Search

Context: Chronic glucocorticoid (GC) overexposure, resulting from endogenous Cushing's syndrome (CS) or exogenous GC therapy, causes several adverse outcomes, including persistent central fat accumulation associated with a low-grade inflammation. However, no previous multiomics studies in visceral adipose tissue (VAT) from patients exposed to high levels of unsuppressed GC during active CS or after remission are available yet., Objective: To determine the persistent VAT transcriptomic alterations and epigenetic fingerprints induced by chronic hypercortisolism., Methods: We employed a translational approach combining high-throughput data on endogenous CS patients and a reversible CS mouse model. We performed RNA sequencing and chromatin immunoprecipitation sequencing on histone modifications (H3K4me3, H3K27ac, and H3K27me3) to identify persistent transcriptional and epigenetic signatures in VAT produced during active CS and maintained after remission., Results: VAT dysfunction was associated with low-grade proinflammatory status, macrophage infiltration, and extracellular matrix remodeling. Most notably, chronic hypercortisolism caused a persistent circadian rhythm disruption in VAT through core clock genes modulation. Importantly, changes in the levels of 2 histone modifications associated to gene transcriptional activation (H3K4me3 and H3K27ac) correlated with the observed differences in gene expression during active CS and after CS remission., Conclusion: We identified for the first time the persistent transcriptional and epigenetic signatures induced by hypercortisolism in VAT, providing a novel integrated view of molecular components driving the long-term VAT impairment associated with CS., (© The Author(s) 2021. Published by Oxford University Press on behalf of the Endocrine Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Introduction: Urine free cortisol measurements are routinely performed to evaluate hypercortisolism. Despite their analytical inaccuracy, immunoassay-based methods are frequently used. Advances in liquid chromatography–high-resolution mass spectrometry (LC-HRMS) facilitate the incorporation of powerful diagnostic tools into clinical laboratories. In addition to its high analytical specificity and simultaneous analysis of different metabolites, accurate mass measurement allows for untargeted compound identification, which may help to identify clinically relevant metabolites or drugs. Methods: The present study aimed to validate a simple routine LC–HRMS method to quantify cortisol, cortisone, 6β-hydroxycortisol, and 18-hydroxycortisol simultaneously in human urine. Additionally, the study also validated a GC-MS method for the same steroids, evaluated their cross-reactivity with commercial cortisol immunoassays, and quantified the 24 h urine excretion in patients under clinical suspicion or follow-up for hypercortisolism. Results: The LC-HRMS method involved liquid–liquid extraction using dichloromethane, micro-LC for chromatographic separation and detection using the accurate masses of the steroids, and simultaneous high-resolution full scan acquisition. The method presented acceptable linearity, precision, and accuracy. Significant interference from 6β-hydroxycortisol and cortisone was demonstrated in the cortisol immunoassays, which impacted their reliability in the follow-up of patients with hypercortisolism and significant changes in these cortisol metabolites (i.e., due to drug-induced changes in CYP3A4 activity). Conclusion: A rapid and accurate routine LC-HRMS method was validated, which is useful for the evaluation of hypercortisolism and other disorders of glucocorticoid and mineralocorticoid metabolism. [ABSTRACT FROM AUTHOR]

Glucocorticoids (GCs) play critical roles in adipose tissue metabolism. Here, we compare in a mouse model the effects of chronic glucocorticoid excess and diet-induced obesity on white adipose tissue mass and distribution, by focusing on visceral adipose tissue (VAT) fatty acid composition changes, the role of de novo lipogenesis (DNL) and the inflammatory state. We used a noninvasive mouse model of hypercortisolism to compare GC-induced effects on adipose tissue with diet-induced obesity [high-fat diet (HFD) 45%] and control mice after 10 wk of treatment. Subcutaneous adipose tissue (SAT) and VAT mass and distribution were measured by nuclear magnetic resonance imaging (NMRI). Fatty acid composition in VAT was analyzed by NMR spectroscopy and gas chromatography. Gene expression of key enzymes involved in DNL was analyzed in liver and VAT. Macrophage infiltration markers and proinflammatory cytokines were measured by gene expression in VAT. HFD or GC treatment induced similar fat mass expansion with comparable distribution between SAT and VAT depots. However, in VAT, GCs induce DNL, higher palmitic acid (PA), macrophage infiltration, and proinflammatory cytokine levels, accompanied by systemic nonesterified fatty acid (NEFA) elevation, hyperinsulinemia, and higher homeostatic model assessment for insulin resistance (HOMA-IR) levels compared with diet-induced obesity. Thus, chronic hypercortisolism induces DNL and fatty acid composition changes toward increased SFA and reduced polyunsaturated fatty acid (PUFA) levels in VAT, promoting macrophage recruitment and proinflammatory cytokines, suggesting a worse cardiometabolic profile even compared with HFD mice. [ABSTRACT FROM AUTHOR]

Excessive and prolonged glucocorticoid (GC) exposure, resulting from either prescribed or endogenous hypercortisolism, is associated with a high cardiovascular and metabolic burden (Cushing's syndrome). Although previous studies in humans and mice have reported heterogeneous data about the persistence of metabolic syndrome features after remission of hypercortisolism, there is still controversy as to whether this is due to the deleterious changes induced by GCs during active disease or the result of various other factors interfering in the recovery period. In order to s tudy metabolic effects after remission, we used a reversible mouse model of corticosterone (CORT) (100 µg/mL) administration in drinking water for 5 weeks, followed by a 10- week recovery period. We compared CORT-induced effects at these time points with a hig h-fat diet-treated group (HFD 45%) and a vehicle group (VEH). Plasma CORT, 11ß-HSD activity, food intake, glucose levels, interscapular brown adiposity, hepatic triglycerides and muscle mass were found altered during CORT treatment but normalized after recovery. Although hyperinsulinemia and insulin resistance were increased during CORT and HFD treatment, insulin homeostasis remained altered following the recovery period only in CORT-treated mice. Subcutaneous and visceral adipose tissues (SAT and VAT) were enlarged during HFD and CORT treatment as measured by MRI. However, increased muscle lipid content, adiposity and macrophage infiltration in VAT were only present in the CORT group following recovery. Taken together, CORT-induced insulin alterations were more potent than HFD-induced ones during the same period of treatment, and also more persistent long term. Moreover, we demonstrated that CORT treatment induces more long-lasting VAT enlargement than HFD. [ABSTRACT FROM AUTHOR]

Glucocorticoids (GC) induce cardiometabolic risk while atherosclerosis is a chronic inflammation involving immunity. GC are immune suppressors, and the adrenocorticotrophic hormone (ACTH) has immune modulator activities. Both may act in atherothrombotic inflammation involving immune cells (IMNC). Aim. To investigate adhesion and activation surface cell markers (CDs) of peripheral IMNC in endogenous Cushing syndrome (CS) and the immune modulator role of ACTH. Material and Methods. 16 ACTH-dependent CS (ACTH-D), 10 ACTH-independent (ACTH-ID) CS, and 16 healthy controls (C) were included. Leukocytes (Leuc), monocytes (MN), lymphocytes (Lym), and neutrophils (N) were analyzed by flow cytometry for atherosclerosis previously associated with CDs. Results. Leuc, N, and MN correlated with CS (p<0.05), WC (p<0.001), WHR (p=0.003), BMI (p<0.001), and hs-CRP (p<0.001). CD14++CD16+ (p=0.047); CD14+CD16++ (p=0.053) MN; CD15+ (p=0.027); CD15+CD16+ (p=0.008) N; and NK-Lym (p=0.019) were higher in CS. CD14+CD16++ MN were higher in ACTH-ID (8.9 ± 3.5%) versus ACTH-D CS (4.2 ± 1.9%) versus C (4.9 ± 2.3%). NK-Lym correlated with c-LDL (r = 0.433, p=0.039) and CD15+ N with hs-CRP (r = 0.446, p=0.037). In multivariate analysis, Leuc, N, and MN depended on BMI (p=0.021), WC (p=0.002), and WHR (p=0.014), while CD15+ and CD15+CD16+ N on hypercortisolism and CS (p=0.035). Conclusion. In CS, IMNC present changes in activation and adhesion CDs implicated in atherothrombotic inflammation. ACTH-IDCS presents a particular IMNC phenotype, possibly due to the absence of the immune modulator effect of ACTH. [ABSTRACT FROM AUTHOR]