| dc.description.abstract | Indolicidin (IL) is a potential cell-penetrating peptide (CPP), however, its high cytotoxicity restricts its clinical application. In this study, we prepared dimeric IL peptides to promote their biocompatibility and investigate their oligodeoxynucleotide (ODN) delivery efficiencies. Cysteine residues were added to the C or N terminals of IL, by which peptides can be linked as dimers through disulfide bond formation, and we denoted them as ILC and CIL, respectively. HPLC and MASS analyses suggested dimers were successfully prepared in both designs. These dimeric peptides can complex with ODN to self-assembly as nanopartiocles. The MTT results suggested that both ILC and CIL exhibited low cytotoxicity compared to the IL group. The DLS results demonstrated that the sizes of formed peptideplex ranged from 500 to 2000 nm, which were suitable for macrophage uptake. Zeta potentials of peptideplex increased with increasing N/P ratio so that these complexes can be positive charged, however, the absorbance of ILC to ODN is inefficient that the surface charges were always negative except N/P 30. The ODN encapsulation efficiencies of peptides were evaluated by gel retardation assay. Both IL and CIL have good complexation ability to load ODN as stable nanocomplex. In contrast, ILC did not stably complex ODN, which probably correlative to its structure. Because the cationic domain of dimeric ILC is in the middle regions, the hydrophobic domains in two ends probably may mask the positive charges and inhibit the electrostatic interaction between peptide and ODN. However, the fluorescent labeling experiment suggested that both ILC and CIL can promote ODN internalization to Raw264.7 cells. We further investigate the internalization pathway by confocal microscopy, and the results suggested ODN delivered in all groups can be internalized through endocytosis. The long-time observation results indicated that the ODN delivered by CIL can escape from endosome to enter cytosol. Finally, the inhibition experiments were performed against the expression of TNF-α. Anti-TNF-α ODN was delivered to Raw264.7 cell by peptide carriers, and the enzyme-linked immunosorbent assay (ELISA) was performed to measure the expression of TNF-α from Raw264.7 stimulated by lipopolysaccharide (LPS). The results demonstrated CIL demonstrated the best silence against TNF-α, and the inhibition duration can be elongated, suggesting CIL is a potential vector for ODN delivery. | en_US |