archives@tulane.edu / During the 2013-2016 West African outbreak, severe gastrointestinal symptoms were common in Ebola patients and associated with poor outcome. The efficient spread of Ebola virus (EBOV) via vomitus and diarrheal fluids, which contain high concentrations of virus, likely contributed to the scale of the outbreak. The delta-peptide is a conserved product of post-translational processing of the abundant EBOV soluble glycoprotein (sGP). Here, the murine ligated ileal loop model, which is well-established for the study of diarrheal disease, was used to demonstrate that delta-peptide is a potent enterotoxin. Dramatic intestinal fluid accumulation peaked at 9-12 hours following injection of biologically relevant amounts of delta-peptide into ileal loops, along with gross destruction of the villous architecture, loss of goblet cell polysaccharides, and secretion of pro-inflammatory cytokines. Transcriptomic analyses showed that delta-peptide triggers immune and cell survival responses. Delta-peptide may contribute greatly to EBOV-induced gastrointestinal pathology in humans. These findings demonstrate that the EBOV delta-peptide is an enterotoxic viroporin and may be categorized as a novel virulence factor. We then hypothesized that the delta-peptide may also be a druggable target to explore a new avenue for novel EBOV therapeutics, which are direly needed. An unconventional coupling strategy was employed to conjugate a modified version of the 23-residue delta-peptide to a carrier protein Keyhole Limpet Hemocyanin (KLH) in order to immunize rabbits so that they may generate high-affinity binding antibodies against the delta-peptide. Antisera was collected from the rabbits after regular immunizations and the IgG fraction of the antisera demonstrated binding and recognition against several delta-peptide variants confirmed by Western blotting and ELISAs. We then used this knowledge to determine therapeutic index in vitro by testing the antibody against the delta-peptide in the context of synthetic PC-PG vesicles and CHO cells. The purified IgG was then tested against the peptide in vivo to determine therapeutic efficacy by returning to the mouse model of diarrheal pathology mentioned previously. In a small pilot experiment, we were able to successfully block the enterotoxic activity of the delta-peptide and did not observe gastrointestinal distress in the mice that were treated with the peptide and antibody together versus just the peptide alone, signifying that the delta-peptide is a druggable target and may reveal a new therapeutic avenue against EBOV pathogenesis. / 1 / Shantanu Guha
| Identifer | oai:union.ndltd.org:TULANE/oai:http://digitallibrary.tulane.edu/:tulane_119728 |
| Date | January 2020 |
| Contributors | Guha, Shantanu (author), Wimley, William (Thesis advisor), School of Medicine Biomedical Sciences Graduate Program (Degree granting institution) |
| Publisher | Tulane University |
| Source Sets | Tulane University |
| Language | English |
| Detected Language | English |
| Type | Text |
| Format | electronic, pages: 187 |
| Rights | No embargo, Copyright is in accordance with U.S. Copyright law. |
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