Determination Cytotoxic effect of Thymoquinone and Expression analyses of Apoptotic and Autophagic related genes with Thymoquinone treatment in Colon Cancer Cells

Abstract

Thymoquinone (Tq; 2-isopropyl-5-methyl-1,4 benzoquinone) is the main ingredient that presents in the volatile oil of Nigella sativa (black seed). Recently, it has been reported that, Tq was inhibited cell proliferation of many cancer cell lines including ovarian, lung, breast, osteosarcoma, pancreatic, fibrosarcoma, lung cancer, squamous cell carcinoma and myeloblastic leukemias. In this study, the anti-cancer effect of Tq was investigated in human Colon Cancer Cells (HT-29) with MTT assay. Positive control was achieved with 5-fluorouracil (5-FU). Also, the invasion capability of HT-29 cells was determined in presence of Tq and 5-FU with scratch analyses. In addition, the autophagic and apoptotic effect of Tq were investigated by qRT-PCR method to find out the pathway of anti-proliferative effect in Tq-treated HT-29 cells. As a result, Tq was inhibited the cell proliferation of HT-29 cells in time dependent manner, with 118 µM for 24 h and 84 µM for 48 h, respectively. A concentration of 150 µM of Tq was able to significantly reduced the invasiveness of HT-29 cells approximately 8 fold compared with not treated cells at 6h. ATG-12, ATG-7 and LC3-II were significantly downregulated in presence of Tq. Conversely, the expression of all the autophagy genes were downregulated in presence of 5-FU. Moreover, pro-apoptotic gene Bax was significantly upregulated nearly 15 fold whereas the expressions of Bcl-2 and Bcl-XL (pro-survival genes) were decreased in presence of Tq. On the other hand, in 5-FU treated HT-29 cells, the expression of Bax, Bcl-2 and Bcl-XL genes and autophagy related genes were significantly downregulated. © 2020, Sakarya University. All rights reserved.