Molecular Biology and Genetics / Moleküler Biyoloji ve Genetik
Permanent URI for this collectionhttps://hdl.handle.net/11147/9
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Article Blank Frame and Intensity Variation Distortion Detection and Restoration Pipeline for Phase-Contrast Microscopy Time-Lapse Images(Aves, 2024) Ucar, Mahmut; Iheme, Leonardo O.; Onal, Sevgi; Pesen-Okvur, Devrim; Yalcin-Ozuysal, Ozden; Toreyin, Behcet U.; Unay, DevrimIn this study, we propose a preprocessing pipeline for the detection and correction of distorted frames in time-lapse images obtained from phase-contrast microscopy. The proposed pipeline employs the average intensities of frames as a foundational element for the analysis. In order to evaluate the degree of correction required for intensity variance, a normalization technique is applied to the difference between the average intensity of a specific frame and the median average intensity of all frames within the study. Our restoration method increases the histogram similarity between the distorted and non-distorted frames, preserves trans-passing pixels in regions of interest, and mitigates the development of additional distortions. The efficacy of the proposed method was evaluated using 15 395 time-lapse image frames from 27 experiments using our own dataset and 830 time-lapse images from four different experiments obtained from the cell tracking challenge. The results of the validation demonstrate a high degree of numerical and visual accuracy of the proposed pipeline.Article Epithelial-Mesenchymal Transition as a Potential Route for Dapt Resistance in Breast Cancer Cells(Walter de Gruyter GmbH, 2023) Tellı, Kubra; Ozuysal, Ozden Yalcın; Telli, Kübra; Yalçın Özuysal, ÖzdenObjectives: Notch is a conserved pathway involved in cell- fate determination and homeostasis. Its dysregulation plays a role in poor prognosis and drug resistance in breast cancer. Targeting Notch signaling via inhibition of the gamma- secretase complex is in the spotlight of modern cancer treat- ments. Gamma-secretase inhibitors (GSI) have shown suc- cessful clinical activity in treating cancers, yet the possible resistance mechanism remains unstudied. Modeling the resistance and understanding culprit molecular mechanisms can improve GSI therapies. Accordingly, the aim of this study is to generate and analyze GSI-resistant breast cancer cells. Methods: Gradually increasing doses of DAPT, a well-known GSI, were applied to MCF-7 breast cancer cell lines to generate resistance. Cell viability, migration and gene expressions were assessed by MTT, wound healing and qRT-PCR analyses. Results: DAPT-resistant MCF-7 cells exhibited abnormal expression of Notch receptors, Notch targets (HES1, HES5, HEY1), and epithelial-mesenchymal transition (EMT) markers (E-cadherin, ZO-1, SNAIL2, N-cadherin) to overcome the continuous increase in DAPT toxicity by increased migration through mesenchymal transition. Conclusions: This study prospects into the role of EMT in the potential resistance mechanism against DAPT treatment for breast cancer cells. Complementary targeting of EMT should be investigated further for a possible effect to potentiate DAPT’s anti-cancer effects.Article Citation - WoS: 1Pasif Tümör Hedefli İnorganik İlaç Nanotaşıyıcıların Akciğer Sağlıklı ve Kanser Hücreleri Üzerindeki Uzun Dönemli Etkisi(Gazi Üniversitesi, 2020) Dağlıoğlu, Cenk; Kacı, Fatma Necmiyeİlaç nanotaşıyıcıları, kontrollü ve sürekli ilaç salım özellikleri ile kanser tedavisinde büyük bir potansiyele sahiptir. Bu nanotaşıyıcılar pasif veya aktif hedefli olarak ilaç taşınımı sağlayabilmektedir, ancak aktif hedefli muadillerine göre, pasif hedefli nanotaşıyıcılar tümörlü dokularda daha yavaş ve düşük düzeyde ilaç birikimi sağladığından kanserli hücrelerin yanında sağlıklı hücrelerde uzun süre bu nanotaşıyıcılara maruz kalmaktadır. Bu nedenle, bu çalışmada, pasif hedefli ilaç nanotaşıyıcıların insan akciğer epitel BEAS-2B hücreleri ve insan akciğer kanser A549 hücreleri üzerindeki uzun dönem etkileri araştırıldı. Bunun için, görüntüleme ve tedavi edici özellikleri bir arada barından Fe3O4@SiO2(FITC)-DOX formülasyonuna sahip ilaç nanotaşıyıcıları kullanıldı ve hücresel birikim, sitotoksisite ve apoptoz üzerindeki etkileri araştırıldı. Hücresel alım ve sitotoksisite deneyleri, pasif hedefli nanotaşıyıcıların kanser hücresi canlılığının etkin bir şekilde azalttığını gösterirken, 24 saatlik inkübasyon sürecinde sağlıklı hücreler üzerinde kayda değer bir etki görülmedi. Ancak 96 saatlik uzun inkübasyon sürecinde, sağlıklı BEAS-2B hücreleri makul seviyelerde nanotaşıyıcı alımı gerçekleştirirken, A549 kanser hücrelerine kıyasla düşük düzeylerde ilaç-aracılı sitotoksisite sergiledi. Ayrıca, nanotaşıyıcılar A549 hücrelerindeki apoptoz seviyelerini önemli ölçüde artırırken, BEAS-2B hücrelerinde 96 saat sonunda dahi apoptotik etki göstermedi. Bu sonuçlar, pasif hedefli inorganik ilaç nanotaşıyıcıların, sağlıklı hücreleri ihmal edilebilir düzeyde etkileyerek, antikanser ilaçların kemoterapötik etkilerini artırmada umut verici olduğunu göstermektedir.Article Citation - WoS: 2Citation - Scopus: 2Antiviral Microrna Expression Signatures Are Altered in Subacute Sclerosing Panencephalitis(Wolters Kluwer Medknow Publications, 2021) Tüfekçi, Kemal Uğur; Allmer, Jens; Çarman, Kürşat Bora; Bayram, Erhan; Topçu, Yasemin; Hız, Semra; Genç, Şermin; Yiş, UluçBackground: Subacute sclerosing panencephalitis (SSPE) is a chronic, progressive disease caused by a persistent infection of the measles virus. Despite extensive efforts, the exact neurodegeneration mechanism in SSPE remains unknown. MicroRNAs (miRNAs) have emerged as an essential part of cellular antiviral defense mechanisms and can be modulated by antiviral cytokines Such as interferon-beta (IFN-beta). Aims and Objectives: In this study, we aimed to elucidate the role of antiviral miRNAs in the pathogenesis of SSPE and analyze the interaction between host antiviral miRNAs and virus genes. Materials and Methods: Thirty-seven patients who were followed with SSPE and age-matched healthy children were included in the study. Peripheral blood mononuclear cell levels of miR-196b, miR-296, miR-431, and miR-448 were analyzed using quantitative polymerase chain reaction. Target predictions and pathway constructions of deregulated miRNAs were assessed. Results: Here, we showed that IFN-beta-modulated miR-196b, miR-296, and miR-431 were significantly upregulated in patients with SSPE compared with healthy controls. Besides, sequence complementarity analysis showed that miR-296 and miR-196b predicted binding regions in measles virus genomic RNA. Conclusion: Our findings suggest that antiviral miRNAs are upregulated in patients with SSPE, which could be a part of the host antiviral defense mechanism. </p>Article Citation - WoS: 8Citation - Scopus: 8Simple High-Performance Liquid Chromatographic Method for Determination of Donepezil Hcl in Pharmaceutical Formulations(ACG Publications, 2020) Bulduk, İbrahim; Aydın, Beyza SultanDonepezil HCl is a hydrochloride salt of a piperidine derivative acetylcholinesterase inhibitor and, it uses in treatment demantia of Alzheimer’s disease. In this study, a sensitive and rapid HPLC-UV method was developed and validated for determination of Donepezil HCl in API and tablet dosage forms. Chromatographic separation was performed using a Ace 5 C18 (5 ?m, 250 x 4.6 mm) by using isocratic phosphate buffer at pH:2.0 and acetonitrile (55:45, v/v) mobile phase was used at the rate of 1.2 mL/min. The column temperature was set at 30 ?C and the UV detection was recorded at 268 nm. The method was validated with respect to specificity, precision, accuracy, linearity, repeatability and reproducibility parameters in a concentration range of 25-125 µg/mL. The limit of detection (LOD) and limit of quantification (LOQ) were determined as 1.40 and4.20 µg/mL, respectively. The uncertainty budget of the measurement for Donepezil HCl was estiamted as 5.80 % at 95% confidence level (k = 2).Article Citation - WoS: 2Citation - Scopus: 2Identification of Cytoplasmic Sialidase Neu2-Associated Proteins by Lc-ms/Ms(Türk Biyokimya Derneği, 2019) Akyıldız Demir, Seçil; Seyrantepe, VolkanBackground: Cytoplasmic sialidase (NEU2) plays an active role in removing sialic acids from oligosaccharides, gly-copeptides, and gangliosides in mammalian cells. NEU2 is involved in various cellular events, including cancer metabolism, neuronal and myoblast differentiation, proliferation, and hypertrophy. However, NEU2-interacting protein(s) within the cell have not been identified yet. Objective: The aim of this study is to investigate NEU2 interacting proteins using two-step affinity purification (TAP) strategy combined with mass spectrometry analysis. Methods: In this study, NEU2 gene was cloned into the pCTAP expression vector and transiently transfected to COS-7 cells by using PEI. The most efficient expression time of NEU2- tag protein was determined by real-time PCR and Western blot analysis. NEU2-interacting protein(s) were investigated by using TAP strategy combined with two different mass spectrometry experiment; LC-MS/MS and MALDI TOF/TOF. Results: Here, mass spectrometry analysis showed four proteins; a-actin, beta-actin, calmodulin and histone H1.2 proteins are associated with NEU2. The interactions between NEU2 and actin filaments were verified by Western blot analysis and immunofluorescence analysis. Conclusions: Our study suggests that association of NEU2 with actin filaments and other protein(s) could be important for understanding the biological role of NEU2 in mammalian cells.Article Citation - WoS: 1Investigation of Potential Anticarcinogenic Effects of Corilagin in Lung Cancer Cells(Marmara Üniversitesi, 2019) Rencüzoğulları, Çağla; Çinçin, Zeynep Birsu; İplik, Elif Sinem; Baran, Yusuf; Çakmakoğlu, BediaObjective: Lung cancer (LC) is the most extensive reason of cancer associated deaths in men and women in the world. LC categorizes into two main groups due to their molecular clinicopathological features and therapeutic responses. Non-small cell lung cancer (NSCLC) is the main subgroup that consists of nearly 85% of all lung cancer types. Corilagin, a biologically active ellagitannin, could be extracted from Phyllanthus species which are known as Chinese medicinal plant. It has been recently shown that Corilagin could exert anti-inflammatuar and antioxidative effects in different experimental cancer models. However, the molecular effects of Corilagin in NSCLC remain unclear. Methods: In this study, the antiproliferative and apoptotic effects of Corilagin were identified by WST-1 cell proliferation test, caspase-3 and mitochondrial membrane potential (MMP). Results: We found that Corilagin significiantly suppressed the proliferation of NSCLC cells. Furthermore, we also showed that Corilagin could contribute apoptosis by inducing activity of caspase-3 molecule and loss of MMP. Conclusion: Taken together, our study first showed that Corilagin could be a new treatment method for NSCLC after verifying its effects with in vivo and clinical studies.Article Alteration of Protein Localization and Intracellular Calcium Content Due To Connexin26 D50a and A88v Mutations(Türk Biyokimya Derneği, 2017) Aypek, Hande; Meşe, GülistanIntroduction: Connexins (Cx) play essential roles in cellular homeostasis by forming gap junctions and non-junctional hemichannels. In vitro characterization of Cx26 mutations causing keratitis-ichthyosis-deafness (KID) syndrome, were shown to form leaky hemichannels. The molecular/ cellular mechanisms affected by aberrant hemichannels have recently been elucidated. Here, we further wanted to characterize Cx26 KID syndrome mutations, D50A and A88V, which were shown to form aberrant hemichannels and remained unaddressed in the literature. Methods: Neurobiotin uptake assay in HeLa and N2A cells transfected with Cx26-WT, D50A or A88V verified the presence of aberrant hemichannels and immunofluorescent staining with fluorescent microscopy determined cellular localization of Cx26. Finally, intracellular calcium content was examined by using calcium indicator, Fluo-3AM, and flow cytometer. Results: Cx26-D50A and A88V mutations prevented the formation of gap junction plaques at cell-cell appositions and mutant proteins were observed to localize to the Golgi apparatus. Further, comparison of intracellular calcium content showed an increase in calcium amount in cells containing Cx26-D50A and A88V relative to Cx26-WT. Conclusion: Retention of Cx26 in the Golgi apparatus and alteration in the intracellular calcium content due to KID syndrome mutations may influence various cellular processes that might contribute to development of epidermal phenotypes.