Intestinal mucositis is one of the major troublesome side effects of cancer chemotherapy leading to poor patient compliance. In this study we address the role of the new apolipoprotein E (ApoE) COG 133 mimetic peptide in Swiss mice challenged with 5-fluorouracil (5-FU) and IEC-6 cell monolayers. Experiments were also performed in C57BL6J ApoE knockout mice to evaluate the effects of apoE peptide treatment.
The experimental groups were the following: unchallenged controls, mice challenged with 5-FU (450 mg/kg, IP) with or without the ApoE peptide (0.3, 1, and 3 μM, administered twice daily ip for 4 days ). Mice were sacrificed 3 days after the 5-FU challenge. Proximal small intestine samples were collected for molecular biology and histological processing. We perform ELISA and RT-PCR assays to target IL-1β, TNF-α, IL-10, iNOS, and myeloperoxidase (MPO) to assess intestinal inflammation.
Cell death and NF-κB assays were also performed in apoE knockout mice. In our in vitro models, IEC-6 cells were exposed to 1 mM 5-FU in medium without glutamine with or without the ApoE peptide (0.02, 0.2, 2, 5, 10, and 20 µM). We investigated the proliferation and migration of IEC-6 cells, 24 h after challenge with 5-FU. Furthermore, IEC-6 apoptotic cells were measured by Tunnel and flow cytometry. Equimolar doses of the ApoA-I (D4-F) peptide was also used in some experiments for comparative studies.
Villus attenuation and intense inflammatory infiltrate were observed in the 5-FU challenged group, findings that were partially ameliorated by the ApoE peptide. We found an increase in intestinal MPO and pro-inflammatory levels of IL-1β and TNF-α, and transcripts of TNF-α and iNOS, and reduction of IL-10 after treatment with 5-FU, each of which was partially abrogated by the peptide. Improvements in apoptosis and migration of IEC-6 cells were also found after treatment with ApoE and D-4F.
Taken together, these findings suggest that the novel ApoE COG 133 mimetic peptide may reduce 5-FU-induced intestinal changes and potentially benefit mucositis.
Keywords: mucositis, apolipoprotein E, 5-fluorouracil, inflammation, cytokines