Assembly of Bioreducible Layer-By-Layer Films for Sequential and Dual Stage DNA Delivery

Polyelectrolyte layer-by-layer (LbL) thin films made by cationic and anionic polymers or bioactive molecules are promising biomaterials in biomedical applications. The layer structure enables control of the amount and location of DNA in the film to be released at different times. The LbL method of assembling polyelectrolyte multilayers (PEMs) has become one of the most promising coating methods capable of mimicking cellular microenvironments and releasing therapeutic agents from the surface of biomedical devices. Our research focuses on the study of the relationship between film assembly and disassembly for potential gene delivery applications. Such biomedical applications generally require the LbL films to disassemble in physiological conditions in order to release the bioactive molecules embedded in the film. We fabricated the LbL film with bioreducible poly(amido amine)s (PAAs) containing the disulfide bond and non-bioreducible polycations in addition to DNA. We show that by combining the two types of polycations, we can maintain the well-defined layer structure in the film and achieve sustained DNA release. The DNA release is based on the bioreducible property of the disulfide bond in PAA. The periodic insertion of non-bioreducible polymer such as poly(ethylenimine) (PEI) as a barrier layer can slow down the release rate to obtain a sequential DNA release pattern. We studied DNA release sequence from the LbL film by atomic force microscopy, fluorescence spectroscopy, and dynamic light scattering. We changed the LbL film disassembly behavior from bulk to sustained release by inserting the PEI barrier layer. The LbL film gene delivery property was demonstrated by transfection experiments in vitro using human embryonic kidney 293 cells and osteoblast precursor cells. By using two different DNA plasmids, we observed sequential DNA release and successful cell transfection. We will present results that directly link the LbL film structure with DNA release dynamics and cell transfection efficiency. Our work demonstrates a simple method for the design of LbL films for sequential, sustained, and dual stage DNA release for biomedical applications.