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Öğe A Study of the Effects of Metformin, a Biguanide Derivative, on Annulus Fibrosus and Nucleus Pulposus Cells(Turkish Neurosurgical Soc, 2020) Kaya, Yasin Emre; Karaarslan, Numan; Yılmaz, İbrahim; Şirin, Duygu Yaşar; Akalan, Hande; Özbek, HanefiAIM: To investigate the effects of metformin, a drug used widely for the treatment of type 2 diabetes mellitus, on human primary cell cultures prepared from uninjured segment of disc material intervertebral disk tissues. MATERIAL and METHODS: Primary cell cultures were prepared using the tissues of six patients (three males and three females) who had undergone lumbar microdiscectomy and sequestrectomy. Untreated samples served as the control group, and metformintreated samples served as the experimental group. All the samples were evaluated using an inverted light microscope, acridine orange/propidium iodide staining (AO/PI), and a fluorescence microscope. The cytostatic and cytotoxic effects of metformin, which was administered to the samples using a commercial MTT assay kit, were also evaluated. The data obtained were statistically assessed, and the alpha significance value was accepted as less than 0.05. In addition, for the groups' changes in the expressions of chondroadherin (CHAD), cartilage oligomeric matrix protein (COMP), interleukin-1 beta (IL-1 beta) matrix metalloproteinase 7 (MMP-7), and matrix metalloproteinase 19 (MMP-19), genes related to the extracellular matrix synthesis and degradation were determined using gene-specific TaqMan Gene Expression Assays. RESULTS: The administration of the drug adversely affected nucleus pulposus (NP)/annulus fibrosus (AF) cells and extracellular matrix-like structures. This was statistically significant (p<0.05). CONCLUSION: Clinicians should not disregard the adverse effects of metformin, which is used widely in clinical practice, on the components of intervertebral disk tissues.Öğe A Study on the Effects of Direct Factor Xa Inhibitors and Direct Thrombin Inhibitors on Human Primary Chondrocyte Cultures(2019) Kaya, Yasin Emre; Akalan, Hande; Yılmaz, İbrahim; Karaarslan, Numan; Şirin, Duygu Yaşar; Özbek, Hanefi; Ateş, ÖzkanAim:This study investigates the effects of two direct factor Xa inhibitors, apixaban and rivaroxaban, and a direct thrombin inhibitor, dabigatran, on human primary chondrocytecultures.Materials and Methods:Monolayer cultured chondrocytes were prepared. Cell cultures were treated with dabigatran, apixaban, and rivaroxaban. Cultures without drug treatments served as the control group. Using an inverted light microscope, the cell surface morphology was examined. Cell viability and the toxicity of drugs were evaluated using a commercial assay kit, and the results were confirmed using two nucleic acid binding dyes, acridine orange and propidium iodide. The expressions of cartilage oligomeric protein, matrix metalloproteinase-7, and matrix metalloproteinase-19 were assessed using the real-time polymerase chain reaction analysis. All the analyses were performed within 21 days. The data obtained were statistically evaluated.Results:The administration of the three drugs changed the cell viability, proliferation, and expressions of cartilage oligomeric protein, matrix metalloproteinase-7, and matrix metalloproteinase-19. The results were statistically significant (P<0.05).Conclusion:Results obtained from in vitro studies may not provide accurate and reliable insight for clinical practices. However, clinicians should know that drugs used for the prevention or treatment of diseases may suppress chondrocyte proliferation and damage the extracellular matrix formation.Öğe Does Nimodipine, a Selective Calcium Channel Blocker, Impair Chondrocyte Proliferation or Damage Extracellular Matrix Structures?(Bentham Science Publ Ltd, 2019) Kaplan, Necati; Yılmaz, İbrahim; Karaarslan, Numan; Kaya, Yasin Emre; Şirin, Duygu Yaşar; Özbek, HanefiBackground: The study aimed to investigate the effects of the active ingredient, nimodipine, on chondrocyte proliferation and extracellular matrix (ECM) structures in cartilage tissue cells. Methods: Chondrocyte cultures were prepared from tissues resected via surgical operations. Nimodipine was then applied to these cultures and molecular analysis was performed. The data obtained were statistically calculated. Results: Both, the results of the (3-(4,5 dimethylthiazol2-yl)-2,5-diphenyltetrazolium (MTT) assay and the fluorescence microscope analysis [a membrane permeability test carried out with acridine orange/propidium iodide staining (AO/PI)] confirmed that the active ingredient, nimodipine, negatively affects the cell cultures. Conclusion: Nimodipine was reported to suppress cellular proliferation; chondroadherin (CHAD) and hypoxia-inducible factor-1 alpha (HIF-1 alpha) expression thus decreased by 2.4 and 1.7 times, respectively, at 24 hrs when compared to the control group (p < 0.05). Furthermore, type II collagen (COL2A1) expression was not detected (p < 0.05). The risk that a drug prescribed by a clinician in an innocuous manner to treat a patient by relieving the symptoms of a disease may affect the proliferation, differentiation, and viability of other cells and/or tissues at the molecular level, beyond its known side effects or adverse events, should not be forgotten.Öğe Does transcription factor, induced by daptomycin and vancomycin, affect HIF-1?, Chondroadherin, and COL2A1?(2018) Karaarslan, Numan; Yılmaz, İbrahim; Şirin, Duygu Yaşar; Özbek, Hanefi; Kaya, Yasin Emre; Akyuva, Yener; Doğan, Mustafa; Erdem, İlknurAim: In this study, it was firstly aimed to investigate the effect of Daptomycin (DAP) on the proliferation in Vancomycin (VCM)-administered primary chondrocyte cultures and non-drug-administered primary chondrocyte cultures. Our second objective was to investigate the effects of DAP and VCM on the NP-specific marker protein chondroadherin (CHAD), which is associated with spinal cord and dorsal column growth, on the transcription factor-1 alpha (HIF-1?), which is induced by hypoxia, and on a type II collagen (COL2A1), which is also known to play a significant role in the development of extracellular matrix, at the pharmaco-molecular level.Material and Methods: Standard human primary chondrocyte cultures were established. DAP and VCM were added to the samples. In all groups, molecular analysis was performed at 0th, 24th and 48th hours. In addition, the surface morphology of the cells was evaluated.Results: Changes in cell morphology and cell death in cultures were observed 24 hours after administration of antibiotics to cell cultures. It was observed that drug administration was associated with the cell viability and that cell viability rate for two antibiotics was similar at the 0th and 48th hours. The expression of three genes decreased at the 24th hour in the experimental group where DAP was administered.Conclusion: Thanks to this molecular-based research, it should not be forgotten that DAP and VCM active pharmacological agents, especially used in the treatment of Methicillin-resistant Staphylococcus aureus induced surgical infections, have a negative effect on human chondrocyte and ECM components.Öğe Evaluation of the expression and proliferation of degenerative markers in primary cell cultures obtained from human intervertebral disc tissue(2020) Kaya, Yasin Emre; Akalan, Hande; Yılmaz, İbrahi?m; Karaarslan, Numan; Şirin, Duygu Yaşar; Özbek, HanefiAim: A major cause of low back pain is disc degeneration. Nevertheless, no specific and reliable markers of the degeneration of thenucleus pulposus (NP) are available. This presented study aimed to examine changes in the expressions of genes in primary cellcultures isolated from intact and degenerated tissues to give insights into the biopathogenesis of intervertebral disc (IVD) tissue.Material and Methods: Tissues of eight patients (n = 8; average age: 41.74 ± 9.86 years) were resected through microdiscectomy,and primary cell cultures were prepared using degenerated disc tissue. The cultured degenerated tissues served as the study group.The samples in the control group comprised the intact tissues of patients (n = 8; average age: 38.68 ± 7.91 years) resected followinga trauma. Morphology of the cell surface were evaluated using an inverted light/fluorescent microscopy at 0 and 24 h on days 10and 21. The expressions of the chondroadherin (CHAD), cartilage oligomeric matrix protein (COMP), interleukin-1 beta (IL-1 beta),and matrix metalloproteinase (MMP)-7 and MMP-19 genes were evaluated using the reverse transcription-quantitative polymerasechain reaction (RT-qPCR). The data obtained were statistically analyzed.Results: The four genes investigated, except COMP (P > 0.05), changed significantly in primary cell cultures isolated from degenerativeIVD tissues. This result was statistically significant (P < 0.05). The gene expressions in the samples derived from intact IVD tissueschanged markedly and these changes were associated with proliferation (P < 0.05).Conclusion: Analyzing the changes in gene expression levels associated with IVD should contribute to future studies on theprevention and treatment of such pathologies. The data obtained from the present study will shed light on cellular-based personaltargeted therapies through which genetic information can be transmitted to cells.Öğe In-vitro evaluation of the effects of tigecycline on annulus firosus and nucleus pulposus(2020) Kaya, Yasin Emre; Yılmaz, İbrahi?m; Karaarslan, Numan; Bilir, Bülent; Şirin, Duygu Yaşar; Özbek, HanefiAim: This study aimed to examine the effects of tigecycline on primary cell cultures established using intervertebral disc tissues.Material and Methods: Primary Annulus Fibrosus and Nucleus Pulposus cultures were obtained from human intervertebral disctissue. Untreated samples served as the control group, and treated samples served as the study group. Treated and untreatedsamples were statistically evaluated using an alpha signifiance value of < 0.05.Results: Proliferation and gene expression decreased in the tigecycline administered cultures when compared to the control groupsamples (P < 0.05).Conclusion: Drugs used in clinics may have side effects other than those indicated in their package insert.Öğe Investigation of the Effects of Methylphenidate, an Amphetamine Derivative, on Intervertebral Disc Tissue Cell Cultures and Matrix Structures(Turkish Neurosurgical Soc, 2019) Kaya, Yasin Emre; Karaarslan, Numan; Yasar Şirin, Duygu; Özbek, Hanefi; Kaplan, Necati; Yılmaz, İbrahimAIM: To investigate the effects of methylphenidate (MPH), on intervertebral disc tissue (IVD) cell cultures and extracellular matrix structures. Changes in the expression of some important marker genes involved in anabolic and catabolic mechanisms of IVD extracellular matrix formation were also evaluated. MATERIAL and METHODS: Primary cultures of nucleus pulposus cells (NPCs) and annulus fibrosus cells (AFCs) were isolated from tissues obtained from the operated patients. Cell viability and proliferation were tested, and the cell surface morphologies were evaluated by microscopy. The expressions of the chondroadherin (CHAD), cartilage oligomeric matrix protein (COMP), interleukin-1 beta (IL-1 beta) and matrix metalloproteinase (M MP) -7 and M MP-19 genes were evaluated using the quantitative real-time polymerase chain reaction (qRT-PCR). A value of p<0.05 was considered statistically significant. RESULTS: The viability and proliferation of intervertebral disc tissue cells decreased in response to MPH treatment and the expression of the investigated genes also changed. CONCLUSION: The data obtained from in-vitro studies may not directly adaptable to clinical applications. However, the fact that the central nervous system stimulant MPH can suppress proliferation of cells derived from IVD tissue should be considered carefully by clinicians.Öğe Investigation of the effects of tigecycline, a semi-synthetic derivative of minocycline, on chondrocyte cultures(2020) Kaya, Yasin Emre; Karaarslan, Numan; Yılmaz, İbrahim; Şirin, Duygu Yaşar; Yaman, Onur; Özbek, Hanefi; Ateş, ÖzkanAbstractAim: Although antibiotics are generally well-tolerated, they may have cytotoxic effects. The present randomized, double-blind, in vitro study aimed to investigate the effects of tigecycline on cartilage tissue cells and the extracellular matrix.Material and Methods: Cartilage tissues of patients (n = 8) were used for the preparation of primary cell cultures. Tigecycline-treated cell cultures served as the study group. Non-treated cell cultures served as the control group. Analyses were performed at 0, 24, 48, and 72 h in both groups. The results obtained were statistically evaluated. The alpha significance value was determined to be 0.05.Results: Proliferation remained unchanged in the tigecycline-treated cell cultures. The gene expressions of the markers involved in anabolic pathways increased in the tigecycline-treated cell cultures. The results obtained were statistically significant (P < 0.05).Conclusion: Although tigecycline had no toxic effect on the chondrocyte cell cultures and caused no damage to the extracellular matrix, the present study was performed in an in vitro environment.Öğe Iopromide - and gadopentetic acid-derived preparates used in MR arthrography may be harmful to chondrocytes(Bmc, 2017) Öznam, Kadir; Şirin, Duygu Yaşar; Yılmaz, İbrahim; Kaya, Yasin Emre; İşyar, Mehmet; Gümüştaş, Seyit Ali; Mahiroğulları, MahirBackground: Magnetic resonance arthrography, a procedure through which contrast agents containing gadolinium and/or iopromide are administered intra-articularly, has become a useful tool in musculoskeletal diagnosis. Nevertheless, despite being considered safe for systemic use, certain tissue toxicities have been identified for both drugs. In this study, the effects of short-term exposure of human primary chondrocyte cell cultures to gadolinium and/or iopromide contrast agents were examined by assaying for stage-specific embryonic antigen-1 (SSEA-1) protein expression (a chondrogenic differentiation marker), cell viability, toxicity, and proliferation. Methods: Human articular chondrocytes were grown in monolayer culture and were exposed to iopromide and/or gadolinium diethylenetriamine-pentaacetate (Gd-DPT) for 2 and 6 h. Cell cultures with no drug exposure were used as the control group. Cell differentiation status was assessed according to SSEA-1 protein expression. Contrast agent effects on cell viability and proliferation were analyzed using MTT analysis. Further, changes in cell morphology in relation to the control group were evaluated using inverted light microscopy, environmental scanning electron microscopy (ESEM), and 3-tesla magnetic resonance imaging. The obtained data were statistically compared. Results: When compared with the control group, both SSEA-1 protein expression and cell proliferation were lowest in the Gd-DPT group (P = 0.000). There was a statistically significant correlation between SSEA-1 expression and MTT results (rho = 0.351; P = 0.003). Conclusions: Nevertheless, the data obtained from in vitro experiments may not directly correspond to clinical applications. However, the mere fact that a drug used solely for diagnostic purposes may repress chondrocyte cell proliferation should be carefully considered by clinicians.Öğe Pregabalin treatment for neuropathic pain may damage intervertebral disc tissue(Spandidos Publ Ltd, 2018) Karaarslan, Numan; Yılmaz, İbrahim; Şirin, Duygu Yaşar; Özbek, Hanefi; Kaplan, Necati; Kaya, Yasin Emre; Ateş, ÖzkanThe aim of the present study was to determine whether pharmaceutical preparations with pregabalin (PGB) as an active ingredient, which are widely prescribed by clinicians, exert toxic effects on human primary nucleus pulposus (NP) and annulus fibrosis (AF). Primary human cell cultures were obtained from intact (n=6) and degenerated (n=6) tissues resected from the two groups of patients. Different doses of PGB were applied to these cultures and cells were subjected to molecular analyses at 0, 24 and 48 h. Cell vitality, toxicity and proliferation were assessed using a spectrophotometer. The expression of chondroadherin (CHAD), a (member of the NP-specific protein family), hypoxia-inducible factor-1 alpha (HIF-1 alpha) and type II collagen (COL2A1) was measured using reverse transcription-quantitative polymerase chain reaction. The results revealed that cell intensity increased in a time-dependent manner and cell vitality continued in the cultures without pharmaceuticals. Cell proliferation was suppressed in the PGB-treated cultures independent from the dose and duration of application. PGB was demonstrated to suppress the expression of CHAD and HIF-1 alpha. In contrast, COL2A1 gene expression was not revealed in any experimental group. The present study utilized an in vitro model and the PGB active ingredient used herein may not be representative of clinical applications; however, the results demonstrated that PGB has a toxic effect on NP/AF cell cultures containing primary human intervertebral disc tissue. In summary, the use of pharmacological agents containing PGB may suppress the proliferation and differentiation of NP/AF cells and/or tissues, which should be considered when deciding on an appropriate treatment regime.
