ABSTRACT
Over the last two decades, a multitude of gain-of-function studies have been conducted on genes that encode antioxidative enzymes, including one of the key enzymes, manganese superoxide dismutase (SOD2). The results of such studies are often contradictory, as they strongly depend on many factors, such as the gene overexpression level. In this study, the effect of altering the ectopic expression level of major transcript variants of the SOD2 gene on the radioresistance of HEK293T cells was investigated using CRISPRa technology. A significant increase in cell viability in comparison with the transfection control was detected in cells with moderate SOD2 overexpression after irradiation at 2 Gy, but not at 3 or 5 Gy. A further increase in the level of SOD2 ectopic expression up to 22.5-fold resulted in increased cell viability detectable only after irradiation at 5 Gy. Furthermore, a 15-20-fold increase in SOD2 expression raised the clonogenic survival of cells after irradiation at 5 Gy. Simultaneous overexpression of genes encoding SOD2 and Catalase (CAT) enhanced clonogenic cell survival after irradiation more effectively than separate overexpression of both. In conjunction with the literature data on the suppression of the procarcinogenic effects of superoxide dismutase overexpression by ectopic expression of CAT, the data presented here suggest the potential efficacy of simultaneous overexpression of SOD2 and CAT to reduce oxidative stress occurring in various pathological processes. Moreover, these results illustrate the importance of selecting the degree of SOD2 overexpression to obtain a protective effect.
Subject(s)
Oxidative Stress , Superoxide Dismutase , Humans , HEK293 Cells , Superoxide Dismutase/metabolism , TransfectionABSTRACT
20-Hydroxyecdysone (20E) is an arthropod hormone which is synthesized by some plants as part of their defense mechanism. In humans, 20E has no hormonal activity but possesses a number of beneficial pharmacological properties including anabolic, adaptogenic, hypoglycemic, and antioxidant properties, as well as cardio-, hepato-, and neuroprotective features. Recent studies have shown that 20E may also possess antineoplastic activity. In the present study, we reveal the anticancer properties of 20E in Non-Small Cell Lung Cancer (NSCLC) cell lines. 20E displayed significant antioxidant capacities and induced the expression of antioxidative stress response genes. The RNA-seq analysis of 20E-treated lung cancer cells revealed the attenuation of genes involved in different metabolic processes. Indeed, 20E suppressed several enzymes of glycolysis and one-carbon metabolism, as well as their key transcriptional regulators-c-Myc and ATF4, respectively. Accordingly, using the SeaHorse energy profiling approach, we observed the inhibition of glycolysis and respiration mediated by 20E treatment. Furthermore, 20E sensibilized lung cancer cells to metabolic inhibitors and markedly suppressed the expression of Cancer Stem Cells (CSCs) markers. Thus, in addition to the known beneficial pharmacological activities of 20E, our data uncovered novel antineoplastic properties of 20E in NSCLC cells.
