Background: Brain tumors can be serious and life-threatening when they are treated effectively. Many therapeutic approaches, such as chemotherapy, radiotherapy, and surgery have been used to treat brain tumors. In this regard, selenium supplements have been reported effective.Methods: Selenium Nanoparticles (SeNPs) were produced in two forms of synthetic and biogenic to evaluate their cytotoxicity on brain glioblastoma cell lines. A-172 cell line was cultured in DMEM medium. The cytotoxicity of the synthetic and biogenic SeNP was assessed by MTT assay. Results: There was a significant difference between the group treated with biogenic and synthetic SeNP compared with non-treated cells after 24, 48, and 72 h. Both biogenic and synthetic SeNP increased Bax gene and decreased Bcl-2 gene expression. Conclusion: It seems that biogenic SeNP was more lethal than its synthetic form. Therefore, it should be considered that the method of NP construction may be an important parameter for its bioactivity.