Bakır (II) metal-karışık ligand kompleksinin [Cu (5-nitro-1-10-fenantrolin) (L-asparajin)] CIO4)n sitotoksik ve DNA hasar etkilerinin çeşitli hücre hatlarında in vitro belirlenmesi
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Date
2016-10-17
Authors
Oral, Neylan
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Publisher
Uludağ Üniversitesi
Abstract
Bu çalışmada hazır besiyerinde (RPMI medium + Fetal Bovine Serum + L-glutamin + Sodyum pirüvat + Penisilin-Streptomisin) çözünen yeni sentezlenmiş polimerik yapılı bir bakır kompleksinin[Cu (5-nitro-1-10-fenantrolin) (L-asparajin)] CIO4)n, A549 insan akciğer kanseri hücre hattı, Beas-2B insan sağlıklı bronş epitel hücre hattı, MCF7 insan meme kanseri hücre hattı ve Caco-2 insan kolon kanseri hücre hattında sitotoksik etkisi XTT testi ile incelenmiştir. Bu testte hücre hatları 0,1'den 25.6 μM arasında değişen konsantrasyonlarda bakır kompleksi ile 24 saat muamele edilip sonucunda yüzde canlılık eğrisi elde edilerek IC12,5, IC25, IC50 ve IC75 değerleri bulunmuştur. Hücrelerin canlılığını sonlandıran mekanizmayı aydınlatmak için ise bir genotoksisite testi olan komet testinden yararlanılmıştır. Hücre hatlarının ortalama kuyruk uzunluğu, kuyruk %DNA ve Olive kuyruk momenti ölçümleri değerlendirilmiştir. Hücre içi ROS düzeyleri DCF-DA maddesi ile yapılan ROS testi ile ölçülmüştür. XTT testi sonuçları, düşük dozlarla muamele görmüş hücrelere oranla yüksek dozlarla muamele görmüş hücrelerin yüzde canlılık değerinin düştüğünü göstermiştir. Komet testi sonuçları, bu bakır kompleksine maruz kalan hücrelerde DNA fragmentasyonu gerçekleştiğini göstermiştir, ROS testi ise bu fragmentasyonların oksidatif hasar sebebiyle oluştuğuna işaret etmiştir. Sonuçlar bu bakır kompleksinin Caco-2 insan kolon kanseri hücre hattı ve MCF7 insan meme kanseri hücre hattı üzerinde daha etkili olduğunu göstermiştir. Tüm metaller arasında bakırın her canlı organizmanın fizyolojisi ve biyokimyasında merkezi rol oynaması, kanser tedavisinde sıklıkla kullanılan cisplatinin yapısındaki platin metalinin aksine endojen bir metal olması, bu sebeple daha az toksik yan etki göstermesi ve nükleik asit kırabilme yeteneğiyle DNA'da hasar yaratması kanser tedavisinde onu başarılı bir alternatif haline getirmiştir.
In this study cytotoxic effect of the newly synthesized copper complex [Cu (5-nitro -1-10 - phenanthroline)(L- asparagine)] ClO4)n which dissolves in the stock medium (RPMI medium + fetal bovine serum + L- glutamine + sodium pyruvate + Penicillin-Streptomycin) is analyzed in A549 human lung cancer cell line, Beas-2B human healthy bronchial epithelial cell line, MCF7 human breast cancer cell line and Caco-2 human colon cancer cell line by XTT assay. The cell lines in this test are treated with the copper complex at concentrations of from 0.1 to 25.6 µM for 24 hours obtaining from the results a percentage vitality curve and so IC12,5, IC25, IC50 and IC75 values are found. To elucidate the mechanisms that ends the vitality of the cells, comet which is a genotoxicity assay is utilized. Average tail length, tail % DNA and olive tail moment of cell lines measurements are evaluated. Intracellular Reactive Oxygen Species levels is measured by the assay which is made with DCF-DA. XTT assay results are revealed that percentage vitality curve decreases in high doses by comparison low doses. Comet assay results are revealed that DNA fragmentations are formed in the cells treated with this copper complex and according to ROS assay results these fragmentations are occured by oxidative damage. Results of the assays are presented that this copper complex is more effective in Caco-2 human colon cancer cell line and MCF7 human breast cancer cell line than the other cell lines. Among all metals, copper plays a central role in every living organism's biochemistry and physiology, it is an endogenous metal in comparison with cisplatin which is frequently used in cancer treatments and for this reason it causes less toxic side effects, it causes DNA damage with nucleic acid break ability and so copper is a successful alternative in cancer treatment because of these properties.
In this study cytotoxic effect of the newly synthesized copper complex [Cu (5-nitro -1-10 - phenanthroline)(L- asparagine)] ClO4)n which dissolves in the stock medium (RPMI medium + fetal bovine serum + L- glutamine + sodium pyruvate + Penicillin-Streptomycin) is analyzed in A549 human lung cancer cell line, Beas-2B human healthy bronchial epithelial cell line, MCF7 human breast cancer cell line and Caco-2 human colon cancer cell line by XTT assay. The cell lines in this test are treated with the copper complex at concentrations of from 0.1 to 25.6 µM for 24 hours obtaining from the results a percentage vitality curve and so IC12,5, IC25, IC50 and IC75 values are found. To elucidate the mechanisms that ends the vitality of the cells, comet which is a genotoxicity assay is utilized. Average tail length, tail % DNA and olive tail moment of cell lines measurements are evaluated. Intracellular Reactive Oxygen Species levels is measured by the assay which is made with DCF-DA. XTT assay results are revealed that percentage vitality curve decreases in high doses by comparison low doses. Comet assay results are revealed that DNA fragmentations are formed in the cells treated with this copper complex and according to ROS assay results these fragmentations are occured by oxidative damage. Results of the assays are presented that this copper complex is more effective in Caco-2 human colon cancer cell line and MCF7 human breast cancer cell line than the other cell lines. Among all metals, copper plays a central role in every living organism's biochemistry and physiology, it is an endogenous metal in comparison with cisplatin which is frequently used in cancer treatments and for this reason it causes less toxic side effects, it causes DNA damage with nucleic acid break ability and so copper is a successful alternative in cancer treatment because of these properties.
Description
Keywords
Bakır (II) karışık ligand kompleksleri, Antikanser etki, XTT testi, ROS testi, Komet testi, Copper (II) mixed ligand complexes, Anticancer effects, XTT assay, ROS assay, Comet assay
Citation
Oral, N. (2016). Bakır (II) metal-karışık ligand kompleksinin [Cu (5-nitro-1-10-fenantrolin) (L-asparajin)] CIO4)n sitotoksik ve DNA hasar etkilerinin çeşitli hücre hatlarında in vitro belirlenmesi. Yayınlanmamış yüksek lisans tezi. Uludağ Üniversitesi Fen Bilimleri Enstitüsü.