BackgroundIntestinal mucositis is one of the major troublesome side effects of anticancer chemotherapy leading to poor patient compliance. In this study we addressed the role of the novel apolipoprotein E (ApoE) COG 133 mimetic peptide in 5-fluorouracil (5-FU)-challenged Swiss mice and IEC-6 cell monolayers. Experiments were also conducted in C57BL6J ApoE knock-out mice to assess the effects of apoE peptide treatment.MethodsExperimental groups were as follows: unchallenged controls, 5-FU-challenged mice (450 mg/kg, i.p) with or without the ApoE peptide (0.3, 1, and 3 μM, given twice daily i.p. for 4 days). Mice were sacrificed 3 days after 5-FU challenge. Proximal small intestinal samples were harvested for molecular biology and histological processing. We conducted ELISA assays and RT-PCR to target IL-1β, TNF-α, IL-10, iNOS, and myeloperoxidase (MPO) to assess intestinal inflammation. Cell death and NF-κB assays were also conducted in apoE knock-out mice. In our in vitro models, IEC-6 cells were exposed to 1 mM of 5-FU in glutamine free media with or without the ApoE peptide (0.02, 0.2, 2, 5, 10, and 20 μM). We investigated IEC-6 cell proliferation and migration, 24 h after the 5-FU challenge. Additionally, apoptotic IEC-6 cells were measured by Tunel and flow cytometry. Equimolar doses of the ApoA-I (D4-F) peptide were also used in some experiments for comparative studies.ResultsVillus blunting and heavy inflammatory infiltrates were seen in the 5-FU-challenged group, findings that were partially ameliorated by the ApoE peptide. We found increased intestinal MPO and pro-inflammatory IL-1β and TNF-α levels, and TNF-α and iNOS transcripts, and reduction of IL-10 following 5-FU treatment, each of which were partially abrogated by the peptide. Improvements were also found in IEC-6 cell apoptosis and migration following ApoE and D-4F treatment.ConclusionAltogether, these findings suggest that the novel ApoE COG 133 mimetic peptide can reduce 5-FU-induced intestinal changes and potentially benefit mucositis.

Background. Clostridium difficile is an anaerobic bacterium that causes antibiotic-associated diarrhea. It produces toxin A and toxin B (TcdB), which cause injury to the gut epithelium. Glutamine is a fundamental fuel for enterocytes, maintaining intestinal mucosal health. Alanyl-glutamine (AQ) is a highly soluble dipeptide derivative of glutamine. We studied whether administration of AQ ameliorates the effects of TcdB in the intestinal cells and improves the outcome of C. difficile infection in mice. Methods. WST-1 proliferation and cell-wounding-migration assays were assessed in IEC-6 cells exposed to TcdB, with or without AQ. Apoptosis and necrosis were assessed using Annexin V and flow cytometry. C57BL/6 mice were infected with VPI 10463 and treated with either vancomycin, AQ, or vancomycin with AQ. Intestinal tissues were collected for histopathologic analysis, apoptosis staining, and determination of myeloperoxidase activity. Results. AQ increased proliferation in intestinal cells exposed to TcdB, improved migration at 24 and 48 hours, and reduced apoptosis in intestinal cells challenged with TcdB. Infected mice treated with vancomycin and AQ had better survival and histopathologic findings than mice treated with vancomycin alone. Conclusions. AQ may reduce intestinal mucosal injury in C. difficile-infected mice by partially reversing the effects of TcdB on enterocyte proliferation, migration, and apoptosis, thereby improving survival from C. difficile infection.