Objective
To comprehensively characterize androgens and androgen precursors in classic 21-hydroxylase deficiency (21OHD) and to gain insight to the mechanisms of their formation.
Design
Serum samples were obtained from 38 patients (19 men) with classic 21OHD, age 3-59, and 38 sex- and age-matched controls; 3 patients with 11β-hydroxylase deficiency; 4 patients with adrenal insufficiency; and 16 patients (8 men) undergoing adrenal vein sampling. Paraffin-embedded normal (n=5) and 21OHD adrenal tissue (n=3) was used for immunohistochemical studies.
Methods
We measured 11 steroids in all sera using liquid chromatography-tandem mass spectrometry. Immunofluroescence localized 3β-hydroxysteroid dehydrogenase type 2 (HSD3B2) and cytochrome b5 (CYB5A) within the normal and 21OHD adrenals.
Results
Four 11-oxygenated 19-carbon (11oxC19) steroids were significantly higher in male and female 21OHD patients than in controls: 11β-hydroxyandrostenedione, 11-ketoandrostenedione 11β-hydroxytestosterone, and 11-ketotestosterone (3-4-fold, p< 0.0001). For 21OHD patients, testosterone and 11-ketotestosterone were positively correlated in females, but inversely correlated in males. All 11oxC19 steroids were higher in adrenal vein than in inferior vena cava samples from men and women and rose with cosyntropin stimulation. Only trace amounts of 11oxC19 steroids were found in sera from patients with 11β-hydroxylase deficiency and adrenal insufficiency, confirming their adrenal origin. HSD3B2 and CYB5A immunoreactivities were sharply segregated in the normal adrenal glands, whereas areas of overlapping expression were identified in the 21OHD adrenals.
Conclusions
All four 11oxC19 steroids are elevated in both men and women with classic 21OHD. Our data suggest that 11oxC19 steroids are specific biomarkers of adrenal-derived androgen excess.
Pancreatic β-cells couple the oxidation of glucose to the secretion of insulin. Apart from the canonical KATP-dependent glucose-stimulated insulin secretion (GSIS), there are important KATP-independent mechanisms involving both anaplerosis and mitochondrial GTP (mtGTP). How mtGTP that is trapped within the mitochondrial matrix regulates the cytosolic calcium increases that drive GSIS remains a mystery. Here we have investigated whether the mitochondrial isoform of phosphoenolpyruvate carboxykinase (PEPCK-M) is the GTPase linking hydrolysis of mtGTP made by succinyl-CoA synthetase (SCS-GTP) to an anaplerotic pathway producing phosphoenolpyruvate (PEP). Although cytosolic PEPCK (PEPCK-C) is absent, PEPCK-M message and protein were detected in INS-1 832/13 cells, rat islets, and mouse islets. PEPCK enzymatic activity is half that of primary hepatocytes and is localized exclusively to the mitochondria. Novel 13C-labeling strategies in INS-1 832/13 cells and islets measured substantial contribution of PEPCK-M to the synthesis of PEP. As high as 30% of PEP in INS-1 832/13 cells and 41% of PEP in rat islets came from PEPCK-M. The contribution of PEPCK-M to overall PEP synthesis more than tripled with glucose stimulation. Silencing the PEPCK-M gene completely inhibited GSIS underscoring its central role in mitochondrial metabolism-mediated insulin secretion. Given that mtGTP synthesized by SCS-GTP is an indicator of TCA flux that is crucial for GSIS, PEPCK-M is a strong candidate to link mtGTP synthesis with insulin release through anaplerotic PEP cycling.
Comprehensive NGS of CYP11B2-expressing adrenal tumors identified somatic mutations in aldosterone-driving genes in 88% of APAs, a higher rate than in previous studies using conventional approaches.
Synopsis
Adrenal steroidogenesis is a dynamic process, reliant on de novo synthesis from cholesterol, under the stimulation of ACTH and other regulators. The syntheses of mineralocorticoids, glucocorticoids and adrenal androgens occur in separate adrenal cortical zones, each expressing specific enzymes. Congenital adrenal hyperplasia (CAH) encompasses a group of autosomal recessive enzymatic defects in cortisol biosynthesis. 21-hydroxylase (21OHD) deficiency accounts for over 90% of CAH cases and when milder or nonclassic forms are included, 21OHD is one of the most common genetic diseases. This review discusses in detail the epidemiology, genetics, diagnostic, clinical aspects and management of 21OHD.
Findings on MRI remain key in guiding the diagnosis of pituitary stalk lesions, particularly when used in conjunction with other clinical clues. There are no good imaging predictors for hypopituitarism, making clinical evaluation of all patients with pituitary stalk lesions crucial.
Tg-IAs remain the method of choice for Tg quantitation in TgAb- patients. In TgAb+ patients with undetectable Tg by immunometric assay, the Tg-MS will detect Tg in up to 20% additional cases. The Tg-RIA will detect Tg in approximately 35% cases, but a significant proportion of these will be clinical false-positive results. The undetectable Tg-MS seen in approximately 40% of TgAb+ cases in patients with disease need further evaluation.
Our findings suggest that 11KT is the dominant bioactive androgen in children during adrenarche and PremA. Its androgenic capacity suggests that it may be responsible for the phenotypic changes seen in these phenomena.
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