Persistent hypercalcaemia associated with two pathogenic variants in the CYP24A1 gene and a parathyroid adenoma-a case report and review.

Introduction: 24-Hydroxylase, encoded by the CYP24A1 gene, is a crucial enzyme involved in the catabolism of vitamin D. Loss-of-function mutations in CYP24A1 result in PTH-independent hypercalcaemia with high levels of 1,25(OH)2D3. The variety of clinical manifestations depends on age, and underlying genetic predisposition mutations can lead to fatal infantile hypercalcaemia among neonates, whereas adult symptoms are usually mild. Aim of the study: We report a rare case of an adult with primary hyperparathyroidism and loss-of-function mutations in the CYP24A1 gene and a review of similar cases. Case presentation: We report the case of a 58-year-old woman diagnosed initially with primary hyperparathyroidism. Preoperatively, the suspected mass adjoining the upper pole of the left lobe of the thyroid gland was found via ultrasonography and confirmed by 99mTc scintigraphy and biopsy as the parathyroid gland. The patient underwent parathyroidectomy (a histopathology report revealed parathyroid adenoma), which led to normocalcaemia. After 10 months, vitamin D supplementation was introduced due to deficiency, and the calcium level remained within the reference range. Two years later, biochemical tests showed recurrence of hypercalcaemia with suppressed parathyroid hormone levels and elevated 1,25(OH)2D3 concentrations. Further investigation excluded the most common causes of PTH-independent hypercalcaemia, such as granulomatous disease, malignancy, and vitamin D intoxication. Subsequently, vitamin D metabolites were measured using LC-MS/MS, which revealed high levels of 25(OH)D3, low levels of 24,25(OH)2D3 and elevated 25(OH)2D3/24,25(OH)2D3 ratios, suggesting a defect in vitamin D catabolism. Molecular analysis of the CYP24A1 gene using the NGS technique revealed two pathogenic variants: p.(Arg396Trp) and p.(Glu143del) (rs114368325 and rs777676129, respectively). Conclusions: The diagnostic process for hypercalcaemia becomes complicated when multiple causes of hypercalcaemia coexist. The measurement of vitamin D metabolites using LC-MS/MS may help to identify carriers of CYP24A1 mutations. Subsequent molecular testing may contribute to establishing the exact frequency of pathogenic variants of the CYP24A1 gene and introducing personalized treatment.

Musculoskeletal complications of Cushing syndrome.

Prolonged exposure to an excess of glucocorticosteroids (GCs), both endogenous and exogenous, leads to a wide range of comorbidities, including cardiovascular, metabolic, psychiatric, and musculoskeletal disorders. The latter comprise osteopenia and osteoporosis leading to skeletal fractures and myopathy. Although endogenous hypercortisolemia is a rare disorder, GCs are among the most frequently prescribed drugs, often administered chronically and despite multiple side effects, impossible to taper off due to therapeutic reasons. The pathophysiology of the effect of GC excess on bone often leads to fractures despite normal or low-normal bone mineral density and it includes direct (mainly disturbance in bone formation processes, through inactivation of the Wnt/ß-catenin signalling pathway) and indirect mechanisms (through suppressing the gonadal and somatotrophic axis, and also through antagonizing vitamin D actions). Glucocorticosteroid-induced fast-twitch, glycolytic muscles atrophy occurs due to increased protein catabolism and impaired synthesis. Protein degradation is a result of activation of the ubiquitin proteasome and the lysosomes stimulated through overexpression of several atrogenes (such as FOXO-1 and atrogin-1). This review will discuss pathophysiology, clinical presentation, prevention, and management of GC-induced osteoporosis (including calcium and vitamin D supplementation, and bisphosphonates) and myopathy associated with GC excess.

Determination of vitamin D status in singleton and twin gestations using CLIA and LC-MS/MS.

Objective: Vitamin D plays an important role during pregnancy. The aim was to compare vitamin D status in a group of singleton (SP) and twin pregnancies (TP) using two diagnostic methods: chemiluminescence immunoassay (CLIA) and liquid chromatography with tandem mass spectrometry (LC-MS/MS). Design: This is a cross-sectional study. Methods: The study was conducted in the population of SP and TP at the gestational age above 20 + 0 at the Bielanski Hospital in Warsaw, Poland, between October 2020 and January 2023. All patients had their venous blood samples collected and were given an original survey containing questions on demography and vitamin D supplementation. Results: The study group included 53 Caucasian women with SP and 78 with TP aged from 21 to 47. Considering LC-MS/MS, patients with TP had lower concentrations of 25-hydroxyvitamin D (25(OH)D) than patients with SP. However, no significant difference was observed in the frequency of the occurrence of vitamin D deficiency (25(OH)D < 30 ng/mL). In both groups, the levels obtained with CLIA were significantly lower than in case of LC-MS/MS, however, strongly correlated. The intermethod agreement accounted for 52.4% and the Cohen's kappa coefficient was 0.142. Conclusions: The concentration of 25(OH)D in pregnant women depends on the type of gestation (SP/TP) and on the diagnostic methods used (CLIA/LC-MS/MS). Based on LC-MS/MS, the incidence of vitamin D deficiency was low in our group and no differences occurred in its frequency between SP and TP. The intermethod agreement between CLIA and LC-MS/MS on the detection of vitamin D deficiency was low. Significance statement: This is the first study to compare the concentration of 25(OH)D levels between SP and TP using two methods: CLIA and the gold standard - LC-MS/MS. Based on LC-MS/MS, a low incidence of vitamin D deficiency was observed in our group, in which the vast majority of patients took cholecalciferol supplements. Moreover, there were no differences in its frequency between SP and TP. However, the 25(OH)D level was significantly lower in TP. The intermethod agreement between CLIA and LC-MS/MS on the detection of vitamin D deficiency was low, which is associated with substantial clinical implications.

Escherichia albertii as a Potential Enteropathogen in the Light of Epidemiological and Genomic Studies.

Escherichia albertii is a new enteropathogen of humans and animals. The aim of the study was to assess the prevalence and pathogenicity of E. albertii strains isolated in northeastern Poland using epidemiological and genomic studies. In 2015-2018, a total of 1154 fecal samples from children and adults, 497 bird droppings, 212 food samples, 92 water samples, and 500 lactose-negative E. coli strains were tested. A total of 42 E. albertii strains were isolated. The PCR method was suitable for their rapid identification. In total, 33.3% of E. albertii isolates were resistant to one antibiotic, and 16.7% to two. Isolates were sensitive to cefepime, imipenem, levofloxacin, gentamicin, trimethoprim/sulfamethoxazole, and did not produce ESBL ß-lactamases. High genetic variability of E. albertii has been demonstrated. In the PFGE method, 90.5% of the strains had distinct pulsotypes. In MLST typing, 85.7% of strains were assigned distinct sequence types (STs), of which 64% were novel ST types. Cytolethal distending toxin (CDT) and Paa toxin genes were found in 100% of E. albertii isolates. Genes encoding toxins, IbeA, CdtB type 2, Tsh and Shiga (Stx2f), were found in 26.2%, 9.7%, 1.7%, and 0.4% of E. albertii isolates, respectively. The chromosome size of the tested strains ranged from 4,573,338 to 5,141,010 bp (average 4,784,003 bp), and at least one plasmid was present in all strains. The study contributes to a more accurate assessment of the genetic diversity of E. albertii and the potential threat it poses to public health.

Risk of malignancy in adrenal tumors in patients with a history of cancer.

Purpose: Adrenal gland is a common site of metastasis and on the other hand, metastases are the most frequent malignant adrenal tumors. The aim of this study was to estimate the risk of malignancy in suspicious adrenal mass in patients with a history of cancer. Methods: This is a single-center retrospective analysis of patients with adrenal tumors treated previously for different types of cancers. Between 2004 and 2021 a hundred and six such patients were identified. Mean age of patients was 62.6 years (30-78), and mean time from oncologic treatment was 55.8 months (0-274). The most common primary cancer was kidney (RCC): 29 (27.4%), colon/rectum (CRC): 20 (18.9%) and lung (NSCLC): 20 (18.9%). Results: Of 106 patients, 12 had hormonally active (HA) (11,3%) and 94 (88,7%) non active (HNA) tumors In group of patients with HA tumours 4 had hypercortisolaemia and 8 had elevation of urinary metanephrines. In the first group of HA patients pathology confirmed preoperative diagnosis of adrenocortical cancer and no metastasis was found. In all patients from the second group pheochromocytomas were confirmed. Primary (PM) and secondary (SM) malignancies were found in 50 patients (47.2%). In hormone inactive group only SM - 46/94 (48.9%) were diagnosed. The odds that adrenal lesion was a metastasis were higher if primary cancer was RCC (OR 4.29) and NSCLC (OR 12.3). Metastases were also more likely with high native tumor density, and bigger size in CT. The cut-off values for tumor size and native density calculated from receiver operating characteristic (ROC) curves were 37mm and 24, respectively. Conclusion: Risk of malignancy of adrenal mass in a patient with a history of cancer is high (47,2%), regardless of hormonal status. 47,2% risk of malignancy. In preoperative assessment type of primary cancer, adrenal tumour size and native density on CT should be taken into consideration as predictive factors of malignancy. Native density exceeding 24 HU was the strongest risk factor of adrenal malignancy (RR 3.23), followed by history of lung or renal cancer (RR 2.82) and maximum tumor diameter over 37 mm (RR 2.14).

Evaluation of plasma ACTH in the metyrapone test is insufficient for the diagnosis of secondary adrenal insufficiency.

Objective: To determine whether a single measurement of ACTH instead of less available in daily practice 11-deoxycortisol assay is sufficient to rule out or confirm secondary adrenal insufficiency (SAI) in the short Metyrapone test. Design: A retrospective analysis of diagnostic tests (Metyrapone and Synacthen tests) performed at our Center between 2016 and 2018 in patients with suspicion of secondary adrenal insufficiency. Material and methods: In 103 patients short metyrapone test was performed with assessment of 11-deoxycortisol and ACTH concentration after Metyrapone administered at midnight. In 89 of them short Synacthen (SST) test was also done (1 or/and 250 mcg 1-24ACTH). ROC curves have been performed to evaluate the diagnostic performance of ACTH level in metyrapone test as the predictor of secondary adrenal insufficiency (SAI) analysing sensitivity and specificity for various possible thresholds proposed in literature. Results: 40 (39%) of examined subjects were diagnosed as SAI, basing on post-Metyrapone 11-deoxycortisol concentration below 70 µg/l. In this group ACTH concentration was 128.1 ng/l (95% CI 96.8-159.4) versus 289.9 ng/l (95% CI 249.1-330.9) in patients with proper adrenal response. There was only a moderate positive correlation between ACTH and 11-deoxycortisol concentrations (r=0.5; p<0.05). The best cut off value of ACTH in relation to 11-deoxycortisol serum concentrations was 147 ng/l - with sensitivity of 73.2% and specificity 83.9%. However, plasma ACTH was>200ng/ml (the highest threshold proposed in literature) in 8 cases (20%) with positive diagnosis of SAI made on the basis of low 11-deoxycortisole and confirmed in short Synacthen test. Conclusion: Our results indicate that for a valuable evaluation of the results of the metyrapone test, the more readily available plasma ACTH assay cannot replace the measurement of 11-deoxycortisol concentrations.

The Effect of a Single High Dose of Vitamin D on Serum Levels of Its Metabolites in the Elderly.

BACKGROUND: Vitamin D is a dietary micronutrient responsible for calcium and phosphorus metabolism and multiple extraskeletal actions. The assessment of vitamin D status is commonly based on measurement of 25(OH)D total concentration in serum. However, the usage of liquid chromatography with tandem mass spectrometry (LC-MS/MS) technique allows to reliably assess a panel of vitamin D metabolites in serum or plasma, which may help to investigate the metabolic paths of vitamin D, especially in populations at risk of deficiency. METHODS: A randomized, two-arms, open study was conducted on 58 patients (28 female and 30 male; aged from 61 to 96 years old). The primary aim was to assess the effects of a single, high, oral dose of vitamin D3 (120,000 IU) on serum 25(OH)D3, 25(OH)D2, 24,25(OH)2D3, 3-epi-25(OH)D3, 1,25(OH)2D3, 24,25(OH)2D3/25(OH)D3 ratio, and 25(OH)D3/3-epi-25(OH)D3 ratio concentration (measured by LC-MS/MS) at baseline, 3 days and 7 days after administration, compared to control group. The secondary aim was assessment of influence of percentage of fat tissue on serum metabolites of vitamin D and their changes after bolus dose. RESULTS: 56.6% study group attained a serum 25(OH)D3 concentration >30 ng/mL. All subjects, except for one patient achieved a serum 25(OH)D3 concentration >20 ng/mL after administration. No one exceed reference value of vitamin D (30-50 ng/mL). Among participants who received vitamin D3 there were significant increase in 25(OH)D3, 3-epi-25(OH)D3, 1,25(OH)2D3, 24,25(OH)2D3 on 3rd day after administration. 24,25(OH)2D3 concentration gradually grew, achieving the highest concentration on 7th day. The percentage increase of 25(OH)D3 was negatively correlated with baseline 25(OH)D3 (r = -0.688, p = 0.001). Positive correlation between percentage increase in 25(OH)D3 and a percentage increase serum concentration of 24,25(OH)2D3 (r = 0.954, p < 0.001), 3-epi-25(OH)D3 (r = 8.03, p < 0.001) and 1,25(OH)2D3 (r = 0.789, p <0.001) were found. None of the study participants developed hypercalcemia. The baseline concentration of analyzed metabolites of vitamin D in serum and their percentage increase were neither dependent on BMI nor percentage of fat tissue. CONCLUSIONS: High dose of vitamin D rapidly increases 25(OH)D3 concentration in the elderly patients. The response to the bolus of vitamin D includes activation of 3-epimerase, followed by production of 24,25(OH)2D3, which protects from excessive increase of active form of vitamin D.

Adrenal "nonadenoma" - clinical characteristics and risk of malignancy.

INTRODUCTION: There is an increasing number of adrenal tumours discovered incidentally during imaging examinations performed for many different indications. Based on imaging results, it is possible to differentiate benign from malignant adrenal masses, although there is still a number of equivocal imaging findings. Our study presents 77 cases of adrenal tumours in which imaging was inconclusive and the final diagnosis was stated only after surgery and histopathological examination. MATERIAL AND METHODS: Retrospective data analysis: 77 cases of indeterminate adrenal tumours with a presumptive diagnosis of "nonadenoma" in patients operated within a 16-year period (2004-2019). None of the patients had a history of malignancy, and all tumours were hormonally inactive. On contrast-enhanced computed tomography (CT), the native density of all tumours was higher than 10 Hounsfield Units (HU), and the absolute percentage washout (APW) and relative percentage washout (RPW) were lower than 60% and 40%, respectively. RESULTS: The most common findings were adrenal adenoma (25.9%), macronodular adrenal hyperplasia (16.9%), ganglioneuroma (15.6%), and haemorrhage with posthaemorrhagic changes (13%). In total, there were 12 various histopathological diagnoses in this group. There were only 2 (2.6%) malignant (adrenal cancer and leiomyosarcoma) and 3 (3.9%) potentially malignant (pheochromocytoma) lesions in this group. CONCLUSIONS: It is often impossible to make a correct diagnosis in a clinical setting until it is histologically verified. "Nonadenoma" adrenal tumours constitute a heterogeneous group including very rare pathologies. The risk of malignancy in indeterminate adrenal tumours is relatively low.

Therapeutic plasma exchange with albumin as a valuable method of preparing thyrotoxic patients for a life-saving thyroidectomy.

Hyperthyroidism affects approximately 1.2% of the population and its routine treatment includes antithyroid drugs (ATDs), radioiodine and surgery. Management of patients with resistance or contraindications to ATDs who require thyroidectomy may be challenging. We present the experience of our department in preparing thyrotoxic patients for life-saving thyroidectomy by using therapeutic plasma exchange (TPE) with albumin: one patient with Graves' disease and previous history of agranulocytosis and cholestatic jaundice after ATDs and two patients with amiodarone-induced thyrotoxicosis. Five to six TPEs were applied to each patient resulting in a decrease of fT3 by 57% to 83%, fT4 by 21% to 60% and decrease/normalization of total thyroid hormones. All patients underwent surgery successfully. In case of drug-resistant thyrotoxicosis or contraindications to ATDs, TPE can be a valuable tool in preparing patients for surgery. Albumin used as a replacement fluid appears to be effective in ameliorating clinical and laboratory symptoms of thyrotoxicosis.

Obesity in menopause - our negligence or an unfortunate inevitability?

Numerous concerns about menopause exist among women, and fear of an increase in body weight is one of the most important of them. This paper presents an overview of current knowledge concerning the etiology of obesity related to menopause and about the mechanisms of its development, with particular regard to the hormonal changes that occur during this period of life. The role of estrogens in the regulation of energy balance and the effect of sex hormones on metabolism of adipose tissue and other organs are presented. The consequence of the sharp decline in the secretion of estrogens with subsequent relative hyperandrogenemia is briefly discussed. The main intention of this review is to clarify what is inevitable and what perhaps results from negligence and unhealthy lifestyles. In the last part of the paper the possibilities of counteracting the progress of adverse changes in body composition, by promoting beneficial lifestyle modifications and the use of hormonal substitution treatment, in cases where it is reasonable and possible, are described.