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The adrenal gland provides an important function by integrating neuronal, immune, vascular, metabolic and endocrine signals under a common organ capsule. It is the central organ of the stress response system and has been implicated in numerous stress-related disorders. While for other diseases, regeneration of healthy organ tissue has been aimed at such approaches are lacking for endocrine diseases - with the exception of type-I-diabetes. Moreover, adrenal tumor formation is very common, however, appropriate high-throughput applications reflecting the high heterogeneity and furthermore relevant 3D-structures in vitro are still widely lacking. Recently, we have initiated the development of standardized multidimensional models of a variety of endocrine cell/tissue sources in a new multiwell-format. Firstly, we confirmed common applicability for pancreatic pseudo-islets. Next, we translated applicability for spheroid establishment to adrenocortical cell lines as well as patient material to establish spheroids from malignant, but also benign adrenal tumors. We aimed furthermore at the development of bovine derived healthy adrenal organoids and were able to establish steroidogenic active organoids containing both, cells of cortical and medullary origin. Overall, we hope to open new avenues for basic research, endocrine cancer and adrenal tissue-replacement-therapies as we demonstrate potential for innovative mechanistic insights and personalized medicine in endocrine (tumor)-biology., (© 2022. The Author(s).)

Background: Rats exposed to chronic predator scent stress mimic the phenotype of complex post-traumatic stress disorder (PTSD) in humans, including altered adrenal morphology and function. High- and low-anxiety phenotypes have been described in rats exposed to predator scent stress (PSS). This study aimed to determine whether these high- and low-anxiety phenotypes correlate with changes in adrenal histomorphology and corticosteroid production., Methods: Rats were exposed to PSS for ten days. Thirty days later, the rats' anxiety index (AI) was assessed with an elevated plus-maze test. Based on differences in AI, the rats were segregated into low- (AI ≤ 0.8, n = 9) and high- (AI > 0.8, n = 10) anxiety phenotypes. Plasma corticosterone (CORT) concentrations were measured by ELISA. Adrenal CORT, desoxyCORT, and 11-dehydroCORT were measured by high-performance liquid chromatography. After staining with hematoxylin and eosin, adrenal histomorphometric changes were evaluated by measuring the thickness of the functional zones of the adrenal cortex., Results: Decreased plasma CORT concentrations, as well as decreased adrenal CORT, desoxyCORT and 11-dehydroCORT concentrations, were observed in high- but not in low-anxiety phenotypes. These decreases were associated with increases in AI. PSS led to a significant decrease in the thickness of the zona fasciculata and an increase in the thickness of the zona intermedia . The increase in the thickness of the zona intermedia was more pronounced in low-anxiety than in high-anxiety rats. A decrease in the adrenal capsule thickness was observed only in low-anxiety rats. The nucleus diameter of cells in the zona fasciculata of high-anxiety rats was significantly smaller than that of control or low-anxiety rats., Conclusion: Phenotype-associated changes in adrenal function and histomorphology were observed in a rat model of complex post-traumatic stress disorder.

The adrenal gland consists of two tissues, cortex and medulla, united under one capsule. Adrenal stem/progenitor cells play a key role in development and homeostasis. Here, we describe a protocol for generating primary cultures of adrenal cells from mice. We describe techniques for separating the cortex and medulla, generating spheroid cultures containing stem- and progenitor cells, and for the differentiation into steroidogenic and chromaffin cells, respectively. This protocol enables analysis of various treatments before, during, or after differentiation. For complete details on the use and execution of this protocol, please refer to Rubin de Celis et al. (2015), Steenblock et al. (2018), and Werdermann et al. (2021)., Competing Interests: The authors declare no competing interests., (© 2021 The Author(s).)

The adrenal gland is important for many physiological and pathophysiological processes, but studies are often restricted by limited availability of sample material. Improved methods for sample preparation are needed to facilitate analyses of multiple classes of adrenal metabolites and macromolecules in a single sample. A procedure was developed for preparation of chromaffin cells, mouse adrenals, and human chromaffin tumors that allows for multi-omics analyses of different metabolites and preservation of native proteins. To evaluate the new procedure, aliquots of samples were also prepared using conventional procedures. Metabolites were analyzed by liquid-chromatography with mass spectrometry or electrochemical detection. Metabolite contents of chromaffin cells and tissues analyzed with the new procedure were similar or even higher than with conventional methods. Catecholamine contents were comparable between both procedures. The TCA cycle metabolites, cis -aconitate, isocitate, and α-ketoglutarate were detected at higher concentrations in cells, while in tumor tissue only isocitrate and potentially fumarate were measured at higher contents. In contrast, in a broad untargeted metabolomics approach, a methanol-based preparation procedure of adrenals led to a 1.3-fold higher number of detected metabolites. The established procedure also allows for simultaneous investigation of adrenal hormones and related enzyme activities as well as proteins within a single sample. This novel multi-omics approach not only minimizes the amount of sample required and overcomes problems associated with tissue heterogeneity, but also provides a more complete picture of adrenal function and intra-adrenal interactions than previously possible., Competing Interests: The authors declare that they have no conflict of interest., (The Author(s). This is an open access article published by Thieme under the terms of the Creative Commons Attribution-NonDerivative-NonCommercial-License, permitting copying and reproduction so long as the original work is given appropriate credit. Contents may not be used for commercial purposes, or adapted, remixed, transformed or built upon. (https://creativecommons.org/licenses/by-nc-nd/4.0/).)

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was originated in November-December 2019 in Wuhan, China, and has rapidly spread around the world causing severe health and socioeconomical damage to the entire civilization. The key feature of coronavirus disease 2019 (COVID-19), caused by SARS-CoV-2, is upper respiratory tract infection, which may be complicated by bilateral pneumonia. Angiotensin converting enzyme 2 (ACE2) has been identified as a key host factor, required for virus entry into cells. Interestingly, ACE2 is expressed not only in the respiratory system, but also in the other organs and systems including adrenal glands. Here we provide the first description of the pathomorphological changes in adrenal glands in patients with severe COVID-19 characterized by perivascular infiltration of CD3+ and CD8+ T-lymphocytes. Due to the central role of the adrenals in the stress response of the organism, this finding is of potential clinical relevance, because infection with the SARS-CoV-2 virus might critically impair adrenal function under pathophysiological conditions., Competing Interests: The authors declare that they have no conflict of interest., (© Georg Thieme Verlag KG Stuttgart · New York.)

Adrenal insufficiency requires lifelong corticoid replacement therapies. However, current therapies are not able to replace the physiological circadian pattern of the adrenal cortex and are associated with many metabolic, vascular, neuroendocrine, and mental perturbations. Therefore, regenerative and more curative strategies would be desirable. In the current perspective, we describe emerging new regenerative therapies for the treatment of adrenal insufficiency. In particular, we discuss gene therapy and cell replacement strategies. Furthermore, we discuss how adrenal cells might be used as a source for regenerative therapies of nonadrenal neurodegenerative diseases such as Parkinson disease., (© Endocrine Society 2020.)

Human individuals are constantly confronted to various kinds of stressors and the body's response and adaptation is essential for human health. The adrenal gland as the main producer of stress hormones plays a major role in the response to physiological challenges and is able to adapt to these physiological needs. Proper adaptation is of particular importance since dysregulation of the stress system is the cause of various human diseases including obesity, depression, Parkinson's disease, and post-traumatic stress disorder. Therefore, it is fundamental to understand the physiological, cellular, and molecular underpinnings of the stress adaptation in humans. Because of ethical reasons it is problematic to study the plasticity of the human gland in stress. Hence, various experimental models have been established for the analysis of the functional and cellular role of the adrenal gland adaptation on a translational approach. Here, we summarize the insights of stress-induced adrenal plasticity gained from these models and discuss their relevance to clinical observations., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Diseases of the adrenal gland are as important for the general practitioner as for the endocrine specialist. The high prevalence of some adrenal endocrinopathies, such as adrenal incidentalomas (1-2% of the population) and primary aldosteronism (6% of hypertensives), which affect millions of patients, makes adrenal diseases a relevant health issue. The high morbidity and mortality of some of the rarer adrenal diseases, i. e., Addison's disease and Cushing's syndrome (Table 1), make early detection and appropriate treatment such a challenge for the health care system., Competing Interests: No conflict of interest has been declared by the authors., (© Georg Thieme Verlag KG Stuttgart · New York.)

Necroinflammation is defined as the inflammatory response to necrotic cell death. Different necrotic cell death pathways exhibit different immune reponses, despite a comparable level of intracellular content release (referred to as damage associated molecular patterns or DAMPs). In addition to DAMP release, which is inevitably associated with necrotic cell death, the active production of pro/anti-inflammatory cytokines characterizes certain necrotic pathways. Necroptosis, ferroptosis and pyroptosis, therefore, are immunogenic to a different extent. In this review, we discuss the clinical relevance of necroinflammation highlighting potential human serum markers. We focus on the role of the adrenal glands and the lungs as central organs affected by systemic and/or local DAMP release and underline their role in intensive care medicine. In addition, data from models of acute kidney injury (AKI) and kidney transplantation have significantly shaped the field of necroinflammation and may be helpful for the understanding of the potential role of dialysis and plasma exchange to treat ongoing necroinflammation upon intensive care unit (ICU) conditions. In conclusion, we are only beginning to understand the importance of necroinflammation in diseases and transplantation, including xenotransplantation. However, given the existing efforts to develop inhibitors of necrotic cell death (ferrostatins, necrostatins, etc), we consider it likely that interference with necroinflammation reaches clinical routine in the near future.

The main treatment algorithm for adrenal insufficiency is hormonal replacement, however, inadequate hormone substitution often leads to severe side effects. Adrenal cell transplantation could be a more effective alternative but would require life-long immune suppressive therapy. PreImplantation Factor (PIF) is an endogenous peptide secreted by viable human embryos that leads to maternal tolerance without immunosuppression. PIF could be effective for xenogeneic cell transplantation such as of bovine adrenocortical cells (BAC), which are used for bioartificial adrenal gland development that may more effectively restore complex adrenal functions. We report here that PIF exerts a dual regulatory effect on BAC by targeting mostly hyper-activated cells to specifically reduce adrenocorticotropic hormone (ACTH)-stimulated cortisol secretion. Reverse transcription real time PCR analysis revealed that PIF modulates the expression of two genes in the cortisol synthesis pathway, Steroidogenic Factor 1 (SF1), an activator of steroidogenesis, and the downstream steroidogenic enzyme Cytochrome P450 17A1 (CYP17A1). PIF increased basal expression of SF1 and CYP17A1 regardless of the activation level of the adrenocortical cells. In contrast, following ACTH stimulation, PIF reduced SF1 expression and induced expression of the immune suppressing anti-inflammatory cytokine IL10 only in the hyper-activated cells, suggesting both a protective and immune tolerant function. In conclusion, PIF regulates stress-induced adrenal steroidogenesis and immune tolerance in BAC, supporting a potential clinical application to reduce rejection by the host's immune response following xenotransplantation., Competing Interests: The authors declare that they have no conflict of interest. ERB is CSO of BioIncept, LLC., (© Georg Thieme Verlag KG Stuttgart · New York.)

The adrenal gland is a key component of the stress system in the human body. Multiple direct and paracrine interactions between different cell types and their progenitors take place within the adrenal gland microenvironment. These unique interactions are supported by high vascularization and the adrenal cortex extracellular matrix. Alterations in the adrenal gland microenvironment are known to influence the progression of several pathological conditions, such as obesity and sepsis, and to be influenced by these disorders. For example, it has been suggested that activation of immune-adrenal crosstalk during sepsis induces elevated adrenal glucocorticoid levels, whereas crosstalk between adrenocortical cells and sonic hedgehog responsive stem cells was found to contribute to the increased size of the adrenal cortex during obesity. By contrast to sepsis, where activation of adrenal glucocorticoid production has protective effects, chronic exposure to high levels of glucocorticoids induces adverse effects, typically manifested in patients with Cushing syndrome, such as increased body weight, dyslipidemia, glucose intolerance, and hypertension. Therefore, a better understanding of factors involved in the regulation of the adrenal gland microenvironment is crucial. This review highlights bidirectional interactions occurring between the adrenal gland microenvironment and systemic responses during obesity and sepsis. Furthermore, it presents and discusses recent advancements and challenges in attempts to restore or regenerate adrenal gland function, including the use of oxygenated immune-isolating devices.

Objectives: Sepsis and septic shock are commonly present in the ICU and accompanied by significant morbidity, mortality, and cost. The frequency of secondary adrenal insufficiency in sepsis remains open to debate and a challenge to identify and treat appropriately. Animal models of sepsis using genetic or surgical initiation of adrenal insufficiency resulted in increased mortality, but the mechanisms are still unclear. The present study investigates the impact of adrenal inflammation in septic mice challenged with cecal ligation and puncture., Design: Prospective experimental study., Setting: University laboratory., Subjects: C57BL/6N wild-type mice., Interventions: Sepsis, induced by cecal ligation and puncture for 24 and 48 hours., Measurements and Main Results: Both septic and control mice were carefully monitored (every 30 min) for up to 48 hours and divided into survivors and nonsurvivors. We observed a significant and massive increase of interleukin-6, interleukin-1β, and tumor necrosis factor-α in adrenal protein extracts of nonsurvivors compared with sham animals and survivors. This pattern was partly reflected in liver and lung but not in plasma samples. Notably, a significant increase in nonsurvivors compared with survivors was only found for lung interleukin-6. In line with these findings, we detected a higher degree of leukocyte infiltration and hemorrhage in the adrenal glands of deceased mice. Evaluation of the hypothalamic-pituitary-adrenal axis response in these animals revealed an increase of adrenocorticotropic hormone, which was only partly reflected in the corticosterone level. Notably, using the adrenocorticotropic hormone stimulation test, we found an impaired adrenocorticotropic hormone response, particularly in nonsurvivors, which significantly correlated with the number of infiltrated leukocytes., Conclusions: Cecal ligation and puncture-induced murine sepsis induces a strong inflammatory response in the adrenal glands, which is accompanied by cell death and hemorrhage. Our data suggest that mortality and adrenal incapacitation are associated with the degree of adrenal inflammation, thereby underscoring the importance of adrenal function on survival.

Sepsis and septic shock in response to bacterial or viral infections remain the major health problem worldwide. Despite decades of intensive research and improvements in medical care, severe sepsis is associated with high mortality. Rapid activation of the adrenal gland glucocorticoid and catecholamine production is a fundamental component of the stress response and is essential for survival of the host. However, in many critically ill patients this homeostatic function of the adrenal gland is often impaired. In these patients, plasma levels of adrenocorticotropic hormone (ACTH) and cortisol are often dissociated. This has been attributed to the stimulatory action of non-ACTH factors within the adrenal gland such as cytokines, and recently with decreased cortisol metabolism and suppressed ACTH synthesis. Regulation of the hypothalamus-pituitary-adrenal (HPA) axis function during sepsis is a complex process which involves various immune and neuroendocrine interactions occurring at the levels of the central nervous system (CNS) and the adrenal gland. A coordinated interaction of numerous cell types and systems within the adrenal gland is involved in the sustained adrenal glucocorticoid production. This review article describes and discusses recent experimental findings regarding the role of adrenal gland microenvironment including the adrenal vasculature and the immune-adrenal crosstalk in the disregulated HPA axis during sepsis conditions. In summary, in addition to the reduced cortisol breakdown and related ACTH suppression, sepsis-mediated chronic activation of the immune-adrenal crosstalk and vascular dysfunction may contribute to the HPA axis dysregulation found in septic patients., (Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.)

Adrenal blood flow (ABF) is closely coupled to steroid hormone release. ACTH and angiotensin (Ang) II stimulate cortisol and aldosterone secretion; however, their effects on ABF remain poorly defined. We used the laser-Doppler technique to measure rat ABF. Anesthetized male Sprague-Dawley rats were cannulated for mean arterial pressure (MAP) measurement and drug infusion. The left adrenal gland was exposed for ABF measurement. ABF and MAP changes to ACTH and Ang II were determined. Bolus injections of Ang II (0.01-1000 ng/kg) increased ABF (maximal increase = 110 ± 18 perfusion units at 1000 ng/kg) and increased MAP at doses greater than 10 ng/kg (basal, 99.2 ± 1.4 mm Hg; 1000 ng/kg Ang II, 149.7 ± 3.9 mm Hg). ACTH (0.1-1000 ng/kg) increased ABF (maximum increase = 158 ± 33 perfusion units) without increasing MAP. ABF increases induced by Ang II and ACTH were ablated by the cytochrome 450 inhibitor miconazole (2 mg/kg). Bolus injections of endothelin-1 (1-1000 ng/kg) increased ABF only at 1 ng/kg and increased MAP at 1000 ng/kg. Bolus injections of sodium nitroprusside increased ABF at 1 and 10 μg/kg and decreased MAP at 10 μg/kg. Thus, laser-Doppler flowmetry is a useful tool for understanding ABF regulation by peptides that stimulate steroid hormone release. Our results demonstrate that Ang II and ACTH increases in ABF are mediated by a cytochrome P450 metabolite.

Adrenal vein sampling is used to establish the origins of excess production of adrenal hormones in primary aldosteronism. Correct catheter positioning is confirmed using adrenal vein measurements of cortisol, but this parameter is not always reliable. Plasma metanephrine represents an alternative parameter. The objective of our study was to determine the use of plasma metanephrine concentrations to establish correct catheter positioning during adrenal vein sampling with and without cosyntropin stimulation. We included 52 cosyntropin-stimulated and 34 nonstimulated sequential procedures. Plasma cortisol and metanephrine concentrations were measured in adrenal and peripheral venous samples. Success rates of sampling, using an adrenal to peripheral cortisol selectivity index of 3.0, were compared with success rates of metanephrine using a selectivity index determined by receiver operating characteristic curve analysis. Among procedures assessed as selective using cortisol, the adrenal to peripheral vein ratio of metanephrine was 6-fold higher than that of cortisol (94.0 versus 15.5; P<0.0001). There were significant positive relationships between adrenal to peripheral vein ratios of cortisol and metanephrine for cosyntropin-stimulated samplings but not for nonstimulated samplings. Receiver operating characteristic curve analysis indicated a plasma metanephrine selectivity index cutoff of 12. Using this cutoff, concordance in sampling success rates determined by cortisol and metanephrine was substantially higher in cosyntropin-stimulated than in nonstimulated samplings (98% versus 59%). For the latter procedures, sampling success rates determined by metanephrine were higher (P<0.01) than those determined by cortisol (91% versus 56%). In conclusion, metanephrine provides a superior analyte compared with cortisol in assessing the selectivity of adrenal vein sampling during procedures without cosyntropin stimulation.

Background: Metabolic syndrome is a common metabolic disorder that is associated with an increased risk of type 2 diabetes and cardiovascular diseases. Disturbances in adrenal steroid hormone production significantly contribute to the development of this disorder. Therefore, it is extremely important to fully understand the mechanisms governing adrenal gland function, both in physiological and pathological conditions., Results: Recently, Sonic hedgehog has emerged as an important regulator of adrenal development, with a possible role in adult gland homeostasis. Recent work of our group shows that lipoproteins are important regulators of Hedgehog signaling; they act as carriers for the spread of Hedgehog proteins, but also contain lipid(s) that inhibit the pathway., Conclusions: We propose that lipoproteins may affect Sonic hedgehog signaling in the adult adrenal gland at multiple levels. Understanding the interplay between lipoprotein metabolism and adrenal Hedgehog signaling may improve our understanding of how adrenal gland disorders contribute to the metabolic syndrome., (© 2013 Stichting European Society for Clinical Investigation Journal Foundation. Published by John Wiley & Sons Ltd.)

Systemic administration of endotoxin, which closely mimics the bacteria-induced systemic inflammatory response syndrome (SIRS) can ultimately lead to organ failure. Adrenal gland insufficiency is frequently diagnosed in critically ill patients; however, the underlying mechanisms are still unclear. In the present study, we studied comprehensively the characteristics of adrenal gland dysregulation, including inflammation, leukocyte infiltration and cell death in the adrenal glands in the course of LPS-induced systemic inflammation in mice. LPS enhanced expression of many proinflammatory cytokines, chemokines and adhesion molecules, which resulted in rapid recruitment of leukocytes into the adrenal gland. Furthermore, LPS-mediated inflammation was associated with increased apoptosis of adrenocortical and chromaffin cells. Our results performed in mice, suggest that LPS-induced adrenal gland inflammation and cell death might be mechanisms potentially involved in the adrenal gland dysfunction in patients with sepsis., (Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.)

Inflammation in the course of systemic inflammatory response syndrome (SIRS) or sepsis often results in dysregulation of the hypothalamic-pituitary-adrenal axis; however, the underlying mechanisms are not well understood. The adrenal gland is highly vascularized; thus, we hypothesized that endothelial dysfunction may actively participate in inflammation-related adrenal insufficiency. To address this hypothesis, we used the properties of developmental endothelial locus-1 (Del-1), which is an endothelial-derived anti-inflammatory factor that antagonizes integrin-dependent leukocyte adhesion. Here we identified that Del-1 is expressed in the adrenal gland and that its expression was down-regulated upon SIRS induction by systemic lipopolysaccharide administration. Furthermore, we observed increased leukocyte accumulation, inflammation, and higher apoptosis in the adrenal glands of Del-1-deficient mice as compared with wild-type mice. Strikingly, Del-1 deficiency was also associated with reduced corticosterone and ACTH levels 24 hours after lipopolysaccharide administration. Together, these data suggest that Del-1 may act as a gatekeeper of adrenal gland inflammation and may regulate the integrity of the hypothalamic-pituitary-adrenal axis stress response, thereby modulating adrenal (dys)function in the course of SIRS.

Patients with type 2 diabetes (T2D) manifest significant abnormalities in lipoprotein structure and function. The deleterious impact of oxidative and glycoxidative modifications on HDL-mediated atheroprotective, antiinflammatory, and antioxidative phenomena has been well established. However, the biological effects of modified HDL on adrenal steroidogenesis-which could reveal a pathophysiological link to the overactivity of the renin-angiotensin-aldosterone system and its adverse cardiovascular consequences often observed in T2D-are not well delineated. We studied the role of modified HDL on aldosterone release from adrenocortical carcinoma cells (NCI-H295R). In vitro modifications of native HDL were performed in the presence of glucose for glycoxidized HDL (glycoxHDL) and sodium hypochlorite for oxidized HDL. Angiotensin II (AngII)-sensitized H295R cells were treated with lipoproteins for 24 h, and supernatant was used to measure aldosterone release. Both native and modified HDL augmented the steroid release from AngII-sensitized cells, with glycoxHDL having the greatest impact. Both the modified forms of HDL induced a significant increase in scavenger receptor expression and employed protein kinase C as well as extracellular signal-regulated kinase as downstream effectors of aldosterone release. Native HDL and modified HDL required Janus kinase-2 for combating increased demand in steroidogenesis. Therefore, our data support the hypothesis that diabetes-induced modification of HDL may promote adrenocortical aldosterone secretion via different signal transduction pathways. This significant influence on multiple signaling mechanisms could be targeted for future research to implement novel therapeutic trials.

Our objective is to outline the utility of adrenal venous sampling (AVS) with measurements of metanephrine to normetanephrine ratios for diagnostic localization of phaeochromocytoma in a patient with normal plasma levels of catecholamines. A 53-year-old-woman was referred for evaluation of recurrent pheochromocytoma following a right adrenalectomy 14 years earlier. Diagnosis of recurrent disease was established from elevations in plasma metanephrines with normal levels of catecholamines. Magnetic resonance imaging indicated two 1-2 cm masses in the right surgical bed and another 1-1.5 cm mass in the left adrenal. These masses were negative on (123)I-metaiodobenzylguanidine scintigraphy. There was no evidence of a hereditary syndrome. We, therefore, carried out two investigational approaches to identify the tumor masses. Hybrid positron emission tomography/computed tomography with (68)Ga-DOTATOC ((68)Ga-DOTATOC-PET/CT) confirmed the presence of recurrent disease in the right surgical bed and also suggested additional left adrenal involvement. Normal plasma catecholamines precluded the use of adrenal venous sampling (AVS) with catecholamine measurements. Hence, we performed AVS with measurements of plasma metanephrines, which were 4- to 7-fold higher in the left adrenal vein than in central venous plasma. We observed a reversal of the normally high metanephrine to normetanephrine ratio (mean value ± SD 5.28 ± 1.86; range 3.36-8.84, n = 13) to 0.73, establishing the presence of a left adrenal pheochromocytoma. Surgical pathology confirmed bilateral disease. This case highlights a scenario where a combination of (68)Ga-DOTATOC-PET/CT and AVS with measurement of the metanephrine to normetanephrine ratio was crucial for the preoperative assessment of a patient with bilateral pheochromocytoma.

Context: Pheochromocytomas and paragangliomas (PPGLs) are diagnosed earlier in patients with hereditary than sporadic disease. Whether other factors influence age at diagnosis is unclear., Objective: We examined ages at which PPGLs were diagnosed according to different catecholamine phenotypes and locations of tumors., Design & Setting: Retrospective multicenter study., Patients: Patients with PPGLs included 172 with and 183 without identified germline mutations or hereditary syndromes. BIOCHEMICAL MEASUREMENTS: Differences in plasma concentrations of metanephrine, a metabolite of epinephrine, were used to distinguish epinephrine-producing tumors from those lacking epinephrine production., Results: Patients with epinephrine-producing tumors were diagnosed 11 yr later (P < 0.001) than those with tumors lacking appreciable epinephrine production. Among patients without evidence of a hereditary condition, those with and without epinephrine-producing tumors had respective mean ± se ages of 50 ± 2 and 42 ± 2 yr (P < 0.001) at diagnosis. Patients with multiple endocrine neoplasia type 2 and neurofibromatosis type 1, all with epinephrine-producing tumors, were similarly diagnosed with disease at a later age than patients with tumors that lacked appreciable epinephrine production secondary to mutations of von Hippel-Lindau and succinate dehydrogenase genes (40 ± 2 vs. 31 ± 1 yr, P < 0.001). Among the latter patients, those with multifocal tumors were diagnosed earlier than those with solitary tumors (19 ± 3 vs. 34 ± 2 yr, P < 0.001)., Conclusions: The variations in ages at diagnosis associated with different tumor catecholamine phenotypes and locations suggest origins of PPGLs from different chromaffin progenitor cells with variable susceptibility to disease causing mutations. Different optimal age cut-offs for mutation testing are indicated for patients with and without epinephrine-producing tumors (44-49 vs. 30-35 yr, respectively).

The adrenal gland is formed by the adrenal medulla and the adrenal cortex. Both tissues descend from different origins during embryonal development. While the chromaffin cells are derived from the neural crest, the adrenocortical cells stem from a cell condensation in the celomic epithelium. Already during adrenal organogenesis, close interactions between the two tissue types are necessary for the differentiation, morphogenesis and survival of the adrenal gland. Moreover, the communication between the chromaffin and adrenocortical cells ensures a regular function of the adult adrenal gland including the regulation of hormone synthesis and responses to stress. This is even more important since the cortical-chromaffin crosstalk is also relevant for the pathogenesis of different diseases. In the past decade, significant progress in the understanding of the cortical-chromaffin communication has been made. Here, we summarize the insights gained from in vitro studies, from animal models and from clinical observations., (Copyright © 2011 S. Karger AG, Basel.)

Background: Serum aldosterone is a causative factor for various metabolic and cardiovascular disorders. Low-density lipoprotein (LDL) is a major cholesterol source for aldosterone steroidogenesis; however, the effect of oxidative modification of LDL on aldosterone release is not known. We studied the effect of hypochlorite-oxidized LDL (oxLDL) on adrenal aldosterone secretion., Methods: LDL (native LDL (natLDL)) was obtained from healthy volunteers and oxidatively modified in vitro. NCI-H295R cells were stimulated with natLDL and oxLDL, and the aldosterone release was quantified by radioimmunoassay. Molecular changes were studied with western blot analysis and quantitative RT-PCR analysis., Results: NatLDL and oxLDL caused dose-dependent increase in aldosterone release up to threefold. However, the stimulatory effects of modified LDL on aldosterone secretion decreased with increasing degree of LDL oxidation. 24-h incubations with natLDL, mild- and medium-oxidized LDL sensitized the adrenocortical cells to subsequent angiotensin II (Ang II) stimulations by 2.9-, 2.8-, and 2.5-folds, respectively. Heavily oxidized LDL did not sensitize the cells to Ang II stimulations to a similar extent. At the molecular level, the ERK pathway was activated within a minute by both natLDL and oxLDL; however, oxLDL showed a stronger (2.75-fold at 1 and 15 min) and longer (15 min) activation of ERK than natLDL (twofold)., Conclusions: This study demonstrates the following: (i) both natLDL and hypochlorite-oxidized LDL utilize ERK pathway to mediate aldosterone release; (ii) mildly oxidized LDL sensitizes the adrenocortical cells to further stimulations by Ang II similar to natLDL that may have a role in pathological processes; (iii) extensive LDL oxidation counteracts adrenocortical aldosterone release.

Orthotopic cell transplantation models are important for a complete understanding of cell-cell interactions as well as tumor biology. In published studies of orthotopic transplantation in the mouse adrenal gland, human neuroblastoma cells have been shown to invade and occupy the adrenal, but in these investigations a true orthotopic model was not established. Here we show an orthotopic model in which transplanted cells are retained within the adrenal gland by formation of a fibrin clot. To establish an appropriate technique, we used brightly fluorescent 10 microm polystyrene microspheres injected into the mouse adrenal gland. In the absence of fibrinogen/thrombin for clot formation, much of the injected material was extruded to the outside of the gland. When the microspheres were injected in a fibrinogen/thrombin mixture, fluorescence was confined to the adrenal gland. As a model neoplastic cell originating from the cortex of the gland, we used a tumorigenic bovine adrenocortical cell line. When 3 x 10(5) cells were implanted orthotopically, by 16 days the cell mass had expanded and had invaded the cortex, whereas when 1 x 10(5) cells were used, tumor masses were much smaller. We therefore subsequently used 3 x 10(5) cells. When mice were sacrificed at different time points, we found that tumor growth resulting was progressive and that by 26 days cells there was extensive invasion into the cortex or almost complete replacement of the cortex with tumor cells. As a model neoplastic cell of neural crest origin, we used SK-N-AS human neuroblastoma cells. Orthotopic transplantation of 3 x 10(5) cells resulted in extensive invasion and destruction of the gland by 26 days. In summary, the present orthotopic model for intra-adrenal cell transplantation is valuable for investigation of growth of neoplastic cells of both cortical and medullary origin and should be useful for future studies of cortex-medulla interactions.

The adrenal gland contains a well-organized network of blood vessels, and adrenocortical cells are situated in close proximity to endothelial cells. Recently, several new mechanisms have been characterized that control the release of aldosterone by adrenocortical cells, including the involvement of endothelial-cell-derived factors. Interestingly, a CBP/p300-interacting transactivator with ED-rich tail 2 (CITED2), which is necessary for adrenal development, has been linked to aldosterone synthesis. We have therefore examined the effects of endothelial-cell-conditioned medium (ECCM), as produced during the incubation of human umbilical vein endothelial cells for 24 h, on the promoter activity and mRNA and protein expression of CITED2 in adrenocortical cells as represented by the NCI-H295R cell line. We have found a dose-dependent effect of ECCM on CITED2 promoter activity; this peaks at 480%. Activation of the CITED2 promoter occurs in parallel to an increase in CITED2 messenger RNA (as quantified by real-time polymerase chain reaction) and protein. The stimulatory effect of ECCM can be reversed by blocking mitogen-activated protein kinase activity with the MEK1-inhibitor PD98059. We conclude that products secreted by endothelial cells control not only steroidogenesis, but also factors that are important for adrenocortical development, thereby highlighting the role of cellular interactions within adrenocortical development and physiology.

Decrease in connexin (Cx) expression and loss of gap junctional intercellular communication (GJIC) have been associated with aberrant cell growth and enhanced neoplastic phenotype. We studied the expression of Cx26, Cx32, Cx43, and Cx50 in chromaffin cells of 10 normal human adrenal glands, 10 benign, and 13 malignant pheochromocytomas. Immunohistochemistry showed that Cx50 expression seemed to be the predominant form of Cx expressed in human chromaffin cells, whereas Cx43 immunoreactivity was the most prominent form found in the adrenal cortex. However, Western blotting revealed that nonphosphorylated and singly and doubly phosphorylated forms of Cx43 were present within the malignant adrenomedullary cells at the same time. Cx26 and Cx32 were distributed inhomogeneously with no emphasis of expression in the types of tissues studied. Cx50 expression was diminished in malignant pheochromocytomas. We conclude that there is a differential expression of Cx in human adrenals. Immunohistological testing for Cx expression does not however allow differentiation of benign from malignant pheochromocytomas.

Sepsis and septic shock are leading killers in the noncoronary intensive care unit, and they remain worldwide health concerns. The initial host defense against bacterial infections involves Toll-like receptors (TLRs), which detect and respond to microbial ligands. In addition, a coordinated response of the adrenal and immune systems is crucial for survival during severe inflammation. Previously, we demonstrated a link between the innate immune system and the endocrine stress response involving TLR-2. Like TLR-2, TLR-4 is also expressed in human and mouse adrenals. In the present work, by using a low dose of LPS to mimic systemic inflammatory response syndrome, we have revealed marked cellular alterations in adrenocortical tissue and an impaired adrenal corticosterone response in TLR-4-/- mice. Our findings demonstrate that TLR-4 is a key mediator in the crosstalks between the innate immune system and the endocrine stress response. Furthermore, TLR polymorphisms could contribute to the underlying mechanisms of impaired adrenal stress response in patients with bacterial sepsis.

Ascorbic acid (vitamin C) is a cofactor required in catecholamine synthesis for conversion of dopamine to norepinephrine by dopamine beta-hydroxylase. Mutant mice lacking the plasma membrane ascorbic acid transporter (SVCT2) have severely reduced tissue levels of ascorbic acid and die after birth. We therefore investigated whether these mice might have impaired synthesis of catecholamines. Levels of catecholamines in brain were unaffected by SVCT2 deficiency. In heart, the only evidence for impaired dopamine beta-hydroxylase activity was a twofold increase in tissue dopamine. An influence of the deficiency on tissue catecholamines was most prominent in the adrenals where norepinephrine was decreased by 50% and epinephrine, by 81%. On the ultrastructural level, adrenal chromaffin cells in SVCT2 null mice showed depletion of catecholamine storage vesicles, increased amounts of rough endoplasmic reticulum, signs of apoptosis, and increased glycogen storage. Decreased plasma levels of corticosterone indicated additional effects of the deficiency on adrenal cortical function. These data show that deranged catecholamine system function in SVCT2 null mice is largely restricted to the adrenal medulla and cannot account for the lethality in these animals. The data, however, establish a crucial role for ascorbic acid in adrenal chromaffin cell function.

The hypothalamic-pituitary-adrenal (HPA) axis is integrated in the human stress system and controls the metabolism of many cell systems in the body. Therefore, hypofunction or hyperfunction of the HPA axis potentially threatens the life of the whole organism. Noncontrolled overproduction of its key regulators, CRH and ACTH, causes dysfunction of the stress system. Ectopic secretion of these compounds may be part of extraadrenal paraneoplastic syndromes caused by various benign or malignant tumors. However, ectopic ACTH and CRH may originate from the adrenal itself. A local CRH/ACTH system exists in the normal human adrenal medulla. Overproduction of CRH and ACTH has been documented in pheochromocytomas causing Cushing's syndrome. Finally, ectopic production of ACTH causing Cushing's syndrome has also been demonstrated in adrenocortical cells. This suggests a marked plasticity within the HPA axis and the neuroendocrine cell system., (Copyright 2003 Wiley-Liss, Inc.)

Adrenocortical carcinoma (ACC) and pheochromocytoma (PCC) are malignancies originating from distinct layers of the adrenal gland. ACCs arise from the adrenal cortex, are often detected at advanced stages and are associated with poor prognosis. PCCs are mostly benign, arise from the adrenal medulla and have a variable prognosis, with 10% of PCCs resulting in metastasis. Genetic background strongly influences metastasis of PCCs, and no reliable biomarkers that predict metastatic behavior exist to date. Current therapeutic strategies for both ACCs and PCCs are overall limited. Thus, novel preclinical models and drug screening approaches need to be established to aid in the identification of more promising drugs and treatment schemes. In this review, we summarize the currently available human and murine cell lines for both tumor entities. [ABSTRACT FROM AUTHOR]

Intraportal islet transplantation in patients with type 1 diabetes enables restoration of glucose-regulated insulin secretion. However, several factors hamper a widespread application and long-term success: chronic hypoxia, an inappropriate microenvironment and suppression of regenerative and proliferative potential by high local levels of immunosuppressive agents. Therefore, the identification of alternative and superior transplant sites is of major scientific and clinical interest. Here, we aim to evaluate the adrenal as an alternative transplantation site. The adrenal features a particular microenvironment with extensive vascularization, anti-apoptotic and pro-proliferative, anti-inflammatory and immunosuppressive effects. To validate this novel transplantation site, an in vitro co-culture system of adrenal cells and pancreatic islets was established and viability, islet survival, functional potency and antioxidative defense capacity were evaluated. For in vivo validation, an immune-deficient diabetic mouse model for intra-adrenal islet transplantation was applied. The functional capacity of intra-adrenally grafted islets to reverse diabetes was compared to a standard islet transplant model and measures of engraftment such as vascular integration were evaluated. The presence of adrenal cells positively impacted on cell metabolism and oxidative stress. Following transplantation, we could demonstrate enhanced islet function in comparison to standard models with improved engraftment and superior re-vascularization. This experimental approach allows for novel insights into the interaction of endocrine systems and may open up novel strategies for islet transplantation augmented through the bystander effect of other endocrine cells or the active factors secreted by adrenal cells modulating the microenvironment. [ABSTRACT FROM AUTHOR]

Primary adrenal insufficiency is a life‐threatening disorder, which requires lifelong hormone replacement therapy. Transplantation of xenogeneic adrenal cells is a potential alternative approach for the treatment of adrenal insufficiency. For a successful outcome of this replacement therapy, transplanted cells should provide adequate hormone secretion and respond to adrenal physiological stimuli. Here, we describe the generation and characterization of primary porcine adrenal spheroids capable of replacing the function of adrenal glands in vivo. Cells within the spheroids morphologically resembled adult adrenocortical cells and synthesized and secreted adrenal steroid hormones in a regulated manner. Moreover, the embedding of the spheroids in alginate led to the formation of cellular elongations of steroidogenic cells migrating centripetally towards the inner part of the slab, similar to zona Fasciculata cells in the intact organ. Finally, transplantation of adrenal spheroids in adrenalectomized SCID mice reversed the adrenal insufficiency phenotype, which significantly improved animals' survival. Overall, such adrenal models could be employed for disease modeling and drug testing, and represent the first step toward potential clinical trials in the future. [ABSTRACT FROM AUTHOR]

In a number of adult tissues, Nestin-positive stem cells/progenitors have been identified and shown to be involved in maintenance and remodeling. Various studies have shown that under stressful conditions, quiescent Nestin-positive progenitor cells are activated. Thereby, they migrate to their target location and differentiate into mature cells. In the current paper, we discuss if Nestin-positive progenitors in the hippocampus and adrenal gland belong to unique cell populations that are responsive to stress. Furthermore, we speculate about the mechanism behind their activation and the clinical importance of this stress-response. [ABSTRACT FROM AUTHOR]

Bacterial sepsis is a serious threat to the body homeostasis and is often associated with high mortality in non-coronary intensive stations. In order to survive sepsis, rapid activation of the hypothalamus-pituitary-adrenal gland axis and sympathomedullary system is necessary. In many patients with sepsis, the function of those two arms of the stress system is dysregulated with underlying mechanisms remaining unknown. In our previous experimental studies, we have demonstrated that LPS-induced systemic inflammation and CLP-induced peritonitis can result in adrenal gland damage. Histological and transcriptomic analysis revealed a potential involvement of the adrenal microvascular endothelium in this process. However, our knowledge about the function of adrenal microvascular cells during sepsis is scarce. In the present study, we have characterized transcriptomic alterations in isolated mouse adrenal microvascular endothelial cells induced by systemic administration of bacterial LPS. Our results revealed that LPS induced a distinct transcriptomic profile in the adrenal microvascular cells, including multiple genes regulating inflammation, activation of the coagulation cascade and vascular permeability. Activation of those genes may be potentially involved in the damage to the microvascular endothelium and altogether contribute to the sepsis-mediated adrenal dysregulation. [ABSTRACT FROM AUTHOR]

The adrenal gland is a master regulator of the human body during response to stress. This organ shows constant replacement of senescent cells by newly differentiated cells. A high degree of plasticity is critical to sustain homeostasis under different physiological demands. This is achieved in part through proliferation and differentiation of adult adrenal progenitors. Here, we report the isolation and characterization of a Nestin+ population of adrenocortical progenitors located under the adrenal capsule and scattered throughout the cortex. These cells are interconnected with progenitors in the medulla. In vivo lineage tracing revealed that, under basal conditions, this population is noncommitted and slowly migrates centripetally. Under stress, this migration is greatly enhanced, and the cells differentiate into steroidogenic cells. Nestin+ cells cultured in vitro also show multipotency, as they differentiate into mineralocorticoid and glucocorticoid-producing cells, which can be further influenced by the exposure to Angiotensin II, adrenocorticotropic hormone, and the agonist of luteinizing hormone-releasing hormone, triptorelin. Taken together, Nestin+ cells in the adult adrenal cortex exhibit the features of adrenocortical progenitor cells. Our study provides evidence for a role of Nestin+ cells in organ homeostasis and emphasizes their role under stress. This cell population might be a potential source of cell replacement for the treatment of adrenal insufficiency. [ABSTRACT FROM AUTHOR]

Discusses advances in the diagnosis and clinical management of adrenocortical tumors. Intercellular communication in the adrenal gland and tumors; Molecular genetics of the tumors; Endocrine testing; Imaging procedures; Laparoscopic adrenalectomy; Surgical resection.

A letter to the editor is presented regarding "Adrenal Lymphocytic Infiltration and Adrenocortical Tumors in a Patient with 21-Hydroxylase Deficiency," by Deborah P. Merke, Stefan R. Bornstein, Demetrios Braddock and George P. Chrousos.

The mammalian adrenal gland is composed of two distinct tissue types in a bidirectional connection, the catecholamine-producing medulla derived from the neural crest and the mesoderm-derived cortex producing steroids. The medulla mainly consists of chromaffin cells derived from multipotent nerve-associated descendants of Schwann cell precursors. Already during adrenal organogenesis, close interactions between cortex and medulla are necessary for proper differentiation and morphogenesis of the gland. Moreover, communication between the cortex and the medulla ensures a regular function of the adult adrenal. In tumor development, interfaces between the two parts are also common. Here, we summarize the development of the mammalian adrenal medulla and the current understanding of the cortical-medullary interactions under development and in health and disease. • Development of the adrenal medulla is influenced by the cortex. • Chromaffin cells in the adrenal medulla are derived from Schwann cell precursors. • Adrenal pathologies often involve both the adrenal cortex and medulla. [ABSTRACT FROM AUTHOR]

• RNA-Seq analysis was used to investigate adrenal gland specific transcriptomic signatures altered by LPS. • LPS strongly upregulates the gene sets involved in inflammation, hypoxia and steroidogenesis. • LPS highly inhibits the pathways involved in Wnt/Shh signaling in the adrenal gland. • LPS-mediated alterations in the adrenal gland transcriptome found by RNA-seq were validated in CLP-induced sepsis model. • RNA-Seq analysis is a useful method to study the novel pathways of sepsis-induced adrenal dysfunction. Activation of the adrenal gland stress response is of utmost importance to survive sepsis. Experimental and clinical evidence exists demonstrating that adrenal gland often develops functional and structural damage due to sepsis with mechanisms remaining largely unknown. In the present study, we have used RNA Sequencing (RNA-Seq) technology to analyze changes in adrenal transcriptome elucidated by bacterial LPS. We aimed to find particularly alterations in genes that were previously not reported to be involved in the adrenal gland dysregulation in contexts of sepsis. Our results demonstrate that systemic administration of LPS significantly altered expression of 8458 genes as compared to saline injected animals. The subsequent quality and functional analysis of these gene signatures revealed that LPS-induced highly homogenous transcriptional response in total upregulating 4312 and downregulating 4146 genes. Furthermore, functional annotation analysis together with gene enrichment set analysis (GSEA) clearly demonstrated that adrenal response to LPS involved alterations in multiple pathways related to the inflammatory response along with previously unexplored activation of the hypoxia pathway. In addition, LPS strongly downregulated genes involved in the adrenal homeostasis, development, and regeneration. Those alterations were subsequently verified in clinically relevant cecal ligation and puncture (CLP)-induced sepsis model. Collectively, our study demonstrates that RNA-seq is a very useful method that can be applied to search for new unexplored pathways potentially involved in adrenal gland dysregulation during sepsis. [ABSTRACT FROM AUTHOR]