The relevance of estrogen-regulated expression of pS2 and cathepsin D proteins in primary operable breast carcinoma

Karcinom dojke je najzastupljenije maligno oboljenje i jedan od vodećih uzroka smrtnosti od malignih oboljenja među ženama u industrijski-razvijenim zemljama. Tok bolesti ranog karcinoma dojke u prve 3 godine praćenja toka bolesti se karakteriše pojavom maksimalnog broja udaljenih metastaza koji je nezavisan od primenjene terapije. Identifikacija karakteristika tumora/domaćina tumora koje bi preciznije ukazale na pacijente sa niskim/visokim rizikom za pojavu metastatske bolesti predstavlja važan predmet istraživanja. Cilj ovog istraživanja je bio određivanja granične vrednosti za definisanje povišene ekspresije i, posledično, statusa molekulskih biomarkera, pS2 i katepsina D, kao i identifikovanje grupe pacijentkinja sa niskim/visokim rizikom za relaps, u zavisnosti od statusa biomarkera, u ranom karcinomu dojke. Analiza povezanosti klasičnih i biohemijskih faktora prognoze/predikcije toka bolesti sa koncentracijama pS2 i katepsina D je omogućila definisanje graničnih vrednosti, za ekspresiju pS2 (15 ng/mg) i za ekspresiju katepsina D (39 pmol/mg) u karcinomu dojke, koje imaju i biološki značaj, u smislu definisanja estrogen-zavisne naspram estrogen-nezavisne ekspresije. Analiza toka bolesti pacijentkinja na osnovu statusa katepsina D, u periodu od 3 godine nakon hirurške terapije, je ukazala na biološki značaj, tj. na ulogu proteina u procesima proliferacije i apoptoze, i omogućila da se odredi njegov klinički značaj, tj. da se definišu visoko-rizične grupe (pN+ pT2 CD-, pN+ III CD-, I CD+), kao i nisko-rizična grupa pacijentkinja (I CD-), bez obzira na primenjenu terapiju. Analiza toka bolesti pacijentkinja na osnovu statusa pS2, u periodu od 3 godine nakon hirurške terapije, je ukazala na biološki značaj proteina, tj. na njegovu anti-proliferativnu i anti-apoptotsku, kao i ulogu ovog proteina u rasejavanju malignih ćelija, i omogućila da se odredi njegov klinički značaj, tj. da se definišu visoko-rizične (post pN+ pS2-, pT2 III pS2-, pre II ER+ PR+ pS2+) i nisko-rizične grupe pacijentkinja (post II ER+ PR+ pS2+, pre II ER+ PR+ pS2-), bez obzira na primenjenu terapijuBreast carcinoma is the most common malignancy and one of the leading causes of death from cancer among women in industrial countries. During the first 3 years of follow-up, early breast carcinoma is characterized by the maximum number of distant metastases, independent of applied therapy. Identification of the tumor/tumor-host characteristics that would specifically indicate patients with low/high risk for metastatic disease represents an important research goal. pS2 and cathepsin D were investigated in order to determine cut-off values for defining elevated expression and the status of biomarkers, providing the basis to identify groups of patients with low/high risk for relapse in early breast carcinoma. The relationship between classical and biochemical prognostic/predictive factors and concentrations of pS2 and cathepsin D allowed us to define the cut-off values for the expression of pS2 (15 ng/mg) and cathepsin D (39 pmol/mg) in breast carcinoma which have a biological relevance, in terms of defining the estrogen-dependent vs. estrogen-independent expression. Analysis of the first 3 years of follow-up on the basis of cathepsin D status indicated biological relevance, i.e. the role of the protein within the processes of proliferation and apoptosis, and allowed us to determine its clinical significance, i.e. to define high-risk groups (pN + pT2 CD-, pN + III CD-, I CD +) and low-risk group of patients (I CD-), regardless of applied therapy. Analysis of the first 3 years of follow-up on the basis of pS2 status indicated biological relevance, i.e. the anti- roliferative and anti-apoptotic as well as the role of this protein in dissemination of malignant cells, and allowed us to determine its clinical significance, i.e. to define high-risk (post pN + pS2-, pT2 III pS2-, pre II ER+ PR+ pS2+) and low-risk groups of patients (post II ER+ PR+ pS2+, pre II ER+ PR+ pS2-), regardless of applied therapy

Chemical Composition and Protective Possibilities of Juglans Nigra Leaves and Green Husks Extracts: DNA Binding and Micronucleus Assay in Human Lymphocytes

To better understand the mechanism of action of the compounds in the ethanolic extracts of J. nigra leaves and green husks, their binding to CT-DNA was investigated. This study was conducted to elucidate the in vitro protective effect of extracts against chromosomal damage in mitogen-induced human lymphocytes and investigate the possible application of selec+ted extracts as a natural source of polyphenolic compounds. Using HPLC-MS analysis, 103 different compounds were identified as having a higher number of active species, which is consistent with their activity. The frequency of micronuclei (MN) was scored in binucleated cells, and the nuclear proliferation index was calculated. Cyclic voltammetry experiments demonstrate that the nature of the interaction between extracts and CT-DNA is a synergy of electrostatic and intercalative modes, where leaves extracts showed a higher ability to bind to DNA. Extracts showed excellent antioxidant activity. At a concentration of only 4 μg/mL, extract of J. nigra leaves and the green husks reduced the incidence of MN by 58.2% and 64.5%, respectively, compared to control cell cultures

Application of Screen Printed Diamond Electrode, Coupled with “Point-of-Care” Platform, for Nanomolar Quantification of Phytonutrient Pterostilbene in Dietary Supplements: An Experimental Study Supported by Theory

Herein, a screen–printed diamond electrode (SPDE) coupled with a “point-of-care” platform (30 µL-drop concepts, single-drop-detection approach) was successfully applied for the electrochemical determination of pterostilbene (PTS). Cyclic voltammetry identified irreversible oxidation of PTS, where oxidation peak was shown to be strongly dependent on the pH of the working environmental. Although the proposition of the detailed electrochemical oxidation mechanism of PTS goes out of the scope of the present research, we have determined the most probable reactive site of our analyte, by utilizing DFT-based reactivity descriptors (Fukui functions). For electrochemical quantification of PTS, oxidation peak at 0.32 V (vs. Ag/AgCl) was followed in presence of 0.5 mol L−1 of Briton–Robinson buffer solution (pH = 9). Coupled with the optimized parameters of differential pulse voltammetry (DPV), SPDE detected PTS in two linear ranges (first range was from 0.011 to 0.912 µmol L−1; second range was from 0.912 to 4.420 µmol L−1), providing the LOD and LOQ on a nanomolar level (3.1 nmol L−1 and 10.0 nmol L−1, respectively). The selectivity of the optimized DPV method was found to be excellent, with the current changes of less than 7%, in the presence of ten times higher concentrations of the certain interferences. The practical applicability of the SPDE and single-drop-detection approach in dietary supplements (with a declared PTS content of 50 mg/tablet), with the recovery values ranging from 95 to 102%, shows that the developed method has high potential for precise and accurate PTS detection, as well as exceptional miniaturization possibilities of relevant equipment for on-site sensing

Application of Screen Printed Diamond Electrode, Coupled with “Point-of-Care” Platform, for Nanomolar Quantification of Phytonutrient Pterostilbene in Dietary Supplements: An Experimental Study Supported by Theory

Herein, a screen–printed diamond electrode (SPDE) coupled with a “point-of-care” platform (30 µL-drop concepts, single-drop-detection approach) was successfully applied for the electrochemical determination of pterostilbene (PTS). Cyclic voltammetry identified irreversible oxidation of PTS, where oxidation peak was shown to be strongly dependent on the pH of the working environmental. Although the proposition of the detailed electrochemical oxidation mechanism of PTS goes out of the scope of the present research, we have determined the most probable reactive site of our analyte, by utilizing DFT-based reactivity descriptors (Fukui functions). For electrochemical quantification of PTS, oxidation peak at 0.32 V (vs. Ag/AgCl) was followed in presence of 0.5 mol L−1 of Briton–Robinson buffer solution (pH = 9). Coupled with the optimized parameters of differential pulse voltammetry (DPV), SPDE detected PTS in two linear ranges (first range was from 0.011 to 0.912 µmol L−1; second range was from 0.912 to 4.420 µmol L−1), providing the LOD and LOQ on a nanomolar level (3.1 nmol L−1 and 10.0 nmol L−1, respectively). The selectivity of the optimized DPV method was found to be excellent, with the current changes of less than 7%, in the presence of ten times higher concentrations of the certain interferences. The practical applicability of the SPDE and single-drop-detection approach in dietary supplements (with a declared PTS content of 50 mg/tablet), with the recovery values ranging from 95 to 102%, shows that the developed method has high potential for precise and accurate PTS detection, as well as exceptional miniaturization possibilities of relevant equipment for on-site sensing

Application of Screen Printed Diamond Electrode, Coupled with “Point-of-Care” Platform, for Nanomolar Quantification of Phytonutrient Pterostilbene in Dietary Supplements: An Experimental Study Supported by Theory

Herein, a screen–printed diamond electrode (SPDE) coupled with a “point-of-care” platform (30 µL-drop concepts, single-drop-detection approach) was successfully applied for the electrochemical determination of pterostilbene (PTS). Cyclic voltammetry identified irreversible oxidation of PTS, where oxidation peak was shown to be strongly dependent on the pH of the working environmental. Although the proposition of the detailed electrochemical oxidation mechanism of PTS goes out of the scope of the present research, we have determined the most probable reactive site of our analyte, by utilizing DFT-based reactivity descriptors (Fukui functions). For electrochemical quantification of PTS, oxidation peak at 0.32 V (vs. Ag/AgCl) was followed in presence of 0.5 mol L−1 of Briton–Robinson buffer solution (pH = 9). Coupled with the optimized parameters of differential pulse voltammetry (DPV), SPDE detected PTS in two linear ranges (first range was from 0.011 to 0.912 µmol L−1; second range was from 0.912 to 4.420 µmol L−1), providing the LOD and LOQ on a nanomolar level (3.1 nmol L−1 and 10.0 nmol L−1, respectively). The selectivity of the optimized DPV method was found to be excellent, with the current changes of less than 7%, in the presence of ten times higher concentrations of the certain interferences. The practical applicability of the SPDE and single-drop-detection approach in dietary supplements (with a declared PTS content of 50 mg/tablet), with the recovery values ranging from 95 to 102%, shows that the developed method has high potential for precise and accurate PTS detection, as well as exceptional miniaturization possibilities of relevant equipment for on-site sensing

Angiogenesis: bFGF and VEGF in breast carcinoma

Angiogenesis, or neovascularization, is a complex process leading to formation of new blood vessels from the pre-existing vascular network of the tissue. Angiogenesis plays a central role in various physiological and pathological conditions, including embryonic development, reproduction, inflammation and wound healing, infertility, heart diseases, ulcers, rheumatoid arthritis, diabetic blindness and cancer. It is a multistep process involving EC activation, basement membrane and extracellular matrix (ECM) degradation, EC proliferation, migration and differentiation, synthesis of new basement membrane and maturation of new blood vessels. Tumor vasculature is considered to be of an "immature" nature with series of structural abnormalities. There are reciprocal paracrine interactions between ECs, tumor cells, stroma and ECM. Angiogenesis plays a key role in transformation of normal to malignant cell, tumor progression and metastasis. It is similar to the metastatic process in that it requires EC attachment, proteolysis, and locomotion to proceed. A close relationship exists between the tumor and ECs invasiveness of the tissue. The switch to the angiogenic phenotype involves a change in the local equilibrium between positive and negative regulators of the growth of microvessels. Basic fibroblast growth factor (bFGF) and vas­cular endothelial growth factor (VEGF) are positive regulators of angiogenesis. Intimate cross-talk exists among bFGF and the different members of the VEGF family during angiogenesis, lymphangiogenesis, and vasculogenesis. A substantial body of experimental evidence supports the hypothesis that angiogenesis and angiogenic factors may be strong prognostic and predictive factors in breast carcinoma. This article reviews the current knowledge on angiogenesis and its positive regulators: bFGF and VEGF.

The importance of simultaneous determination of breast cancer biomarkers

It is shown that steroid hormone receptors by themselves are not sufficiently strong prognostic factors in management of breast cancer. For that reason, simultaneous consideration of different biomarkers seems to be more appropriate for clinical use, i.e. selections of patients with high/inter-mediate/low risk of disease outcome. However, the amount of tumor material available from breast carcinoma can preclude determination of estrogen- regulated biomarkers together with estrogen receptor and progesterone receptor. The aim of this study was to assess the possibility of estrogen receptor and progesterone receptor determination by a single-point instead of five-point biochemical method. Our results demonstrated that the correlation between measurements of estrogen and progesterone receptor contents obtained by the five-point and single-point assay in the total population was very high. Consequently, we could use the single-point assay instead of five-point assay for estrogen receptor and progesterone receptor determination, thus making possible determination of other molecular biomarkers from the same breast carcinoma

Application of Screen Printed Diamond Electrode, Coupled with “Point-of-Care” Platform, for Nanomolar Quantification of Phytonutrient Pterostilbene in Dietary Supplements: An Experimental Study Supported by Theory

Herein, a screen–printed diamond electrode (SPDE) coupled with a “point-of-care” platform (30 µL-drop concepts, single-drop-detection approach) was successfully applied for the electrochemical determination of pterostilbene (PTS). Cyclic voltammetry identified irreversible oxidation of PTS, where oxidation peak was shown to be strongly dependent on the pH of the working environmental. Although the proposition of the detailed electrochemical oxidation mechanism of PTS goes out of the scope of the present research, we have determined the most probable reactive site of our analyte, by utilizing DFT-based reactivity descriptors (Fukui functions). For electrochemical quantification of PTS, oxidation peak at 0.32 V (vs. Ag/AgCl) was followed in presence of 0.5 mol L−1 of Briton–Robinson buffer solution (pH = 9). Coupled with the optimized parameters of differential pulse voltammetry (DPV), SPDE detected PTS in two linear ranges (first range was from 0.011 to 0.912 µmol L−1; second range was from 0.912 to 4.420 µmol L−1), providing the LOD and LOQ on a nanomolar level (3.1 nmol L−1 and 10.0 nmol L−1, respectively). The selectivity of the optimized DPV method was found to be excellent, with the current changes of less than 7%, in the presence of ten times higher concentrations of the certain interferences. The practical applicability of the SPDE and single-drop-detection approach in dietary supplements (with a declared PTS content of 50 mg/tablet), with the recovery values ranging from 95 to 102%, shows that the developed method has high potential for precise and accurate PTS detection, as well as exceptional miniaturization possibilities of relevant equipment for on-site sensing

Optimization of radioprotective dose of Juglans nigra leaf extract using response surface methodology

J. nigra leaf contains mixture of various pharmacologically active compounds and it is assumed that they may have the potential radioprotective effect. The purpose of this work was to predict radioprotective doses by correlating changes in organ distribution of radiopharmaceuticals with extract dose levels and rat body weight using response surface methodology (RSM) based on a second-order polynomial equation. The extract was administered daily by oral gavage to rats at dose of 6.9, 10.3, or 13.7 mg kg−1 body weight (bw) day−1 for 10 days. On the eleventh day, 0.1 ml of the one radiopharmaceutical (Na99mTcO4, 99mTc-dimercaptosuccinic acid and 99mTc-tin colloid) was injected into the tail vein. The statistical parameters: the coefficient of determination (0.81–0.95), the coefficient of variation (3.05–11.1), the adequate precision (8.82–19.12) and the mean relative percentage deviation (± 2.3–8.2) were indicated the precision and reliability of RSM. Using RSM, the predicted daily dose of leaf extract ranging from 11.19 to 11.99 mg kg−1 bw may be considered as an optimal daily radiopotective dose for protection of organs from radiation in cases of planned radiation exposures