Search

Ribonucleotide Reductase (RNR) is a two-subunit (RRM1, RRM2) enzyme, responsible for the conversion of ribonucleotides to deoxyribonucleotides required for DNA replication. To evaluate RNR as a biomarker of response to 5-azacytidine, we measured RNR mRNA levels by a quantitative real-time PCR in bone marrow samples of 98 patients with myelodysplastic syndrome (MDS) treated with 5-azacytidine with parallel quantification of the gene promoter's methylation. Patients with low RRM1 levels had a high RRM1 methylation status (p = 0.005) and a better response to treatment with 5-azacytidine (p = 0.019). A next-generation sequencing for genes of interest in MDS was also carried out in a subset of 61 samples. Splicing factor mutations were correlated with lower RRM1 mRNA levels (p = 0.044). Our results suggest that the expression of RNR is correlated with clinical outcomes, thus its expression could be used as a prognostic factor for response to 5-azacytidine and a possible therapeutic target in MDS.

Copper-based bactericides have appeared as a new tool in crop protection and offer an effective solution to combat bacterial resistance. In this work, two copper nanoparticle products that were previously synthesized and evaluated against major bacterial and fungal pathogens were tested on their ability to control the bacterial spot disease of tomato. Growth of Xanthomonas campestris pv. vesicatoria, the causal agent of the disease, was significantly suppressed by both nanoparticles, which had superior function compared to conventional commercial formulations of copper. X-ray fluorescence spectrometry measurements in tomato leaves revealed that bioavailability of copper is superior in the case of nanoparticles compared to conventional formulations and is dependent on synthesis rather than size. This is the first report correlating bioavailability of copper to nanoparticle efficacy.

It is known that children are more sensitive to the effects of medical treatments and environment than adults. Today there is limited information regarding the differences in genotoxic effects in children. The micronucleus assay is a method that is used to monitor genotoxicity, and it was validated several years before. Today there is international interest for exfoliated buccal cells. Most of the micronuclei studies in children have been performed with the analyses of lymphocytes. However, there is vast interest in using exfoliated cells from the oral cavity. The reason is that other type of cells are acquired non-invasively, this is an important issue in paediatric cohorts. Unfortunately a limitation of measuring micronuclei frequency is that it has been observed to be low in newborns and on the other hand there are a large number of patients and cell sample counts. It has been observed that radiation exposure and environmental pollutants increase the micronuclei frequency in newborn and children. Regarding the medical treatments, there is little data and several studies are needed to optimise the doses. There is the need to observe if there is a relationship between micronuclei in lymphocytes and exfoliated cells and to identify the baseline of the micronuclei levels. Moreover, we evaluate the changes in response to the toxic agents. Prospective cohorts studies will clarify the predictive value of micronuclei for cancer and chronic diseases for both children and adults. Novel molecular technologies will assist in the elucidation of different biological pathways and molecular mechanisms connected with the micronulcei levels in newborn and children.

BACKGROUND: Stenting of infrainguinal lesions can be rather challenging due to the mechanical stress applied on the arteries during motion. We assessed the short-term and midterm safety and efficacy of Supera interwoven nitinol stent for the treatment of infrainguinal arterial disease. EVIDENCE ACQUISITION: We conducted a systematic review for articles published from December 2011 up to May 2021 regarding studies assessing the safety and efficacy of Supera interwoven nitinol stents for the treatment of infra-inguinal peripheral arterial disease. Studies that involved synchronous application of the Supera stent and drug delivering devices, or any alternative endoprosthesis were excluded. Pooled Kaplan-Meier survival curves and smoothed hazard estimates were generated. Data were meta-analyzed using a random effects model. Primary endpoints included primary patency and freedom from clinically driven target lesion revascularization (TRL). Secondary endpoints included technical success and major amputation at 1 year post intervention. EVIDENCE SYNTHESIS: Seventeen studies with 2,015 patients (65.3% males) and a mean lesion length of 137.2 mm were included. Of the total treated lesions, 44.9% involved femoropopliteal artery and 37.4% the popliteal artery. Chronic total occlusions made up 49% of the treated lesions. There were no stent fractures reported. The pooled technical success rate is 99.84% (95% CI: 99.26-100). Pooled major amputation rate at 1 year is 1.48% (95% CI: 0.47-2.87). Pooled primary patency and freedom from TLR rates at 1 year are 83.5% (95% CI: 80.24-86.54) and 90.32% (95% CI: 88.75-91.79), respectively. Pooling of individual patient data produced primary patency and freedom from TLR rates of 84.48% (95% CI: 82.66-86.11) and 90.81% (95% CI: 88.64-92.58) respectively. According to the smoothed hazard ratio estimate the risk for losing primary patency peaked between four and five months while the risk for TLR peaked between seven and eight months after the intervention. CONCLUSIONS: This review and meta-analysis indicated the safety and efficacy of Supera stents for the treatment of challenging infrainguinal lesions in the short-term and mid-term periods, with acceptable primary patency and freedom from TLR rates. Clinicians should be aware that between four and five months patients face a higher risk for event occurrence.

Σκοπός: Οι αθηρωματικές πλάκες των στεφανιαίων αγγείων που προκαλούν οξέα στεφανιαία σύνδρομα, είναι συχνά μη στενωτικές και δεν προκαλούν συμπτώματα. Αυτές οι ευάλωτες πλάκες χαρακτηρίζονται από λεπτή ινώδη κάψα που περιβάλλει ένα μεγάλο νεκρωτικό λιπιδικό πυρήνα και επεκτατική αναδιαμόρφωση [(μη-στενωτικά ινωδοαθηρωματα με λεπτή ινώδη κάψα (TCFA)] και συνηθέστερα δεν απεικονίζονται με την στεφανιογραφία. Ο συνδυασμός της μελέτης λειτουργικών χαρακτηριστικών των αθηρωματικών πλακών [Ενδοθηλιακής Διατμητικής Τάσης (ΕΔΤ)] με ανατομικά και απεικονιστικά ευρήματα (ινώδης κάψα, λιπώδης πυρήνας) τα οποία μπορούν να εκτιμηθούν με μεγάλη ακρίβεια με τη χρήση ενδοστεφανιαίας οπτικής συνεκτικής τομογραφίας (OCT) μπορεί να συμβάλλει στην πρώιμη ανίχνευση των ευάλωτων αθηρωματικών πλακών.Μέθοδοι και αποτελέσματα: Μελετήθηκαν 35 στεφανιαίες αρτηρίες από 30 ασθενείς οι οποίοι και υποβλήθηκαν σε κλινικά ενδεδειγμένο καρδιακό καθετηριασμό και μελέτη με OCT. Σε συγκεκριμένο αριθμό ασθενών αυτών των ασθενών (ν=10) και αρτηριών (ν=10) πραγματοποιήθηκε και κλινικά ενδεδειγμένη μελέτη με IVUS. Αναπτύχθηκε μεθοδολογία τρισδιάστατης ανακατασκευής των στεφανιαίων αρτηριών με τη χρήση OCT και διεπίπεδης στεφανιογραφίας (3D OCT) και η ακρίβειά της επικυρώθηκε έναντι της μεθόδου εκλογής 3D IVUS. Κάθε μια από τις στεφανιαίες αρτηρίες ανακατασκευάστηκε τρισδιάστατα στο χώρο, μελετήθηκε η μορφολογία τους και προσομοιώθηκε η αιματική ροή εντός αυτών προς υπολογισμό της ΕΔΤ. Αναπτύχθηκε το shear stress score, ένα μέτρο εκτίμησης της βαρύτητας και της έκτασης της χαμηλής ΕΔΤ σε τμήματα κάθε στεφανιαίας αρτηρίας με παρουσία αθηροσκλήρωσης. Η χαμηλή ΕΔΤ παρουσιάζει συσχέτιση με τα μορφολογικά χαρακτηριστικά υψηλού κινδύνου-ευάλωτων πλακών. Τα μη-στενωτικά ινωδοαθηρωματα με λεπτή ινώδη κάψα παρατηρούνται κυρίως σε περιοχές με χαμηλή ΕΔΤ ενώ τα μη-στενωτικά ινωδοαθηρωματα με παχύτερη ινώδη κάψα κυρίως σε με μέτρια ΕΔΤ. Το shear stress score παρουσιάζει θετική συσχέτιση με τον αριθμό των TCFA σε κάθε δεδομένη αρτηρία.Συμπεράσματα:. H τοπική χαμηλή ΕΔΤ, το υψηλό shear stress score (που αντανακλά την έκταση του ενδοθηλίου όπου διαπιστώνεται χαμηλή ΕΔΤ), η επεκτατική αναδιαμόρφωση του αρτηριακού αυλού, το πάχος της ινώδους κάψας και ο όγκος του λιπιδικού πυρήνα είναι ευρήματα που μπορούν να μετρηθούν αξιόπιστα. Τα στοιχεία αυτά αποτελούν αντικειμενικούς δείκτες για την εκτίμηση της βαρύτητας της στεφανιαίας νόσου και του ενδεχόμενου καρδιαγγειακού κινδύνου, καθώς συνδέονται με τον σχηματισμό και την εξέλιξη ευάλωτων αθηρωματικών πλακών.

Στην παρούσα διδακτορική διατριβή έγινε η προσπάθεια να διασαφηνιστεί η δομή και ο ρόλος των ιδιοβλαστών που σχηματίζουν κυτταρικά έγκλειστα οξαλικού και ανθρακικού ασβεστίου στα φυτά Vitis vinifera (ευρωπαϊκή άμπελοςι), Amaranthus hybridus (βλήτο) και Parietaria judaica (περδικάκι) και η πιθανή συμβολή τους στη φωτοσύνθεση. Στη διεθνή βιβλιογραφία δεν υπάρχουν επαρκή πειραματικά δεδομένα σχετικά με το ρόλο των εγκλείστων αυτών. Η αποθήκευση και η ρύθμιση του Ca, η φυτοπροστασία και η αποτοξίνωση βαρέων μετάλλων είναι μερικοί από τους ρόλους που έχουν προταθεί. Ωστόσο, πειραματικά δεδομένα της ερευνητικής μας ομάδας έχουν δείξει ότι τα κυτταρικά έγκλειστα αποτελούν δυναμικές δομές και όχι τελικά προϊόντα του μεταβολισμού. Ως εκ τούτου διατυπώθηκε η υπόθεση εργασίας σύμφωνα με την οποία τα κυτταρικά έγκλειστα διαδραματίζουν σημαντικό ρόλο στη διαχείριση του άνθρακα των φωτοσυνθετικών ιστών και στη απόδοση CO2 για την πραγματοποίηση της φωτοσύνθεσης σε συνθήκες πενίας άνθρακα («φωτοσύνθεση κινδύνου»). Επίσης διατυπώθηκε η υπόθεση ότι τα έγκλειστα σχηματίζονται κατά τα πρώτα στάδια ανάπτυξης των φύλλων, όταν τα στομάτια δεν είναι πλήρως λειτουργικά. Λαμβάνοντας υπόψη τα παραπάνω πραγματοποιήθηκε πείραμα κατά το οποίο συλλέχθηκαν, και από τα τρία πειραματόφυτα, φύλλα διαφορετικής ηλικίας, εκφραζόμενης από την επιφάνειά τους, και στη συνέχεια μετρήθηκαν και συσχετίστηκαν οι παράμετροι: ρυθμός φωτοσύνθεσης, ρυθμός διαπνόης, υποστοματική συγκέντρωση CO2, στοματική αγωγιμότητα, πυκνότητα κυτταρικών εγκλείστων και συνολικός αριθμός κυτταρικών εγκλείστων ανά φύλλο. Τα αποτελέσματα έδειξαν πως τα κυτταρικά έγκλειστα σχηματίζονται από τα πρώτα κιόλας στάδια έκτπυξης των φύλλων. Η πυκνότητά τους είναι αρχικά υψηλή και στη συνέχεια μειώνεται και σταθεροποιείται. Αντίθετη πορεία ακολουθεί ο συνολικός αριθμός τους ο οποίος αυξάνεται αλλά φτάνει σύντομα σε μέγιστη τιμή όπου και σταθεροποιείται. Παράλληλα, τα φύλλα φωτοσυνθέτουν σε μικρό βαθμό από τα πρώτα κιόλας στάδια της έκπτυξης τους χωρίς ωστόσο να έχουν τη δυνατότητα ανταλλαγής αερίων.Σε επόμενο πείραμα ελέγχθηκε η μεταβολή του όγκου των κυτταρικών εγκλείστων στο βλήτο και το περδικάκι σε συνθήκες πενίας άνθρακα. Συνθήκες πενίας άνθρακα επικρατούν in planta όταν τα στομάτια κλείνουν είτε λόγω υδατικής καταπόνησης είτε λόγω επικράτησης δυσμενών καιρικών συνθηκών. Ένας αριθμός φύλλων κατηγοριοποιήθηκε σε δύο ομάδες χειρισμών: φύλλα control και φύλλα που αναπτύχθηκαν σε συνθήκες πενίας άνθρακα (υδατική καταπόνηση, εφαρμογή ΑΒΑ ή/και παραμονή στο σημείο αντιστάθμησης για το CO¬¬2). Μετά από παραμονή του κάθε χειρισμού κάτω από συνθήκες συνεχούς φωτισμού για 24 και 48 ώρες μετρήθηκε ο όγκος των κυτταρικών εγκλείστων. Τα αποτελέσματα έδειξαν πως τα φύλλα στα οποία εφαρμόστηκε πενία άνθρακα εμφάνισαν σημαντικά μικρότερο όγκο κυτταρικών εγκλειστων (40-50 %) σε σχέση με τα φύλλα control. Στο περδικάκι, η προσθήκη ανθρακικών στο θρεπτικό διάλυμα των φύλλων της πενίας άνθρακα αντέστρεψε το φαινόμενο και οι κυστόλιθοι επανέκτησαν τις αρχικές τιμές του όγκου τους σε μικρό χρονικό διάστημα.Το ανθρακικό ασβέστιο διαλύεται ελεύθερα στο υδατικό περιβάλλον, ενώ το οξαλικό οξύ εικάζεται πως οξειδώνεται ενζυμικά προς CO2 και Η2Ο2 με τη δράση του ενζύμου οξαλική οξειδάση. Για την επιβεβαίωση της λειτουργίας του μηχανισμού αυτού πραγματοποιήθηκε πείραμα ιστοχημικού εντοπισμού της οξαλικής οξειδάσης αλλά και της καταλάσης σε φύλλα control και φύλλα στα οποία εφαρμόστηκε πενία άνθρακα. Η οξαλική οξειδάση εντοπίστηκε στα κύτταρα του σπογγώδους παρεγχύματος των φύλλων και κυρίως στις κυτταρικές μεμβράνες ή τα κυτταρικά τοιχώματα. Μάλιστα σε κάθε χειρισμό δεν εμφανίστηκε χρώση στα κύτταρα του κολεού της δέσμης. Παρόμοια ήταν και τα αποτελέσματα σχετικά με την θέση της καταλάσης στους ιστούς των φύλλων. Ο βαθμός κατά τον οποίο αυξήθηκε η δραστικότητα της οξαλικής οξειδάσης ελέγθηκε με ενζυμική δοκιμή. Τα αποτελέσματα, αφενός επιβεβαίωσαν τον ιστοχημικό εντοπισμό αφού δραστικότητα εμφανίστηκε μόνο στο κλάσμα των μεμβρανών-κυτταρικών τοιχωμάτων, αφετέρου έδειξαν πως η δραστικότητα του ενζύμου στα φύλλα της πενίας άνθρακα ήταν 2 με 3 φορές υψηλότερη από ό,τι στα φύλλα control.Στους ίδιους χειρισμούς πραγματοποιήθηκε ιστοχημικός εντοπισμός των ελέυθερων κατίοντων ασβεστίου. Το ασβέστιο, τόσο στο περδικάκι όσο και στο βλήτο, εντοπίστηκε στους ιδιοβλάστες και στα γειτονικά τους κύτταρα. Στο περδικάκι χρώση εμφανίστηκε και στους δύο χειρισμούς, ενώ αντιθέτως στο βλήτο μόνο στα φυτά της πενίας άνθρακα.Συνοψίζοντας, το μέγεθος των κυτταρικών εγκλείστων μεταβάλλεται σε συνθήκες πενίας άνθρακα. Για τον έλεγχο του βαθμού στον οποίο συμβάλλουν στη φωτοσύνθεση πραγματοποιήθηκε, και στα τρία φυτικά είδη, μέτρηση των παραμέτρων της φωτοσύνθεσης με τη χρήση φθορισμομετρίας της χλωροφύλλης σε μεταβαλλόμενες συνθήκες φωτισμού και σύστασης της ατμόσφαιρας, τόσο σε φύλλα control όσο και σε φύλλα πενίας άνθρακα. Από τα αποτελέσματα προέκυψε πως αφενός τα φύλλα της πενίας άνθρακα διατηρούν έναν ικανοποιητικό φωτοσυνθετικό ρυθμό ακόμα και στις συνθήκες αυτές, αφετέρου, όταν παύσουν οι συνθήκες που προκαλούν την πενία άνθρακα, είναι σε θέση να αποκτήσουν υψηλό ρυθμό φωτοσυνθετικής δραστηριότητας πολύ συντομότερα από ότι τα control φύλλα.Υπάρχουν συνεπώς σαφείς ενδείξεις πως τα κυτταρικά έγκλειστα συμβάλλουν στη φωτοσυνθετική διαδικασία. Εκτιμάται πως στην περίπτωση του P. judaica οι κυστόλιθοι έχουν τη δυνατότητα να υποστηρίξουν ρυθμό φωτοσύνθεσης 14,27 μmol CO2 m-2 s-1 για 30 min περίπου, και στο A. hybridus ρυθμό φωτοσύνθεσης 13,01 μmol CO2 m-2 s-1 για 70 min.Τα αποτελέσματα επιβεβαιώνουν την υπόθεση εργασίας για τη λειτουργία των κυτταρικών εγκλείστων ως αποθεμάτων φωτοσυνθετικού άνθρακα. Η λειτουργία αυτή θα μπορούσε να χαρακτηριστεί ως «φωτοσύνθεση κινδύνου» η οποία επάγεται από την πενία άνθρακα και είναι ζωτικής σημασίας για την επιβίωση σε συνθήκες έλλειψης νερού. Εκτός αυτού, η φωτοσύνθεση κινδύνου δίδει τη δυνατότητα ανάκτησης του CO2 το οποίο απελευθερώνεται μέσω της νυκτερινής αναπνοής (του υπέργειου, αλλά και του υπόγειου τμήματος του φυτού) και επομένως εξοικονομούνται σκελετοί άνθρακα. Αποτρέπει την άνοδο της συγκέντρωσης του CO2 σε τοξικά επίπεδα στα κύτταρα, τουλάχιστον στη διάρκεια της νύκτας, όταν τα στόματα παραμένουν κλειστά και δεν υπάρχει δυνατότητα φωτοσυνθετικής αφομοίωσης του εκλυόμενου αναπνευστικού CO2. Επίσης δίνει τη δυνατότητα τροφοδοσίας των φύλλων με ανόργανο άνθρακα από το υπόγειο τμήμα του φυτού ή από το έδαφος, με ενδιάμεσο σταθμό ελέγχου της διάθεσής του τα κύτταρα που σχηματίζουν τους κρυστάλλους.

The recently reported 'alarm photosynthesis' acts as a biochemical process that assimilates CO2 derived from the decomposition of calcium oxalate crystals. This study examined whether CaCO3 cystoliths could also serve as CO2 pools, fulfilling a similar role. Shoots of Parietaria judaica were subjected to carbon starvation, abscisic acid (ABA), or bicarbonate treatments, and the volume of cystoliths and the photochemical parameters of photosystem II (PSII) were determined. The size of cystoliths was reduced under carbon starvation or ABA treatments, whereas it was restored by xylem-provided bicarbonate. Under carbon starvation, ABA, or bicarbonate treatments, the photochemical efficiency of PSII was higher, while non-photochemical quenching, representing the safe dissipation of excess PSII energy due to lack of electron sinks, was lower in treated samples compared with controls. This observation suggests the involvement of ABA or other carbon starvation cues in the release of subsidiary CO2 for photosynthesis, inevitably from an internal source, which could be the cystoliths. Carbon remobilized from cystoliths can be photosynthetically assimilated, thus acting as a safety valve under stress. Together with alarm photosynthesis, these results show a tight link between leaf carbon deposits and photosynthesis.

Background Redo coronary artery bypass grafting (CABG) can be performed with either the off-pump (OPCAB) or the on-pump (ONCAB) technique. Method Following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA), this meta- analysis compared the safety and efficacy of OPCAB versus ONCAB redo CABG. Results Twenty-three (23) eligible studies were included (OPCAB, n=2,085; ONCAB, n=3,245). Off-pump CABG significantly reduced the risk of perioperative death (defined as in-hospital or 30-day death rate), myocardial infarction, atrial fibrillation, and acute kidney injury. The two treatment approaches were comparable regarding 30-day stroke and late all-cause mortality. Conclusions Off-pump redo CABG resulted in lower perioperative death and periprocedural complication rates. No difference was observed in perioperative stroke rates and long-term survival between the two techniques.

In many plant species, carbon-calcium inclusion (calcium oxalate crystals or cystoliths containing calcium carbonate) formation is a fundamental part of their physiology even necessary for normal growth and development. Despite the long-standing studies on carbon-calcium inclusions, the alterations in their properties during leaf development and their possible association with the maturation of the photosynthetic machinery have not been previously examined. In order to acquire more insights into this subject, we examined three of the most common species bearing abundant inclusions of different types, i.e., Amaranthus hybridus, Vitis vinifera, and Parietaria judaica. Results of our study showed that, irrespective of species and type of inclusion, similar patterns in the alterations of their properties are observed during leaf maturation, except for some differences in cell differentiation and distribution between raphides and druses in Vitis vinifera. As expected, inclusion formation has taken place at very early developmental stages and maximum density was observed in very young leaves. Inclusion properties are changing in a coordinated way with leaf area and these modifications are compatible with the concept that each idioblast or lithocyst "services" a finite number/area of adjacent cells. This tight coordination is also evident at the whole leaf level. Moreover, we observed an association of the properties of carbon-calcium inclusions and gas exchange, suggesting a possible implication of these structures in photosynthesis.

Calcium oxalate crystals are widespread among animals and plants. In land plants, crystals often reach high amounts, up to 80% of dry biomass. They are formed within specific cells, and their accumulation constitutes a normal activity rather than a pathological symptom, as occurs in animals. Despite their ubiquity, our knowledge on the formation and the possible role(s) of these crystals remains limited. We show that the mesophyll crystals of pigweed (Amaranthus hybridus) exhibit diurnal volume changes with a gradual decrease during daytime and a total recovery during the night. Moreover, stable carbon isotope composition indicated that crystals are of nonatmospheric origin. Stomatal closure (under drought conditions or exogenous application of abscisic acid) was accompanied by crystal decomposition and by increased activity of oxalate oxidase that converts oxalate into CO2 Similar results were also observed under drought stress in Dianthus chinensis, Pelargonium peltatum, and Portulacaria afra Moreover, in A. hybridus, despite closed stomata, the leaf metabolic profiles combined with chlorophyll fluorescence measurements indicated active photosynthetic metabolism. In combination, calcium oxalate crystals in leaves can act as a biochemical reservoir that collects nonatmospheric carbon, mainly during the night. During the day, crystal degradation provides subsidiary carbon for photosynthetic assimilation, especially under drought conditions. This new photosynthetic path, with the suggested name "alarm photosynthesis," seems to provide a number of adaptive advantages, such as water economy, limitation of carbon losses to the atmosphere, and a lower risk of photoinhibition, roles that justify its vast presence in plants.

Background and Objectives Somatic mutations in the calreticulin gene (CALR) are detected in approximately 70% of patients with essential thrombocythemia (ET) and primary or secondary myelofibosis (MF), lacking the JAK2and MPLmutations. To determine the prevalence of CALRframeshift mutations in a population of MPN patients of Greek origin, we developed a rapid low-budget PCR-based assay and screened samples from 5 tertiary Haematology units. This is a first of its kind report of the Greek patient population that also disclosed novel CALRmutants. Methods MPN patient samples were collected from different clinical units and screened for JAK2and MPLmutations after informed consent was obtained. Negative samples were analyzed for the presence of CALRmutations. To this end, we developed a modified post Real Time PCR High Resolution Melting Curve analysis (HRM-A) protocol. Samples were subsequently confirmed by Sanger sequencing. Results Using this protocol we screened 173 MPN, JAK2and MPLmutation negative, patients of Greek origin, of whom 117 (67.63%) displayed a CALRexon 9 mutation. More specifically, mutations were detected in 90 out of 130 (69.23%) essential thrombocythaemia cases (ET), in 18 out of 33 (54.55%) primary myelofibrosis patients (pMF) and in 9 out of 10 (90%) cases of myelofibrosis secondary to ET (post-ET sMF). False positive results were not detected. The limit of detection (LoD) of our protocol was 2%. Furthermore, our study reavealed 6 rare novel mutations which are to be added in the COSMIC database. Conclusions Overall, our method could rapidly and cost-effectively detect the mutation status in a representative cohort of Greek patients; the mutation make-up in our group was not different from what has been published for other national groups.

Background: Recent findings show that a number of single nucleotide polymorphisms (SNPs) within the promoter region of the annexin A5-gene (ANXA5) reduce the expression of the reporter gene...

Significant progress in the molecular pathology of melanocytic tumors have revealed that benign neoplasms, so-called nevi, are initiated by gain-of-function mutations in one of several primary oncogenes, such as BRAF in acquired melanocytic nevi, NRAS in congenital nevi or GNAQ/GNA11 in blue nevi, with consequent MAPK and PI3K/AKT/mTOR activation. Secondary genetic alterations overcome tumor suppressive mechanisms and allow the progression to intermediate lesions characterized by TERT-p mutation or to invasive melanomas displaying disruption of tumor suppressor genes. Currently, melanoma is molecularly regarded as four different diseases, namely BRAF, NRAS, NF1 and the "triple wild type" subtypes, which are associated with particular clinicopathological features. Melanocytic Spitzoid lesions include benign Spitz nevus, atypical Spitz tumor (AST) and Spitzoid melanoma. This is a challenging diagnostic group, particularly with regard to the distinction between AST and Spitzoid melanoma on clinical and histological grounds. Molecular analysis has identified the presence of HRAS mutation, BAP1 loss (often accompanying by BRAF mutations) or several kinase fusions in distinct categories of Spitz tumors. These aberrations account for the rapid growth characteristic of Spitz nevi. Subsequent growth is halted by various tumor suppressive mechanisms abrogation of which allow the development of AST, now better classified as low-grade melanocytic tumor. Although at present ancillary genetic techniques have not been very helpful in the prediction of biological behavior of AST, they have defined distinct tumor subsets differing with regard to biology and histology. Finally, we discuss how novel molecular markers may assist the differential diagnosis of melanoma, particularly from malignant peripheral nerve sheath tumor (MPNST). It is anticipated that the significant progress in the field of molecular pathology regarding the various types of melanocytic tumors, will eventually contribute to a more accurate histologic categorization, prediction of biologic behavior and personalized treatment.

It has been suggested that myeloid neoplasms with isolated isochromosome 17q [ MN i(17q)] comprise a distinct entity with poor prognosis . However, literature reports show a considerable clinical and molecular heterogeneity. We describe a 58-year-old male patient who was diagnosed as refractory anemia with multilineage dysplasia and ringed sideroblasts with isolated i(17q) . Though he initially responded well to erythropoietin, he gradually progressed to an aggressive form of MDS/MPN refractory to azacytidine and died 29 months after first diagnosis. Notably, in contrast to disease advancement, his karyotype reverted to normal, whereas his mutational profile remained unchanged. To our knowledge this is the first report of karyotype normalization during disease progression in patients with MN i(17q) , suggesting that the i(17q) anomaly is dispensable for the leukemic transformation and highlighting the underlying clinical and molecular complexity which both have to be resolved before the establishment of MN with isolated i(17q) as a distinct entity.

Ribonucleotide Reductase (RNR) is the enzyme that converts ribonucleotides to deoxyribonucleotides required for DNA replication and repair. RNR consists of two subunits, termed subunit 1 (RRM1) and 2 (RRM2). Imbalance in the regulation of RNR activity and control of dNTPs' pool leads to genomic instability and increases mutation rate. The amount of the enzyme present in the cell and allosteric mechanisms with positive and negative effectors control its activity. 5-Azacytidine (AZA) has been found to be a potent RRM2 inhibitor in acute myeloid leukemia. High expression of RNR is associated with chemoresistance and poorer overall survival in several malignancies and antisense inhibitors that bind to the enzyme mRNA have shown promising results. Moreover, RNR overexpression is a potential mechanism for chemoresistance to nucleoside analogs competing for DNA incorporation. These important roles have made RNR an attractive therapeutic target. Aim. We investigated the potential role of RNR as a prognostic factor in patients with MDS and the correlation of bone marrow RNR mRNA levels with treatment response to AZA. Methods. Bone marrow samples were collected from patients with MDS at diagnosis. RNA extraction and reverse transcription were performed using standard protocols. A Taqman based real-time PCR was performed on a CFX96 RT-PCR system (Bio-Rad Laboratories, Hercules, CA, USA). For both the housekeeping and target genes, a Taqman primer/probe mix was used according to the manufacturer's instructions (Applied Biosystems, Foster City, CA, USA). RRM1 and RRM2 mRNA levels were expressed as an RRM1-2/beta-actin ratio. A High-Resolution Melt (HRM) analysis for mutation detection of 8 genes of interest in MDS (IDH1, IDH2, SRSF2, SFEB1, UAF1, ASXL2, DNMT3A, BCOR) and quantification of the methylation levels of RRM1/RRM2 promoters were also carried out in a subset of 63 samples. IBM SPSS statistics, version 23.0 (IBM Corporation, North Castle, NY, USA) was used for the analysis of the results. Results The study included 123 patients diagnosed with MDS per the WHO 2008/2016 classification; 98 of them were treated with AZA. The basic characteristics of the patients are shown in Table 1. The median mRNA levels of RRM1 were 4.2 times higher in non-responders (p=0.019) in comparison to responders. The levels of RRM2 did not differ between the two groups. We found no correlation of any other of the studied factors (MDS type, IPSS, IPSS-R, WPSS, karyotype risk, number of cytopenias, hemoglobin level, WBC and platelet count) to treatment response. The median Overall Survival (OS) of the cohort was 38.6 months. The median survival after treatment with AZA (OST) was 20.4 months. No statistically significant correlations of OS and OST with the levels of RRM1/2 mRNA were found. A statistically significant correlation was found between splicing mutations and lower RRM1 mRNA levels (p=0.044). They were also correlated with higher median age (79.7 vs 69.7, p=0.005) as well as with lower hemoglobin levels (median 9.4 vs 10.3 g/dL, p=0,021). SF3B1 mutation was found to be correlated with higher RRM2 mRNA levels (median 0.2850 vs 0.0241, p=0.023) and with increased platelet and WBC counts. There was no association between the epigenetic mutations and any of the studied variables. The methylation status of RRM1 and RRM2 was also correlated with treatment response. Higher methylation percentages of both RRM1 (median 64.3% vs 25.2%, p=0.004) and RRM2 (median 24.5% vs 11.8%, p=0.046) were found in responders. The methylation status was not correlated to the mutation status of the patients. No correlation was found between OS or OST and the methylation or mutation status. Discussion MDS patients with low RRM1 levels had a better response to AZA-treatment. Moreover, high RRM1 methylation status was observed to responders, thus making RNR a possible biomarker for predicting success of epigenetic treatment. This is theoretically sound, since inhibition of RR leads to deoxyribonucleotide pool reduction, which is fatal for DNA synthesis and repair. RNR activity is related directly to dNTPs production, therefore AZA-mediated inhibition is likely to cause disturbances in DNA synthesis. Although, patients bearing splicing mutations had low RRM1 levels, SF3B1+ patients did not respond to AZA. In conclusion, expression of RNR could be used as a prognostic factor for response to AZA treatment and a possible therapeutic target in MDS. Disclosures Symeonidis: MSD: Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding; Pfizer: Research Funding; Roche: Membership on an entity's Board of Directors or advisory committees, Research Funding; Sanofi: Research Funding; Tekeda: Membership on an entity's Board of Directors or advisory committees, Research Funding; Janssen: Membership on an entity's Board of Directors or advisory committees, Research Funding; Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Gilead: Membership on an entity's Board of Directors or advisory committees, Research Funding. Kotsianidis:Celgene: Research Funding. Pappa:Novartis: Honoraria, Research Funding, Speakers Bureau; Takeda: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Roche: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Celgene / GenesisPharma: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Amgen: Research Funding; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Gilead: Honoraria, Research Funding; Abbvie: Research Funding. Panayiotidis:Bayer: Other: Support of clinical trial.

Land plants face the perpetual dilemma of using atmospheric carbon dioxide for photosynthesis and losing water vapors, or saving water and reducing photosynthesis and thus growth. The reason behind this dilemma is that this simultaneous exchange of gases is accomplished through the same minute pores on leaf surfaces, called stomata. In a recent study we provided evidence that pigweed, an aggressive weed, attenuates this problem exploiting large crystals of calcium oxalate as dynamic carbon pools. This plant is able to photosynthesize even under drought conditions, when stomata are closed and water losses are limited, using carbon dioxide from crystal decomposition instead from the atmosphere. Abscisic acid, an alarm signal that causes stomatal closure seems to be implicated in this function and for this reason we named this path "alarm photosynthesis." The so-far "enigmatic," but highly conserved and widespread among plant species calcium oxalate crystals seem to play a crucial role in the survival of plants.

Introduction Patients with Philadelphia-negative myeloproliferative neoplasms (PN-MPN) namely polycythemia vera (PV), essential thrombocythemia (ET) and primary myelofibrosis (MF) are at a higher risk for arterial and venous thrombosis that constitute a major cause of morbidity and mortality. Global coagulation assays such as thromboelastography, may be more efficient to evaluate the patient's thrombotic risk. The aim of the present study was to examine the hemostatic profile of patients with PN-MPN and correlate it with clinical, laboratory, treatment, and molecular characteristics including mutational analysis of JAK2, MPL, CALR, and polymorphisms of poly(ADP ribose) polymerase (PARP1), since a correlation of specific mutations with PARP1 polymorphisms has been reported in the literature. Materials and methods The study included adult patients with a confirmed diagnosis of PN-MPN according to the revised 2016 WHO classification. A written informed consent was obtained from all patients. The presence of splenomegaly, vascular events, PN-MPN specific therapy, and anticoagulation treatment were recorded. All the patients were assessed with complete blood count, routine coagulation tests [PT, INR, aPTT and fibrinogen, D-Dimers analyzed with the automatic coagulation analyzer Sysmex (Siemens)], platelet function performed with PFA-100 (COL, EPI, ADP), and global hemostatic potential assessed with ROTEM® Tromboelastometry (EXTEM), recording clotting time (CT), clot formation time (CFT), maximum clot firmness (MCF), lysis index at 30 (LI30) and 60 (LI60) minutes, and α angle. Mutation profiles of JAK2, MPL and CALR were defined using peripheral blood DNA. JAK2 and MPL mutations were detected using a standard PCR and CALR mutations using an HRMA-PCR assay. The rs1136410/PARP-1 (V762A) single nucleotide polymorphism (SNP), was detected with an RFLP method using the enzyme AciΙ (New England Biolabs, USA) and the digestion products were evaluated by polyacrylamide gel electrophoresis. Statistical analysis was performed using IBM SPSS statistics, version 23.0 (IBM Corporation, North Castle, NY, USA). Results Seventy-four patients were included in the study (22 PV, 47 ET, 5 MF) with a median age of 63 years (25-87) and 68 healthy controls for the SNP/PARP1 study. At the time of sample collection, 71 (95.9%) patients were under treatment [hydroxyurea (HU), 57 (77.0%); anagrelide, 19 (25.7%); ruxolitinib, 9 (12.2%); interferon alpha, 2 (2.7%); an alkylating agent, 4 (5.4%)]. In terms of anticoagulation, 47 (63.5%) patients were on aspirin, 4 (5.4%) on clopidogrel, 7 (9.5%) on a combination of the two, 3 (4.1%) on a vitamin K antagonist, and 2 (2.7%) on a Xa-inhibitor. Twenty-two (29.7%) patients had abnormally high D-Dimers (>0,5mg/l). Nineteen (25.7%) patients had developed thrombosis after diagnosis (8, ischemic stroke; 4, coronary artery disease (CAD); 3, deep vein thrombosis (DVT); 1, symptomatic carotid stenosis; 2, a combination of stroke and CAD; 1, a combination of CAD and DVT). Among 69 patients who were not receiving anticoagulation, 5 (7,2%) had an abnormal CT, 4 (5,7%) had abnormal CFT, 3 (4,3%) had an increased α angle, and 18 (26%) had an increased MCF value. Women had shorter CFT, higher α angle, and higher MCF (p Patients receiving anagrelide or alkylating agents, had statistically significant shorter CFTs, higher α angles, and higher MCFs compared to the ones receiving HU. Among 54 patients taking aspirin COL-EPI was normal in 10. Among 11 patients taking clopidogrel COL-ADP was normal in 5, implying that the antiplatelet treatment may not be sufficient in certain cases. No correlations were found between PARP1 polymorphic status and any of the studied parameters, nor between patients and healthy controls. Discussion Global assays such as thromboelastography are more useful than conventional hemostatic laboratory tests in depicting the hypercoagulable state in MPN. They may be useful in combination with other parameters such as the mutational status in identifying patients with MPN at higher risk for thrombosis and guide clinicians for the type of treatment (both cytoreductive and anticoagulants). Tests of platelet function assessment may help the clinicians adjust the type and dose of antiplatelet therapy. Disclosures No relevant conflicts of interest to declare.

The coding region determinant-binding protein (CRD-BP) is an RNA binding protein that recognizes c-myc and IGF-II leader 3 mRNAs as well as the oncofetal H19 RNA. CRD-BP exhibits an oncofetal pattern of expression and has been detected in the majority of colon (81%), breast (58.5%) and sarcoma (73%) tumors. The study of CRD-BP expression was extended in brain tumors and Non small cell lung (NSCL) carcinomas and 12/24 malignant, 2/5 benign neuroepithelial tumors and 4/15 of NSCL carcinomas were found positive. All normal matching tissues tested were found negative. The highest frequency (60%) of CRD-BP positive tumors was observed in meningiomas, either benign (11/18) or atypical (3/3). These findings confirm that CRD-BP expression is restricted in tumors; the frequency of its de novo expression may vary according to tumor type and appears to be an early event in the transformation process. Cancer Lett

Concentrations of B-type natriuretic peptide (BNP) are recognised as a reliable marker of ventricular dysfunction in adults. In this study, plasma levels of BNP were determined in children with congenital heart disease (CHD) involving a left-to-right shunt, and were correlated with the shunt volume.Seventy-six children (38 boys/38 girls, mean age 22.4 months) with CHD (Group A: 31 with atrial septal defect [ASD], 23 with ventricular septal defect [VSD], 8 with ASD and VSD, 14 with patent ductus arteriosus [PDA]) and 34 healthy children (group B) were studied. BNP was measured by chemiluminescent microparticle immunoassay in all children. The amount of shunt (the ratio of pulmonary blood flow/systemic blood flow: Qp/Qs) was measured using Doppler velocimetry and two-dimensional echocardiography. A haemodynamically significant left-to-right shunt was defined as Qp/Qs1.5. Correlations were evaluated between all patient groups and healthy subjects and BNP was compared with echocardiographic data reflecting right and left ventricle volume overload.Thirty-four children of group A had Qp/Qs1.5 (group A1) and 42 Qp/Qs1.5 (group A2). BNP levels were higher in group A1 than group A2 (p=0.015), while there were no significant differences in BNP between group A2 and group B (p=0.79). BNP 24.4 pg/ml was determined as the cut-off point to identify patients with Qp/Qs1.5. BNP values were similar among patients with ASD and VSD, but they were significantly higher in patients with PDA. BNP was positively correlated with Qp/Qs (r=0.59, p0.001), and with the pulmonary artery velocity (r=0.27) and gradient (r=0.49), while there was a negative correlation with ejection fraction (r=-0.14). BNP levels were significantly higher in 10 infants with clinical signs of heart failure (p=0.025).These results, which are consistent with previous reports, suggest a possible role of BNP as an early diagnostic marker of the significance of shunt in children with CHD.

Obstructive sleep apnea syndrome (OSAS) is a common disease that is increasingly recognized among pediatric population. The exercise capacity of adults with OSAS has been demonstrated to be impaired, but there are no data about pediatric exercise response.The aim of this study was to evaluate cardiopulmonary response to exercise in children with OSAS and to correlate exercise capacity and severity of OSAS.Twenty-seven children with habitual snoring (Group A) (mean age 10.5 ± 1.8 years) referred for overnight polysomnography and 13 apparently healthy controls (mean age 11 ± 1.5 years) were recruited. According to the apnea hypopnea index (AHI) group A consisted of 15 (55.6%) children with mild OSAS and 12 (44.4%) with moderate-severe OSAS. All children completed a maximal ramping cardiopulmonary exercise test (CPET) on cycle ergometer.According to CPET children with OSAS had significantly lower VO2max (40.3 ± 8.4 ml/kg/min vs. 47.6 ± 7.9 ml/kg/min, P = 0.013) significantly lower VO2max (%) (77.7 ± 15 vs. 92.9 ± 10.5, P = 0.002), lower maximum heart-rate at peak exercise (86.6 ± 8.8 beat/min vs. 90.6 ± 7.2 beat/min) and higher systolic blood pressure level at peak exercise (145 ± 27.4 mmHg vs. 143.92 ± 20 mmHg) compared to control group.The present study demonstrates that young patients with OSAS, even with mild OSAS, had reduced exercise capacity as compared to control group.

Purpose: This study evaluated the feasibility, safety, and efficacy of primary stenting in atherosclerotic stenoses and occlusions of the infrarenal aorta. Methods: Between January 2003 and December 2006, 12 patients (6 men) with a mean age of 66.3 +/- 4.1 years who had infrarenal aortic occlusive disease were treated with primary stenting (aortic stenosis, 8; chronic total aortobiiliac occlusion, 4). Reasons for referral were severe claudication in six patients (50%), ischemic rest pain in four (33.3%), and minor tissue loss in two (16.7%). Three patients (25%) had chronic renal failure and were on dialysis. Follow-up was performed in all 12 patients. Results: Technical success was 91.7% because one patient had a residual stenosis >30% after stent placement and balloon postdilation owing to severe calcification of the aorta. However, clinical and immediate hemodynamic success was achieved in all 12 patients (100%). The preprocedural mean resting ankle-brachial index (ABI) values of 0.56 +/- 0.13 at the right side and 0.59 +/- 0.15 at the left were increased to 0.97 +/- 0.04 and 0.95 +/- 0.06, respectively, after treatment (P

Background Myeloproliferative neoplasms (MPNs) have been associated with a high incidence of thrombosis and bleeding episodes, which significantly contribute to disease-related morbidity and mortality. Clinical data indicate an association of the JAK2V617F mutation, seen in nearly all polycythemia vera (PV) cases and almost 60% of those with essential thrombocythemia (ET) and myelofibrosis (MF). The mutation is also seen in 37% of patients with in splachnic vein thrombosis (SVT) and its presence was associated with an increased risk for SVT. However, the prevalence of JAK2V617 seems to be low in patients with other thromboembolic events in unusual sites such as cerebral sinus, upper limb deep venous thrombosis (DVT). Additionally, activating mutations of MPL gene, seen in 3% of ET and 5% of MF patients, are considered as a significant risk factor for microvessel disturbances and have been associated with an increased risk of arterial thrombosis. Retinal vein occlusion (RVO) is a thrombotic complication in an uncommon site that may result in sight threatening disease. In this study we investigated the prevalence of JAK2V617F and MPLW515L/K mutations in a prospectively assembled cohort of patients with RVO, hypothesizing that some cases may be associated with an underlying undiagnosed MPN. Patients and Methods We studied 52 (23 males and 29 females) consecutive patients with no evidence of an underlying MPN who had been diagnosed with RVO confirmed with fluorangiography from January 2007 to September 2011. The mean age was 70 years (range: 49-85) Twenty eight patients (53.8%) presented with central RVO and 24 patients with branched RVO (46.5%). DNA was extracted from peripheral blood samples by standard procedures. The JAK2V617F mutation was detected using a tetra-primer amplification refractory mutation system (ARMS) polymerase chain reaction (PCR) assay with a sensitivity of 1% and the allele burden was estimated with a semi-quantitative method. MPLW515L/K were detected using allele-specific PCR (AS-PCR) assays with a sensitivity of 1%. Results Overall, MPN associated mutations were detected in 5/52 cases. JAK2V617F was detected in 2/52 cases (3.8%; 95%CI-1.4%-9%), while MPL exon 10 mutations were detected in 3/52 (5.7%; 95%CI-0.6%-12%). The JAK2V617F allele burden in the two positive patients was 45% and 52% respectively. Both patients who carried the JAK2V617F mutation were female. The first patient had been already diagnosed with ET according to the WHO criteria at the time of RVO screening. She was receiving hydroxyurea and aspirin and her platelet count was normal. The second patient who also carried the JAK2V617F mutation had a PLT count of 850.000/μl at the time of screening and was diagnosed with ET within the 3 following months. The patients with MPL mutations presented with normal blood counts. Conclusions Our findings indicate that a latent MPN could underlie RVO even in the absence of conventional diagnostic criteria. Our results represent the first report that MPL mutations could underlie RVO cases and suggest that routine screening of RVO cases for MPN mutations may be useful, especially in older patients. Disclosures: No relevant conflicts of interest to declare.