Διερεύνηση των μεταβολών την ημερήσιας διακύμανσης (κιρκάδιος ρυθμός) των φλοιοεπινεφριδιακών ορμονών και της ιντερλευκίνης-6 σε ασθενείς με οξεία λοίμωξη SARS-CoV-2

Σκοπός: Η ευεργετική επίδραση των γλυκοκορτικοειδών στη λοίμωξη από το νέο κορωνοϊό (COVID-19) έχει τεκμηριωθεί, αλλά δεν έχει αποσαφηνιστεί πλήρως εάν η έκκριση της κορτιζόλης και των άλλων επινεφριδιακών ορμονών διαταράσσονται κατά τη διάρκεια της λοίμωξης με COVID-19. Σκοπός της παρούσας μελέτης, ήταν να διερευνήσουμε την ημερήσια διακύμανση της έκκρισης της κορτιζόλης και του επινεφριδιακού ανδρογόνου δεϋδροεπιανδροστερόνη (DHEA) στο σίελο- όπου μπορεί να μετρηθεί η ελεύθερη βιοδιαθέσιμη μορφή των ορμονών αυτών- σε ασθενείς με οξεία λοίμωξη με COVID-19 με διαφορετική βαρύτητα νόσησης. Μέθοδοι: Η μελέτη αυτή είναι μία συγχρονική μελέτη ασθενών - μαρτύρων και είναι καταχωρημένη στη διεθνή βάση καταγραφής κλινικών μελετών παρατήρησης με κωδικό NCT04988269 (https://clinicaltrials.gov/ct2/show/NCT0498826). Πενήντα δύο ασθενείς με COVID-19 -πριν από την ενδεχόμενη θεραπεία με δεξαμεθαζόνη- που εξετάσθηκαν στο Τμήμα Επειγόντων Περιστατικών του Γενικού Νοσοκομείου Αθηνών ΛΑΙΚΟ την περίοδο μεταξύ 15 Απριλίου και 15 Ιουνίου 2021, συμπεριελήφθησαν στη μελέτη. Τριάντα τρία άτομα αντιστοίχου ηλικίας και φύλου (αντιστοίχιση συχνότητας) χωρίς COVID-19 που εξετάσθηκαν την ίδια χρονική περίοδο στα Τμήμα Εξωτερικών Ιατρείων του Νοσοκομείου συμπεριελήφθησαν στη μελέτη ως μάρτυρες. Μετρήθηκαν και αξιολογήθηκαν τα ημερήσια επίπεδα κορτιζόλης στο σίελο στις κάτωθι ώρες: 8 π.μ., 12 μ.μ., 6 μ.μ. και 10 μ.μ., καθώς επίσης και της κορτικοτροπίνης (ACTH) και της αλδοστερόνης στο πλάσμα, και της ιντερλευκίνης-6 (IL-6) και της C-αντιδρώσας πρωτεΐνης (CRP) στον ορό. Η ημερήσια διακύμανση των επιπέδων της DHEA και της ιντερλευκίνης (IL-6) αξιολογήθηκαν επίσης σε υποομάδες ασθενών και μαρτύρων. Για τη σύγκριση των συνεχών μεταβλητών μεταξύ περισσότερων των 2 ομάδων χρησιμοποιήσαμε τη μέθοδο ANOVA Kruskal-Wallis, αναλόγως την κατανομή των μεταβλητών ακολουθούμενη από δοκιμασία πολλαπλών συγκρίσεων (Bonferroni ή κατά Dunn, αντίστοιχα). Χρησιμοποιήθηκε η μέθοδος ANOVA επαναλαμβανόμενων μετρήσεων με Bonferroni post hoc test για πολλαπλές συγκρίσεις μεταξύ διαφορετικών χρονικών σημείων στις μετρήσεις κορτιζόλης, DHEA και IL-6 σιέλου σε κάθε ομάδα. Η συνολική ημερήσια έκκριση κορτιζόλης και DHEA υπολογίστηκε χρησιμοποιώντας την περιοχή κάτω από την καμπύλη (area under the curve, AUC) με την τραπεζοειδή μέθοδο. Η πολυμεταβλητή ανάλυση γραμμικής παλινδρόμησης χρησιμοποιήθηκε για διόρθωση των αναλύσεων σχετικά με τον παράγοντα βαρύτητας της νόσου εντός της ομάδας ασθενών με COVID-19. Αποτελέσματα Οι διάμεσες τιμές των CRP και IL-6 ήταν περίπου 6 φορές υψηλότερες στους ασθενείς από ό,τι στους μάρτυρες (p<0.001). Τα πρωινά επίπεδα κορτιζόλης στο σίελο δεν διέφεραν σημαντικά μεταξύ των δύο ομάδων, ωστόσο, οι ασθενείς εμφάνισαν σημαντικά υψηλότερα επίπεδα κορτιζόλης σιέλου τις βραδινές ώρες σε σύγκριση με τους μάρτυρες [0.391 (min 0.054, max 0.663) v.s. 0.081 (min 0.054, max 0.243) μg/dl, p<0.001 και 0.183 (min 0.090, max 0.834) vs. 0.054 (min 0.052, max 0.054) μg/dl, αντίστοιχα]. H συνολική ημερήσια έκκριση της κορτιζόλης, [όπως αυτή προσδιορίστηκε από τον υπολογισμό της περιοχής κάτω από την καμπύλη των μετρήσεων, AUC] ήταν επίσης σημαντικά υψηλότερη στους ασθενείς συγκριτικά με τους μάρτυρες (4.81 ± 2.46 vs. 2.75 ± 0.810, αντίστοιχα, p<0.001), ανεξάρτητα από τη βαρύτητα της νόσου στους ασθενείς με COVID-19 (β = 0.132, 95% CI; -0.8, 2.0). Τα επίπεδα, ωστόσο, των λοιπών ορμονών ACTH, DHEA και αλδοστερόνης στην κυκλοφορία ήταν παρόμοια σε ασθενείς και μάρτυρες. Τα επίπεδα της IL-6 στον ορό, αλλά όχι τα επίπεδα της ACTH, συσχετίστηκαν σημαντικά με τα επίπεδα της κορτιζόλης στο σίελο τις νυχτερινές ώρες (rho=0.555, p<0,001) στους ασθενείς. Συμπεράσματα Τα επίπεδα της κορτιζόλης κυρίως τις βραδινές ώρες, αλλά όχι η πρωϊνή έκκριση της κορτιζόλης βρέθηκαν σημαντικά υψηλότερα στους ασθενείς με COVID-19 λοίμωξη ακόμα και σε ήπιας βαρύτητας νόσηση. Στο πλαίσιο της οξείας ιογενούς λοίμωξης (όπως η λοίμωξη COVID-19), η IL-6 διεγείρει την έκκριση της κορτιζόλης από τα επινεφρίδια, αντικαθιστώντας εν μέρει τη δράση της ορμόνης ACTH που παράγεται από την υπόφυση και αποτελεί τον φυσικό ρυθμιστή της έκκρισης των επινεφριδίων. Οι λοιπές επινεφριδιακές ορμόνες, ωστόσο, δεν φαίνεται να επηρεάζονται σημαντικά από τα αυξημένα επίπεδα της IL-6 στα πλαίσια οξείας ιογενούς λοίμωξης, όπως ο COVID-19. Η χορήγηση θεραπευτικών σκευασμάτων που περιέχουν γλυκοκορτικοειδή έχουν ρόλο στην θεραπευτική προσέγγιση της λοίμωξης με COVID-19, καθώς φαίνεται να συμμετέχουν στην ανοσολογική απόκριση του οργανισμού έναντι του ιού.Purpose: The beneficial effect of glucocorticoids in coronavirus disease (COVID-19) is established, but whether adrenal cortisol secretion is impaired in COVID-19 is not fully elucidated. In this case-control study we investigated the diurnal free bioavailable salivary cortisol secretion in COVID-19 patients. Methods: Fifty-two consecutive COVID-19 patients -before dexamethasone treatment- recruited between April 15th -June15th-2021, (NCT04988269) at Laikon Athens University-Hospital, and 33 age- and sex-matched (frequently matched with patients) apparently healthy volunteers who visited the Outpatient Clinic of Laikon General Hospital of Athens during the same time-period were also included in the analysis and served as controls. Only volunteers with negative PCR test for SARS-CoV-2 and without history of a contact with a confirmed COVID-19 case at the time of the study were included. Diurnal salivary cortisol (8am, 12, 6, and 10pm), plasma adrenocorticotropin (ACTH) and aldosterone, and serum interleukin-6 (IL-6) and C-reactive protein (CRP) levels were assessed. Diurnal salivary dehydroepiandrosterone (DHEA) and IL-6 were also assessed in subgroups of patients. Results Median CRP and IL-6 measurements were about 6-fold higher in patients than controls (both p<0.001). Morning salivary cortisol levels did not differ between the two groups, but patients exhibited higher median levels of evening and nocturnal salivary cortisol compared to controls [0.391 (min: 0.054, max: 0.663) vs. 0.081 (min: 0.054, max: 0.243) μg/dl, p<0.001 and 0.183 (min: 0.090, max: 0.834) vs. 0.054 (min: 0.054, max: 0.332) μg/dl, p<0.001, respectively], resulting in higher time-integrated area under the curve (AUC) (4.81 ± 2.46 vs. 2.75 ± 0.810, respectively, p<0.001). Circulating ACTH, DHEA, and aldosterone levels were similar in patients and controls. Serum IL-6, but not ACTH levels, was strongly correlated with nocturnal cortisol salivary levels (rho=0.555, p<0.001) in patients. Conclusion Increased evening and nocturnal but not morning cortisol secretion occur in even clinically mild COVID-19. In the context of acute viral infection (COVID-19), IL-6 may partially replace ACTH as a stimulus of the glucocorticoid-secreting adrenal zona-fasciculata without influencing the secretion of DHEA and aldosterone from the zona-reticularis and zona-glomerulosa respectively

Serum sclerostin levels in Paget's disease and prostate cancer with bone metastases with a wide range of bone turnover

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Hypercalcitoninaemia in pseudohypo-parathyroidism type 1A and type 1B

Pseudohypoparathyroidism (PHP) is a heterogeneous group of rare endocrine disorders characterised by normal renal function and renal resistance to the action of the parathyroid hormone. Type 1A (PHP1A), which is the most common variant, also include developmental and skeletal defects named as Albright hereditary osteodystrophy (AHO). We present two cases, a 54- and a 33-year-old male diagnosed with PHP who were referred to us for persistently high levels of serum calcitonin. AHO and multinodular goitre were present in the 54-year-old male, while the second patient was free of skeletal deformities and his thyroid gland was of normal size and without nodular appearance. We performed GNAS molecular analysis (methylation status and copy number analysis by MS-MLPA) in genomic DNA samples for both patients. The analysis revealed a novel missense variant c.131T>G p.(Leu44Pro) affecting GNAS exon 1, in the patient with the clinical diagnosis of PHP1A. This amino acid change appears to be in accordance with the clinical diagnosis of the patient. The genomic DNA analysis of the second patient revealed the presence of the recurrent 3-kb deletion affecting the imprinting control region localised in the STX16 region associated with the loss of methylation (LOM) at the GNAS A/B differentially methylated region and consistent with the diagnosis of an autosomal dominant form of PHP type 1B (PHP1B). In conclusion, hypercalcitoninaemia may be encountered in PHP1A and PHP1B even in the absence of thyroid pathology. Learning points: We describe a novel missense variant c.131T>G p.(Leu44Pro) affecting GNAS exon 1 as the cause of PHP1A. Hypercalcitoninaemia in PHP1A is considered an associated resistance to calcitonin, as suggested by the generalised impairment of Gs\u3b1-mediated hormone signalling. GNAS methylation defects, as in type PHP1B, without thyroid pathology can also present with hypercalcitoninaemia

Role of Vanadium in Cellular and Molecular Immunology: Association with Immune-Related Inflammation and Pharmacotoxicology Mechanisms

Over the last decade, a diverse spectrum of vanadium compounds has arisen as anti-inflammatory therapeutic metallodrugs targeting various diseases. Recent studies have demonstrated that select well-defined vanadium species are involved in many immune-driven molecular mechanisms that regulate and influence immune responses. In addition, advances in cell immunotherapy have relied on the use of metallodrugs to create a “safe,” highly regulated, environment for optimal control of immune response. Emerging findings include optimal regulation of B/T cell signaling and expression of immune suppressive or anti-inflammatory cytokines, critical for immune cell effector functions. Furthermore, in-depth perusals have explored NF-κB and Toll-like receptor signaling mechanisms in order to enhance adaptive immune responses and promote recruitment or conversion of inflammatory cells to immunodeficient tissues. Consequently, well-defined vanadium metallodrugs, poised to access and resensitize the immune microenvironment, interact with various biomolecular targets, such as B cells, T cells, interleukin markers, and transcription factors, thereby influencing and affecting immune signaling. A synthetically formulated and structure-based (bio)chemical reactivity account of vanadoforms emerges as a plausible strategy for designing drugs characterized by selectivity and specificity, with respect to the cellular molecular targets intimately linked to immune responses, thereby giving rise to a challenging field linked to the development of immune system vanadodrugs

Co-Expression of c-Fos with Oestradiol Receptor alpha or Somatostatin in the Arcuate Nucleus, Ventromedial Nucleus and Medial Preoptic Area in the Follicular Phase of Intact Ewes: Alteration after Insulin-Induced Hypoglycaemia

The aim of this study was to investigate how acute insulin‐induced hypoglycaemia (IIH) alters the activity of cells containing oestradiol receptor α (ERα) or somatostatin (SST) in the arcuate nucleus (ARC) and ventromedial nucleus (VMN), and ERα cells in the medial preoptic area (mPOA) of intact ewes. Follicular phases were synchronized with progesterone vaginal pessaries. Control animals were killed at 0 h or 31 h (n = 5 and 6, respectively) after progesterone withdrawal (PW; time zero). At 28 h, five other animals received insulin (INS; 4 iu/kg) and were subsequently killed at 31 h. Hypothalamic sections were immunostained for ERα or SST each with c‐Fos, a marker of neuronal transcriptional activation. Insulin did not alter the percentage of activated ERα cells in the ARC; however, it appeared visually that two insulin‐treated animals (INS responders, with no LH surge) had an increase in the VMN (from 32 to 78%) and a decrease in the mPOA (from 40 to 12%) compared to no increase in the two INS non‐responders (with an LH surge). The percentage of activated SST cells in the ARC was greater in all four insulin‐treated animals (from 10 to 60%), whereas it was visually estimated that activated SST cells in the VMN increased only in the two insulin responders (from 10 to 70%). From these results, we suggest that IIH stimulates SST activation in the ARC as part of the glucose‐sensing mechanism but ERα activation is unaffected in this region. We present evidence to support a hypothesis that disruption of the GnRH/LH surge may occur in insulin responders via a mechanism that involves, at least in part, SST cell activation in the VMN along with decreased ERα cell activation in the mPOA

Role of PACAP and VIP Signalling in Regulation of Chondrogenesis and Osteogenesis

Pituitary adenylate cyclase activating polypeptide (PACAP) and vasoactive intestinal peptide (VIP) are multifunctional proteins that can regulate diverse physiological processes. These are also regarded as neurotrophic and anti-inflammatory substances in the CNS, and PACAP is reported to prevent harmful effects of oxidative stress. In the last decade more and more data accumulated on the similar function of PACAP in various tissues, but its cartilage- and bone-related presence and functions have not been widely investigated yet. In this summary we plan to verify the presence and function of PACAP and VIP signalling tool kit during cartilage differentiation and bone formation. We give evidence about the protective function of PACAP in cartilage regeneration with oxidative or mechanically stress and also with the modulation of PACAP signalling in vitro in osteogenic cells. Our observations imply the therapeutic perspective that PACAP might be applicable as a natural agent exerting protecting effect during joint inflammation and/or may promote cartilage regeneration during degenerative diseases of articular cartilage

Circulating microRNAs as potential diagnostic biomarkers for osteoporosis

Osteoporosis is the most common age-related bone disease worldwide and is usually clinically asymptomatic until the first fracture happens. MicroRNAs are critical molecular regulators in bone remodelling processes and are stabilised in the blood. The aim of this project was to identify circulatory microRNAs associated with osteoporosis using advanced PCR arrays initially and the identified differentially-expressed microRNAs were validated in clinical samples using RT-qPCR. A total of 161participants were recruited and 139 participants were included in this study with local ethical approvals prior to recruitment. RNAs were extracted, purified, quantified and analysed from all serum and plasma samples. Differentially-expressed miRNAs were identified using miRNA PCR arrays initially and validated in 139 serum and 134 plasma clinical samples using RT-qPCR. Following validation of identified miRNAs in individual clinical samples using RT-qPCR, circulating miRNAs, hsa-miR-122-5p and hsa-miR-4516 were statistically significantly differentially-expressed between non-osteoporotic controls, osteopaenia and osteoporosis patients. Further analysis showed that the levels of these microRNAs were associated with fragility fracture and correlated with the low bone mineral density in osteoporosis patients. The results show that circulating hsa-miR-122-5p and hsa-miR-4516 could be potential diagnostic biomarkers for osteoporosis in the future

Expression of Circulating MicroRNAs Linked to Bone Metabolism in Chronic Kidney Disease-Mineral and Bone Disorder

The pathophysiology of chronic kidney disease–mineral and bone disorder (CKD-MBD) is complex and multifactorial. Recent studies have identified a link between microRNAs (miRNAs) and bone loss. In this study, we investigated the expression of miRNAs in CKD-MBD. In this case-control study, we included thirty patients with CKD-MBD (cases) and 30 age- and gender-matched healthy individuals (controls). Bone mineral density (BMD) and trabecular bone score (TBS) evaluation was performed with dual X-ray absorptiometry. The selected panel of miRNAs included: hsa-miRNA-21-5p; hsa-miRNA-23a-3p; hsa-miRNA-24-2-5p; hsa-miRNA-26a-5p; hsa-miRNA-29a-3; hsa-miRNA-124-3p; hsa-miRNA-2861. The majority of cases had low BMD values. The relative expression of miRNA-21-5p was 15 times lower [fold regulation (FR): −14.7 ± 8.1, p = 0.034), miRNA-124-3p, 6 times lower (FR: −5.9 ± 4, p = 0.005), and miRNA-23a-3p, 4 times lower (FR: −3.8 ± 2.0, p = 0.036) in cases compared to controls. MiRNA-23a-3p was significantly and inversely correlated with TBS, adjusted for calcium metabolism and BMD values (beta = −0.221, p = 0.003, 95% CI −0.360, −0,081) in cases. In a receiver operating characteristic (ROC) analysis, expression of miRNA-124-3p demonstrated 78% sensitivity and 83% specificity in identifying CKD patents with osteoporosis. Serum expression of miRNAs related to osteoblasts (miRNA-23a-3p) and osteoclasts (miRNA-21-5p, miRNA-124-3p) is significantly altered in patients with CKD-MBD