Polyethyleneimine-capped Silver Nanoclusters For MicroRNA Oligonucleotide Delivery And Bacterial Inhibition

Background&Aims:In recent years,with the development of modern molecular biology techniques,gene therapy has become a new biological technology.Especially as microRNA were found,there have been many studies proved that its important role in the occurrence and development of disease.And the carrier used in gene therapy system determines whether the exogenous gene can express efficient and stable in target cells.Viral vectors are used for gene delivery with wide transfection spectrum,high transfection efficiency and exogenous gene can be fully integrated in target cells,but the potential immunogenicity and infectivity of those viral vectors has been a primary disadvantage for their effective utilization.Because of the limitation of viral vectors,a variety of nonviral gene carriers with clinical potential and ease of production have been designed to transfect,and it has become a hot area.Due to their superior quality such as easy to synthesis,remarkable biocompatibility and photochemical stability,more and more researchs about silver clusters have been reported in the field of biological research.As a new macromolecule material,polyethyleneimine(PEI)is not only used as a transfection material,but also widely used as a template for metal nanoclusters synthesis.At present,polyethyleneimine-capped silver nanoclusters(PEI-AgNCs)have been widely reported to use in biological detection and bioimaging,but its research in cell transfection and antibacterial is still a blank.In this paper,we futher optimization the synthesis conditions of PEI-AgNCs on the basis of predecessors' work,which reduced the time of synthesis and made the synthesis process easier.PEI-AgNCs was researched as a nonviral vector based on the potential of its characterization,the nucleic acid loading efficiency,cell toxicity,and the ability of DNA and microRNAs transfection.Moreover,antibacterial activity and cell imaging of PEI-AgNCs was also studied.Methods&results:First of all,the synthesizing conditions for the highly fluorescent and water-soluble PEI-AgNCs were optimized through the fluorescence intensity of it,and we synthesized poper PEI-AgNCs in accordance with the best synthetic conditions.Subsequently,the resultant PEI-AgNCs were characterized by a photoluminescence assay and transmission electron microscopy.Upon excitation at 380 nm,PEI-AgNCs displayed an intense emission band with a maximum at?500 nm,and the cluster with an average diameter of?2 nm.Then,an agarose gel electrophoresis assay was used to investigate the ability of PEI-AgNCs to condense DNA/RNA at different complex ratios,we find that PEI-AgNCs have great capacity to absorb DNA/RNA,and showed greater ability to absorb DNA.Next,MTT assay was used to investigate the cytotoxicity of PEI-AgNCs,compared to PEI,PEI-AgNCs exhibited less toxicity.Through the analysis of these results,PEI-AgNCs have the potential to act as efficient transfection carrier in the process of gene transfer.To assess the transfection efficiency of DNA mediated by PEI-AgNCs,we performed in vitro transfection-activity assays using HepG2 and 293A cells.In pEGFP-C1transfection experiment,the optimal transfection efficiency for HepG2 cells at a N/P value of 10.8,and for 293A cells was 4.8.The results demonstrated that PEI-AgNCs is an efficient carrier in gene delivery.However,DNA transfection data cannot readily be implemented for other genes,such as small RNAs.Next,we further verify the transfection ability of the PEI-AgNCs to miRNA inhibitors and miRNA mimics,and found that PEI-AgNCs can transfect miR-21/221 inhibitors effectively.More importantly,PEI-AgNCs were confirmed to transfect miRNA mimics more effectively,and this fascinating result provided an attractive chance for tumor treatment that depends on miRNA replacement.To prevent bacterial contamination in the process of transfection,we explored the antimicrobial activities of PEI-AgNCs.We found that PEI-AgNCs exhibited higher antibacterial activity than PEI alone,which made it possible to prevent bacterial contami-nation in the process of transfection.Because AgNCs showed the fascinating optical properties and the superiority of easy synthesis by designing different ligands as stabilizing agents,we employed PEI-AgNCs for biological imaging,and the cells revealed an extremely intense green fluorescence.Compared to PEI,this characteristic make PEI-AgNCs having great potential to be an ideal fluorescent probe to track transfection behavior.Conclusions:In conclusion,motivated by the appropriate size,the capacity to absorb nucleic acid and the appropriate particle size of complexes,PEI-AgNCs have great potential to act as gene carriers.PEI-AgNCs have been proved to be a new and great gene carrier through transfected with DNA,the transfection of miRNAs further demonstrated its potential in gene delivery.More importantly,PEI-AgNCs transfected miRNA mimics more effectively than PEI,this may provided an attractive chance for tumor treatment that depends on miRNA replacement.Simultaneously,compared to PEI,PEI-AgNCs showed the lower cytotoxicity,the higher antibacterial activity,and AgNCs endow PEI with strong fluorescence,which make PEI-AgNCs as a more advantageous gene carrier than PEI.All in all,PEI-AgNCs can act as a promising and novel nonviral vectors for gene delivery.