Investigation of the Effect of Carbonic Anhydrase-IX Enzyme Inhibition with Acetazolamide on Glutathione Reductase and Glutathione Peroxidase Activities

Öz

Oxidative stress is one of the important parameters for renal cancer. The antioxidant system plays a prominent role and resists oxidative stress in the development of renal cancer. The endogenous antioxidants GPx and GR, are defined as crucial enzymes in the antioxidant system of the kidneys. The main purpose of this study is to determine the effect of AZA, a carbonic anhydrase classical enzyme inhibitor in the renal cancer cell line CAKI-2 to evaluate the GPx and GR enzyme activities. The optimal dose of AZA was found to be 8.65 μM at the 48th hour by WST-1 cytotoxicity test. ELISA test was performed to determine the effect of AZA on CA-IX enzyme in CAKI-2 cells. All measurements were performed at 340 nm by Epoch™ Microplate Spectrophotometer using “Glutathione Reductase Assay Kit” and “Glutathione Peroxidase Activity Assay Kit”. After AZA application, an increase was observed in GR and GPx enzyme activities (p≤0.05). As a consequence, AZA may be an important therapeutic agent for the glutathione mechanism related to renal cancer. Our study makes a valuable contribution to the literature due to the importance of the CA-IX enzyme related to renal cancer. The investigation of other endogenous antioxidant enzymes in renal cancer may be designed for further prospective studies.

Anahtar Kelimeler

• Acetazolamide
• Carbonic anhydrase
• Glutathione peroxidase
• Glutathione reductase
• Renal Cancer

Karbonik Anhidraz-IX Enziminin Asetazolamid ile İnhibisyonunun Glutatyon Redüktaz ve Glutatyon Peroksidaz Aktiviteleri Üzerine Olan Etkisinin İncelenmesi

Öz

Oksidatif stres, renal kanser için önemli parametrelerden birisidir. Antioksidan sistem renal kanser oluşumunda devreye girerek oksidatif strese karşı koyar. Endojen antioksidanlar olarak tanımlanan GR ve GPx, böbreklerin antioksidan sistemindeki önemli enzimlerdir. Renal kanserdeki önemli parametrelerden biri olan CA-IX, bir pH pompası olarak görev yaparak tümör mikroçevresinin asidifikasyonuna sebep olur ve karsinogenezde rol oynar. Çalışmamızın temel amacı, bir karbonik anhidraz enzim inhibitörü olan AZA’nın glutatyon mekanizması üzerine olan etkisinin renal kanserde incelenmesidir. Deneysel çalışmalarda öncelikle renal kanser hücre hattı olan CAKI-2 çoğaltılarak WST-1 sitotoksisite testi ile AZA’nın uygun dozu 48. saatte 8.65 μM olarak bulunmuştur. AZA’nın CAKI-2 hücrelerinde CA-IX enzimi üzerine olan etkisi belirlenmek için ELISA testi yapılmıştır. CAKI-2 hücrelerine AZA uygulandıktan sonra GR ve GPx üzerine olan etkisini belirlemek için “Glutathione Reductase Assay Kit” ve “Glutathione Peroxidase Assay Kit” kullanılarak Epoch™ Microplate Spectrophotometer cihazında 340 nm’de ölçüm yapılmıştır. AZA uygulaması sonrası GR ve GPx enzim aktivitelerinde artış görülmüştür (p≤0.05). Çalışmanın sonucu olarak AZA inhibisyonunun, renal kanserde glutatyon mekanizmasının devreye girmesi için önemli bir ajan olabileceği söylenebilir. Renal kanserde hem CA-IX enziminin önemli bir terapötik biyobelirteç olması hem de glutatyon mekanizmasının bu kanser türündeki öneminden dolayı çalışmamız literatüre önemli bir katkı sağlamaktadır. Bu çalışmanın devamı niteliğinde olması planlanan diğer endojen antioksidan enzim aktivitelerinin renal kanserde araştırılması yeni terapötik yaklaşımların geliştirilmesini sağlayacaktır.

Anahtar Kelimeler

• Asetazolamid
• Glutatyon peroksidaz
• Glutatyon redüktaz
• Karbonik anhidraz
• Renal Kanser

Kaynakça

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