Nonfunctioning Adrenocortical Adenomas Research Articles

FRI178 The Clinicopathological Distinction And Malignant Potential In Patients With Lipid-rich and Lipid-poor Non-functioning Adrenal Adenoma

Abstract Disclosure: P. Pengkhum: None. S. Lertlam: None. P. Cheunsuchon: None. V. Suvannarerg: None. S. Sirinvaravong: None. T. Wannachalee: None. Introduction: Adrenal incidentaloma (AI) is defined as an adrenal mass detected during imaging performed for conditions unrelated to suspected adrenal diseases. The latest evidence shows that most AIs are non-functioning adrenocortical adenomas (NFA). Of these, 70% are lipid-rich adenomas (LRA) and 30% are lipid-poor adenomas (LPA). Current guidelines do not recommend further follow-up imaging in patients with an adrenal mass size <4 cm with clear benign features. But some non-functioning lipid-poor lesions may carry malignant potential and proper management including surgical resection should be offered. However, the natural history and optimal specific follow-up surveillance of non-functioning LPA have not been well characterized. Objectives: We aimed to evaluate the clinical characteristics, natural courses, and histopathological data in NFA, and compared the malignant potential between LRA and LPA. Methods: We retrospectively studied all patients with AI between Jan 2005 to June 2022. Only patients with NFA who underwent adrenalectomy or were followed up by imaging for at least 6 months were included. The demographic, radiographic, and follow-up data were evaluated in all patients. Available adrenal tissues were performed for pathological assessment by using the Weiss score, Ki-67, and reticulin algorithm to identify the malignant potential. Results: Of 303 patients with AIs, 146 patients (48%) with NFAs were recruited, including 79 masses in LRA group and 77 masses in LPA group. Of these, the median age was 61 years, 43% were men, and the median follow-up period was 47 months. The initial median adenoma size was 1.6 cm [1.2-2.1]. During the follow-up period, only 3 LPAs developed significant interval enlargement. However, none of these were neither treated by surgical resection nor demonstrated malignant transformation based on imaging appearance. Overall, there was no difference in tumor growth between both groups by radiological assessment. Furthermore, 27 patients of NFAs underwent adrenalectomy; 8 masses in LRA group and 19 masses in LPA group. Of these, no difference in histological assessment in both groups. However, two large LPAs which size ≥4 cm revealed a Weiss score ≥3 with the presence of reticulin algorithm indicating potential malignant behavior. In contrast to LRA. Regardless of the adrenal mass size, the histopathological results revealed no suggestive malignant features. Conclusions: There was no significant difference in characteristics and tumor growth in patients with non-functioning LRA and LPA across 4-year interval follow-up data. Significant enlargement of adenomas was observed in few small LPAs, therefore periodic follow-up imaging should be offered in small LPAs with evidence of interval enlargement. According to our histopathological data, surgical resection should be considered in the large LPA (≥4 cm) due to its malignant potential. Presentation: Friday, June 16, 2023

European Society of Endocrinology Clinical Practice Guidelines on the management of adrenal incidentalomas, in collaboration with the European Network for the Study of Adrenal Tumors.

Adrenal incidentalomas are adrenal masses detected on imaging performed for reasons other than suspected adrenal disease. In most cases, adrenal incidentalomas are non-functioning adrenocortical adenomas, but may also require therapeutic intervention including that for adrenocortical carcinoma, pheochromocytoma, hormone-producing adenoma or metastases. Here, we provide a revision of the first international, interdisciplinary guidelines on incidentalomas. We followed the GRADE (Grading of Recommendations Assessment, Development and Evaluation) system and updated systematic reviews on four predefined clinical questions crucial for the management of incidentalomas: A) How to assess risk of malignancy? ; B) How to define and manage mild autonomous cortisol secretion? ; C) Who should have surgical treatment and how should it be performed? ; D) What follow-up is indicated if the adrenal incidentaloma is not surgically removed? Selected Recommendations: 1) Each adrenal mass requires dedicated adrenal imaging. Recent advances now allow discrimination between risk categories: Homogeneous lesions with HU ≤ 10 on unenhanced CT are benign and do not require any additional imaging independent of size. All other patients should be discussed in a multidisciplinary expert meeting, but only lesions >4 cm that are inhomogeneous or have HU >20 have sufficiently high risk of malignancy that surgery will be the usual management of choice. 2) Every patient needs a thorough clinical and endocrine work-up to exclude hormone excess including the measurement of plasma or urinary metanephrines and a 1-mg overnight dexamethasone suppression test (applying a cutoff value of serum cortisol ≤50 nmol/l (≤1.8 µg/dl)). Recent studies have provided evidence that most patients without clinical signs of overt Cushing's syndrome but serum cortisol levels post dexamethasone >50 nmol/l (>1.8 µg/dl) harbor increased risk of morbidity and mortality. For this condition, we propose the term 'mild autonomous cortisol secretion' (MACS). 3) All patients with MACS should be screened for potential cortisol-related comorbidities that are potentially attributably to cortisol (e.g. hypertension and type 2 diabetes mellitus), to ensure these are appropriately treated. 4) In patients with MACS who also have relevant comorbidities surgical treatment should be considered in an individualized approach. 5) The appropriateness of surgical intervention should be guided by the likelihood of malignancy, the presence and degree of hormone excess, age, general health and patient preference. We provide guidance on which surgical approach should be considered for adrenal masses with radiological findings suspicious of malignancy. 6) Surgery is not usually indicated in patients with an asymptomatic, non-functioning unilateral adrenal mass and obvious benign features on imaging studies. Furthermore, we offer recommendations for the follow-up of non-operated patients, management of patients with bilateral incidentalomas, for patients with extra-adrenal malignancy and adrenal masses, and for young and elderly patients with adrenal incidentalomas. Finally, we suggest ten important research questions for the future.

Ang–Tie Angiogenic Pathway Is Distinctively Expressed in Benign and Malignant Adrenocortical Tumors

The differential diagnosis between adrenocortical adenomas (ACAs) and adrenocortical carcinomas (ACCs) relies on unspecific clinical, imaging and histological features, and, so far, no single molecular biomarker has proved to improve diagnostic accuracy. Similarly, prognostic factors have an insufficient capacity to predict the heterogeneity of ACC clinical outcomes, which consequently lead to inadequate treatment strategies. Angiogenesis is a biological process regulated by multiple signaling pathways, including VEGF and the Ang–Tie pathway. Many studies have stressed the importance of angiogenesis in cancer development and metastasis. In the present study, we evaluated the expression of VEGF and Ang–Tie pathway mediators in adrenocortical tumors (ACTs), with the ultimate goal of assessing whether these molecules could be useful biomarkers to improve diagnostic accuracy and/or prognosis prediction in ACC. The expression of the proteins involved in angiogenesis, namely CD34, VEGF, VEGF-R2, Ang1, Ang2, Tie1 and Tie2, was assessed by immunohistochemistry in ACC (n = 22), ACA with Cushing syndrome (n = 8) and non-functioning ACA (n = 13). ACC presented a significantly higher Ang1 and Ang2 expression when compared to ACA. Tie1 expression was higher in ACC with venous invasion and in patients with shorter overall survival. In conclusion, although none of these biomarkers showed to be useful for ACT diagnosis, the Ang–Tie pathway is active in ACT and may play a role in regulating ACT angiogenesis.

Izazovi u laboratorijskoj dijagnostici autonomne sekrecije kortizola kod pacijenata sa adrenalnim incidentalomima

Adrenal incidentalomas are unexpectedly identified adrenal masses without prior suspicion of the existence of adrenal disease. The majority of adrenal incidentalomas are benign adrenal adenomas (80-90%), and they are non-functioning adrenocortical adenomas in more than 70% of cases. Mild hypercortisolemia is the most common finding in hormonally active adrenal incidentalomas. It is defined as autonomous cortisol secretion in up to 15% of patients. Despite a lot of research that anticipated higher cardiometabolic risk in patients with autonomous cortisol secretion, there is still no clear consensus on biochemical criteria for an autonomous cortisol secretion diagnosis in patients with adrenal incidentalomas. This review delineates the advantages and limitations of different laboratory tests recommended for the diagnosis of autonomous cortisol secretion in adrenal incidentalomas.

Adrenal incidentaloma - differential diagnosis and management

Adrenal masses are common incidental findings on imaging procedures.In most cases, adrenal incidentalomas are nonfunctioning adrenocortical adenomas, but in up to 20 % they require therapeutic intervention. The aim of this article is to provide essential guidance for clinicians regarding clinical management of patients with adrenal incidentalomas based on the respective current clinical guidelines. The following main questions are addressed: How to distinguish a nonfunctioning incidentaloma from a functioning one and how to assess risk of malignancy? How to define and manage low-level autonomous cortisol secretion, formerly called "subclinical" Cushing's syndrome? Which patients have to be treated surgically and which approach should be used? What follow-up is indicated if the adrenal incidentaloma is not surgically removed?

Study of urine steroid profiles by gas chromatography-mass spectrometry in patients with adrenocortical cancer in the course of treatment

Background: Adrenocortical carcinoma (ACC) is a rare and aggressive disease. There are only few studies evaluating the diagnostic value of gas chromatography-mass spectrometry (GC-MS) for detection of ACC recurrence after surgery. It is necessary to conduct an in-depth study to search for the most informative markers of the disease relapse.Aim: To study urine steroid metabolism by GC-MS during treatment to identify early signs of metastatic disease and relapse.Materials and methods: Thirty nine (39) ACC patients were examined before and after surgery, in the early postoperative period (< 1 year) and late postoperative period (at 2 to 5 years). Ten (10) patients were disease-free at less than 1 year after surgery. Twenty nine (29) patients had metastases in lungs and other organs: 14, within 1 year after surgery, and 15, at 2 to 5 years. The control group included 25 patients with nonfunctioning adrenocortical adenomas (NAA) without malignant characteristics at histological examination. Urine steroid profiles were assessed with a gas chromatograph-mass spectrometer Shimadzu GCMS-QP2020.Results: As assessed by GC-MS, 16 major ACC biomarkers were found before surgery, including etiocholanolone, dehydroepiandrosterone (DHEA) and its metabolites, pregnanediol, pregnanetriol, 5-ene-pregnenes, and tetrahydro-11-deoxycortisol (THS). Their urine excretion was increased compared to that in the patients with NAA (р < 0.002). A non-classic 5-ene-pregnene, 3β,16,20-pregnenetriol (3β,16,20-dP3), was identified, with its urine excretion of > 500 mcg/day that was typical for ACC patients. After surgery, decreased urinary excretion of THS (р < 0.0001) and 3β,16,20-dP3 (р < 0.0001), increased 3α,16,20-dP3/3β,16,20-dP3 ratio (р = 0.003), compared to those before surgery, were indicative of the absence of any metastases. No difference of urine THS excretion and 3α,16,20-dP3/3e,16,20-dP3 ratio from the corresponding values before surgery (p > 0.05) is a sign of metastatic diseases in the ACC patients at less than 1 year after the surgery, of the disease relapse at 2 to 5 years, and of the disease relapse after chemotherapy. In addition, in the ACC patients with metastatic disease within 1 year after surgery, increased progestogen urine excretion was found. Urine excretion of DHEA and its metabolites in the patients with the disease relapse after chemotherapy was not different from those in the ACC patients before surgery (p > 0.05).Conclusion: Determination of urine excretion of THS, DHEA and its metabolites, etiocholanolone, 5-ene-pregnenes, 3β,16,20-dP3, and 3α,16,20-dP3/3β,16,20-dP3 ratio by GC-MS is of utmost importance in the monitoring of treatment for ACC and early diagnosis of the disease progression.

Hypertension Editors' Picks: Hyperaldosteronism.

Hypertension Editors' Picks: Hyperaldosteronism.

Demographic and Clinicopthologic Properties of Patients Operated for Adrenal Masses

The prevalence of adrenal incidentaloma is nearly 4 to 10% and increases with age. Most adrenal masses are nonfunctioning adrenocortical adenomas and found as an adrenal incidentaloma in 80% of cases, but also be conditions requiring therapeutic intervention. We presented our patients with adrenal masses and their management.

The Potential of Steroid Profiling by Mass Spectrometry in the Management of Adrenocortical Carcinoma.

Radiological and endocrinological work up of adrenal neoplasms is aimed at distinguishing between frequent non-functioning adenomas and rare but very aggressive adrenocortical carcinoma (ACC). Relevant research has addressed the identification of molecular, genetic and hormonal markers that could have clinical significance for malignancy, as well as a prognostic value. Regarding endocrine aspects, attention has been paid to the pattern of steroid secretion that can be affected by altered steroidogenic pathway in ACC. The advent of mass spectrometry techniques has overcome many limitations usually associated with immunoassays, allowing the determination of both common and rarely measured steroids in a single analysis with high specificity and sensitivity. Indeed, mass spectrometry strategies may be able to identify an individualized steroid profile of ACC, allowing a rapid diagnosis and a specific follow-up. In this review, insights, strengths and limitations of mass spectrometry-based approaches in steroid profiling, as well as of immunoassay in steroid measurements, will be specifically discussed. Moreover, the latest findings on steroid profiling by mass spectrometry-based techniques, the most promising analytical tool, will be summarized to evaluate if steroid profiling might be the clue for solving the clinical dilemma in differentiating ACC from non-functioning adrenocortical adenomas (ACA).

Endoplasmic Reticulum Chaperone Calmegin Is Upregulated in Aldosterone-Producing Adenoma and Associates With Aldosterone Production.

The endoplasmic reticulum (ER) plays a pivotal role in syntheses of proteins and steroid hormones and regulation of intracellular Ca2+ level. We aimed to investigate ER-associated genes in aldosterone-producing adenomas (APAs) and clarify their effect on aldosterone production. Microarray analysis targeting 288 ER-associated genes was conducted using nonfunctioning adrenocortical adenomas (n=5) and APAs (n=19). Immunohistochemistry and quantitative polymerase chain reaction analyses were performed with 13 nonfunctioning adrenocortical adenoma and 48 APA samples. Functional studies were performed with human adrenocortical carcinoma (HAC15) cells, some of which were genetically modified using lentiviruses. The ER chaperone calmegin (CLGN) was the most highly expressed ER-associated gene in APAs relative to nonfunctioning adrenocortical adenomas. Analysis with quantitative polymerase chain reaction revealed CLGN to be 9.5-fold upregulated in APAs relative to nonfunctioning adrenocortical adenomas. There were no differences among different APA genotypes affecting aldosterone production. Immunohistochemistry analysis revealed that CLGN was strongly expressed in APAs and aldosterone-producing cell clusters. Angiotensin II stimulation or KCNJ5 T158A overexpression in HAC15 cells did not affect CLGN mRNA levels. CLGN overexpression in HAC15 cells increased aldosterone levels but did not stimulate CYP11B2 mRNA levels. Pathway and gene ontology analyses using RNA sequencing results showed that tRNA aminoacyl metabolism was the most enriched pathway in CLGN-overexpressing cells. CYP11B2 (aldosterone synthase) and HSD3B2 (3 beta-hydroxysteroid dehydrogenase/delta 5->4-isomerase type 2) protein expression were more abundant in CLGN-overexpressing cells. CLGN knockdown using CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/clustered regularly interspaced short palindromic repeat-associated 9) method in HAC15 cells that carry the KCNJ5 mutation did not affect aldosterone production. To summarize, CLGN was upregulated and associated with aldosterone production via translational regulation of CYP11B2 in APAs.

Does IGF-1 play a role in the etiopathogenesis of non-functioning adrenocortical adenoma?

The aim of this study was to investigate the possible association of insulin-like growth factor-1 (IGF-1) with the pathogenesis of non-functioning adrenocortical adenomas (NFAs). This study included 50 female patients (mean age 54years) with NFAs, 55 patients (mean age 48years; 20 male, 35 female) with acromegaly and 38 female control subjects (mean age 58years). Body mass index (BMI) and waist circumference (WC) of the subjects were recorded and blood samples for IGF-1 were taken. Insulin resistance was calculated using the homeostatic model assessment (HOMA) score. Since most of the acromegaly patients had been using medicine that could have effected insulin resistance, HOMA scores were calculated only in patients with NFAs and the controls. Computerized tomography or magnetic resonance imaging was taken of the acromegalics and controls to detect adrenal mass frequency. The mean age was similar among the groups. As expected, the serum IGF-1 levels were significantly higher in patients with acromegaly than in patients with NFAs and the controls (p < 0.001). Although BMI, WC, and serum IGF-1 levels were significantly higher (p < 0.001) in patients with NFAs, the HOMA scores were similar between patients with NFAs and control groups. Although none of the control subjects had adrenal masses, NFAs were detected in 14 (25%) out of 55 acromegalic patients. Higher serum IGF-1 levels in patients with NFAs compared to the control group and an increased prevalence of NFAs in acromegaly patients compared to control subjects and the general population suggest an association of IGF-1 with the etiopathogenesis of NFA.

Calneuron 1 Increased Ca2+ in the Endoplasmic Reticulum and Aldosterone Production in Aldosterone-Producing Adenoma.

Aldosterone production is initiated by angiotensin II stimulation and activation of intracellular Ca2+ signaling. In aldosterone-producing adenoma (APA) cells, the activation of intracellular Ca2+ signaling is independent of the renin-angiotensin-aldosterone systems. The purpose of our study was to clarify molecular mechanisms of aldosterone production related to Ca2+ signaling. Transcriptome analysis revealed that the CALN1 gene encoding calneuron 1 had the strongest correlation with CYP11B2 (aldosterone synthase) among genes encoding Ca2+-binding proteins in APA. CALN1 modulation and synthetic or fluorescent compounds were used for functional studies in human adrenocortical carcinoma (HAC15) cells. CALN1 expression was 4.4-fold higher in APAs than nonfunctioning adrenocortical adenomas. CALN1 expression colocalized with CYP11B2 expression as investigated using immunohistochemistry in APA and zona glomerulosa of male rats fed by a low-salt diet. CALN1 expression was detected in the endoplasmic reticulum (ER) by using GFP-fused CALN1, CellLight ER-RFP, and the corresponding antibodies. CALN1-overexpressing HAC15 cells showed increased Ca2+ in the ER and cytosol fluorescence-based studies. Aldosterone production was potentiated in HAC15 cells by CALN1 expression, and dose-responsive inhibition with TMB-8 showed that CALN1-mediated Ca2+ storage in ER involved sarcoendoplasmic reticulum calcium transport ATPase. The silencing of CALN1 decreased Ca2+ in ER, and abrogated angiotensin II- or KCNJ5 T158A-mediated aldosterone production in HAC15 cells. Increased CALN1 expression in APA was associated with elevated Ca2+ storage in ER and aldosterone overproduction. Suppression of CALN1 expression prevented angiotensin II- or KCNJ5 T158A-mediated aldosterone production in HAC15 cells, suggesting that CALN1 is a potential therapeutic target for excess aldosterone production.

Management of adrenal incidentalomas: European Society of Endocrinology Clinical Practice Guideline in collaboration with the European Network for the Study of Adrenal Tumors.

: By definition, an adrenal incidentaloma is an asymptomatic adrenal mass detected on imaging not performed for suspected adrenal disease. In most cases, adrenal incidentalomas are nonfunctioning adrenocortical adenomas, but may also represent conditions requiring therapeutic intervention (e.g. adrenocortical carcinoma, pheochromocytoma, hormone-producing adenoma or metastasis). The purpose of this guideline is to provide clinicians with best possible evidence-based recommendations for clinical management of patients with adrenal incidentalomas based on the GRADE (Grading of Recommendations Assessment, Development and Evaluation) system. We predefined four main clinical questions crucial for the management of adrenal incidentaloma patients, addressing these four with systematic literature searches: (A) How to assess risk of malignancy?; (B) How to define and manage low-level autonomous cortisol secretion, formerly called 'subclinical' Cushing's syndrome?; (C) Who should have surgical treatment and how should it be performed?; (D) What follow-up is indicated if the adrenal incidentaloma is not surgically removed? SELECTED RECOMMENDATIONS: (i) At the time of initial detection of an adrenal mass establishing whether the mass is benign or malignant is an important aim to avoid cumbersome and expensive follow-up imaging in those with benign disease. (ii) To exclude cortisol excess, a 1mg overnight dexamethasone suppression test should be performed (applying a cut-off value of serum cortisol ≤50nmol/L (1.8µg/dL)). (iii) For patients without clinical signs of overt Cushing's syndrome but serum cortisol levels post 1mg dexamethasone >138nmol/L (>5µg/dL), we propose the term 'autonomous cortisol secretion'. (iv) All patients with '(possible) autonomous cortisol' secretion should be screened for hypertension and type 2 diabetes mellitus, to ensure these are appropriately treated. (v) Surgical treatment should be considered in an individualized approach in patients with 'autonomous cortisol secretion' who also have comorbidities that are potentially related to cortisol excess. (vi) In principle, the appropriateness of surgical intervention should be guided by the likelihood of malignancy, the presence and degree of hormone excess, age, general health and patient preference. (vii) Surgery is not usually indicated in patients with an asymptomatic, nonfunctioning unilateral adrenal mass and obvious benign features on imaging studies. We provide guidance on which surgical approach should be considered for adrenal masses with radiological findings suspicious of malignancy. Furthermore, we offer recommendations for the follow-up of patients with adrenal incidentaloma who do not undergo adrenal surgery, for those with bilateral incidentalomas, for patients with extra-adrenal malignancy and adrenal masses and for young and elderly patients with adrenal incidentalomas.

18F-Fluorodeoxyglucose Positron Emission Tomography for the Diagnosis of Adrenocortical Tumors: A Prospective Study in 77 Operated Patients

Most adrenal incidentalomas are nonfunctioning adrenocortical adenomas (ACAs). Adrenocortical carcinomas (ACCs) are rare but should be recognized at an early stage. The objective of the study was to evaluate the usefulness of (18)F-fluorodeoxyglucose positron emission tomography ((18)F-FDG PET) to predict malignancy in patients without a previous history of cancer. This was a prospective, multicenter study from 2001 to 2006. The study was conducted at a network of seven university hospitals in Paris. Seventy-seven patients were included. All underwent surgery because of hypersecretory and/or growing benign lesions (n = 18), obvious ACCs (n = 21), or radiologically indeterminate lesions (n = 38). The degree of (18)F-FDG PET uptake [maximum standardized uptake value (maxSUV)] was related to the pathological findings serving as a reference, and its diagnostic value was compared with that of computerized tomography (CT) scan. Pathology eventually diagnosed 43 ACAs, 22 ACCs, and 12 nonadrenocortical lesions. Using a cutoff value above 1.45 for adrenal to liver maxSUV ratio, the sensitivity and specificity to distinguish ACAs from ACCs were, respectively, 1.00 (95% confidence interval 0.85-1.00) and 0.88 (95% confidence interval 0.75-0.96). Among the 38 indeterminate lesions at CT scan, we could analyze a subgroup of 16 adrenocortical tumors with high unenhanced density (>10 HU) and an inappropriate washout: (18)F-FDG PET correctly predicted the benignity in 13 of 15 ACAs. In a multidisciplinary team approach, (18)F-FDG PET helps to manage suspicious CT scan lesions. An adrenal to liver maxSUV ratio less than 1.45 is highly predictive of a benign lesion.

The role of 21-hydroxylase in the pathogenesis of adrenal masses: Review of the literature and focus on our own experience

An exaggerated response of 17- hydroxyprogesterone (17-OHP) to exogenous ACTH stimulation has been found in 30 to 70% of patients with incidentally discovered adrenal tumors, supporting the concept that congenital 21- hydroxylase deficiency may be a predisposing factor for adrenocortical tumorigenesis. Decreased expression of 21-hydroxylase gene has been observed in sporadic non-functioning adrenocortical adenomas and adrenocortical carcinomas, in agreement with the reduced steroidogenic activity found in these types of tumors. Screening studies for the presence of mutations in CYP21A2 gene, encoding 21-hydroxylase, in patients with sporadic adrenocortical tumors yielded discordant results. Overall, a higher frequency of germline 21-hydroxylase mutation carriers has been found among patients with adrenal tumors, including incidentalomas, than in the general population. However, the presence of mutations did not correlate with endocrine test results and tumor mass features, suggesting that 21-hydroxylase deficiency does not represent a relevant mechanism in adrenal tumorigenesis. Mechanisms leading to reduced 21-hydroxylase expression and activity are still unknown.

Posterior Retroperitoneoscopic Adrenalectomy: A Comparison between the Initial Experience in the Invention Phase and Introductory Phase of the New Surgical Technique

Today, the posterior retroperitoneoscopic technique has become a standard procedure in adrenal surgery. The procedure allows direct access to the adrenal glands, but it seems to be difficult because of the uncommon anatomic view. This study compares the learning period of the new procedure of "posterior retroperitoneoscopic adrenalectomy" in the primary invention phase and the secondary introductory phase in a different hospital 10 years later. The analysis included 100 posterior retroperitoneoscopic adrenalectomies (PRA) and involved 50 procedures in each center. Group A consisted of 44 patients (14 males, 30 females; age: 48.7 +/- 14.5 years) undergoing surgery between 07/1994 and 8/1996 (24 right, 26 left; 8 Cushing adenomas, 14 Conn adenomas, 11 pheochromocytomas, 7 nonfunctioning adrenocortical adenomas, 10 ACTH-dependent adrenal hyperplasias). Group B consisted of 50 patients (12 males, 38 females; mean age 59.3 +/- 10.7 years) operated between 01/2004 and 01/2006 (28 right, 22 left tumors; 5 Cushing adenomas, 12 Conn adenomas, 4 pheochromocytomas, 29 nonfunctioning adrenocortical adenomas). All PRAs were performed with the patient in the prone position with 3-4 trocars placed caudally in the region of the 11th and 12th ribs. In group A, the surgical team developed the technique of PRA themselves. Before their first PRA, the surgical team of group B was introduced to the technique by the group A surgeons and afterwards were supervised continuously. No serious intraoperative or postoperative complication occurred in either group. Group A experienced 7 conversions to open surgery, whereas group B had one conversion and one early reoperation due to bleeding (P = 0.03; chi(2)-test). The mean operative time was 117 +/- 41 minutes versus 83 +/- 35 minutes (group A and B respectively; P < 0.001; t-test). Estimated blood loss was similar in the two groups (47.2 +/- 46.2 ml versus 54 +/- 16.3 ml, group A versus B, respectively; P = 0.36; t-test). The study demonstrates the feasibility, safety, and reproducibility of the new surgical method of PRA both when it is employed in the early phase of invention, as well as when performed by surgeon-learners. After comprehensive training, the operative time and conversion rate are dramatically reduced, allowing for a short learning period.

High Plasma Levels of Human Chromogranin a and Adrenomedullin in Patients with Pheochromocytoma

The aim of our study was to investigate the plasma chromogranin A (CgA) and adrenomedullin (AM) levels in patients with pheochromocytomas. We collected blood samples for measurement of plasma CgA and AM in 21 patients with pheochromocytomas, 43 healthy subjects and 26 patients with solid non-functioning adrenocortical adenomas. In 11 patients with pheochromocytomas plasma CgA and AM were measured again four weeks after tumor removal. CgA and AM were measured by means of a novel solid-phase two-site immunoradiometric assay based on monoclonal antibodies (CgA-RIA CT, CIS bio international) and by a specific radioimmunoassay (RIA, Phoenix Pharm. Inc.), respectively. The mean plasma CgA level (+/- SD) in patients with pheochromocytomas (204 +/- 147.9 ng/mL) was significantly higher (P < 0.001) than that in healthy subjects (41.6 +/- 10.7 ng/mL) and in patients with non-functioning adrenocortical adenomas (47.3 +/- 17.6 ng/mL). The mean plasma AM concentration (+/- SD) in patients with pheochromocytomas (27.5 +/- 10.4 pg/mL) was significantly higher (P < 0.001) than that in HS (13.8 +/- 4.5 pg/mL) and in patients with non-functioning adrenocortical adenomas (16.6 +/- 7.3 pg/mL). Plasma CgA levels correlated with plasma AM levels (r = 0.501; P < 0.02) and with plasma metanephrine levels (r = 0.738; P < 0.0001) in patients with pheochromocytomas. In 11 patients with pheochromocytomas plasma CgA and AM concentrations significantly decreased after tumor removal (P < 0.001 for both). Circulating CgA and AM had a sensitivity of 76.2% and 81%, a specificity of 97.7% and 90.7%, and an accuracy of 91% and 88%, respectively. This study demonstrates that circulating CgA and AM levels are increased in pheochromocytoma patients compared with healthy subjects and patients with non-functioning adrenocortical adenomas. Moreover, at the time of diagnosis plasma CgA levels correlated with plasma AM levels and with plasma metanephrine levels in all patients with pheochromocytomas. In conclusion, plasma CgA and AM concentrations may represent additional biochemical parameters for clinical monitoring of patients with pheochromocytomas.

25 Reduction of organelles and produce of precursor substances without hormone activity in non-functioning adrenocortical adenomas

25 Reduction of organelles and produce of precursor substances without hormone activity in non-functioning adrenocortical adenomas

Metabolism of glucocorticoids and mineralocorticoids in patients with adrenal incidentalomas.

Adrenal incidentalomas are mostly nonfunctioning adrenocortical adenomas (NFI). However, in 5%-12% of the patients a preclinical Cushing's syndrome (PCS) with autonomous cortisol production by the tumour is present. Since urinary free cortisol excretion is not sensitive enough to determine subclinical hypercortisolism, in the present study more sensitive indicators of daily cortisol production were measured. (1) Tetrahydrocortisol, tetrahydrocortisone, urinary free cortisone together with urinary free cortisol were measured in 35 patients with adrenal incidentalomas (29 NFI and six PCS) and in 35 healthy controls. (2) Since little is known about daily aldosterone production, aldosterone metabolite excretions were measured. (3) As recently reported, ACTH stimulation revealed an increased response of precursors of the glucocorticoid and mineralocorticoid pathway. To find out which steroidogenic enzymes have altered activities, a 1-24ACTH stimulation test (250 microg i.v.) was carried out in 25 patients and 18 healthy controls with determination of multiple steroids. (4) Finally, since it was assumed that 21-hydroxylase deficiency or even 11b-hydroxylase deficiency may be involved in adrenal tumourigenesis, the prevalence of germline CYP21B and CYP11B1 mutations were studied in the same patients, who had underwent the ACTH stimulation test. (1) Glucocorticoid metabolites were within the normal range in all but three patients with NFI. As a group, the patients had subtle alterations in cortisol metabolism. Tetrahydrocortisol excretion was elevated in NFI and PCS compared with normal subjects (2.1 +/- 0.2 and 2.5 +/- 0.5 vs. 1.5 +/- 0.1 mg 24 h(-1); P < 0.05). Accordingly, the twofold elevation of the tetrahydrocortisol/free cortisol ratio indicates an increased 5beta-reduction of cortisol in the liver. (2) Tetrahydroaldosterone and aldosterone-18-glucuronide excretions were not different to controls. (3) In patients with incidentalomas an increased response to ACTH was seen for 17-hydroxyprogesterone (595 +/- 133 vs. 160 +/- 25 ng dL(-1)), 21-desoxycortisol (105 +/- 25 vs. 29 +/- 9 ng dL(-1)) and 11-desoxycortisol (401 +/- 40 vs. 293 +/- 17 ng dL(-1)). (4) In only one of 25 patients, a heterozygous deletion in exon 3 of the CYP21B gene was detected. (1) In conclusion, even the excretion of the main glucocorticoid metabolites is not a marker sensitive enough to distinguish between NFI and PCS. However, it is also possible that alterations in cortisol secretion are qualitative rather than quantitative. (2) Zona glomerulosa function is not influenced. (3) The elevation of 21-desoxycortisol argues against an impairment of 11beta-hydroxylase and favours a decreased activity of 21-hydroxylase. All others had wild-type sequences of both genes. (4) In conclusion, neither 21-hydroxylase deficiency nor 11beta-hydroxylase deficiency are predisposing factors for adrenal tumourigenesis.

Evidence for androgen receptor gene expression and growth inhibitory effect of dihydrotestosterone on human adrenocortical cells.

Evidence for the expression of the canonic androgen receptor (AR) in human adrenal cortex has not been provided so far. The aim of the present study was to demonstrate the expression of the AR gene in normal and neoplastic adrenocortical human tissues and in the human adrenocortical cancer cell line, NCI-H295, and then to evaluate the effect of dihydrotestosterone (DHT) on human adrenocortical cell growth. An AR cDNA fragment with the expected size of 262 bp was detected by using reverse transcription (RT)-PCR in normal and neoplastic adrenocortical human tissues and in the neoplastic cell line, demonstrating that the gene for AR is indeed expressed in human adrenal cells. In the human adrenocortical cancer cell line NCI-H295, DHT at physiological concentrations produced a significant reduction in cell proliferation and inhibition of colony formation in soft agar. The inhibitory effect on adrenocortical cell growth was evident after both 24 and 48 h of treatment. The antiandrogens, cyproterone acetate and hydroxyflutamide, were capable of reversing the effects exerted by DHT. The androgen-induced growth inhibitory effect was also detected in primary culture of three non-functioning adrenocortical adenomas. These findings show that the canonic AR is present in human adrenocortical cells and that androgens may have a role in the adrenal cortex by reducing cell proliferation.