Nanosheet transfection: effective transfer of naked DNA on silica glass

Abstract

Existing gene delivery technologies using nonviral vectors or physical stimulation are limited by cytotoxicity. Here, we reveal an easy and new method to fabricate silica upright nanosheets, which can be easily scaled up and used for gene delivery. We demonstrate that naked DNA can be transferred into difficult-to-transfect cells (for example, stem cells) by plating cells on silica glass with the upright nanosheets without using any vector. Gene entry is probably achieved through the integrin-FAK-Rho signaling axis, which can be activated by the cytoskeleton rearrangement on nanosheets in a limited time frame (‘transfection window’). Transfecting naked GATA4-binding protein 4 plasmids into stem cells can upregulate the other two important cardiac marker genes myocyte enhancer factor 2C and T-box 5. The transfected mesenchymal stem cells express the cardiac marker proteins at 7 days after transfection, which confirms the success of the innovative gene delivery approach. The vector-free silica nanosheet-induced transfection is simple and effective but faster and safer than the conventional technologies. ‘Naked’ DNA strands can directly transfer into host cells using surfaces embedded with freestanding silica nanosheets, finds a new study. Gene therapy has the potential to elimate disease by altering a patient's genetic code, but getting designer DNA plasmids past cellular membranes is difficult. Now, Shan-hui Hsu from the National Taiwan University and co-workers in Japan have developed a strategy to improve gene delivery with a procedure that elimates the need to use viruses or other biocompatible molecules to carry nucleic acids into cells. Adding naked DNA plasmid onto glass substrates covered with rough nanosheet textures enhanced entry of cardiac marker gene (GATA4) into stem cells by mediating cell-membrane receptors. This low-cost strategy introduced genes to 70% of starting cells within 24 hours, making it attractive for scalable in vivo gene delivery with limited cytotoxicity side effects. A nanosheet structure on a glass plate can transfer naked DNA into difficult-to-transfect cells (for example, stem cells) in limited time after contact (‘transfection window’) without any vector. The transfected cells express the specific proteins a week later. The gene uptake and expression are associated with the cytoskeleton arrangement on the silica nanosheets by activating the cell-membrane integrin receptor.