The effect of oral microbes on the proliferation, healing, and immune response of intestinal epithelial cells, IEC, treated with SN-, metabolite of irinotecan

Abstract

Introduction: Gastrointestinal mucositis is a toxicity of chemotherapy. Gut microbial composition has been shown to be altered following chemotherapy, and therefore may affect proliferation and healing properties of intestinal cells. Toll-like receptors (TLRs) are immune regulators triggered by microbes that may contribute to the initiation of mucositis. The aim was to investigate effects of microbes on intestinal epithelial cells (IEC-6) following SN-38, and whether microbes affects TLR signaling. Methods: IEC-6 cells were seeded and cultured in 24-well Transwell TM plates until confluent. Next, monolayers were wounded by means of a sterile pipette tip and treated with 500 muM SN-38 or DMSO (control). In parallel, microbes from an oral swab were transferred onto an agar/mucin layer in the TranswellTM inserts and inserts were placed on top of the wounded monolayers. Wound areas were calculated at baseline and after 24 h of exposure to the microbes. Proliferation of the cells at 24 h was evaluated using an SRB assay. TLRs were measured with real time PCR. Results: In the presence of microbes, SN-38 significantly inhibited wound healing at 24 h (p < 0.05), while SN-38 as such had no effect. In contrast, cell proliferation was significantly inhibited by SN-38 and more pronounced when microbes were present (p < 0.05). At 48 h, TLR4 expression significantly increased in the presence of microbes, irrespective of the presence of SN-38 (p < 0.05). Results are pending for TLR2, 5 and 9. Conclusions: Cellular activities such as proliferation and migration (wound healing) are clearly impacted by the presence of microbes. The increased expression levels of TLR4 following exposure to microbes suggests a mechanistic role for this receptor in these events.