Bacteroides fragilis Induce Apoptosis and subG1/G1 Arrest Via Caspase and Nrf2 Signaling Pathways in HT-29 Cell Line

Abstract

Background: Although the link between the human microbiome and colorectal cancer has been elucidated over recent years, its underlying mechanisms for various bacteria such as nontoxigenic Bacteroides fragilis (NTBF) are still unclear. Methods: HT29 cells were treated with Non-toxigenic B. fragilis Sonicated Extract (NBF-SE), strain ATCC-23745, in the dose (10 8 and 10 9 CFU/ml) -and time (24 and 72 hours)-dependent manners. To investigate the cytotoxic impact of NBF-SE on HT29 along with apoptosis-induction and cell cycle distribution, MTT assay and flow cytometry was carried out. Changes in the expression level of Keap1/NRF2/Caspase3/caspase10/GCLM proposed signaling pathway were conducted by real time PCR analyses. Results: We found that the NBF-SE had an antiproliferative effect on HT29 cells. In addition, NBF-SE induced sub-G1 cell cycle cessation and an increment in cell apoptosis, all in a dose-and time-dependent manner. Further, NBF-SE significantly up-regulated Caspase10 and Caspase3 genes, correlated to increased apoptosis. However, Kelch-like ECH-associated protein 1 (KEAP1) and Nuclear factor erythroid 2-related factor 2 (Nrf2) as well as Glutamate-cysteine ligase modulator (GCLM)mediated antioxidation and anti-apoptosis pathway genes were upregulated, which was in contradiction with apoptosis activation. Conclusions: We revealed NTBF as a new inducer of cell death confirmed by cytotoxic activity, cell cycle arrest and, increased apoptosis together with NTBF's greater potency in the activation of caspase apoptotic signaling pathway over NRF2/ GCLM antioxidant response. Moreover, NTBF strain ATCC-23745 may be a hidden probiotic and be used for further studies in CRC treatment.