic and transcriptomic analysis of 30 high-risk neuroblastoma cases. Based on the expression profiling of RNA sequencing, neuroblastoma was classified into Mesenchymal (MES; n = 5) and Noradrenergic (ADRN; n = 25) clusters, as previously reported in the super-enhancer landscape. The expression patterns in MES-cluster cases were similar to normal adrenal glands, with enrichment in secretion-related pathways, suggesting chromaffin cell-like features built from NCC-derived Schwann cell precursors (SCPs). In contrast, neuron-related pathways were enriched in the ADRN-cluster, indicating sympathoblast features reported to originate from NCC but not via SCPs. Thus, MES- and ADRN-clusters were assumed to be corresponding to differentiation pathways through SCP and sympathoblast, respectively. ADRN-cluster cases were further classified into MYCN- and ATRX-clusters, characterized by genetic alterations, MYCN amplifications and ATRX alterations, respectively. MYCN-cluster cases showed high expression of ALDH18A1, encoding P5CS related to proline production. As reported in other cancers, this might cause reprogramming of proline metabolism leading to tumor specific proline vulnerability candidate for a target therapy of metabolic pathway. In ATRX-cluster, SLC18A2 (VMAT2), an enzyme known to prevent cell toxicity due to the oxidation of dopamine, was highly expressed and VMAT2 inhibitor (GZ-793A) represented significant attenuation of cell growth in NB-69 cell line (high SLC18A2 expression, no MYCN amplification) but not in IMR-32 cell line (MYCN amplification). In addition, the correlation of VMAT2 expression with metaiodobenzylguanidine (MIBG) avidity suggested a combination of VMAT2 inhibitor and MIBG radiation for a novel potential therapeutic strategy in ATRX-cluster cases. Thus, targeting the characteristics of unique neuroblastomas may prospectively improve prognosis.

Dielectric Characterization of Ex Vivo Ovine and Human Adrenal Glands for Microwave Thermal Ablation Applications

Historically, adrenal glands diseases causing hypertension, such as Primary Aldosteronism (PA), have been treated through pharmacotherapy or surgical resection. Given the shortcomings of the available treatment options, the interest in alternative and less invasive treatment modalities such as microwave ablation (MWA), has increased. In order to develop and optimize this novel electromagnetic-based therapy, an accurate knowledge of the dielectric properties of human adrenal glands, as well as preclinical animal models, is crucial. In particular, ovine models represent a feasible animal model to test the safety and performances of MWA. In this study, the dielectric properties of ovine adrenal glands and of normal and diseased human adrenal glands are characterized <italic xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink">ex vivo</i> in the microwave frequency range. The dielectric properties of the two functional tissues (cortex and medulla) composing ovine adrenal glands are measured using the open-ended coaxial probe technique and represented with a two pole Cole-Cole model in the frequency range from 0.5 GHz to 8 GHz. This paper presents the first dielectric data of normal and diseased human adrenal tissues, including a functioning adenoma responsible for PA and it compares the human data with data from the animal model.

Feline abdominal ultrasonography: what's normal? what's abnormal? The adrenal glands.

Abdominal ultrasound plays a vital role in the diagnostic work-up of many cats presenting to general and specialist practitioners. Ultrasound examination of the adrenal glands can provide important information pertaining to several conditions including hyperaldosteronism and hyperadrenocorticism. Despite ultrasonography being a commonly used modality, many practitioners are not comfortable performing an ultrasound examination or interpreting the resulting images. Even for the experienced ultrasonographer, differentiating between incidental findings, such as adrenal mineralisation, and clinically significant pathological changes can be challenging. This review, part of an occasional series on feline abdominal ultrasonography, discusses the ultrasonographic examination of the normal and diseased adrenal glands. Aimed at general practitioners who wish to improve their knowledge of and confidence in feline abdominal ultrasound, this review is accompanied by high-resolution images and videos available online as supplementary material. Ultrasound facilities are readily available to most practitioners, although the use of ultrasonography as a diagnostic tool is highly dependent on operator experience. Information provided in this article is drawn from the published literature and the author's own clinical experience.

Evaluation of Normal Adrenal Gland Volume and Morphometry and Relationship with Waist Circumference in an Adult Population Using Multidetector Computed Tomography.

Objectives:This study aims to determine the normal range of values of the right, left, and total volume and shape of the adrenal gland (AG) and to evaluate its relationship with gender, age, height, weight, body mass index (BMI), and waist circumference (WC) in multidetector computed tomography (MDCT) images.Methods:The study included 115 MDCT scans, of which 56 were men and 59 were women. For volume measurement, the outlines of the AG were drawn semi-automatically for all patients. Then, collecting the area in each slice, the volumes were automatically measured. The intraclass correlation coefficient (ICC) test was used to analyze intraobserver reliability for repeated measurements with a 95% confidence interval. Participant’s age, gender, weight, height, BMI, and WC were obtained. p<0.05 was considered statistically significant.Results:The mean age of participants was 49.5±17.7 (19–81). The average right AGV (RAGV), left AGV (LAGV), and total AGV were 3.47±1.33, 4.77±1.33, and 8.25±2.74, respectively. The ICC values for all measurements were >0.80–0.90, indicating good and excellent agreement. LAGV was measured as higher than the RAGV. A positive moderate correlation of the AGVs with BMI and WC was observed.Conclusion:The increase in BMI and WC, which are indicators of obesity, correlates with the increase in AGV, we think that the findings will be valuable in evaluating the pathophysiology of the hypothalamic-pituitary-adrenal axis.

Pathology of Aldosterone Biosynthesis and its Action.

Aldosterone plays pivotal roles in renin-angiotensin-aldosterone system in order to maintain the equilibrium of liquid volume and electrolyte metabolism. Aldosterone action is mediated by both mineralocorticoid receptor and 11β-hydroxysteroid dehydrogenase type 2 (11β-HSD2). Its excessive actions directly induced tissue injuries in its target organs such as myocardial and vascular fibrosis in addition to chronic kidney diseases. Excessive aldosterone actions were also reported to be involved in unbalanced electrolyte metabolism in inflammatory bowel disease and development of pulmonary diseases. Hyperaldosteronism is tentatively classified into primary and secondary types. Primary aldosteronism is more frequent and has been well known to result in secondary hypertension with subsequent cardiovascular damages. Primary aldosteronism is also further classified into distinctive subtypes and among those, aldosterone-producing adenoma is the most frequent one accounting for the great majority of unilateral primary aldosteronism cases. In bilateral hyperaldosteronism, aldosterone-producing diffuse hyperplasia and aldosterone-producing micronodules or nodules are the major subtypes. All these aldosterone-producing lesions were reported to harbor somatic mutations including KCNJ5, CACNA1D, ATP1A1 and ATP2B3, which were all related to excessive aldosterone production. Among those mutations above, somatic mutation of KCNJ5 is the most frequent in aldosterone-producing adenoma and mostly composed of clear cells harboring abundant aldosterone synthase expression. In contrast, CACNA1D-mutated aldosterone-producing micronodules or aldosterone-producing nodules were frequently detected not only in primary aldosteronism patients but also in the zona glomerulosa of normal adrenal glands, which could eventually lead to an autonomous aldosterone production resulting in normotensive or overt primary aldosteronism, but their details have remained unknown.

Ultrasonographic evaluation of adrenal gland size in two body weight categories of healthy adult cats.

Adrenal gland size and its association with body weight have been rarely evaluated in cats. This study was undertaken to assess the association between feline body weight and adrenal gland thickness, and to propose reference intervals (RIs) for adrenal gland thickness in healthy cats. This was a cross-sectional study in which 39 healthy cats were included. The cats were divided into two weight categories, classified as ⩽4.0 kg and >4-8 kg of ideal body weight (with 13 and 26 cats in each group, respectively), which took into consideration the body condition score of the cats. All cats underwent an ultrasound examination that was taken from a subcostal position. Maximum dorsoventral thicknesses of the left (MTL) and right (MTR) adrenal glands were measured in a sagittal plane. RIs were obtained for the maximum thickness (MT), which included the MTLs and MTRs of each cat. RIs with the 90% confidence intervals were calculated according to American Society for Veterinary Clinical Pathology guidelines on RIs. No statistical differences for adrenal gland thickness were observed between the left and right (P = 0.543) adrenal glands or between male and female cats (P = 0.943). Mean MT was significantly greater in the group of cats weighing >4-8 kg compared with the group of cats weighing ⩽4 kg (3.7 ± 0.6 vs 3.2 ± 0.4 mm; P <0.005). The lower limit of the RI for MT was 2.4 mm (range 2.2-2.6 mm) in the group weighing ⩽4 kg and 2.6 mm (range 2.4-2.8 mm) in the group weighing >4-8 kg. The upper limit of the RI for MT was 3.9 mm (range 3.7-4.1 mm) in the group of cats weighing ⩽4 kg and 4.8 mm (range 4.6-5.1 mm) in the group of cats weighing >4-8 kg. The use of RIs based on two group sizes allows for a more accurate ultrasonographic evaluation of adrenal gland thickness in cats. The maximum normal adrenal gland thickness is lower in smaller cats (3.9 mm for those weighing ⩽4 kg and 4.8 mm for those weighing >4-8 kg).

Impact of the Chemokine Receptors CXCR4 and CXCR7 on Clinical Outcome in Adrenocortical Carcinoma.

Chemokine receptors have a negative impact on tumor progression in several human cancers and have therefore been of interest for molecular imaging and targeted therapy. However, their clinical and prognostic significance in adrenocortical carcinoma (ACC) is unknown. The aim of this study was to evaluate the chemokine receptor profile in ACC and to analyse its association with clinicopathological characteristics and clinical outcome. A chemokine receptor profile was initially evaluated by quantitative PCR in 4 normal adrenals, 18 ACC samples and human ACC cell line NCI-H295. High expression of CXCR4 and CXCR7 in both healthy and malignant adrenal tissue and ACC cells was confirmed. In the next step, we analyzed the expression and cellular localization of CXCR4 and CXCR7 in ACC by immunohistochemistry in 187 and 84 samples, respectively. These results were correlated with clinicopathological parameters and survival outcome. We detected strong membrane expression of CXCR4 and CXCR7 in 50% of ACC samples. Strong cytoplasmic CXCR4 staining was more frequent among samples derived from metastases compared to primaries (p=0.01) and local recurrences (p=0.04). CXCR4 membrane staining positively correlated with proliferation index Ki67 (r=0.17, p=0.028). CXCR7 membrane staining negatively correlated with Ki67 (r=−0.254, p=0.03) but positively with tumor size (r=0.3, p=0.02). No differences in progression-free or overall survival were observed between patients with strong and weak staining intensities for CXCR4 or CXCR7. Taken together, high expression of CXCR4 and CXCR7 in both local tumors and metastases suggests that some ACC patients might benefit from CXCR4/CXCR7-targeted therapy.

Adrenal Insufficiency due to Total Primary Empty Sella Syndrome

A 64-year-old woman was transported suffering from persistent lower abdominal pain, vomiting, and low-grade fever. Magnetic resonance imaging revealed an empty sella (ES) and hormone tests revealed a disappearance of diurnal variation of cortisol, low cortisol and adrenocorticotropic hormone (ACTH) secretion especially in the morning, and poor ACTH-cortisol axis reaction, as well as normal hypothalamus-pituitary gland-thyroid or adrenal gland axis hormone reaction. The cause of ES remained unclear; however, based on a diagnosis as adrenal insufficiency due to inappropriate ACTH secretion caused by total primary ES syndrome, we started hydrocortisone (15 mg/day). Afterwards, she immediately became symptom-free and was discharged. J Endocrinol Metab. 2020;10(5):144-153 doi: https://doi.org/10.14740/jem686

Adropin Stimulates Proliferation and Inhibits Adrenocortical Steroidogenesis in the Human Adrenal Carcinoma (HAC15) Cell Line.

Adropin is a multifunctional peptide hormone encoded by the ENHO (energy homeostasis associated) gene. It plays a role in mechanisms related to increased adiposity, insulin resistance, as well as glucose, and lipid metabolism. The low adropin levels are strongly associated with obesity independent insulin resistance. On the other hand, overexpression or exogenous administration of adropin improves glucose homeostasis. The multidirectional, adropin-related effects associated with the regulation of metabolism in humans also appear to be attributable to the effects of this peptide on the activity of various elements of the endocrine system including adrenal cortex. Therefore, the main purpose of the present study was to investigate the effect of adropin on proliferation and secretory activity in the human HAC15 adrenal carcinoma cell line. In this study, we obtained several highly interesting findings. First, GPR19, the main candidate sensitizer of adrenocortical cells to adropin, was expressed in HAC15 cells. Moreover, GPR19 expression was relatively stable and not regulated by ACTH, forskolin, or adropin itself. Our findings also suggest that adropin has the capacity to decrease expression levels of steroidogenic genes such as steroidogenic acute regulatory protein (StAR) and CYP11A1, which then led to a statistically significant inhibition in cortisol and aldosterone biosynthesis and secretion. Based on whole transcriptome study and research involving transforming growth factor (TGF)-β type I receptor kinase inhibitor we demonstrated that attenuation of steroidogenesis caused by adropin is mediated by the TGF-β signaling pathway likely to act through transactivation mechanism. We found that HAC15 cells treated with adropin presented significantly higher proliferation levels than untreated cells. Using specific intracellular inhibitors, we showed that adropin stimulate proliferation via ERK1/2 and AKT dependent signaling pathways. We have also demonstrated that expression of GPR19 is elevated in adrenocortical carcinoma in relation to normal adrenal glands. High level of GPR19 expression in adrenocortical carcinoma may constitute a negative prognostic factor of disease progression.

CME Sonography 94/Answers: Adrenal Glands

CME Sonography 94/Answers: Adrenal Glands Abstract. Adrenal glands are often not examined in abdominal ultrasound because it is thought that the adrenal glands cannot be visualized. The normal right adrenal gland can be represented in most cases and the left one very often. Sonography makes an important contribution to the diagnosis of adrenal diseases. In addition to bleeding and tuberculosis, there are benign tumors (adrenal adenomas), pheochromocytomas, and adrenal carcinomas. There are also adrenal metastases and lymphomas.

Ultrasonography of Normal Adrenal Glands in Adult Holstein-Friesian Cows: A Pilot Study.

Simple SummaryKnowledge of the normal appearance of the abdominal organs of animals is essential for accurate ultrasound evaluation in daily clinical routine in veterinary practice. In this paper we describe, for the first time, a systematic protocol for ultrasonographic examination of the adrenal glands of cows and provide preliminary reference values for healthy adult Holstein–Friesian cows.Ultrasonographic reference values for the adrenal glands of cattle have not been reported to date. Adrenal glands can be affected by different pathologies, such as hyperplasia, neoplasia and atrophy (either primary or secondary). The present findings indicate that the right adrenal gland can be easily characterized by transabdominal ultrasound in adult Holstein–Friesian cows, with no apparent influence of age or weight. The right adrenal gland (mean length 3.86 ± 1.39 cm; and mean thickness 1.39 ± 0.26 cm) was consistently and mainly located in the 12th intercostal space. The left adrenal gland was more difficult to locate due to its more medial position, and to the presence of gas in the gastrointestinal tract, so it could not be visualized in most animals (18/25). Its mean length was 3.72 ± 0.95 cm, and mean thickness was 1.36 ± 0.33 cm, in the sagittal section. This is the first report of the ultrasonographic appearance of the adrenal glands of cows and of the corresponding reference preliminary values.

Isolated Paraganglioma in a Patient with VHL P.L163F Mutation

Paragangliomas (PGLs) are one of the many neoplasms associated with von Hippel-Lindau (VHL) disease. VHL disease type 2C is a unique subtype characterized by the presence of a PGL or pheochromocytoma without other VHL-associated neoplasms. This report describes a rare germline mutation in the VHL gene in a patient with isolated PGL. The clinical presentation, urinary metanephrines and normetanephrines, computed tomography scan, meta-iodobenzylguanidine scintiscan, surgical pathology, and genetic testing of a patient with PGL and a rare VHL gene mutation are described. A literature review is also presented. A 23-year-old, Indian woman was incidentally found to have an indeterminate 4.2 × 3.6 × 3.2-cm mass adjacent to the liver. A 36-year-old first cousin was recently diagnosed with a PGL. Her 24-hour urinary metanephrines were 6,886 μg/g creatinine (reference range is 81 to 330 μg/g creatinine) and normetanephrines were 6,810 μg/g creatinine (reference range is 20 to 158 μg/g creatinine). Surgical pathology revealed a PGL adjacent to a normal adrenal gland. Genetic testing revealed a mutation in VHL p.L163F. Surveillance for other tumors associated with VHL disease has been negative thus far. Her cousin has not undergone genetic testing despite recommendations to do so. We present the first reported case of PGL in a patient with VHL disease caused by a missense mutation in VHL p.L163F. To date, reports of this rare mutation have only involved patients with pheochromocytoma and without other tumors associated with VHL disease, suggesting that VHL p.L163F mutation may cause a VHL disease type 2C phenotype.

Expression of unfolded protein response markers in the pheochromocytoma with Waardenburg syndrome: a case report

BackgroundIt is clinically emergent to further understand the pathological mechanism to advance therapeutic strategy for endocrine tumors. A high amount of secretory protein with tumorigenic triggers are thought to induce unfolded protein response in endoplasmic reticulum in endocrine tumors, but its evidence is limited.Case presentationA 40-year-old woman had an approximately 10-year history of intermittent headaches. After the incidental detection of a mass in her right adrenal gland by CT scan, she was admitted to our hospital. She had been diagnosed as type 1 Waardenburg syndrome with the symptoms of dystopia canthorum, blue iris, and left sensorineural hearing loss. Urinary catecholamine levels were markedly elevated. 123I-MIBG scintigraphy showed uptake in the mass in her adrenal gland. After the adrenalectomy, her headaches disappeared and urinary catecholamine levels decreased to normal range within 2 weeks. Genome sequencing revealed germline mutation of c.A175T (p.Ile59Phe) in transcription factor PAX3 gene and somatic novel mutation of c.1893_1898del (p. Asp631_Leu633delinsGlu) in proto-oncogene RET in her pheochromocytoma. RNA expression levels of RET were increased 139 times in her pheochromocytoma compared with her normal adrenal gland. Those of unfolded protein response markers, Bip/GRP78, CHOP, ATF4, and ATF6, were also increased in the pheochromocytoma.ConclusionWe report a rare case of pheochromocytoma with type 1 Waardenburg syndrome. This is the first case to show the activation of unfolded protein response in the pheochromocytoma with the novel somatic mutation in RET gene. Our findings may support that unfolded protein response is activated in endocrine tumors, which potentially could be a candidate of therapeutic target.

MON-LB042 A Rare Case of Adrenocortical Carcinoma Arising From Ectopic Adrenal Tissue in the Mesentery

Background: ACTH independent Cushing’s syndrome usually arises from benign or malignant tumors of the adrenal gland. Ectopic adrenal tissue can undergo malignant transformation resulting in the development of adrenocortical carcinoma (ACC) with normal adrenal glands. We present a unique case of ACTH independent Cushing’s syndrome from a cortisol and androgen producing ACC arising from the mesentery. Clinical Case: A 72-year-old woman presented with a 6-month history of progressive weakness, lower extremity edema, worsening hypertension and uncontrolled DM. She had moon facies, hirsutism, multiple bruises, and proximal muscle weakness. Labs revealed hypokalemia (2.6 mmol/L, N 3.5 -5.1 nmol/L), elevated random cortisol (51.3 mcg/dL, Nl- 5-23), suppressed ACTH (<1 pg/mL, Nl-7.2-63.3) abnormal 8 mg dexamethasone suppression test (58.1 mcg/dL, N <1.8 mcg/dL), elevated DHEAS (538mcg/dL, N 10-90 mcg/dL), elevated testosterone (590.2 ng/dL, N <75 ng/dL) and elevated 11-deoxycortisol (4650 ng/dL N-<32 ng/dL). 24 hour urinary free cortisol was 2810mcg/mL (<45mcg/dL). MRI of pituitary was normal. CT scan of abdomen/pelvis showed normal bilateral adrenal glands and innumerable enhancing masses throughout the abdomen with the largest mass near the distal ileum and cecum. Biopsy of right lower abdominal mass revealed adrenocortical morphology with immunohistochemical staining positive for inhibin, synaptophysin and calretinin. Ki-67 index was 10-15%, suggestive of low-grade adrenocortical carcinoma. A CT scan done one and half years prior noted a 4.4 cm soft tissue mass in the right lower mesentery supporting origin of the tumor from the mesentery. Hypercortisolism was controlled with Metyrapone 250 mg BID. Mitotane 1000 mg bid was initiated but patient developed peritoneal carcinomatosis within 1 month. Conclusions: Our case is remarkable for the development of a metastatic ACC from an ectopic adrenal tissue with normal bilateral adrenal glands. Ectopic ACC is very rare with only a handful of cases reported in the literature. This is the first reported case of ACC arising from the mesentery. Ectopic adrenal tissue can be found close to the adrenal glands, along the path from gonads to adrenal glands or in association with the gonads. In the setting of ACTH independent Cushing’s syndrome with normal adrenal glands, physicians should direct their search to a functioning ectopic adrenocortical tissue. Concomitant DHEAS secretion suggests ectopic ACC. If surgery is not an option due to metastatic disease, a multidisciplinary approach should be adopted to control tumor growth and associated symptoms. In such cases, control of the hypercortisolemia can be achieved with adrenolytic medications such as Metyrapone, Ketoconazole or Mitotane. Adjuvant chemotherapy (Mitotane and combination of cytotoxic drugs) might be considered for metastatic ACC treatment.

SAT-553 Use of Optimal Cutting Temperature Compound (OCT)-Embedded Adrenal Tumor Tissue for Intratumoral Steroid Hormone Profiling

Background: Primary aldosteronism (PA) is the most common cause of secondary hypertension, accounting for 5-8% of all hypertension. PA is most commonly attributed to an aldosterone-producing adenoma (APA) or to bilateral hyperaldosteronism (BHA). Mutations in the inward-rectifying K+ channel (mKCNJ5), which increase autonomous aldosterone production, are most frequently detected in APAs. APAs with mKCNJ5 display aberrant expression of aldosterone synthase (CYP11B2) and 17α-hydroxylase (CYP17A1), which are involved in aldosterone and cortisol synthesis, respectively. Co-expression of these enzymes results in the production of a set of “hybrid” steroids, which have been proposed as serum biomarkers. The relative production of hybrid steroids in adrenal tumors vs. adjacent normal adrenal (NA) tissue has not been investigated. Objectives: To determine the utility of OCT-embedded adrenal tumor tissue for steroid profiling. To use immunohistochemistry (IHC)-guided OCT tumor capture for intratumoral hybrid steroid profiling in mKCNJ5 APA and NA tissue. Methods: OCT-embedded adrenal tissue from 9 patients (8 women, Age 45.9 ± 3.3 years) with APAs harboring known KCNJ5 mutations were used for the study. Where available OCT-embedded normal adrenal (NA) tissue adjacent to APAs were used as controls (n=4). IHC was performed for CYP11B2 and CYP17A1 on OCT tissue allowing guided APA capture from serial sections. Steroids were extracted from APA and adjacent NA tissue, and quantified by liquid chromatography/tandem mass spectrometry. Steroids measured were normalized to the protein content of the extracted tissue. Results: Compared to NA, APA tissue demonstrated 23-, 5.6- and 6.4-fold higher levels of aldosterone, 11-deoxycorticosterone, and 18-hydroxycorticosterone, respectively (P<0.05). In addition, the “hybrid” steroid products, 18-oxocortisol and 18-hydroxycortisol, were significantly elevated in APA vs. NA (P<0.01). Conversely, the adrenal androgens dehydroepiandrosterone and 11-hydroxyandrostenedione were lower in APA as compared with NA (P<0.05). All mKCNJ5 APAs were also found to co-express CYP11B2 and CYP17A1. Conclusion: IHC-guided mKCNJ5 APA capture and steroid extraction identified a distinct intratumoral hybrid steroid signature that associated with co-expression of CYP11B2 and CYP17A1.These findings also demonstrate that OCT-embedded tissue can be used to accurately define intra-tissue steroid profiles, which will have application for steroid-producing and steroid-responsive tumors.

OR19-02 Luteinizing Hormone/Human Chorionic Gonadotropin Receptor Protein Expression in Adrenocortical Progenitor Cells, Aldosterone Producing Cell Clusters and Adrenal Adenomas Derived from Postmenopausal Women

Objective/BackgroundAdrenal pathologies are more common in women than men. Embryologically the adrenals and gonads develop from the adrenogenital ridge with differential migration and differentiation. We hypothesized that in adult females there are adrenocortical progenitor cells that express the LH/hCG-R and proliferate in response to elevated LH. Indeed, several case reports demonstrated LH/hCG-R expression in adrenal secretory tumors in postmenopausal and pregnant females. In aging adults, nests of cells known as aldosterone-producing cell clusters (APCCs) that may be precursors to aldosterone producing adenomas are frequently detected. We retrospectively studied the immunohistochemical expression of LH/hCG-R in normal adrenals, adrenal adenomas and APCCs in archival specimens derived from post-menopausal women.MethodsArchival specimens from adrenal adenomas derived from 23 women >55 years of age were examined. Clinical data was obtained in a blinded fashion and hormonal data was available in 9/23 cases; 6/9 were secreting cortisol and 3/9 adenomas were secreting aldosterone. In addition, 6 samples derived from a repository of normal adrenal tissues from deceased kidney donors (1 male, and 5 postmenopausal females) were studied. All specimens were immunostained for LH/hCG-R and the adrenal stem cell marker DLK1 that facilitates the maintenance of an undifferentiated phenotype. The normal adrenal tissues were also stained for aldosterone synthase (CYP11B2) to detect APCCs. The slides were reviewed and graded by a pathologist in a blinded fashion.ResultsExpression of LH/hCG-R was demonstrated in both normal and adenomatous tissues in all 23 specimens. The staining in adenomas was heterogeneous, with clusters of densely stained LH/hCG-R positive cells in all specimens. There were less densely stained clusters in normal adjacent adrenocortical tissue that was most prominent in the subcapsular, zona glomerulosa region, an area where the putative adrenal cortical stem cells are found as well as the zona reticularis. Double staining for the stem cell marker DLK1 and LH/hCG-R confirmed that these cells represent adrenocortical progenitor cells. CYP 11B2 immunohistochemistry of normal adrenals demonstrated cell foci dipping from the capsule into the zona fasciculata classified as APCCs that co-expressed cytoplasmic LH/hCGR.ConclusionAdrenal adenomas and APCCs derived from postmenopausal women exhibited heterogeneous but strong immunohistochemical expression of LH/hCG-R in all samples. Interestingly, DLK1-positive adrenocortical stem cells in the subcapsular zone also expressed LH/hCG-R. These data may provide insights into the female predominance of adrenal pathologies, particularly in postmenopausal women with high LH levels. The LH/hCG-R may be a viable target for treatment of adrenal adenomas in postmenopausal women.

SAT-179 Increased Telomere Length in Adrenocortical Tumors Is Associated with Abnormal Expression of Chromatin Remodelling Factors

Background: The pathogenesis of adrenocortical tumors (ACTs) in the pediatric population is partially known, and few prognostic factors have been identified in this age group. Recently, ATRX and DAXX have been implicated in the pathogenesis and prognosis of a variety of cancers. Their altered function has been shown to affect telomere length through a telomerase-independent mechanism. Objective: To investigate ATRX and DAXX gene expression, ATRX and DAXX protein expression, and telomere length, as well as their clinical significance, in ACT samples from pediatric patients. Methods: The records of 110 pediatric patients with available ACT samples were reviewed. ATRX, DAXX, TERT and TERC gene expression was assessed by qPCR (n = 100 ACTs; n = 12 normal adrenals). ATRX and DAXX protein expression was assessed by IHC (n = 45 ACTs). Telomere length was assessed by qPCR (n = 64 ACTs). For survival analysis, Kaplan-Meier curves were obtained. For association analysis, simple linear regression models were adjusted. Results: Most patients were female (70.9%) and harbored germline TP53 mutations (90.2%). Median age at diagnosis was 21.1 months (2.1 – 199). Younger patients (< 3 years) had better survival (p < 0.01), while those with metastasis at diagnosis and carcinomas (classified by the Wieneke score) had worse survival (p < 0.01). ATRX gene expression was decreased (p < 0.01), while DAXX gene expression was increased (p < 0.01) in ACTs, compared to normal adrenals. ATRX gene expression was even lower in the context of the germline TP53 (R337H) mutation (p < 0.01). TERT expression was not detected in ACTs or normal adrenals, and TERC expression was not altered (p = 0.69). ATRX protein expression was lost in the majority of ACTs (95.6%), while DAXX was lost in a minority (21.1%). There was no association between gene or protein expression and disease-free or overall survival. There was a significant association between decreased ATRX and DAXX gene expression and increased telomere length (p < 0.01 and p = 0.03, respectively). Conclusion: In pediatric ACTs, decreased ATRX and DAXX gene expression was associated with increased telomere length, independently of TERT or TERC expression. In these tumors, ATRX gene expression was decreased and ATRX protein expression was overall lost, while DAXX gene expression was increased and DAXX protein expression was overall retained. No significant association between these alterations and prognosis was found in this cohort. These findings suggest that ATRX and DAXX altered function may be more involved in the pathogenesis of pediatric ACTs than in the prognosis of the affected patients.

SAT-LB90 Angiotensin II and ACTH Receptor Expression in Aldosterone-Producing Adenomas

Background: The mechanisms leading to elevated aldosterone synthesis in aldosterone-producing adenomas (APAs) remain an area of active research. Aldosterone-driver somatic gene mutations that allow inappropriate intracellular calcium entrance have been identified in most APAs. Cell-based studies of such mutations indicate that responses to physiologic stimuli, such as angiotensin II or ACTH, are increased. Little is known, however, regarding possible variations in response to hormonal stimuli between APAs with different aldosterone-driver mutations. Herein, we analyzed the transcript expression of the ACTH receptor (MC2R), the melanocortin 2 receptor accessory protein (MRAP) and the type 1 angiotensin II receptor (AGTR1) in APAs with known aldosterone-driver somatic mutations. Methods: RNA was isolated from normal adrenal glands (n=8), and from APAs with mutations in: KCNJ5 (n=14), ATP1A1 (n=14), CACNA1D (n=14), and ATP2B3 (n=5). The gene expressions of MC2R, MRAP, AGTR1 and aldosterone synthase (CYP11B2) were quantified using qPCR and normalized to β-actin. Results: All APA mutation groups had significantly higher transcript levels of CYP11B2, MC2R and AGTR1 as compared to whole normal adrenals. While MRAP and AGTR1 transcripts were comparable between tumor mutation groups, MC2R expression was significantly lower in KCNJ5-mutated APAs compared to other APAs. Overall, CYP11B2 expression demonstrated positive correlations with MC2R (R=0.728, p<0.0001) and AGTR1 (R=0.397, p=0.006) in APAs. These correlations were strongest in APAs harboring ATP1A1 mutations, and weakest in KCNJ5-mutated APAs. Conversely, CYP11B2 did not correlate with MC2R and AGTR1 in the normal adrenals. Conclusions: ACTH and angiotensin II receptors are expressed in all APAs, regardless of the underlying aldosterone-driver somatic mutations. Further research to clarify the effects of ACTH and posture on aldosterone production from APAs could provide additional insight into developing diagnostic and subtyping tools for primary aldosteronism.