Bioactive, functional and edible film-forming properties of isolated hazelnut (Corylus avellana L.) meal proteins

This study aimed characterization of bioactive, functional and edible film making properties of isolated proteins from untreated (HPI), hot extracted (HPI-H), acetone washed (HPI-AW), and acetone washed and hot extracted (HPC-AW-H) hazelnut meals. The most bioactive protein extract was HPC-AW-H, followed by HPI-AW, HPI-H and HPI, based on antioxidant activity (TEAC and ORAC: 158-461mmolTrolox/kg), iron chelation (60.7-126.7mmolEDTA/kg), angiotensin-converting enzyme inhibition (IC50: 0.57-1.0mg/mL) and antiproliferative activity on colon cancer cells (IC50: 3.0-4.6mg/ml). Protein contents of HPI, HPI-H and HPI-AW (93.3-94.5%) were higher than that of HPC-AW-H (86.0%), but HPC-AW-H showed the best pH-solubility profile. The extracts showed good oil absorption (7.4-9.4g/g) and foaming, but limited water holding and gelling capacities, and emulsion stability. The protein extracts gave transparent, yellowish to brownish and reddish colored and water soluble edible films. The HPI gave the lightest colored films with acceptable mechanical properties (elongation up to 144% and tensile strength up to 4.9MPa). 1-D and 2-D electrophoresis clearly showed the molecular and isoelectric profiles of hazelnut proteins. The overall results of this study showed that the bioactive, solubility and gelation properties of hazelnut proteins could be improved by simple processes like acetone washing and/or heat treatment. The hazelnut proteins are valuable as multipurpose food ingredients.Foundation of Izmir Institute of Technology (IZTECH

Fisetin, hesperetin ve viteksinin kronik miyeloid lösemi ve akut miyeloid lösemi hücreleri üzerine terapötik potansiyelleri

Thesis (Doctoral)--Izmir Institute of Technology, Molecular Biology and Genetics, Izmir, 2015Includes bibliographical references (leaves: 86-101)Text in English; Abstract: Turkish and Englishxi, 101 leavesFisetin, hesperetin and vitexin are plant-derived flavonoids. This thesis study aims to investigate therapeutic potentials of them on human HL60 APL and K562 CML cells since there are no studies on these cells. The effects of these compounds on APL and CML cells have been considered in terms of cytotoxicity, apoptosis and cell cycle progression. In this study, genome-wide microarray analysis has been also performed for APL and CML cells to identify the genes and networks that are responsible for fisetin and hesperetin-induced effects. In summary, we intented to explain the molecular mechanisms and global gene expression patterns related with the effects of these flavonoids on both APL and CML for the first time. There were decreases in the viability/proliferation of K562 and HL60 cells treated with fisetin, hesperetin and vitexin. Fisetin was the most effective flavonoid for the induction of apoptosis in both cells. Fisetin, hesperetin and vitexin have been found to affect cell cycle progression at different phases of the cell cycle in both CML and AML cells, thus having cytostatic effects. In conclusion, the results of this study indicated that especially fisetin and hesperetin may have therapeutic potential in APL and CML cells due to induction of apoptosis, inhibition of cell proliferation and cell cyle arrest. Moreover, the genetic networks derived from this study illuminate some of the biological pathways affected by fisetin and hesperetin treatment while providing a proof of principle for identifying candidate genes that might be targeted for CML and APL therapy

Purification and biochemical characterization of an extracellular lipase from psychrotolerant Pseudomonas fluorescens KE38

An extracellular lipase producing bacterium was isolated from a soil sample, and identified as a strain of Pseudomonas fluorescens by 16S rRNA gene sequencing. It was named Pseudomonas fluorescens KE38. KE38 showed psychrotolerant properties with an optimum growth temperature of 25 °C. The lipase enzyme secreted by KE38 was purified 41.13-fold with an overall yield of 54.99%, and a specific activity of 337.3 U/mg. The molecular mass of purified lipase was estimated to be approximately 43 kDa by SDS-PAGE. Although the lipase was active at a temperature range of 15-65 °C, it exhibited maximum activity at 45 °C, at pH 8.0. The enzyme exhibited high stability retaining 100% and 70% of its activity after an incubation period of 45 and 100 min at 45 °C and pH 8.0 respectively. It also showed a broad substrate specificity acting on p-nitrophenyl esters with C8-C18 acyl groups as substrates and was activated by Ca2+ and Ni2+ at 1 mM. While the enzyme retained its activity levels in the presence of a variety of organic solvents, DMSO and dimethylformamide enhanced this. High stability, broad substrate specificity and activity at cold temperatures in the presence of organic solvents, and metal ions make the extracellular lipase of KE38 a candidate for industrial applications.State Planning Agency of Turke

Resveratrol and quercetin-induced apoptosis of human 232B4 chronic lymphocytic leukemia cells by activation of caspase-3 and cell cycle arrest

Chronic lymphocytic leukemia (CLL), defined by accumulation of pathogenic B cells, has a very complex biology due to various factors such as inherited, host, and enviromental factors. Recently, finding new therapeutic agents or development of novel treatment strategies have been paid attention. Resveratrol and quercetin, important phytoalexins found in many plants, have been reported to have cytotoxic effects on various types of cancer. In this study, we examined cytotoxic, cytostatic, and apoptotic effects of these two important phenolic compounds on 232B4 human CLL cells. Cytotoxic effects of resveratrol and quercetin were determined by MTT cell proliferation assay. Changes in caspase-3 enzyme activity were measured using caspase-3 colorimetric assay. Annexin V-FITC/PI double staining was performed to measure apoptotic cell population. Effects of resveratrol and quercetin on cell cycle profiles of CLL cells were investigated by flow cytometry. Treatment of CLL cells with resveratrol and quercetin caused dose dependent inhibition of cell proliferation and increased apoptotic cell population through induction of caspase-3 activity. Cell cycle analysis displayed cell cycle arrest mainly in G0/G1 for both polyphenols. Our data, in total, showed for the first time that resveratrol and quercetin might block CLL growth through inducing apoptosis and cell cycle arrest.Turkish Academy of Sciences Outstanding Young Investigator Programm

Yeni doğal bir ürün olan KL-21 kronik lenfositik lösemi hücrelerinin çoğalmasını inhibe etmekte ve apoptozu indüklemektedir

Objective: The aims of this study were to examine the cytotoxic and apoptotic effects of KL-21, a novel plant product (produced by naturin natural Products, İzmir, Turkey), on 232B4 chronic lymphocytic leukemia (CLL) cells and to determine the cytotoxic effects on healthy BEAS-2B human bronchial epithelial cells. Materials and Methods: The cytotoxic effect of KL-21 was determined by MTT cell proliferation assay. Changes in caspase-3 enzyme activity were measured using the caspase-3 colorimetric assay. Changes in mitochondrial membrane potential were determined using the JC-1 dye-based method. Annexin V-FITC/PI double staining was performed to measure the apoptotic cell population. Effects of KL-21 on cell cycle profiles of CLL cells were investigated by flow cytometry. Results: We detected time- and concentration-dependent increases in the cytotoxic effect of KL-21 on 232B4 CLL cells. However, we also showed that, especially at higher concentrations, KL-21 was less cytotoxic towards BEAS-2B healthy cells than towards CLL cells. Annexin-V/PI double staining results showed that the apoptotic cell population increased in 232B4 cells. Increasing concentrations of KL-21 increased caspase-3 enzyme activity and induced loss of mitochondrial membrane potential. KL-21 administration resulted in small increases in the percentage of the cells in the G0/G1 phase while it decreased the S phase cell population up to 1 mg/mL. At the highest concentration, most of the cells accumulated in the G0/G1 phase. Conclusion: KL-21 has a growth-inhibitory effect on 232B4 CLL cells. KL-21 causes apoptosis and cell cycle arrest at G0/G1.Amaç: Bu çalışmanın amacı yeni bitkisel bir ürün olan KL-21’in [Naturin doğal ürünler şirketi (İzmir, Türkiye) tarafından üretilen] 232B4 kronik lenfostik lösemi (KLL) hücreleri üzerindeki sitotoksik ve apoptotik etkilerinin araştırılmasıdır. Ayrıca, KL-21’in BEAS-2B sağlıklı insan bronşial epitelyum hücreleri üzerindeki sitotoksik etkisine de bakılmıştır. Gereç ve Yöntemler: KL-21’in sitotoksik etkisine MTT hücre çoğalma testiyle bakılmıştır. Kaspaz-3 enzim aktivitesindeki ve mitokondri membran potansiyelindeki değişimlere sırasıyla kaspaz-3 kolorimetrik testi ve JC-1 boyasına dayalı bir yöntem kullanılarak bakılmıştır. Apoptotik hücre popülasyonunu belirlemek amacıyla Anneksin 5-FITC/PI ikili boyama yöntemi kullanılmıştır. KL-21’in KLL hücrelerinin hücre siklusu üzerindeki etkilerine akım sitometresi ile bakılmıştır. Bulgular: KL-21’in KLL hücrelerinin çoğalması üzerine etkisi zamana ve doza bağımlı olarak artmıştır. Bununlar beraber, KL- 21’in özellikle yüksek konsantrasyonlarda KLL hücreleri ile karşılaştırıldığında BEAS-2B hücreleri üzerinde daha az sitotoksik etki gösterdiği saptanmıştır. Anneksin-5/PI ikili boyaması 232B4 hücrelerinde apoptotik hücre popülasyonunun arttığını göstermiştir. KL-21’in artan konsantrayonları kaspaz-3 enzim aktivitesini arttırmış ve mitokondri membran potansiyelindeki kayıpları indüklemiştir. KL-21 1 mg/ml konsantrasyonuna kadar G0/G1 fazındaki hücrelerin yüzdesinde küçük artışlara ve S fazındaki hücre popülasyonunda ise azalmalara neden olmaktadır. En yüksek konsantrasyonda ise hücrelerin büyük bir çoğunluğu G0/G1 fazında birikmiştir. Sonuç: Elde edilen sonuçlar, KL-21’in 232B4 KLL hücreleri üzerinde büyümeyi inhibe edici bir etkisi olduğunu göstermiştir. Ayrıca, K-21 apoptozu indüklemekte ve G0/G1 fazında hücre siklusunun tutulumuna neden olmaktadır

Molecular mechanisms of drug resistance and its reversal in cancer

Chemotherapy is the main strategy for the treatment of cancer. However, the main problem limiting the success of chemotherapy is the development of multidrug resistance. The resistance can be intrinsic or acquired. The resistance phenotype is associated with the tumor cells that gain a cross-resistance to a large range of drugs that are structurally and functionally different. Multidrug resistance arises via many unrelated mechanisms, such as overexpression of energy-dependent efflux proteins, decrease in uptake of the agents, increase or alteration in drug targets, modification of cell cycle checkpoints, inactivation of the agents, compartmentalization of the agents, inhibition of apoptosis and aberrant bioactive sphingolipid metabolism. Exact elucidation of resistance mechanisms and molecular and biochemical approaches to overcome multidrug resistance have been a major goal in cancer research. This review comprises the mechanisms guiding multidrug resistance in cancer chemotherapy and also touches on approaches for reversing the resistance

Novel agents targeting bioactive sphingolipids for the treatment of cancer

Sphingolipids are a class of lipids that have important functions in a variety of cellular processes such as, differentiation, proliferation, senescence, apoptosis and chemotherapeutic resistance. The most widely studied bioactive shingolipids include ceramides, dihydroceramide (dhCer), ceramide-1-phosphate (C1P), glucosyl-ceramide (GluCer), sphingosine and sphingosine-1-phosphate (S1P). Although the length of fatty acid chain affects the physiological role, ceramides and sphingosine are known to induce apoptosis whereas C1P, S1P and GluCer induce proliferation of cells, which causes the development of chemoresistance. Previous studies have implicated the significance of bioactive shingolipids in oncogenesis, cancer progression and drug- and radiation-resistance. Therefore, targeting the elements of sphingolipid metabolism appears important for the development of novel therapeutics or to increase the effectiveness of the current treatment strategies. Some approaches involve the development of synthetic ceramide analogs, small molecule inhibitors of enzymes such as sphingosine kinase, acid ceramidase or ceramide synthase that catalyze ceramide catabolism or its conversion to various molecular species and S1P receptor antagonists. These approaches mainly aim to up-regulate the levels of apoptotic shingolipids while the proliferative ones are down-regulated, or to directly deliver cytotoxic sphingolipids like short-chain ceramide analogs to tumor cells. It is suggested that a combination therapy with conventional cytotoxic approaches while preventing the conversion of ceramide to S1P and consequently increasing the ceramide levels would be more beneficial. This review compiles the current knowledge about sphingolipids, and mainly focuses on novel agents modulating sphingolipid pathways that represent recent therapeutic strategies for the treatment of cancer. © 2013 Bentham Science Publishers

Enalapril-induced apoptosis of acute promyelocytic leukaemia cells involves STAT5A

Background: In this study, we aimed at evaluating the cytotoxic and apoptotic effects of enalapril on human HL60 acute promyelocytic leukaemia cells and at clarifying the roles of signal transducers and activator of transcription proteins (STATs) on enalapril-induced cell death. Materials and Methods: Cell viability and cytotoxicity tests were conducted by Trypan blue dye exclusion and 2,3-Bis[2-methoxy-4-nitro-5-sulphophenyl]-2H-tetrazolium-5- carboxanilide inner salt (XTT) assays, respectively. Apoptotic analyses were performed by the AnnexinV-enhanced green fluorescent protein (EGFP) staining method and by fluorescence microscopy. Expression levels of STAT3, -5A and -5B genes were analysed by quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR). Results: The results showed that enalapril reduced viability and proliferation, and induced apoptosis in HL60 cells in a dose-and time-dependent manner as compared to untreated controls. The expression levels of STAT5A gene were significantly reduced in enalapril-treated HL60 cells as compared to untreated controls. Conclusion: Taken together, all data showed for the first time that enalapril has significant anticancer potential for the treatment of acute premyelocytic leukaemia

Nilotinib does not alter the secretory functions of carotid artery endothelial cells in a prothrombotic or antithrombotic fashion

Background: There have been concerns about the possible prothrombotic effects of nilotinib, especially in patients having cardiovascular risk factors. The potential mechanism behind the increased risk of thromboembolic events is still not clear. Objectives: In this study, we aimed to evaluate possible harmful effects of nilotinib on endothelial cells. To this aim, we examined proliferative capacity and secretory functions of healthy human carotid artery endothelial cells (HCtAECs) in response to nilotinib. Methods: 3-(4,5-Dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide (MTT) cell proliferation method was used to determine antiproliferative effects of nilotinib on HCtAECs. The HCtAECs were incubated with 5, 10, and 100 nmol/L doses of nilotinib for 72 hours. Then, in order to assess the endothelial function, levels of nitric oxide (NO), von Willebrand factor (vWF), tissue plasminogen activator, plasminogen activator inhibitor 1 (PAI-1), and endothelin 1 (ET-1) were evaluated using enzyme-linked immunosorbent assay from tissue culture supernatants. Results: There were slight but statistically significant decreases in cell proliferation in response to nilotinib. Nilotinib increased the secretion of t-PA, PAI-1, and vWF in a dose-dependent manner when compared with the untreated control group. The ET-1 secretion was lower in 5 nmol/L and higher in 10 and 100 nmol/L nilotinib-treated cells as compared to untreated cells. Regarding NO secretion, lower levels were observed in 5 and 10 nmol/L, and higher levels were detected in 100 nmol/L nilotinib-treated cells as compared to untreated control group cells. Conclusion: Considering the results obtained in our study, nilotinib does not affect the functions of endothelial cells either in a prothrombotic or an antithrombotic fashion, despite a dose-dependent decline in cell viability

Revealing genome-wide mRNA and microRNA expression patterns in leukemic cells highlighted “hsa-miR-2278” as a tumor suppressor for regain of chemotherapeutic imatinib response due to targeting STAT5A

BCR-ABL oncoprotein stimulates cell proliferation and inhibits apoptosis in chronic myeloid leukemia (CML). For cure, imatinib is a widely used tyrosine kinase inhibitor, but developing chemotherapeutic resistance has to be overcome. In this study, we aimed to determine differing genome-wide microRNA (miRNA) and messenger RNA (mRNA) expression profiles in imatinib resistant (K562/IMA-3 μM) and parental cells by targeting STAT5A via small interfering RNA (siRNA) applications. After determining possible therapeutic miRNAs, we aimed to check their effects upon cell viability and proliferation, apoptosis, and find a possible miRNA::mRNA interaction to discover the molecular basis of imatinib resistance. We detected that miR-2278 and miR-1245b-3p were most significantly regulated miRNAs according to miRNome array. Upregulating miR-2278 expression resulted in the inhibition of resistant leukemic cell proliferation and induced apoptosis, whereas miR-1245b-3p did not exhibit therapeutic results. Functional analyses indicated that AKT2, STAM2, and STAT5A mRNAs were functional targets for miR-2278 as mimic transfection decreased their expressions both at transcriptional and translational level, thus highlighting miR-2278 as a tumor suppressor. This study provides new insights in discovering the mechanism of imatinib resistance due to upregulating the tumor-suppressor hsa-miR-2278 which stands for a functional therapeutic approach, inhibited leukemic cell proliferation, induced apoptosis, and regain of chemotherapeutic drug response in CML therapy. © 2015, International Society of Oncology and BioMarkers (ISOBM).Ege University Research Fund (APAK 2013-TIP/083