부산대학교 도서관

Endotoxin, also identified as lipopolysaccharide (LPS), is considered the pathogenic factor of septic shock triggered by Gram-negative bacteria and generates inflammatory responses. Synthetic peptides have attracted increasing attention from researchers for the blocking of LPS and treatment of sepsis. The aim of the study was to design a novel synthetic anti-endotoxin peptide and evaluate its effect in vitro. To design a new peptide, anti-endotoxin peptides were extracted from the APD3 site. The physicochemical features, secondary structure content, and tertiary structure type of each residue were determined by ProtParam, GOR IV, pep-fold, and I-TASSER. Hemolytic activity and cytotoxicity of the peptide on RAW264.7 cells were assessed by human RBC hemolysis test and MTT assay, respectively. Real-time PCR and western blot were used to evaluate the gene expression of IL-1β, TNF-α, IL-6, IL-10, iNOS, and TLR4, as well as the protein expression of NF-KB(P65), correspondingly. The designed peptide has 13 amino acid residues (GRRWWRFKKWWKF). The second structure of the peptide had 46.15% random coil and 53.85% extended strand. The results of the prediction of the tertiary structure demonstrated the peptide forms an alpha helix structure. It possesses low hemolytic activity and low cytotoxicity against RAW264.7 cells. This peptide remarkably restored LPS-induced TLR4 overexpression, and reduced gene expression of IL-1β, IL-6, iNOS, and TNF-α, whereas increased IL-10. This peptide significantly reduced the protein expression of NF-KB (P65). These findings imply that this peptide with low toxicity, hemolytic activity, and LPS-neutralizing activity merits more research as a possible anti-LPS agent for managing septic shock. [ABSTRACT FROM AUTHOR]