Six-cell penetrating peptide-based fusion proteins for siRNA delivery.
The siRNA deliveries, for the siRNA's high negative charge, short serum half life, poor cellular internalization, etc, are still the key barriers for its application in clinic. In this study, several cell penetrating peptide (CPP) and dsRNA binding domain (dsRBD)-based fusion proteins have been developed and screened as the siRNA vector. The siRNA binding ability was measured by the agarose gel retardation, the cell uptaking was characterized under fluorescence microscopy, and further more RNAi effect was evaluated on the endogenous (GAPDH, western blot) and exogenous (GFP, flow cytometry analysis) genes in HeLa cell. Finally, the cytotoxicity was assessed on HeLa cells using cell counting kit-8. The efficiency of siRNA delivery by the CPP-dsRBD fusion protein was the CPP and the dsRBD dependent. Three fusion proteins showed similar efficiency of siRNA delivery when comparing to Lipofectamine RNAi Max as the siRNA carrier. These results indicated that these CPP-dsRBD-based fusion proteins were promising candidates as siRNA carriers.
Department of General, Visceral and Thoracic Surgery, University Medical Center Hamburg-Eppendorf , Hamburg , Germany and.
Hua
H
Li
TungYu
T
Tsui
D015153:Blotting, Western; D018929:Cell Culture Techniques; D002470:Cell Survival / Q000187:drug effects / Q000235:genetics; D057846:Cell-Penetrating Peptides / Q000737:chemistry* / Q000633:toxicity; D004337:Drug Carriers / Q000737:chemistry* / Q000633:toxicity; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics; D005434:Flow Cytometry; D055785:Gene Knockdown Techniques; D006367:HeLa Cells; D006801:Humans; D008856:Microscopy, Fluorescence; D034741:RNA, Small Interfering / Q000008:administration & dosage* / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000737:chemistry* / Q000633:toxicity
D057846:Cell-Penetrating Peptides; D004337:Drug Carriers; D029968:Escherichia coli Proteins; D034741:RNA, Small Interfering; D011993:Recombinant Fusion Proteins
Calcium; RNA interference; cell penetrating peptide; intracellular delivery; siRNA