Etude du metabolism du fer en muscle skeletique

Ο μεταβολισμός του σιδήρου δεν είναι επαρκώς μελετημένος στον σκελετικό ιστό σε αντίθεση με τον ηπατικό. Σκοπός της μελέτης είναι η σύγκριση της έκφρασης των γονιδίων του μεταβολισμού του σιδήρου στον ανθρώπινο σκελετικό και ηπατικό ιστό. Συλλέχτηκαν βιοψίες μυϊκού και ηπατικού ιστού από έξι φυσιολογικά άτομα. Επιλέχθηκαν να μελετηθούν12 γονίδια που εμπλέκονται στο μεταβολισμό του σιδήρου για την εισαγωγή σιδήρου στο κύτταρο, αποθήκευση και εξαγωγή του καθώς και δύο μόρια ρύθμισης της ομοιοστασίας του σιδήρου: εισαγωγή σιδήρου [οι υποδοχείς της τρανσφερρίνης (TfR1 και TfR2), το μόριο HFE, ο μεταφορέας δισθενών μετάλλων (DMT1,DMT1nonIRE), και το σιδεροφόρο μόριο λιποκαλίνη (NGAL)], αποθήκευση σιδήρου [η βαριά αλυσίδα της φερριτίνης (FTH1)] και εξαγωγή σιδήρου [η φερροπορτίνη (IREG1), η ηφαιστίνη (HEPH) και η σερουλοπλασμίνη (CP)] καθώς και δύο μόρια ρύθμισης της ομοιοστασίας του σιδήρου [η εψιδίνη (HAMP) και η αιμοτζουβελίνη (HJV)]. Ακολούθησαν αλυσιδωτές αντιδράσεις της πολυμεράσης, RT-PCR και ημιποσοτικοποίηση των επιπέδων έκφρασης των γονιδίων με τη μέθοδο της πυκνομετρίας (Densitometric Analysis). Τα αποτελέσματα εκφράζονται με βάση το ποσοστό επί τοις εκατό του γονιδίου της β-ακτίνης. Το γονίδιο της β-ακτίνης χρησιμοποιήθηκε για την κανονικοποίηση των επιπέδων έκφρασης, ως γονίδιο αναφοράς της μελέτης. Αναδεικνυόμενες διαφορές συγκριτικής έκφρασης μεγαλύτερες του 20%, των μελετημένων γονιδίων του μεταβολισμού του σιδήρου με ημιποσοτικοποίηση, αναλύθηκαν περαιτέρω με την μέθοδο της PCR σε αληθινό χρόνο (Real time PCR, qPCR) και ποσοτικοποιήθηκαν (LightCycler, Roche). Η στατιστική ανάλυση των αποτελεσμάτων ποσοτικοποίησης έγινε με το one paired t test και τα αποτελέσματα είναι στατιστικά σημαντικά (p70% των επιπέδων έκφρασης της βακτίνης. 2. Εξαίρεση αποτελούν τα HAMP, CP και TfR2 που απουσιάζουν ή παρουσιάζουν ελάχιστη έκφραση (<10% των επιπέδων έκφρασης της βακτίνης) στο σκελετικό μυ αντίστοιχα. 3. Ενώ τα HJV και HEPH παρουσιάζουν μεγαλύτερη έκφραση των επιπέδων mRNA στο σκελετικό μύ συγκριτικά με το ήπαρ (SM/L=2,65±1,1(p<0,05) και SM/L=1,5±0,06(p<0,05 αντίστοιχα στην Q-PCR). (Εικόνα 1). Η εργασία αυτή αφορά φυσιολογικές καταστάσεις ανθρώπινων ιστών, δίνει όμως σημαντικά ερεθίσματα για αντίστοιχες μελέτες του μεταβολισμού του σιδήρου σε παθολογικές καταστάσεις. Υπογραμμίζει δε την σπουδαιότητα του σκελετικού μυϊκού ιστού και την ανάλογη συμμετοχή του στη ομοιοστασία του σιδήρου. Οι ποσοτικές διαφορές που παρατηρούνται στην έκφραση γονιδίων που εμπλέκονται σε διάφορα κυτταρικά μονοπάτια αναδεικνύουν την ανάγκη περαιτέρω έρευνας του κυτταρικού μεταβολισμού στο σκελετικό ιστό

Plasma B-type natriuretic peptide concentration in beta-thalassaemia patients

Background: Plasma B-type natriuretic peptide (BNP) concentration has significant diagnostic accuracy and prognostic value in various forms of heart disease. Whether BNP is also useful in the evaluation and management of thalassaemia heart disease remains to be determined. Methods and results: Eighty three thalassaemia major patients; 8 with acutely decompensated heart failure (New York Heart Association [NYHA] class III or IV, group A), 25 with NYHA class 11 symptoms and impaired systolic left ventricular function (ejection fraction &lt;55% or fractional shortening &lt;30%, group B) and 50 with normal systolic function (group C), as well as 50 healthy controls, were studied. Assessment included history, physical examination, Doppler echocardiography and plasma BNP determination. Mean BNP levels were 431 219 pg/mL (range, 283-890 pg/mL) in group A, 158 +/- 31 pg/mL in group B, 1763 +/- 54 pg/mL in group C and 43 +/- 24 pg/mL in controls. BNP levels were significantly higher in group A (p &lt; 0.001), but did not differ between groups B and C. Moreover, BNP was not correlated with left ventricular end-diastolic diameter, left ventricular mass, right ventricular diameter index, Doppler diastolic indexes (except in group C), the mean 2-year serum ferritin concentration or the peak serum ferritin concentration in any of the three patient groups. Conclusion: A potential deficiency of BNP-related neurohormonal mechanisms may impair its clinical usefulness in thalassaemia major. (C) 2006 European Society of Cardiology. Published by Elsevier B.V All rights reserved

Azasteroid Alkylators as Dual Inhibitors of AKT and ERK Signaling for the Treatment of Ovarian Carcinoma

(1) Background: Previous findings show that lactam steroidal alkylating esters display improved therapeutic efficacy with reduced toxicity. The aim of this study was to evaluate the anticancer activity of two newly synthesized aza-steroid alkylators (ENGA-L06E and ENGA-L08E) against human ovarian carcinoma cells, and consequently, the dual inhibition of RAS/PI3K/AKT and RAS/RAF/MEK/ERK signaling pathways, both of which are closely associated with ovarian cancer; (2) Methods: The in vitro cytostatic and cytotoxic effects of ENGA-L06E and ENGA-L08E were evaluated in a panel of five human ovarian cancer cell lines, as well as in in vivo studies. ENGA-L06E and ENGA-L08E, in addition to another two aniline-mustard alkylators, POPAM and melphalan (L-PAM), were utilized in order to determine the acute toxicity and antitumor efficacy on two human ovarian xenograft models. Also, in silico studies were performed in order to investigate the dual inhibition of ENGA-L06E and ENGA-L08E on RAS/PI3K/AKT and RAS/RAF/MEK/ERK signaling pathways; (3) Results: Both, in vitro and in vivo studies demonstrated that ENGA-L06E and ENGA-L08E were significantly more effective with a lower toxicity profile in comparison to POPAM and L-PAM alkylators. Moreover, in silico studies demonstrated that the two new aza-steroid alkylators could act as efficient inhibitors of the phosphorylation of AKT and ERK1/2 molecules; and (4) Conclusions: Both ENGA-L06E and ENGA-L08E demonstrated high anticancer activity through the inhibition of the PI3K-AKT and KRAS-ERK signaling pathways against human ovarian carcinoma, and thus constituting strong evidence towards further clinical development

Cytocidal Antitumor Effects against Human Ovarian Cancer Cells Induced by B-Lactam Steroid Alkylators with Targeted Activity against Poly (ADP-Ribose) Polymerase (PARP) Enzymes in a Cell-Free Assay

We evaluated three newly synthesized B-lactam hybrid homo-aza-steroidal alkylators (ASA-A, ASA-B and ASA-C) for their PARP1/2 inhibition activity and their DNA damaging effect against human ovarian carcinoma cells. These agents are conjugated with an alkylating component (POPA), which also served as a reference molecule (positive control), and were tested against four human ovarian cell lines in vitro (UWB1.289 + BRCA1, UWB1.289, SKOV-3 and OVCAR-3). The studied compounds were thereafter compared to 3-AB, a known PARP inhibitor, as well as to Olaparib, a standard third-generation PARP inhibitor, on a PARP assay investigating their inhibitory potential. Finally, a PARP1 and PARP2 mRNA expression analysis by qRT-PCR was produced in order to measure the absolute and the relative gene expression (in mRNA transcripts) between treated and untreated cells. All the investigated hybrid steroid alkylators and POPA decreased in vitro cell growth differentially, according to the sensitivity and different gene characteristics of each cell line, while ASA-A and ASA-B presented the most significant anticancer activity. Both these compounds induced PARP1/2 enzyme inhibition, DNA damage (alkylation) and upregulation of PARP mRNA expression, for all tested cell lines. However, ASA-C underperformed on average in the above tasks, while the compound ASA-B induced synthetic lethality effects on the ovarian cancer cells. Nevertheless, the overall outcome, leading to a drug-like potential, provides strong evidence toward further evaluation

Oxidant/Antioxidant Status Is Impaired in Sepsis and Is Related to Anti-Apoptotic, Inflammatory, and Innate Immunity Alterations

Oxidative stress is considered pivotal in the pathophysiology of sepsis. Oxidants modulate heat shock proteins (Hsp), interleukins (IL), and cell death pathways, including apoptosis. This multicenter prospective observational study was designed to ascertain whether an oxidant/antioxidant imbalance is an independent sepsis discriminator and mortality predictor in intensive care unit (ICU) patients with sepsis (n = 145), compared to non-infectious critically ill patients (n = 112) and healthy individuals (n = 89). Serum total oxidative status (TOS) and total antioxidant capacity (TAC) were measured by photometric testing. IL-6, -8, -10, -27, Hsp72/90 (ELISA), and selected antioxidant biomolecules (Ζn, glutathione) were correlated with apoptotic mediators (caspase-3, capsase-9) and the central anti-apoptotic survivin protein (ELISA, real-time PCR). A wide scattering of TOS, TAC, and TOS/TAC in all three groups was demonstrated. Septic patients had an elevated TOS/TAC, compared to non-infectious critically ill patients and healthy individuals (p = 0.001). TOS/TAC was associated with severity scores, procalcitonin, IL-6, -10, -27, IFN-γ, Hsp72, Hsp90, survivin protein, and survivin isoforms -2B, -ΔΕx3, -WT (p p TOS/TAC (0.96 (95% CI = 0.93–0.99)) was higher than AUCTAC (z = 20, p TOS (z = 3.1, p = 0.002) in distinguishing sepsis. TOS/TAC, TOS, survivin isoforms -WT and -2B, Hsp90, IL-6, survivin protein, and repressed TAC were strong predictors of mortality (p < 0.01). Oxidant/antioxidant status is impaired in septic compared to critically ill patients with trauma or surgery and is related to anti-apoptotic, inflammatory, and innate immunity alterations. The unpredicted TOS/TAC imbalance might be related to undefined phenotypes in patients and healthy individuals

Oxidant/Antioxidant Status Is Impaired in Sepsis and Is Related to Anti-Apoptotic, Inflammatory, and Innate Immunity Alterations

Oxidative stress is considered pivotal in the pathophysiology of sepsis. Oxidants modulate heat shock proteins (Hsp), interleukins (IL), and cell death pathways, including apoptosis. This multicenter prospective observational study was designed to ascertain whether an oxidant/antioxidant imbalance is an independent sepsis discriminator and mortality predictor in intensive care unit (ICU) patients with sepsis (n = 145), compared to non-infectious critically ill patients (n = 112) and healthy individuals (n = 89). Serum total oxidative status (TOS) and total antioxidant capacity (TAC) were measured by photometric testing. IL-6, -8, -10, -27, Hsp72/90 (ELISA), and selected antioxidant biomolecules (Zeta n, glutathione) were correlated with apoptotic mediators (caspase-3, capsase-9) and the central anti-apoptotic survivin protein (ELISA, real-time PCR). A wide scattering of TOS, TAC, and TOS/TAC in all three groups was demonstrated. Septic patients had an elevated TOS/TAC, compared to non-infectious critically ill patients and healthy individuals (p = 0.001). TOS/TAC was associated with severity scores, procalcitonin, IL-6, -10, -27, IFN-gamma, Hsp72, Hsp90, survivin protein, and survivin isoforms -2B, -Delta Epsilon x3, -WT (p &lt; 0.001). In a propensity probability (age-sex-adjusted) logistic regression model, only sepsis was independently associated with TOS/TAC (Exp(B) 25.4, p &lt; 0.001). The AUC(TOS/TAC) (0.96 (95% CI = 0.93-0.99)) was higher than AUC(TAC) (z = 20, p &lt; 0.001) or AUC(TOS) (z = 3.1, p = 0.002) in distinguishing sepsis. TOS/TAC, TOS, survivin isoforms -WT and -2B, Hsp90, IL-6, survivin protein, and repressed TAC were strong predictors of mortality (p &lt; 0.01). Oxidant/antioxidant status is impaired in septic compared to critically ill patients with trauma or surgery and is related to anti-apoptotic, inflammatory, and innate immunity alterations. The unpredicted TOS/TAC imbalance might be related to undefined phenotypes in patients and healthy individuals