CAG peptide functionalized graphene quantum dots-cationic polymer composite gene carriers

Abstract

In this study, a targeted graphene quantum dot-cationic polymer composite gene vector with endothelial cell-targeting CAG peptide was successfully designed and prepared. This vector could efficiently bind and deliver the therapeutic gene pZNF580 to endothelial cells (HUVECs). At a concentration of less than 40 μg mL⁻¹, the results of the CCK-8 assay showed that the relative cell viability of each composite gene vector was greater than 80%, and the results of the flow cytometry assay showed that C-GQDs-PEI-PEG-CAG/pZNF580 (88.96%) and N-GQDs-PEI-PLGA-PEG-CAG/pZNF580 (87.70%) treated groups showed significantly higher cell viability than the positive control group Lip2000/pZNF580 (56.76%). The results of in vitro cell transfection and western blot experiments confirmed that the composite gene vector was able to deliver pZNF580 efficiently and enable the high expression of the ZNF580 protein in HUVECs. The results of the EdU assay, wound healing and Transwell experiments indicated that the composite gene vector/pZNF580 nanoparticles (NPs) could significantly promote the proliferation and migration. The results of the EdU method showed that the proliferative ability of C-GQDs-PEI-PLGA/pZNF580 (84.96 ± 1.99%) and N-GQDs-PEI-PLGA/pZNF580 (85.01 ± 1.31%) treatment groups for HUVECs was significantly higher than that of the positive control group Lip2000/pZNF580 (77.89 ± 2.18%). The results of the scratch assay showed that the cell migration rate of C-GQDs-PEI-PLGA-PEG-CAG/pZNF580 (93.08 ± 1.97%) and N-GQDs-PEI-PLGA-PEG-CAG/pZNF580 (91.99 ± 1.52%) groups was significantly higher than that of the positive control group Lip2000/pZNF580 (85.03 ± 2.21%). In addition, the results of the in vitro angiogenesis assay showed that the C-GQDs-PEI-PLGA-PEG-CAG/pZNF580 and N-GQDs-PEI-PLGA-PEG-CAG/pZNF580 groups had significantly higher angiogenesis-promoting ability than the positive control group, Lip2000/pZNF580.The present study provides a highly efficient and low-toxic method to promote endothelial cell migration in the field of regenerative medicine and a low-toxicity strategy to promote endothelial layer formation, which provides new possibilities for future vascular regeneration therapy.