The Effect of Thymoquinone on Expression Levels of Apoptotic, Oxidative and Antioxidant Genes in NRK-52E Cells

Abstract

Thymoquinone (TQ), the activeingredient of Nigella sativa, has manybeneficialeffects, especiallyitsantioxidant and anti-inflammatoryproperties. Thisstudywasplannedtodemonstrate the effects of TQ on apoptotic and oxidativepathways in kidneycells, depending on concentration and time. Forthispurpose, NRK-52E (ATCC® CRL-1571™) ratkidneyepithelialcelllinewasused as material in the study. NRK-52E cellswereproducedbysystematicallypassage of cells in an appropriatemediumunder in vitroconditions. MTT cellviability test wasperformedtodetermine the IC50 value of TQ at 24th and 48th hours. The proliferative (TQp-10µM) and toxic (TQIC50-60µM) concentrationsto be administered in the studyweredetermined. Insamples of theseconcentrationsobtained at 24 and 48 hours, apoptotic CASP3 (caspase 3), CASP8 (caspase 8), CASP9 (caspase 9), BAX (BCL2 associated X), BCL2 (BCL2 apoptosisregulator) and oxidative (GPX1 (glutathioneperoxidase 1)), SOD1 (superoxidedismutase 1), NCF1 (neutrophilcytosolicfactor 1) gene expressionswereperformedby RT-qPCR. The FoldChangewasfoundusing the formula 2^(-delta delta CT). Accordingto the results of this analysis, compared to the control gene, GPX1 at 24 hours at the TQ proliferative concentration was significantly up-regulated at the TQIC50 concentration. It was observed that the oxidative NCF1 gene did not change depending on the concentration at the 24th hour. Apoptotic genes were found to be limited up-regulated at both concentrations at 24 hours. GPX1 was up-regulated (7,215-fold) at the TQp concentration at the 48th hour, while SOD1 was up-regulated (3,623-fold) at the TQIC50 concentration. It was determined that NCF1 gene was significantly up-regulated at high concentration at 48th hour. While apoptotic genes were up-regulated to a limited extent at the TQp concentration at 48th hours, all genes were found to be significantly up-regulated at the TQIC50 concentration. In conclusion, in this thesis study, it was found that GPx was upregulated at the 24th hour and SOD1 was significantly upregulated at the TQIC50 concentration, the NCF gene did not change significantly, the oxidative stress increased at the toxic concentration in the 48th hour, and SOD was used as an antioxidant. It was revealed that the effect of apoptotic pathway in TQ-dependent cell death was limited at the 24th hour, and that cell death at toxic concentration occurred via the external pathway at the 48th hour.

Keywords

• Thymoquinone
• apoptosis
• oxidative
• kidney

References

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