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Öğe Different cortex activation between young and middle-aged people during different type problem-solving: an EEG&fNIRS study(2025) Sarıcaoğlu, Mevhibe; Yücel, Meryem Ayşe; Budak, Miray; Omurtag, Ahmet; Hanoğlu, LütfüProblem-solving strategies vary depending on the type of problem and aging. This study investigated the hemodynamic response measured by the changes in the oxyhemoglobin concentration (HbO), alpha frequency power, and their interrelation during problem-solving in healthy young and middle-aged individuals, employing combined electroencephalography (EEG) and functional near-infrared spectroscopy (fNIRS) recordings. The study included 39 young and 30 middle-aged subjects. The brain activation that occurred while answering different questions was recorded using combined EEG and fNIRS. During the EEG & fNIRS recording, four questions (arithmetic, general knowledge, insight, and basic operation) were used for problem-solving. Alpha power (8–13 Hz) and HbO changes were analyzed. The behavioral results indicated significant differences between age groups in various question types. While the middle-aged group performed better on the general knowledge questions, the older group performed better on the insight and four-process questions. The fNIRS results reveal significant differences in brain activation during problem-solving tasks, particularly in regions like DLPFC/TA, STG, pSSC/Wernicke, and STG/angular gyrus Wernicke's area. The young group with the highest HbO was recorded during arithmetic questions, general knowledge questions, and basic operation questions. In contrast, there was no significant highest HbO during insight questions. Similar findings were observed in the middle-aged group, with the highest HbO recorded during general knowledge questions. However, there was no significant HbO in other channels during the solving of other question types in this group. The alpha power varied across different electrodes for various question types in both young and middle-aged groups. The highest alpha frequency band power for different electrodes was recorded while solving general knowledge questions in the young group and insight questions in the middle-aged group. Finally, the EEG and fNIRS correlation results showed positive correlations between HbO and alpha frequency band power in specific brain regions while solving general knowledge questions, particularly in the middle-aged group. The study reveals age-related differences in behavioral performance, brain activation patterns, and neural correlates during various cognitive tasks, showcasing distinct strengths between middle-aged and young individuals in specific question types.Öğe Computational study of the activation mechanism of wild-type parkin and its clinically relevant mutant(2025) Cinviz, Zeynep Nur; Şensoy, ÖzgeParkinson's disease (PD) is the second most prevalent neurodegenerative disorder. It impairs the control of movement and balance. Parkin mutations worsen the symptoms in sporadic cases and cause the early onset of the disease. Therefore, recent efforts have focused on the rescue of defective parkin by engineered proteins or small-molecule activators to enhance parkin activation. These attempts require holistic understanding of the multistep activation mechanism and molecular effects of disease-associated mutations. Hereby, we provided a comprehensive analysis of the activation mechanism of parkin and a clinically relevant mutant, parkinS167N, using molecular dynamics simulations based on the following crystal structures: (1) parkin, (2) parkin/pUb (phosphorylated Ubiquitin), (3) pparkin/pUb, and (4) pparkin/pUb/UbcH7-Ub. Each of these represents an individual step in the activation process. We showed that the mutation impacted the dynamics of not only the RING0 domain, where it is localized, but also the RING2, Ubl, and IBR domains. We identified residues participating in the allosteric interaction network involved in parkin activation. Some of them are mutated in PD-associated parkin variants. The RING0 domain provides a binding interface with various proteins, so understanding problems associated with the mutation paves the way to the discovery of effective engineered proteins or small molecules that activate mutant parkin.Öğe Temporal microbiome changes in axolotl limb regeneration: stage-specific restructuring of bacterial and fungal communities with a flavobacterium bloom during blastema proliferation(2024) Altın, Hanne; Delice, Büşra; Yıldırım, Berna; Demircan, Turan; Yıldırım, SüleymanThe intricate relationship between regeneration and microbiota has recently gained attention, spanning diverse model organisms. Axolotl (Ambystoma mexicanum) is a critically endangered salamander species and a model organism for regenerative and developmental biology. Despite its significance, a noticeable gap exists in understanding the interplay between axolotl regeneration and its microbiome. Here, we analyse in depth bacterial 16S rRNA amplicon dataset that we reported before as data resource and profile fungal community by sequencing ITS amplicons at the critical stages of limb regeneration (0–1–4–7–30–60 days post amputation, ‘dpa’). Results reveal a decline in richness and evenness in the course of limb regeneration, with bacterial community richness recovering beyond 30 dpa unlike fungi community. Beta diversity analysis reveals precise restructuring of the bacterial community along the three phases of limb regeneration, contrasting with less congruent changes in the fungal community. Temporal dynamics of the bacterial community highlight prevalent anaerobic bacteria in initiation phase and Flavobacterium bloom in the early phase correlating with limb blastema proliferation. Predicted functional analysis mirrors these shifts, emphasising a transition from amino acid metabolism to lipid metabolism control. Fungal communities shift from Blastomycota to Ascomycota dominance in the late regeneration stage. Our findings provide ecologically relevant insights into stage specific role of microbiome contributions to axolotl limb regeneration.Öğe Bioenergetic shift and proteomic signature induced by lentiviral-transduction of gfp-based biosensors(2024) Barakat, Sarah; Çimen, Şeyma; Miri, Seyed Mohammad; Vatandaşlar, Emre; Yelkenci, Hayriye Ecem; San Martín, Alejandro; Beker, Mustafa Çağlar; Kök, Kıvanç; Öztürk, Gürkan; Eroğlu, EmrahFluorescent proteins (FPs) stand as pivotal tools extensively employed across diverse biological research endeavors in various model systems. However, long-standing concerns surround their use due to the numerous side effects associated with their expression. Recent investigations have brought to light the significance of hydrogen peroxide (H2O2) that is associated with the maturation process of green fluorescent protein (GFP) fluorophores. The structural and functional impairments associated with GFP expression are possibly linked to this amount of H2O2. In this study, we assess the impact of the GFP-based HyPer7 biosensor on cellular homeostasis and proteome changes, aiming to identify potential risks related to oxidative stress responses that potentially risks the application of such tools. Cells expressing genome-integrated HyPer7 demonstrated altered mitochondrial membrane potential (MMP), which was alleviated by the addition of antioxidants or culturing cells at physiological normoxia (5 kPa O2). Additionally, HyPer7-expressing cells also exhibited significant impairment in mitochondrial oxidative respiration, suggesting broader mitochondrial dysfunction. Through untargeted proteomics analysis, we identified 26 proteins exhibiting differential expression in HyPer7-expressing cells compared to respective control cells. Functional annotation analysis showed that the list of the delineated proteins is associated with cellular responses to stress and the regulation of antioxidant mechanisms. Our findings underscore the significance of caution and validation in ensuring a thorough comprehension of cellular responses when using fluorescent protein-based tools, thereby enhancing the reliability of the results.Öğe Inhibition of phosphodiesterase 10A mitigates neuronal injury by modulating apoptotic pathways in cold-induced traumatic brain injury(2024) Beker, Mustafa Çağlar; Altıntaş, Mehmet Özgen; Doğan, Enes; Bayraktaroğlu, Çiğdem; Balaban, Buse; Özpınar, Ayşenur; Şengün, Nursena; Altunay, Serdar; Kılıç, ErtuğrulBrain injury develops from a complex series of pathophysiological phases, resulting in acute necrotic or delayed apoptotic cell death after traumatic brain injury (TBI). Inhibition of apoptotic cell death is critical for the treatment of acute neurodegenerative disorders, such as TBI. Here, we investigated the role of phosphodiesterase 10A (PDE10A) in the development of neuronal injury, particularly in apoptotic cell death. Using the PDE10A inhibitor TAK-063, we found that PDE10A inhibition is associated with decreased brain injury, brain swelling, and blood brain barrier disruption 48 h after cold-induced TBI. Furthermore, a particularly notable result was observed with 3 mg/kg TAK-063, which reduced disseminated neuronal injury. Protein abundance analysis revealed that PDE10A inhibition activates survival kinases AKT and ERK-1/-2, which were associated with the decreased activation of MMP-9 and PTEN. Additionally, iNOS and nNOS levels significantly reduced in the TAK-063 group, playing roles in inflammation and apoptosis. A planar surface immunoassay was performed for in-depth analyses of the apoptotic signaling pathways. We observed that inhibition of PDE10A resulted in the decreased expression of TNFRSF1A, TNFRSF10B, and TNFRSF6 receptors, particularly inducing apoptotic cell death. Moreover, these findings correlated with reduced levels of pro-apoptotic proteins, including PTEN, p27, Cytochrome-c, cleaved Caspase-3, Bad, and p53. Interestingly, TAK-063 treatment reduced levels of anti-apoptotic proteins or enzymes, including XIAP, Claspin, and HIF1α, without affecting Bcl-x, MCL-1, SMAC, HO-1, HO-2, HSP27, HSP60, and HSP70. The findings suggest that PDE10A regulates cellular signaling predominantly pro-apoptotic pathways, and inhibition of this protein is a promising approach for the treatment of acute brain injury.Öğe Curcumin-loaded emulsome nanoparticles induces apoptosis through p53 signaling pathway in pancreatic cancer cell line panc-1(2025) Demirci, Züleyha; İşlek, Zeynep; Sığınç, Halime İlhan; Şahin, Fikrettin; Üçışık, Mehmet Hikmet; Bolat, Zeynep BüşraPancreatic cancer is a global health problem with a poor prognosis, limited treatment options and low survival rates of patients. Thus, the exploration of novel treatment approaches is crucial. Curcumin shows promise in pancreatic cancer. Curcumin has anticancer properties promoting apoptosis through the p53 pathway. However, adverse effects and low bioavailability are curcumin's main drawbacks and its delivery by nanoparticles could improve its effectiveness as a treatment option. Curcumin-loaded emulsome nanoparticles (CurEm) have shown promise in colorectal, hepatocellular, and prostate cancers. This study aims to evaluate the anticancer potential of CurEm in pancreatic cancer cell line PANC-1. The cytotoxic effects of CurEm on PANC-1 cells show cytotoxicity in dose and time-dependent manner. The selected dose 30 μM CurEm resulted spheroidal morphology in PANC-1 cells and colony forming and scratch assay conducted demonstrated significant growth inhibition and decrease in migration ability, respectively. Cell cycle analysis shows that CurEm induces G2/M arrest in PANC-1 cells. CurEm-treated PANC-1 cells showed a significant increase in p53 and Caspase 3 genes, while a significant decrease in Bcl-2 genes compared to untreated group. Western blot results showed parallel results to qPCR analysis for Bcl-2 protein levels. Interestingly, we saw low p53 protein levels in CurEm-treated PANC-1 cells. These findings shed light on the potential of CurEm as an effective and stable therapeutic approach for pancreatic cancer.Öğe Improving predictive efficacy for drug resistance in novel hiv-1 protease inhibitors through transfer learning mechanisms(2024) Tunç, Hüseyin; Yılmaz, Sümeyye; Darendeli Kiraz, Büşra Nur; Sarı, Murat; Kotil, Seyfullah Enes; Şensoy, Özge; Durdağı, SerdarThe human immunodeficiency virus presents a significant global health challenge due to its rapid mutation and the development of resistance mechanisms against antiretroviral drugs. Recent studies demonstrate the impressive performance of machine learning (ML) and deep learning (DL) models in predicting the drug resistance profile of specific FDA-approved inhibitors. However, generalizing ML and DL models to learn not only from isolates but also from inhibitor representations remains challenging for HIV-1 infection. We propose a novel drug-isolate-fold change (DIF) model framework that aims to predict drug resistance score directly from the protein sequence and inhibitor representation. Various ML and DL models, inhibitor representations, and protein representations were analyzed through realistic validation mechanisms. To enhance the molecular learning capacity of DIF models, we employ a transfer learning approach by pretraining a graph neural network (GNN) model for activity prediction on a data set of 4855 HIV-1 protease inhibitors (PIs). By performing various realistic validation strategies on internal and external genotype-phenotype data sets, we statistically show that the learned representations of inhibitors improve the predictive ability of DIF-based ML and DL models. We achieved an accuracy of 0.802, AUROC of 0.874, and r of 0.727 for the unseen external PIs. By comparing the DIF-based models with a null model consisting of isolate-fold change (IF) architecture, it is observed that the DIF models significantly benefit from molecular representations. Combined results from various testing strategies and statistical tests confirm the effectiveness of DIF models in testing novel PIs for drug resistance in the presence of an isolate.Öğe Vinpocetine ameliorates neuronal injury after cold-induced traumatic brain injury in mice(2025) Yelkenci, Hayriye Ecem; Değirmenci, Zehra; Koç, Halil İbrahim; Bayırlı, Sevban; Baltacı, Saltuk Buğra; Altunay, Serdar; Öztekin, Nevin; Koçak, Mehmet; Kılıç, Ertuğrul; Beker, Mustafa ÇağlarTraumatic brain injury (TBI), also known as intracranial injury, is a common condition with the highest incidence rate among neurodegenerative disorders and poses a significant public health burden. Various methods are used in the treatment of TBI, but the effects of cold-induced traumatic brain injury have not been thoroughly studied. In this context, vinpocetine (VPN), derived from Vinca minor, exhibits notable anti-inflammatory and antioxidant properties. VPN is known for its neuroprotective role and is generally utilized for treating various neurodegenerative disorders. However, the function of VPN after cold-induced TBI needs to be studied in more detail. This study aims to investigate the neuroprotective effects of VPN at varying doses (5 mg/kg or 10 mg/kg) after cold-induced TBI. C57BL/6 mice were sacrificed 2 or 28 days after cold-induced TBI. Results indicate that VPN administration significantly reduces brain infarct volume, brain swelling, blood–brain barrier disruption, and DNA fragmentation in a dose-dependent manner. Additionally, VPN enhances neuronal survival in the ipsilesional cortex. In the long term, VPN treatment (5 mg/kg/day or 10 mg/kg/day, initiated 48 h post-TBI) improved locomotor activity, cell proliferation, neurogenesis, and decreased whole brain atrophy, specifically motor cortex atrophy. We performed liquid chromatography-tandem mass spectrometry (LC–MS/MS) to elucidate the underlying mechanisms to profile proteins and signaling pathways influenced by prolonged VPN treatment post-TBI. Notably, we found that 192 different proteins were significantly altered by VPN treatment, which is a matter of further investigation for the development of therapeutic targets. Our study has shown that VPN may have a neuroprotective role in cold-induced TBI.Öğe Dual activity of minnelide chemosensitize basal/triple negative breast cancer stem cells and reprograms immunosuppressive tumor microenvironment(2024) Köksalar Alkan, Fulya; Çağlayan, Ahmet Burak; Alkan, Hilmi Kaan; Gündüz, Yunus Emre; Şensoy, Özge; Öztürk, Gürkan; Korkaya, HasanTriple negative breast cancer (TNBC) subtype is characterized with higher EMT/stemness properties and immune suppressive tumor microenvironment (TME). Women with advanced TNBC exhibit aggressive disease and have limited treatment options. Although immune suppressive TME is implicated in driving aggressive properties of basal/TNBC subtype and therapy resistance, effectively targeting it remains a challenge. Minnelide, a prodrug of triptolide currently being tested in clinical trials, has shown anti-tumorigenic activity in multiple malignancies via targeting super enhancers, Myc and anti-apoptotic pathways such as HSP70. Distinct super-enhancer landscape drives cancer stem cells (CSC) in TNBC subtype while inducing immune suppressive TME. We show that Minnelide selectively targets CSCs in human and murine TNBC cell lines compared to cell lines of luminal subtype by targeting Myc and HSP70. Minnelide in combination with cyclophosphamide significantly reduces the tumor growth and eliminates metastasis by reprogramming the tumor microenvironment and enhancing cytotoxic T cell infiltration in 4T1 tumor-bearing mice. Resection of residual tumors following the combination treatment leads to complete eradication of disseminated tumor cells as all mice are free of local and distant recurrences. All control mice showed recurrences within 3 weeks of post-resection while single Minnelide treatment delayed recurrence and one mouse was free of tumor. We provide evidence that Minnelide targets tumor intrinsic pathways and reprograms the immune suppressive microenvironment. Our studies also suggest that Minnelide in combination with cyclophosphamide may lead to durable responses in patients with basal/TNBC subtype warranting its clinical investigation.Öğe Investigation of hemispheric asymmetry in alzheimer’s disease patients during resting state revealed by fnirs(2024) Mızrak, Hazel Gül; Dikmen, Merve; Hanoğlu, Lütfü; Şakul, Bayram UfukAlzheimer's disease (AD) is characterized by the gradual deterioration of brain structures and changes in hemispheric asymmetry. Meanwhile, healthy aging is associated with a decrease in functional hemispheric asymmetry. In this study, functional connectivity analysis was used to compare the functional hemispheric asymmetry in eyes-open resting-state fNIRS data of 16 healthy elderly controls (mean age: 60.4 years, MMSE (Mini-Mental State Examination): 27.3 ± 2.52) and 14 Alzheimer's patients (mean age: 73.8 years, MMSE: 22 ± 4.32). Increased interhemispheric functional connectivity was found in the premotor cortex, supplementary motor cortex, primary motor cortex, inferior parietal cortex, primary somatosensory cortex, and supramarginal gyrus in the control group compared to the AD group. The study revealed that the control group had stronger interhemispheric connectivity, leading to a more significant decrease in hemispheric asymmetry than the AD group. The results show that there is a difference in interhemispheric functional connections at rest between the Alzheimer's group and the control group, suggesting that functional hemispheric asymmetry continues in Alzheimer's patients.Öğe Functional characterization of KCNMA1 mutation associated with dyskinesia, seizure, developmental delay, and cerebellar atrophy(2024) Yücesan, Emrah; Göncü, Beyza; Özgül, Cemil; Kebapçı, Arda; Aslanger, Ayça Dilruba; Akyüz, Enes; Yeşil, GözdeKCNMA1 located on chromosome 10q22.3, encodes the pore-forming α subunit of the ‘Big K+’ (BK) large conductance calcium and voltage-activated K + channel. Numerous evidence suggests the functional BK channel alterations produced by different KCNMA1 alleles may associate with different symptoms, such as paroxysmal non kinesigenic dyskinesia with gain of function and ataxia with loss of function. Functional classifications revealed two major patterns, gain of function and loss of function effects on channel properties in different cell lines. In the literature, two mutations have been shown to confer gain of function properties to BK channels: D434G and N995S. In this study, we report the functional characterization of a variant which was previously reported the whole exome sequencing revealed bi-allelic nonsense variation of the cytoplasmic domain of calcium-activated potassium channel subunit alpha-1 protein. To detect functional consequences of the variation, we parallely conducted two independent approaches. One is immunostaining using and the other one is electrophysiological recording using patch-clamp on wild-type and R458X mutant cells to detect the differences between wild-type and the mutant cells. We detected the gain of function effect for the mutation (NM_001161352.1 (ENST00000286628.8):c.1372C > T;Arg458*) using two parallel approaches. According to the result we found, the reported mutation causes the loss of function in the cell. It should be noted that in future studies, it can be thought that the functions of genes associated with channelopathies may have a dual effect such as loss and gain.Öğe Cellular immunity to nucleoproteins (np) of crimean-congo hemorrhagic fever virus (cchfv) and hazara virus (hazv)(2024) Kalkan Yazıcı, Merve; Karaaslan, Elif; Güler Çetin, Nesibe Selma; Doymaz, Mehmet ZiyaCrimean-Congo Hemorrhagic Fever Virus (CCHFV) is a globally significant vector-borne pathogen with no internationally-licensed preventative and therapeutic interventions. Hazara virus (HAZV), on the other hand, a related Orthonairovirus, has not been reported as a human pathogen. HAZV has been proposed as a surrogate model for studying CCHFV, bisosafety level 4 (BSL-4) agent. Previously, we investigated the humoral immune responses between NPs of these viruses and in this study, we extended the scrutiny to cellular immune responses elicited by NPs of CCHFV and HAZV. Here, mice were immunized with recombinant CCHFV NP and HAZV NP to evaluate the correlates of cell-mediated immunity (CMI). Delayed-type hypersensitivity (DTH) responses were assessed by challenging immunized mice with CCHFV-rNP or HAZV-rNP on the footpad and lymphocyte proliferation assays (LPAs) were performed by stimulating splenocytes in vitro with CCHFV-rNP or HAZV-rNP to compare cellular immune responses. In all test groups, strong DTH and LPA responses were detected against homologous and heterologous challenging antigens. To assess the cytokine response, an RT-qPCR -specific for cytokine mRNAs was utilized. Interestingly, CCHFV NP stimulated groups exhibited a significantly elevated mRNA level of interleukin 17 A (IL-17) compared to HAZV NP, indicating a notable difference in immune responses. This study presents comparison between CMI elicited by NPs of CCHFV and HAZV and contributes to the understanding of a highly pathogenic virus, particularly in the context of the declaration of CCHFV by World Health Organization’s (WHO) as a major viral threat to the world.Öğe Evaluation of the correlation between peripheral inflammatory markers and suicide risk in drug-naive first-episode schizophrenia(2024) Yeşilkaya, Ümit Haluk; Şen, Meltem; Balcıoğlu, Yasin Hasan; Gökçay, Hasan; Çelikkıran, Pınar; Kirlioğlu Balcıoğlu, Simge; Karamustafalioğlu, NesrinIntroduction: Patients with schizophrenia have a higher lifetime prevalence of suicidal behavior (SB) compared to the general population. Therefore, understanding the possible neurobiology of suicide and predicting the risk of suicide in schizophrenia is a solemnly critical issue. Methods: 31 drug-naïve first episode schizophrenia (FES) patients with current SB (FES-S), 69 drug-naive patients with first episode schizophrenia without SB (FES-NS), and 69 drug-naïve non-psychotic patients with current SB (NPS) who were diagnosed according to The Diagnostic and Statistical Manual of Mental Disorders-5 (DSM-5) participated the study. The control group (HC) consisted of 127 individuals matched with the patients. Symptoms at the time of evaluation were assessed using The Positive and Negative Syndrome Scale (PANSS) and Columbia Suicide Severity Rating Scale (CSSRS). Blood samples were collected from all participants to determine White blood cell (WBC), neutrophil, monocyte, albumin, C-reactive protein (CRP), Lymphocyte, and Platelet levels and to measure this protein ratio. Results: The blood levels of WBC, neutrophil, monocyte, albumin, CRP, and Neutrophil/Albumin Ratio (NAR) were higher in all patient groups compared to HC. CRP/Albumin Ratio (CAR) value was observed to be highest in the NPS group. Monocyte/Lymphocyte Ratio (MLR) value was significantly higher in patients with FES compared to HC. There were no significant differences between the FES-S group and the FES-NS and NPS groups. Conclusion: It can be suggested that although inflammation is not a predictor for suicide attempts in schizophrenia, it is associated with the degree of suicide risk in schizophrenia. In addition, the strong relationship between suicide and psychiatric disorders can be the main reason for high peripheral inflammation levels in suicidal patients.Öğe Experimental vitamin d deficiency in rats: clinical chemistry, histopathological, and immunological evaluation(2024) Sitar, Mustafa Erinç; Dönmez Çakıl, Yaprak; Öngen İpek, Belkız; Altıner, Necdet; Aydın, Mehmet Şerif; Karadeniz, AslıBackground: Vitamin D deficiency is a significant global health concern. Experimental models are essential to elucidate the biochemical, histopathological, and immunological consequences of this deficiency. This study established a vitamin D deficiency rat model to mimic insufficient vitamin D intake and examine the resulting health impacts, particularly on liver, kidney, and immune functions. Materials and methods: Sprague-Dawley male rats were randomly assigned to two groups. The control group received a standard rodent diet, while the experimental group was fed a modified diet with reduced vitamin D for three months. Analyses included serum vitamin D levels, clinical chemistry, renal and liver histopathology, and blood immunophenotyping and cytokine analysis for both the control (n=7) and experimental (n=7) groups. Results: Serum vitamin D 25-OH levels were threefold lower in the experimental group (p < 0.001), indicating the induction of vitamin D deficiency. No significant differences in weight gain were observed between the groups. All clinical chemistry parameters remained within reference ranges. However, the experimental group showed significant declines in triglycerides (TG, p=0.0441), alkaline phosphatase (ALP, p=0.0021), and alanine aminotransferase (ALT, p=0.0002). Histopathology revealed normal liver and kidney architecture in the control group, while the experimental group exhibited hepatic cord deterioration, severe vacuolization in the liver, and edema and dilatation in the renal cortex tubular epithelium. Immunophenotyping analysis of lymphocyte subsets and assessment of serum cytokines did not reveal any differences between the two groups. Conclusion: A vitamin D deficiency model without complications such as obesity, parathyroid issues, or mortality was established in rats. This method could be applied in specific disease experimental models.Öğe Effects of platelet-rich plasma and concentrated growth factor on viability of ultra-diced cartilage grafts in a rabbit model(2024) Kulaksız, Yasin; Yenigün, Alper; Aydın, Mehmet Şerif; Doğan, Remzi; Tuğrul, Selahattin; Özturan, OrhanBackground: Although rhinoplastic surgery has progressed considerably in recent years, nasal dorsal irregularities still cause postoperative distress for both surgeons and patients. Purpose: The aim of this study was to measure the association between two biologic graft adjuncts, platelet-rich plasma (PRP) and concentrated growth factor (CGF), and ultra-diced cartilage viability in an animal model. Study Design, Setting, and Sample: This study was designed as a randomized in-vivo study using a rabbit model. Fourteen rabbits were utilized in this investigation. The ultra-diced cartilage was obtained from auricular cartilage. Predictor Variable: The graft biologic adjunct is the predictor variable. There were three treatment groups: graft mixed with PRP or CGF or untreated (control). The grafts were placed in three separate pockets opened on the same rabbit. Grafts were harvested 3 months after insertion for analysis. Main Outcome Variable(s): The primary outcome variable was histopathological and regenerative scores obtained from multiple histopathological parameters indicating the viability of the cartilage. Histopathological score parameters were chondrocyte loss, inflammation, fibrosis, cartilage fragmentation, and calcified area formations in the lacunae. Regenerative score parameters were peripheral cell proliferation in the cartilage tissue, vascularization in the connective tissue, proteoglycan increase in the matrix, and the amount of connective tissue. Covariates: The variables were age, sex, and weight. Analyses: Statistical analysis employed the analysis of variance test, with a significance level of P <.05. Results: The sample was composed of 14 rabbits and 42 samples. The histopathologic scores were 11.93 (±2.49), 8.78 (±2.19), and 6.85 (±1.46) for the control, PRP, and CGF groups, respectively. A statistically significant difference was found in the PRP (P <.0275) and CGF (P <.0001) groups compared to the control group. The regenerative scores were 6.21 (±0.97), 8.85 (±1.70), and 12.07 (±1.26) for the control, PRP and CGF groups, respectively. A statistically significant difference was found in the PRP (P <.0159) and CGF (P <.0001) groups compared to the control group. Conclusion and Relevance: This is the first study investigating the ultra-diced cartilage graft in an experimental animal model. Histopathological examination has shown that mixing ultra-diced cartilage with CGF or PRP increases viability by reducing the histopathological score and increasing the regenerative score.Öğe Electrophysiological and cognitive changes in hard coal miners associated with working underground(2024) Çelik, Samet; Yıldırım, Ebru; Güntekin, BaharMiners working underground face some risk factors that affect the nervous system—such as high noise, dark environment, chronic stress, and exposure to toxic gases. However, it is not known whether these risk factors affect the cognition of miners. In this study, the cognitive changes of miners were examined through event-related oscillations via electroencephalogram (EEG). Twenty underground miners and control groups, equal to each other in age, education level, and working duration, participated in this study. Neuropsychological tests were applied to all participants to examine their cognitive characteristics. Then, 20-channel EEG was recorded for electrophysiological changes during visual oddball paradigm. Event-related power spectrum and phase locking were analyzed in delta (0.5-3.5), theta (4-7), and alpha (8-13?Hz) frequency bands. It was determined that the delta responses that emerged during the target stimulus differed between the two groups in terms of phase locking (p < 0.05). Considering event-related alpha responses, a statistical difference was found regarding power spectrum and phase locking (p < 0.05). Moreover, the alpha power spectrum in the miners was found to be negatively statistically correlated with working duration (p < 0.05). This study determined that the event-related electrophysiological responses of the miners were negatively affected depending on the working conditions. In addition, neuropsychological assessment determined miners had deficiencies in learning and memory skills and many other cognitive functions such as attention, behavioral inhibition, and visual perception.Öğe Development of a dual reporter system to simultaneously visualize ca2+ signals and ampk activity(2024) Erdoğan, Yusuf Ceyhun; Pilic, Johannes; Gottschalk, Benjamin; Yiğit, Esra Nur; Zaki, Asal Ghaffari; Öztürk, Gürkan; Eroğlu, Emrah; Okutan, Begüm; Sommer, Nicole G.; Weinberg, Annelie M.; Schindl, Rainer; Graier, Wolfgang F.; Malli, RolandIn this study, we introduce a new separation of phases-based activity reporter of kinase (SPARK) for AMP-activated kinase (AMPK), named AMPK-SPARK, which reports the AMPK activation by forming bright fluorescent clusters. Furthermore, we introduce a dual reporter system, named GCaMP-AMPK-SPARK, by incorporating a single-fluorescent protein (FP)-based Ca2+ biosensor, GCaMP6f, into our initial design, enabling simultaneous monitoring of Ca2+ levels and AMPK activity. This system offers the essential quality of information by single-channel fluorescence microscopy without the need for coexpression of different biosensors and elaborate filter layouts to overcome spectral limitations. We used AMPK-SPARK to map endogenous AMPK activity in different cell types and visualized the dynamics of AMPK activation in response to various stimuli. Using GCaMP-AMPK-SPARK, we revealed cell-to-cell heterogeneities in AMPK activation by Ca2+ mobilization. We anticipate that this dual reporter strategy can be employed to study the intricate interplays between different signaling networks and kinase activities.Öğe An in silico investigation on the binding site preference of PD-1 and PD-L1 for designing antibodies for targeted cancer therapy(2024) Abdolmaleki, Sarah; Ganjalikhani Hakemi, Mazdak; Ganjalikhany, Mohamad RezaCancer control and treatment remain a significant challenge in cancer therapy and recently immune checkpoints has considered as a novel treatment strategy to develop anti-cancer drugs. Many cancer types use the immune checkpoints and its ligand, PD-1/PD-L1 pathway, to evade detection and destruction by the immune system, which is associated with altered effector function of PD-1 and PD-L1 overexpression on cancer cells to deactivate T cells. In recent years, mAbs have been employed to block immune checkpoints, therefore normalization of the anti-tumor response has enabled the scientists to develop novel biopharmaceuticals. In vivo affinity maturation of antibodies in targeted therapy has sometimes failed, and current experimental methods cannot accommodate the accurate structural details of protein-protein interactions. Therefore, determining favorable binding sites on the protein surface for modulator design of these interactions is a major challenge. In this study, we used the in silico methods to identify favorable binding sites on the PD-1 and PD-L1 and to optimize mAb variants on a large scale. At first, all the binding areas on PD-1 and PD-L1 have been identified. Then, using the RosettaDesign protocol, thousands of antibodies have been generated for 11 different regions on PD-1 and PD-L1 and then the designs with higher stability, affinity, and shape complementarity were selected. Next, molecular dynamics simulations and MM-PBSA analysis were employed to understand the dynamic, structural features of the complexes and measure the binding affinity of the final designs. Our results suggest that binding sites 1, 3 and 6 on PD-1 and binding sites 9 and 11 on PD-L1 can be regarded as the most appropriate sites for the inhibition of PD-1-PD-L1 interaction by the designed antibodies. This study provides comprehensive information regarding the potential binding epitopes on PD-1 which could be considered as hotspots for designing potential biopharmaceuticals. We also showed that mutations in the CDRs regions will rearrange the interaction pattern between the designed antibodies and targets (PD-1 and PD-L1) with improved affinity to effectively inhibit protein-protein interaction and block the immune checkpoint.Öğe Residues in the fructose-binding pocket are required for ketohexokinase-A activity(2024) Ferreira, Juliana C.; Villanueva, Adrian J.; Fadl, Samar; Al Adem, Kenana; Cinviz, Zeynep Nur; Nedyalkova, Lyudmila; Cardoso, Thyago H.S.; Andrade, Mario Edson; Saksena, Nitin K.; Şensoy, Özge; Rabeh, Wael M.Excessive fructose consumption is a primary contributor to the global surges in obesity, cancer, and metabolic syndrome. Fructolysis is not robustly regulated and is initiated by ketohexokinase (KHK). In this study, we determined the crystal structure of KHK-A, one of two human isozymes of KHK, in the apo-state at 1.85 Å resolution, and we investigated the roles of residues in the fructose-binding pocket by mutational analysis. Introducing alanine at D15, N42, or N45 inactivated KHK-A, whereas mutating R141 or K174 reduced activity and thermodynamic stability. Kinetic studies revealed that the R141A and K174A mutations reduced fructose affinity by 2- to 4-fold compared to WT KHK-A, without affecting ATP affinity. Molecular dynamics simulations provided mechanistic insights into the potential roles of the mutated residues in ligand coordination and the maintenance of an open state in one monomer and a closed state in the other. Protein–protein interactome analysis indicated distinct expression patterns and downregulation of partner proteins in different tumor tissues, warranting a reevaluation of KHK's role in cancer development and progression. The connections between different cancer genes and the KHK signaling pathway suggest that KHK is a potential target for preventing cancer metastasis. This study enhances our understanding of KHK-A's structure and function and offers valuable insights into potential targets for developing treatments for obesity, cancer, and metabolic syndrome.Öğe Hexokinase 1 forms rings that regulate mitochondrial fission during energy stress(2024) Pilic, Johannes; Gottschalk, Benjamin; Bourgeois, Benjamin; Habisch, Hansjörg; Koshenov, Zhanat; Oflaz, Furkan Enes; Erdoğan, Yusuf Can; Miri, Seyed Mohammad; Yiğit, Esra Nur; Aydın, Mehmet Şerif; Öztürk, Gürkan; Eroğlu, Emrah; Shoshan Barmatz, Varda; Madl, Tobias; Graier, Wolfgang F.; Malli, RolandMetabolic enzymes can adapt during energy stress, but the consequences of these adaptations remain understudied. Here, we discovered that hexokinase 1 (HK1), a key glycolytic enzyme, forms rings around mitochondria during energy stress. These HK1-rings constrict mitochondria at contact sites with the endoplasmic reticulum (ER) and mitochondrial dynamics protein (MiD51). HK1-rings prevent mitochondrial fission by displacing the dynamin-related protein 1 (Drp1) from mitochondrial fission factor (Mff) and mitochondrial fission 1 protein (Fis1). The disassembly of HK1-rings during energy restoration correlated with mitochondrial fission. Mechanistically, we identified that the lack of ATP and glucose-6-phosphate (G6P) promotes the formation of HK1-rings. Mutations that affect the formation of HK1-rings showed that HK1-rings rewire cellular metabolism toward increased TCA cycle activity. Our findings highlight that HK1 is an energy stress sensor that regulates the shape, connectivity, and metabolic activity of mitochondria. Thus, the formation of HK1-rings may affect mitochondrial function in energy-stress-related pathologies.
