Η ανοσοϊστοχημική μελέτη της Κερατίνης 5/6 σε ουροθηλιακά καρκινώματα της ουροδόχου κύστης: συσχέτιση με περιβαλλοντικούς παράγοντες κινδύνου, την πρόγνωση και ο ρόλος στη μοριακή ταξινόμηση των καρκινωμάτων

Η Κερατίνη 5/6 (KRT 5/6 ή CK5/6) είναι μια επιθηλιακή πρωτεΐνη που σχετίζεται άμεσα με την κυτταρική διαφοροποίηση και την αποπτωτική διαδικασία. Η έκφρασή της χρησιμοποιείται τα τελευταία χρόνια στην μοριακή ταξινόμηση των καρκινωμάτων του μαστού, αποτελώντας βιοδείκτη που χαρακτηρίζει τον βασικό μοριακό υπότυπο. Κατά αντιστοιχία η διερεύνηση της έκφρασης της Κερατίνης 5/6 στο ουροθηλιακό καρκίνωμα της ουροδόχου κύστης αποτελεί βιοδείκτη για την ταξινόμηση των όγκων στη βασική ποικιλία (μοριακό υπότυπο), προσφέροντας χρήσιμα μορφολογικά και προγνωστικά στοιχεία που συμβάλλουν στην επιλογή της κατάλληλης θεραπευτικής αγωγής. Η παρούσα μελέτη στοχεύει στη διερεύνηση της έκφρασης της Κερατίνης 5/6 σε διηθητικά και μη διηθητικά ουροθηλιακά καρκινώματα της ουροδόχου κύστης σε σχέση με τις κλασσικές κλινικοπαθολοανατομικές παραμέτρους (ηλικία, φύλο, βαθμός κακοήθειας και στάδιο της νόσου). Σκοπός είναι η ανάδειξη της αξίας της CK5/6 ως προγνωστικός βιοδείκτης καθώς και η συμβολή για την μοριακή ταξινόμηση των όγκων στο βασικό μοριακό υπότυπο. Εφαρμόστηκε ανοσοϊστοχημική μέθοδος (με τη χρήση πολυμερούς αντισώματος συζευγμένου με HRP) για την ανίχνευση της Κερατίνης 5/6 σε τομές παραφίνης πάχους 4μm που προέρχονται από 120 ασθενείς. Παρατηρήθηκε θετική κυτταροπλασματική εντόπιση της CK5/6 σε 101 ασθενείς (84,2%). Από αυτούς οι 48(40%) εμφάνισαν εστιακή θετικότητα, ενώ οι 53(44,2%) εκτεταμένη θετικότητα. Κατά τη μονοπαραγοντική στατιστική αξιολόγηση των αποτελεσμάτων παρατηρήθηκε θετική συσχέτιση μεταξύ της έκφρασης της Κερατίνης 5/6 (ποσοστό έκφρασης) και του σταδίου της νόσου (p=0.037). Η ίδια θετική συσχέτιση αναδείχθηκε και όταν το στάδιο διακρίθηκε με βάση την παρουσία διήθησης του μυϊκού χιτώνα (pTa/pT1 έναντι pT2) (p=0.014). Επιπλέον παρατηρήθηκε θετική συσχέτιση με το βαθμό κακοήθειας (WHO 1973) (p=0.047). Λαμβάνοντας υπόψη την ένταση της ανοσοϊστοχημικής χρώσης της CK5/6 αναδείχθηκαν οι εξής στατιστικές συσχετίσεις: 1.θετική συσχέτιση με το στάδιο της νόσου (pTa,pT1,pT2) (p=0.001) 2.θετική συσχέτιση με το στάδιο της νόσου (pTa/pT1 έναντι pT2) (p=0.003) και 3.θετική συσχέτιση με το βαθμό κακοήθειας (WHO 1973) (p<0.001). Συμπερασματικά, φαίνεται ότι η CK5/6 σχετίζεται δυσμενώς με την εξέλιξη του ουροθηλιακού καρκινώματος της ουροδόχου κύστης, μέσω των συσχετίσεων με το βαθμό κακοήθειας και το στάδιο της νόσου. Τα ευρήματα αυτά συνάδουν με τη διεθνή βιβλιογραφία. Η CK5/6 μας δίνει τη δυνατότα για ταξινόμηση των θετικών περιπτώσεων στο βασικό μοριακό υπότυπο του ουροθηλιακού καρκινώματος. Ωστόσο, για την ακριβή ταξινόμηση κρίνεται απαραίτητη η συνεκτίμηση με τους λοιπούς δείκτες μοριακής ταξινόμησης που βρίσκονται στο στάδιο διερεύνησης.Κeratin 5/6 (KRT5/6) or Cytokeratin 5/6 (CK5/6) is an epithelial protein which has a significant biological role for cellular differentiation and apoptosis. Its expression is used nowadays for the molecular taxonomy of tumors in breast cancer, since it is a common biomarker of the basal molecular subtype. In the same spectrum, the expression of CK5/6 in urothelial carcinoma of bladder is considered a potent biomarker for the molecular taxonomy of the tumors and represents the basal subtype. The use of expression of CK5/6 is of great prognostic and strategic value for the study of the progression and the best therapeutic plan. This study examines the expression of CK5/6 in muscle invasive and non-muscle invasive urothelial carcinomas of bladder in correlation with the classic clinicopathological and anatomical parameters (age, gender, grade and stage). The aim of this study is to designate the value of CK5/6 as a prognostic biomarker and appoint its use for the characterisation of tumors as of basic subtype in molecular taxonomy. The expression of CK5/6 was found with immunohistochemical (IHC) method (polymer antibody bound with Horse Radish Peroxydase) in paraffin embedded tissue sections, 4nm thick, from 120 patients. Ιn the total pool, it was observed positive expression of CK5/6 in 101 tumors (84.2%) from which the 48 (40%) showed focal positive expression and the 53 (44.2%) showed positive expression. The univariate statistical analysis of the results showed positive correlation between the expression of CK5/6 (% of area stained) and the stage (pTa,pT1,pT2) (p=0.037). The same positive correlation was found between the expression of CK5/6 (% of area stained) and the stage of muscle invasion of the tumors (pTa/pT1, pT2). A positive correlation was also found between the expression of CK5/6 and grade (WHO 1973). From the evalution of the intensity of the CK5/6 IHC stain, it was found that: a) there is a positive correlation between the intensity of the stain and the stage (pTa,pT1,pT2) (p=0.001), b) there is a positive correlation between the intensity of the stain and the muscle invasion (pTa/pT1, pT2) (p=0.003) and c) there is positive correlation between the intensity of the stain and the grade (WHO 1973) (p<0.001). In conclusion, through the correlation of CK5/6 with stage, muscle invasion and grade, it was found that the expression of CK5/6 relates to poor prognosis, tumor aggression and progression in compliance with the latest bibliography. This research also concludes that the expression of CK5/6 can be used combined with other molecular biomarkers for the characterization of the basal molecular subtype in molecular taxonomy of the tumors in bladder

Growth Performance and Environmental Quality Indices and Biomarkers in a Co-Culture of the European Sea Bass with Filter and Deposit Feeders: A Case Study of an IMTA System

This study aimed to evaluate the efficiency of an integrated multi-trophic aquaculture (IMTA) system comprising co-cultured fed fish and organic extractive species representing three distinct trophic levels as well as the impact and potential utilization of two commercially available fish feeds made up of 35% fish meal (FM) and 20% fish meal (LFM) ingredients, using a multi-indicator assessment approach. Significant alterations were observed in growth performance indicators (GPIs), water and sediment quality indices, toxicity tests and biomarkers within the IMTA system. The fish survival, weight gain (WG), and specific growth rate (SGR) were higher in the IMTA system with significantly lower feed conversion ratios (FCRs) and higher feed efficiency (FE) in comparison to the fed fish monoculture system. Yet, organic filter feeders displayed 100% survival, and increased shell growth, while deposit feeders exhibited successful survival and significant weight gain. In the comparison between FM-IMTA and LFM-IMTA, fed fish in FM-IMTA showed higher WG, SGR, and FE with lower FCR. Environmental parameters like temperature, oxygen, and nutrient concentrations fluctuated but generally improved in the IMTA system, indicating lower mesotrophic conditions. Sediment fatty acid profiles differed between systems and toxicity assessments, which suggested a lower impact in IMTA and FM-IMTA systems. The sediment microbial community displayed high similarity within IMTA systems and between FM-IMTA and LFM-IMTA. These findings underscore the potential of IMTA systems for sustainable aquaculture, emphasizing improved growth performance and reduced environmental impact, particularly when using fish meal feeds

Immunohistochemical Study of Bladder Cancer Molecular Subtypes and Their Association with PD-L1 Expression

The significant heterogeneity in clinical outcomes among patients with bladder cancer has highlighted the existence of different biological subtypes of muscle-invasive bladder cancer (MIBC) and non-muscle-invasive bladder cancer (NMIBC). Meanwhile, immune checkpoint proteins and their interference with tumor-related immune-evasive strategies has led to the development of several immunotherapeutic drugs targeting programmed death-1 (PD-1) or programmed death ligand-1 (PD-L1). However, the lack of any known biomarker that could predict responses to immunotherapy has led to a more agnostic therapeutic approach. Here, we present a study conducted in 77 bladder cancer (BC) patients (n = 77), ranging from stages pTa to pT2. Tumor specimens were resected via transurethral resection of bladder tumor (TURBT) and consistuted of 24 low-grade (LG) and 53 high-grade (HG) tumors. Patients’ tumors were then categorized into molecular subtypes, via immunohistochemistry (CK5/6 and GATA3). Furthermore, all tumor specimens were stained with anti-PD-L1 and demonstrated significant correlations with basal immunophenotype, stage pT2 and HG tumors. As such, we attempted to stratify patients into groups of likely-responders and likely-not-responders to immunotherapy with anti-PD-L1, based on their molecular phenotype. Finally, in acknowledging the fact that there is a universal lack of biomarkers associated with predicting BC response to immunotherapeutic drugs, we tested all tumors for deficiency of mismatch repair proteins (MMR)

Immunohistochemical Study of Bladder Cancer Molecular Subtypes and Their Association with PD-L1 Expression

The significant heterogeneity in clinical outcomes among patients with bladder cancer has highlighted the existence of different biological subtypes of muscle-invasive bladder cancer (MIBC) and non-muscle-invasive bladder cancer (NMIBC). Meanwhile, immune checkpoint proteins and their interference with tumor-related immune-evasive strategies has led to the development of several immunotherapeutic drugs targeting programmed death-1 (PD-1) or programmed death ligand-1 (PD-L1). However, the lack of any known biomarker that could predict responses to immunotherapy has led to a more agnostic therapeutic approach. Here, we present a study conducted in 77 bladder cancer (BC) patients (n = 77), ranging from stages pTa to pT2. Tumor specimens were resected via transurethral resection of bladder tumor (TURBT) and consistuted of 24 low-grade (LG) and 53 high-grade (HG) tumors. Patients’ tumors were then categorized into molecular subtypes, via immunohistochemistry (CK5/6 and GATA3). Furthermore, all tumor specimens were stained with anti-PD-L1 and demonstrated significant correlations with basal immunophenotype, stage pT2 and HG tumors. As such, we attempted to stratify patients into groups of likely-responders and likely-not-responders to immunotherapy with anti-PD-L1, based on their molecular phenotype. Finally, in acknowledging the fact that there is a universal lack of biomarkers associated with predicting BC response to immunotherapeutic drugs, we tested all tumors for deficiency of mismatch repair proteins (MMR)