Abstract
BACKGROUND: Colorectal cancer (CRC) remains as one of the leading causes of human morbidity and mortality worldwide. Dysbiosis in the structure of the gut microbiota resulting in increased abundance of harmful bacteria and decreased beneficial bacteria has been frequently associated with CRC development. Recently, probiotics and their derivatives have received great attention as anticancer agents for prevention and treatment of various cancers. This study aimed to assess the potential effects of probiotic Bacillus coagulans Hammer and its derivatives on induction of apoptosis in human colon adenocarcinoma cell lines in vitro. METHODS: Cell viability of HT-29 and Caco-2 cells was examined by MTT assay after exposure to varying concentrations of pasteurized and UV-killed B. coagulans Hammer, its extracellular vesicles (EVs), and cell-free supernatant (CFS). The apoptotic effects of different B. coagulans Hammer derivatives on HT-29 and Caco-2 cells were determined by analyzing the expression level of apoptosis-related genes (Bad, Bax, Bcl-2, Caspase-3, Caspase-9) using RT-qPCR assay. Annexin V-FITC/PI flow cytometry was performed to assess apoptosis. Protein expression of Bax and Bcl-2 was measured by Western blotting. RESULTS: High concentrations of B. coagulans Hammer derivatives significantly (P < 0.05) reduced viability of HT-29 and Caco-2 cells compared to untreated control. The results of RT-qPCR assay showed significant decrease in the expression level of antiapoptotic gene Bcl-2, and notable increase in the expression level of proapoptotic genes Bax, Bad, Caspase-3, and Caspase-9 in HT-29 and Caco-2 cells by B. coagulans Hammer derivatives. Flow cytometry demonstrated a high percentage of apoptotic HT-29 cells compared to untreated cells after exposure to B. coagulans Hammer derivatives. Western blotting analysis confirmed that treatment with EVs, UV-killed cells, and CFS significantly decreased Bcl-2 expression, and increased Bax expression in HT-29 cells. CONCLUSIONS: Our findings suggest that derivatives of probiotic B. coagulans Hammer can exert anticancer activity by induction of apoptosis in colon cancer HT-29 and Caco-2 cells. Further investigations are required to validate the safety and efficacy of B. coagulans Hammer derivatives in CRC prevention and treatment.