Bioengineering / Biyomühendislik
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Article 2’-Methylklavuzon Causes Lipid-Lowering Effects on A549 Non-Small Cell Lung Cancer Cells and Significant Changes on Dna Structure Evidenced by Fourier Transform Infrared Spectroscopy(Elsevier, 2020) Ceylan, Çağatay; Çağır, Ali; Aksoy, Hatice Nurdan; Ceylan, Çağatay; Çağır, Ali; Çetinkaya, Hakkı; 03.08. Department of Food Engineering; 04.01. Department of Chemistry; 03. Faculty of Engineering; 04. Faculty of Science; 01. Izmir Institute of TechnologyVarious chemical agents are used in the treatment of Non-Small Cell Lung Cancer (NSCLC). 2?-methylklavuzon was proposed as a potential chemotherapeutic agent in cancer treatment based on its topoisomerase inhibition activity. In this study the cellular effects of 2?-methylklavuzon was evaluated on A549 cancer cells using FTIR spectroscopy. 2?-methylklavuzon induced significant changes on both the whole cell lyophilizates and the lipid extracts of the A549 lung cancer cells. 2?-methylklavuzon caused significant structural changes in A549 cell DNA structure: T, A and G DNA breathing modes are lost after the drug application indicating the loss of topoisomerase activity. The level of transcription and RNA synthesis was enhanced. 2?-methylklavuzon induced single stranded DNA formation evidenced by the increase in the ratio of asymmetric/symmetric phosphate stretching modes. 2?-methylklavuzon induced band shifts only in the asymmetric mode of phosphate bonds not in the symmetrical phosphate bond stretching. 2?-methylklavuzon induced A form of DNA topography. In addition to the changes in the DNA structure and transcription 2?-methylklavuzon also caused lipid-lowering effect in A549 cancer cells. 2?-methylklavuzon suppressed lipid unsaturation, however, it induced formation of lipids with ring structures. 2?-methylklavuzon suppressed phosphate-containing lipids significantly and decreased carbonyl containing lipids and cholesterol slightly. 2?-methylklavuzon caused increases in the hydrocarbon chain length. Overall, 2?-methylklavuzon can be used as a lipid-lowering compound in the treatment of NSCLC and other cancer therapies. © 2020 Elsevier B.V.Article Citation - WoS: 47Citation - Scopus: 543d Printed Gelatin/Decellularized Bone Composite Scaffolds for Bone Tissue Engineering: Fabrication, Characterization and Cytocompatibility Study(Elsevier, 2022-06) Kara, Aylin; Tıhmınlıoğlu, Funda; Distler, Thomas; Kara, Aylin; Polley, Christian; Schneidereit, Dominik; Seitz, Hermann; Friedrich, Oliver; Tıhmınlıoğlu, Funda; Boccaccini, Aldo R; 03.02. Department of Chemical Engineering; 03.01. Department of Bioengineering; 03. Faculty of Engineering; 01. Izmir Institute of TechnologyThree-dimensional (3D) printing technology enables the design of personalized scaffolds with tunable pore size and composition. Combining decellularization and 3D printing techniques provides the opportunity to fabricate scaffolds with high potential to mimic native tissue. The aim of this study is to produce novel decellularized bone extracellular matrix (dbECM)-reinforced composite-scaffold that can be used as a biomaterial for bone tissue engineering. Decellularized bone particles (dbPTs, ∼100 μm diameter) were obtained from rabbit femur and used as a reinforcement agent by mixing with gelatin (GEL) in different concentrations. 3D scaffolds were fabricated by using an extrusion-based bioprinter and crosslinking with microbial transglutaminase (mTG) enzyme, followed by freeze-drying to obtain porous structures. Fabricated 3D scaffolds were characterized morphologically, mechanically, and chemically. Furthermore, MC3T3-E1 mouse pre-osteoblast cells were seeded on the dbPTs reinforced GEL scaffolds (GEL/dbPTs) and cultured for 21 days to assess cytocompatibility and cell attachment. We demonstrate the 3D-printability of dbPTs-reinforced GEL hydrogels and the achievement of homogenous distribution of the dbPTs in the whole scaffold structure, as well as bioactivity and cytocompatibility of GEL/dbPTs scaffolds. It was shown that Young's modulus and degradation rate of scaffolds were enhanced with increasing dbPTs content. Multiphoton microscopy imaging displayed the interaction of cells with dbPTs, indicating attachment and proliferation of cells around the particles as well as into the GEL-particle hydrogels. Our results demonstrate that GEL/dbPTs hydrogel formulations have potential for bone tissue engineering.Article Citation - WoS: 7Citation - Scopus: 8Absorbance-Based Detection of Arsenic in a Microfluidic System With Push-And Pumping(Elsevier, 2021) Karakuzu, Betül; Tekin, Hüseyin Cumhur; Gülmez, Yekta; Tekin, H. Cumhur; 03.01. Department of Bioengineering; 03. Faculty of Engineering; 01. Izmir Institute of TechnologyRapid and portable analysis of arsenic (As) contamination in drinking water is very important due to its adverse health effects on humans. Available commercial detection kits have shown low sensitivity and selectivity in analysis, and also they can generate harmful by-products. Microfluidic-based approaches allow portable analysis with gold nanoparticles (AuNPs) as labels. However, they need complex surface modification steps that complicate detection protocols. Due to the lack of precise sensing and affordable solution, we focused on developing a microfluidic platform that uses a push-and-pull pumping method for sensitive detection of As. In this detection principle, a sample is introduced in the microfluidic channel modified with -SH functional groups where As can bind. Then, AuNPs are given in the channel and AuNPs bind on free -SH functional groups which are not allocated with As. Absorbance measurements are conducted to detect AuNPs absorbed on the surfaces and the resulting absorbance value is inversely proportional with As concentration. The method enables detection of As down to 2.2 mu g/L concentration levels in drinking water, which is well-below the allowed maximum As concentration of 10 mu g/L in the drinking waters by the World Health Organization (WHO). The paper reveals that multiple push-and-pull pumping of fixed volume of sample and AuNPs with a syringe pump can improve the binding efficiency in the microfluidic channel. With this technique, low amounts of sample (1 mL) and short total assay time (25 min) are sufficient to detect As.Article Citation - WoS: 16Citation - Scopus: 21Adhesive Bonding Strategies To Fabricate High-Strength and Transparent 3d Printed Microfluidic Device(American Institute of Physics, 2020) Keçili, Seren; Tekin, Hüseyin Cumhur; Tekin, Hüseyin Cumhur; 03.01. Department of Bioengineering; 03. Faculty of Engineering; 01. Izmir Institute of TechnologyRecently, the use of 3D printing technologies has become prevalent in microfluidic applications. Although these technologies enable low-cost, rapid, and easy fabrication of microfluidic devices, fabricated devices suffer from optical opaqueness that inhibits their use for microscopic imaging. This study investigates bonding strategies using polydimethylsiloxane (PDMS) and printer resin as interlayer materials to fabricate high-strength optically transparent 3D-printed microfluidic devices. First, we fabricated microfluidic structures using a stereolithography 3D printer. We placed 3D-printed structures on interlayer materials coated surfaces. Then, we either let these 3D-printed structures rest on the coated slides or transferred them to new glass slides. We achieved bonding between 3D-printed structures and glass substrates with UV exposure for resin and with elevated temperature for PDMS interlayer materials. Bonding strength was investigated for different interlayer material thicknesses. We also analyzed the bright-field and fluorescence imaging capability of microfluidic devices fabricated using different bonding strategies. We achieve up to twofold (9.1 bar) improved bonding strength and comparable fluorescence sensitivity with respect to microfluidic devices fabricated using the traditional plasma activated PDMS-glass bonding method. Although stereolithography 3D printer allows fabrication of enclosed channels having dimensions down to similar to 600 mu m, monolithic transparent microfluidic channels with 280 x 110 mu m(2) cross section can be realized using adhesive interlayers. Furthermore, 3D-printed microfluidic chips can be integrated successfully with Protein-G modified substrates using resin interlayers for detection of fluorescent-labeled immunoglobulin down to similar to 30 ng/ml. Hence, this strategy can be applied to fabricate high-strength and transparent microfluidic chips for various optical imaging applications including biosensing.Article Citation - WoS: 17Citation - Scopus: 15Antiproliferative and Apoptotic Effects of Olive Leaf Extract Microcapsules on Mcf-7 and A549 Cancer Cells(American Chemical Society, 2023) Bal, Yıldız; Şanlı Mohamed, Gülşah; Sürmeli, Yusuf; Şanlı Mohamed, Gülşah; 04.01. Department of Chemistry; 04. Faculty of Science; 01. Izmir Institute of TechnologyAlginate microcapsules are a talented means for the delivery of broad curative biomacromolecules. In this study, we immobilized olive leaf extract (OLE) by calcium alginate (CA) and chitosan-coated CA (CCA) and characterized the OLE-loaded CA and CCA. The cytotoxic effect, the cell cycle arrest, and the apoptotic effect of OLE and its microcapsules were investigated against breast adenocarcinoma (MCF-7) and lung carcinoma (A549). As a result, the loading capacity of OLE-CA and OLE-CCA was found to be 80 and 99%, respectively, in optimal conditions. Also, OLE-CA and OLE-CCA were characterized by unique FTIR peaks and morphological display relative to the empty CCA microcapsules. The cytotoxicity analysis showed that the IC50 values of OLE-CA and OLE-CCA were determined to be 312 and 0.94 μg mL-1 against A549, respectively, whereas these were found to be 865.4 and 425.5 μg mL-1 for MCF-7 cells. On the other hand, the OLE microcapsules did not possess in any concentration of cytotoxic influence on the BEAS 2B healthy cell line. Also, the exposure of OLE-CCA to MCF-7 and A549 resulted in the arrest of more MCF-7 and A549 cells at the G0/G1 phase compared to the OLE. A549 and MCF-7 cells were predominantly found in the late apoptosis phase and necrosis phase, respectively. Optical microscopy images confirmed that OLE microcapsules were more effective against MCF-7 and A549 than free OLE. The present work suggested that the OLE microcapsules might be administered as nutrition supplements for cancer therapy. © 2023 The Authors. Published by American Chemical Society.Article Citation - WoS: 3Citation - Scopus: 4Applicability of Low-Intensity Vibrations as a Regulatory Factor on Stem and Progenitor Cell Populations(Bentham Science Publishers, 2020) Baskan, Öznur; Baskan, Öznur; Karadaş, Özge; Meşe Özçivici, Gülistan; Meşe, Gülistan; Özçivici, Engin; Özçivici, Engin; Meşe Özçivici, Gülistan; 03.01. Department of Bioengineering; 01. Izmir Institute of Technology; 04.03. Department of Molecular Biology and Genetics; 03. Faculty of Engineering; 04. Faculty of SciencePersistent and transient mechanical loads can act as biological signals on all levels of an organism. It is therefore not surprising that most cell types can sense and respond to mechanical loads, similar to their interaction with biochemical and electrical signals. The presence or absence of mechanical forces can be an important determinant of form, function and health of many tissue types. Along with naturally occurring mechanical loads, it is possible to manipulate and apply external physical loads on tissues in biomedical sciences, either for prevention or treatment of catabolism related to many factors, including aging, paralysis, sedentary lifestyles and spaceflight. Mechanical loads consist of many components in their applied signal form such as magnitude, frequency, duration and intervals. Even though high magnitude mechanical loads with low frequencies (e.g. running or weight lifting) induce anabolism in musculoskeletal tissues, their applicability as anabolic agents is limited because of the required compliance and physical health of the target population. On the other hand, it is possible to use low magnitude and high frequency (e.g. in a vibratory form) mechanical loads for anabolism as well. Cells, including stem cells of the musculoskeletal tissue, are sensitive to high frequency, low-intensity mechanical signals. This sensitivity can be utilized not only for the targeted treatment of tissues, but also for stem cell expansion, differentiation and biomaterial interaction in tissue engineering applications. In this review, we reported recent advances in the application of low-intensity vibrations on stem and progenitor cell populations. Modulation of cellular behavior with low-intensity vibrations as an alternative or complementary factor to biochemical and scaffold induced signals may represent an increase of capabilities in studies related to tissue engineering.Article Citation - WoS: 30Citation - Scopus: 35Bioactive Sheath/Core Nanofibers Containing Olive Leaf Extract(John Wiley and Sons Inc., 2016-01) Doğan, Gamze; Bayraktar, Oğuz; Başal, Güldemet; Erdoğan, İpek; Bayraktar, Oğuz; Özyıldız, Figen; Uzel, Ataç; Erdoğan, İpek; 04.03. Department of Molecular Biology and Genetics; 03.02. Department of Chemical Engineering; 03. Faculty of Engineering; 04. Faculty of Science; 01. Izmir Institute of TechnologyThis study aimed at producing silk fibroin (SF)/hyaluronic acid (HA) and olive leaf extract (OLE) nanofibers with sheath/core morphology by coaxial electrospinning method, determining their antimicrobial properties, and examining release profiles of OLE from these coaxial nanofibers. Optimum electrospinning process and solution parameters were determined to obtain uniform and bead-free coaxial nanofibers. Scanning electron microscopy and transmission electron microscopy (TEM) were used to characterize the morphology of the nanofibers. The antimicrobial activities of nanofibers were tested according to AATCC test method 100. Total phenolic content and total antioxidant activity were tested using in vitro batch release system. The quality and quantity of released components of OLE were determined by high-performance liquid chromatography. The changes in nanofibers were examined by Fourier-transform infrared spectroscopy. Uniform and bead-free nanofibers were produced successfully. TEM images confirmed the coaxial structure. OLE-loaded nanofibers demonstrated almost perfect antibacterial activities against both of gram-negative and gram-positive bacteria. Antifungal activity against C. albicans was rather poor. After a release period of 1 month, it was observed that ∼70-95% of the OLE was released from nanofibers and it was still bioactive. Overall results indicate that the resultant shell/core nanofibers have a great potential to be used as biomaterials.Article Citation - WoS: 26Citation - Scopus: 28Biocomposite Scaffolds for 3d Cell Culture: Propolis Enriched Polyvinyl Alcohol Nanofibers Favoring Cell Adhesion(John Wiley and Sons Inc., 2021-05) Bilginer, Rumeysa; Arslan Yıldız, Ahu; Özkendir İnanç, Dilce; Yıldız, Ümit Hakan; Yıldız, Ümit Hakan; Arslan Yıldız, Ahu; 04.01. Department of Chemistry; 03.01. Department of Bioengineering; 03. Faculty of Engineering; 04. Faculty of Science; 01. Izmir Institute of TechnologyThe objective of this work is generation of propolis/polyvinyl alcohol (PVA) scaffold by electrospinning for 3D cell culture. Here, PVA used as co-spinning agent since propolis alone cannot be easily processed by electrospinning methodology. Propolis takes charge in maximizing biological aspect of scaffold to facilitate cell attachment and proliferation. Morphological analysis showed size of the electrospun nanofibers varied between 172-523 nm and 345-687 nm in diameter, for non-crosslinked and crosslinked scaffolds, respectively. Incorporation of propolis resulted in desired surface properties of hybrid matrix, where hybrid scaffolds highly favored protein adsorption. To examine cell compatibility, NIH-3T3 and HeLa cells were seeded on propolis/PVA hybrid scaffold. Results confirmed that integration of propolis supported cell adhesion and cell proliferation. Also, results indicated electrospun propolis/PVA hybrid scaffold provide suitable microenvironment for cell culturing. Therefore, developed hybrid scaffold could be considered as potential candidate for 3D cell culture and tissue engineering.Article Citation - WoS: 5Citation - Scopus: 6Bioinspired Multi-Layer Biopolymer-Based Dental Implant Coating for Enhanced Osseointegration(Wiley, 2023-07) Üzülmez, Betül; Can, Özge; Demirsoy, Zeynep; Can, Özge; Gülseren, Gulcihan; 03.01. Department of Bioengineering; 03. Faculty of Engineering; 01. Izmir Institute of TechnologyThe major drawbacks of metal-based implants are weak osseointegration and post-operational infections. These limitations restrict the long-term use of implants that may cause severe tissue damage and replacement of the implant. Recent strategies to enhance the osseointegration process require an elaborate fabrication process and suffer from post-operative complications. To address the current challenges taking inspiration from the extracellular matrix (ECM), the current study is designed to establish enhanced osseointegration with lowered risk of infection. Natural biopolymer pectin, peptide amphiphiles, and enzyme-mimicking fullerene moieties are governed to present an ECM-like environment around the implant surfaces. This multifunctional approach promotes osseointegration via inducing biomineralization and osteoblast differentiation. Application of the biopolymer-based composite to the metal surfaces significantly enhances cellular attachment, supports the mineral deposition, and upregulates osteoblast-specific gene expression. In addition to the osteoinductive properties of the constructed layers, the inherent antimicrobial properties of multilayer coating are also used to prevent infection possibility. The reported biopolymer-artificial enzyme composite demonstrates antimicrobial activity against Escherichia coli and Bacillus subtilis as a multifunctional surface coating.Article Citation - WoS: 26Citation - Scopus: 28Biotransformation of Cyclocanthogenol by the Endophytic Fungus Alternaria Eureka 1e1bl1(Elsevier Ltd., 2018-07) Ekiz, Güner; Duman, Seda; Duman, Seda; Bedir, Erdal; Bedir, Erdal; 03.01. Department of Bioengineering; 01.01. Units Affiliated to the Rectorate; 01. Izmir Institute of Technology; 03. Faculty of EngineeringThe microbial transformation of cyclocanthogenol (CCG), Astragalus sp. originated sapogenin, by the endophytic fungus Alternaria eureka 1E1BL1 isolated from Astragalus angustifolius was investigated. Hydroxylation, oxidation, epoxidation, O-methylation, ring-expansion and methyl migration reactions were observed on the triterpenoid skeleton. As a result, eight metabolites were isolated and the structures of the previously undescribed compounds were established by 1-D, 2-D NMR and HR-MS analyses.Article Citation - WoS: 13Citation - Scopus: 14Biotransformation of Ruscogenins by Cunninghamella Blakesleeana Nrrl 1369 and Neoruscogenin by Endophytic Fungus Neosartorya Hiratsukae(Elsevier Ltd., 2018-08) Özçınar, Özge; Bedir, Erdal; Tağ, Özgür; Yusufoğlu, Hasan; Kıvçak, Bijen; Bedir, Erdal; 03.01. Department of Bioengineering; 03. Faculty of Engineering; 01. Izmir Institute of TechnologyBiotransformation of steroidal ruscogenins (neoruscogenin and ruscogenin) was carried out with Cunninghamella blakesleeana NRRL 1369 and endophytic fungus Neosartorya hiratsukae yielding mainly P450 monooxygenase products together with a glycosylated compound. Fermentation of ruscogenins (75:25, neoruscogenin-ruscogenin mixture) with C. blakesleeana yielded 8 previously undescribed hydroxylated compounds. Furthermore, microbial transformation of neoruscogenin by endophytic fungus N. hiratsukae afforded three previously undescribed neoruscogenin derivatives. While hydroxylation at C-7, C-12, C-14, C-21 with further oxidation at C-1 and C-7 were observed with C. blakesleeana, N. hiratsukae biotransformation provided C-7 and C-12 hydroxylated compounds along with C-12 oxidized and C-1(O) glycosylated derivatives. The structures of the metabolites were elucidated by 1-D (1H, 13C and DEPT135) and 2-D NMR (COSY, HMBC, HMQC, NOESY, ROESY) as well as HR-MS analyses.Article Citation - Scopus: 34Combination of Akt Inhibitor Arq 092 and Sorafenib Potentiates Inhibition of Tumor Progression in Cirrhotic Rat Model of Hepatocellular Carcinoma(Impact Journals, 2018) Macek Jilkova, Zuzana; Zeybek Kuyucu, Ayça; Zeybek Kuyucu, Ayça; Kurma, Keerthi; Tayébéh, Séyédéh; Pour, Ahmad; Roth, Gaël S.; Abbadessa, Giovanni; Yu, Yi; Schwartz, Brian; Sturm, Nathalie; Marche, Patrice N.; Hainaut, Pierre; Decaens, Thomas; 03.01. Department of Bioengineering; 03. Faculty of Engineering; 01. Izmir Institute of TechnologyThe prognosis of patients with advanced hepatocellular carcinoma (HCC) is very poor. The AKT pathway is activated in almost half of HCC cases and in addition, long term exposure to conventional drug treatment of HCC, sorafenib, often results in overactivation of AKT, leading to HCC resistance. Therefore, it is important to assess the safety and the efficacy of selective allosteric AKT inhibitor ARQ 092 (Miransertib) in combination with sorafenib. Here, we demonstrated in vitro that the combination of ARQ 092 with sorafenib synergistically suppressed proliferation, promoted apoptosis, and reduced migration. To test the effect of the combination in vivo, rats with diethylnitrosamine-induced cirrhosis and fully developed HCC were randomized and treated with vehicle, sorafenib, ARQ 092 or the combination of ARQ 092 with sorafenib; (n=7/group) for 6 weeks. Tumor progression, size of tumors and the mean tumor number were significantly reduced by the combination treatment compared to the control or single treatments. This effect was associated with a significant increase in apoptotic response and reduction in proliferation and angiogenesis. Sirius red staining showed a decrease in liver fibrosis. Moreover, treatments improved immune response in blood and in tumor microenvironment. Thus, the combination of ARQ 092 with sorafenib potentiates inhibition of tumor progression and gives the possibility of therapeutic improvement for patients with advanced HCC.Article Citation - WoS: 8Citation - Scopus: 11Cost-Effective and Rapid Prototyping of Pmma Microfluidic Device Via Polymer-Assisted Bonding(Springer, 2021) Sözmen, Alper Baran; Sözmen, Alper Baran; Arslan Yıldız, Ahu; Arslan Yıldız, Ahu; 01. Izmir Institute of Technology; 03.01. Department of Bioengineering; 03. Faculty of EngineeringMicrofluidic systems are relatively new technology field with a constant need of novel and practical manufacturing materials and methods. One of the main shortcomings of current methods is the inability to provide rapid bonding, with high bonding strength, and sound microchannel integrity. Herein we propose a novel method of assembly that overcomes the mentioned limitations. Polymer-assisted bonding is a novel, rapid, simple, and inexpensive method where a polymer is solubilized in a solvent and the constituted solution is used as a bonding agent. In this study, we combined this method with utilization of several phase-changing materials (PCMs) as channel-protective agents. Glauber's salt appeared to be more suitable as a channel-protective agent compared to rest of the salts that have been used in this study. Based on the bonding strength, quality analyses, leakage tests, and SEM imaging, the superior assisting bonding solvent was determined to be dichloromethane with a PMMA concentration of 2.5% (W/V). It showed a bonding strength of 23.794 MPa and a nearly non-visible bonding layer formation of 2.83 mu m in width which is proved by SEM imaging. The said combination of PCM, solvent, and polymer concentration also showed success in leakage tests and an application of micro-droplet generator fabrication. The application was carried out to test the applicability of developed prototyping methodology, which resulted in conclusive outcomes as the droplet generator simulation run in COMSOL Multiphysics version 5.1 software. In conclusion, the developed fabrication method promises simple, rapid, and strong bonding with sharp and clear micro-channel engraving.Article Citation - WoS: 14Citation - Scopus: 15Developing Wound Dressings Using 2-Deoxy To Induce Angiogenesis as a Backdoor Route for Stimulating the Production of Vascular Endothelial Growth Factor(MDPI Multidisciplinary Digital Publishing Institute, 2021-11) Dikici, Serkan; Dikici, Serkan; Yar, Muhammad; Bullock, Anthony J.; Shepherd, Joanna; Roman, Sabiniano; MacNeil, Sheila; 03.01. Department of Bioengineering; 03. Faculty of Engineering; 01. Izmir Institute of Technology2-deoxy-D-Ribose (2dDR) was first identified in 1930 in the structure of DNA and discovered as a degradation product of it later when the enzyme thymidine phosphorylase breaks down thymidine into thymine. In 2017, our research group explored the development of wound dressings based on the delivery of this sugar to induce angiogenesis in chronic wounds. In this review, we will survey the small volume of conflicting literature on this and related sugars, some of which are reported to be anti-angiogenic. We review the evidence of 2dDR having the ability to stimulate a range of pro-angiogenic activities in vitro and in a chick pro-angiogenic bioassay and to stimulate new blood vessel formation and wound healing in normal and diabetic rat models. The biological actions of 2dDR were found to be 80 to 100% as effective as VEGF in addition to upregulating the production of VEGF. We then demonstrated the uptake and delivery of the sugar from a range of experimental and commercial dressings. In conclusion, its pro-angiogenic properties combined with its improved stability on storage compared to VEGF, its low cost, and ease of incorporation into a range of established wound dressings make 2dDR an attractive alternative to VEGF for wound dressing development.Article Citation - WoS: 19Citation - Scopus: 19Development of Adjuvant Nanocarrier Systems for Seasonal Influenza a (h3n2) Vaccine Based on Astragaloside Vii and Gum Tragacanth (aps)(Elsevier, 2019) Yakuboğulları, Nilgün; Yakuboğulları, Nilgün; Genç, Rukan; Bedir, Erdal; Coven, Fethiye; Nalbantsoy, Ayşe; Bedir, Erdal; 01.01. Units Affiliated to the Rectorate; 03.01. Department of Bioengineering; 01. Izmir Institute of Technology; 03. Faculty of EngineeringAdjuvants are chemical/biological substances that are used in vaccines to increase the immunogenicity of antigens. A few adjuvants have been developed for use in human vaccines because of their limitations including lack of efficacy, unacceptable local or systemic toxicity, the difficulty of manufacturing, poor stability, and high cost. For that reasons, novel adjuvants/adjuvant systems are under search. Astragaloside VII (AST-VII), isolated from Astragalus trojanus, exhibited significant cellular and humoral immune responses. The polysaccharides (APS) obtained from the roots of Astragalus species have been used in traditional Chinese medicine and possess strong immunomodulatory properties. In the present study, the immunomodulatory effects of a newly developed nanocarrier system (APNS: APS containing carrier) and its AST-VII containing formulation (ANS: AST-VII + APNS), on seasonal influenza A (H3N2) vaccine were investigated. Inactivated H3N2 alone or its combinations with test compounds/formulations were intramuscularly injected into Swiss albino mice. Four weeks after immunization, the immune responses were evaluated in terms of antibody and cytokine responses as well as splenocyte proliferation. APNS demonstrated Th2 mediated response by increasing IgG1 antibody titers, whereas ANS showed response towards Th1/Th2 balance and Th17 by producing of IFN-gamma, IL-17A and IgG2a. Based on these results, we propose that APNS and ANS are good candidates to be utilized in seasonal influenza A vaccines as adjuvants/carrier systems. (C) 2019 Elsevier Ltd. All rights reserved.Article Citation - WoS: 7Citation - Scopus: 7Development of an Improved Amplex Red Peroxidation Activity Assay for Screening Cytochrome P450 Variants and Identification of a Novel Mutant of the Thermophilic Cyp119(Springer, 2020) Başlar, M. Semih; Güralp, Gülce; Sakallı, Tuğçe; Sürmeli, Nur Başak; Güralp, Gülce; Kestevur Doğru, Ekin; Haklı, Emre; Sürmeli, Nur Başak; 03.01. Department of Bioengineering; 01. Izmir Institute of Technology; 03. Faculty of EngineeringBiocatalysts are increasingly utilized in the synthesis of drugs and agrochemicals as an alternative to chemical catalysis. They are preferred in the synthesis of enantiopure products due to their high regioselectivity and enantioselectivity. Cytochrome P450 (P450) oxygenases are valuable biocatalysts, since they catalyze the oxidation of carbon-hydrogen bonds with high efficiency and selectivity. However, practical use of P450s is limited due to their need for expensive cofactors and electron transport partners. P450s can employ hydrogen peroxide (H2O2) as an oxygen and electron donor, but the reaction with H(2)O(2)is inefficient. The development of P450s that can use H(2)O(2)will expand their applications. Here, an assay that utilizes Amplex Red peroxidation, to rapidly screen H2O2-dependent activity of P450 mutants in cell lysate was developed. This assay was employed to identify mutants of CYP119, a thermophilic P450 fromSulfolobus acidocaldarius, with increased peroxidation activity. A mutant library of CYP119 containing substitutions in the heme active site was constructed via combinatorial active-site saturation test and screened for improved activity. Screening of 158 colonies led to five mutants with higher activity. Among improved variants, T213R/T214I was characterized. T213R/T214I exhibited fivefold higherk(cat)for Amplex Red peroxidation and twofold higherk(cat)for styrene epoxidation. T213R/T214I showed higher stability towards heme degradation by H2O2. While theK(m)for H(2)O(2)and styrene were not altered by the mutation, a fourfold decrease in the affinity for another substrate, lauric acid, was observed. In conclusion, Amplex Red peroxidation screening of CYP119 mutants yielded enzymes with increased peroxide-dependent activity. [GRAPHICS] .Article Citation - WoS: 11Citation - Scopus: 11Development of Tissue-Engineered Vascular Grafts From Decellularized Parsley Stems(Royal Society of Chemistry, 2023) Çevik, Merve; Dikici, Serkan; Dikici, Serkan; 03.01. Department of Bioengineering; 03. Faculty of Engineering; 01. Izmir Institute of TechnologyCardiovascular diseases are mostly associated with narrowing or blockage of blood vessels, and it is the most common cause of death worldwide. The use of vascular grafts is a promising approach to bypass or replace the blocked vessels for long-term treatment. Although autologous arteries or veins are the most preferred tissue sources for vascular bypass, the limited presence and poor quality of autologous vessels necessitate seeking alternative biomaterials. Recently, synthetic grafts have gained attention as an alternative to autologous grafts. However, the high failure rate of synthetic grafts has been reported primarily due to thrombosis, atherosclerosis, intimal hyperplasia, or infection. Thrombosis, the main reason for failure upon implantation, is associated with damage or absence of endothelial cell lining in the vascular graft's luminal surface. To overcome this, tissue-engineered vascular grafts (TEVGs) have come into prominence. Alongside the well-established scaffold manufacturing techniques, decellularized plant-based constructs have recently gained significant importance and are an emerging field in tissue engineering and regenerative medicine. Accordingly, in this study, we demonstrated the fabrication of tubular scaffolds from decellularized parsley stems and recellularized them with human endothelial cells to be used as a potential TEVG. Our results suggested that the native plant DNA was successfully removed, and soft tubular biomaterials were successfully manufactured via the chemical decellularization of the parsley stems. The decellularized parsley stems showed suitable mechanical and biological properties to be used as a TEVG material, and they provided a suitable environment for the culture of human endothelial cells to attach and create a pseudo endothelium prior to implantation. This study is the first one to demonstrate the potential of the parsley stems to be used as a potential TEVG biomaterial. © 2024 The Royal Society of Chemistry.Article Citation - WoS: 62Citation - Scopus: 67Effect of Peg Grafting Density and Hydrodynamic Volume on Gold Nanoparticle-Cell Interactions: an Investigation on Cell Cycle, Apoptosis, and Dna Damage(American Chemical Society, 2016-06-14) Uz, Metin; Alsoy Altınkaya, Sacide; Bulmuş, Volga; Bulmuş Zareie, Volga; Alsoy Altınkaya, Sacide; 03.02. Department of Chemical Engineering; 03.01. Department of Bioengineering; 03. Faculty of Engineering; 01. Izmir Institute of TechnologyIn this study, interactions of polyethylene glycol (PEG)-coated gold nanoparticles (AuNPs) with cells were investigated with particular focus on the relationship between the PEG layer properties (conformation, grafting density, and hydrodynamic volume) and cell cycle arrest, apoptosis, and DNA damage. Steric hindrance and PEG hydrodynamic volume controlled the protein adsorption, whereas the AuNP core size and PEG hydrodynamic volume were primary factors for cell uptake and viability. At all PEG grafting densities, the particles caused significant cell cycle arrest and DNA damage against CaCo2 and PC3 cells without apoptosis. However, at a particular PEG grafting density (∼0.65 chains/nm2), none of these severe damages were observed on 3T3 cells indicating discriminating behavior of the healthy (3T3) and cancer (PC3 and CaCo2) cells. It was concluded that the PEG grafting density and hydrodynamic volume, tuned with the PEG concentration and AuNP size, played an important role in particle-cell interactions.Article Citation - WoS: 32Citation - Scopus: 32Effects of N-Terminal and C-Terminal Polyhistidine Tag on the Stability and Function of the Thermophilic P450 Cyp119(Hindawi Publishing Corporation, 2019) Aslantaş, Yaprak; Sürmeli, Nur Başak; Sürmeli, Nur Başak; 03.01. Department of Bioengineering; 03. Faculty of Engineering; 01. Izmir Institute of TechnologyBiocatalysts are sought-after in synthesis of pharmaceuticals and agrochemicals due to their high regioselectivity and enantioselectivity. Among biocatalysts, heme-containing cytochrome P450 (P450) oxygenases are an attractive target since they catalyze oxidation of "unactivated" carbon-hydrogen bonds with high efficiency. CYP119 is an acidothermophilic P450 from Sulfolobus acidocaldarius, which has the potential to be widely used as a biocatalyst since it shows activity at high temperatures and low pH. Polyhistidine tags (His-tags) are widely used to simplify purification of proteins. However, His-tags can cause changes to protein structure and function. Here, we demonstrate the effects of His-tags on CYP119. To this end, the His-tags were cloned at the N-terminus or C-terminus of the CYP119, and His-tagged proteins were expressed and isolated. The thermostability and peroxidase activity of His-tagged CYP119s were tested and compared to wild type CYP119. Results indicated that while addition of His-tags increased the yield and simplified isolation of CYP119, they also influenced the electronic structure of active site and the activity of the protein. We show that N-terminal His-tagged CYP119 has desirable properties and potential to be used in industrial applications, but mechanistic studies using this protein need careful interpretation since the His-tag affects electronic properties of the active site heme iron.Article Citation - WoS: 9Citation - Scopus: 12An Electromechanical Lab-On Platform for Colorimetric Detection of Serum Creatinine(American Chemical Society, 2022-07) Karakuzu, Betül; Tekin, Hüseyin Cumhur; Tarım, Ergün Alperay; Öksüz, Cemre; Tekin, Hüseyin Cumhur; 03.01. Department of Bioengineering; 03. Faculty of Engineering; 01. Izmir Institute of TechnologyChronic kidney disease (CKD) is a high-cost disease that affects approximately one in ten people globally, progresses rapidly, results in kidney failure or dialysis, and triggers other diseases. Although clinically used serum creatinine tests are used to evaluate kidney functions, these tests are not suitable for frequent and regular control at-home settings that obstruct the regular monitoring of kidney functions, improving CKD management with early intervention. This study introduced a new electromechanical lab-on-a-chip platform for point-of-care detection of serum creatinine levels using colorimetric enzyme-linked immunosorbent assay (ELISA). The platform was composed of a chip containing microreservoirs, a stirring bar coated with creatinine-specific antibodies, and a phone to detect color generated via ELISA protocols to evaluate creatinine levels. An electromechanical system was used to move the stirring bar to different microreservoirs and stir it inside them to capture and detect serum creatinine in the sample. The presented platform allowed automated analysis of creatinine in ~50 min down to ~1 and ~2 mg/dL in phosphate-buffered saline (PBS) and fetal bovine serum (FBS), respectively. Phone camera measurements in hue, saturation, value (HSV) space showed sensitive analysis compared to a benchtop spectrophotometer that could allow low-cost analysis at point-of-care.
