(93f) Controlling Nonviral Gene Delivery through the Cell-Biomaterial Interface

The use of gene delivery in therapeutic applications, including gene therapy to treat genetic deficiencies or tissue engineering matrices for the treatment of organ loss and failure, has been limited due to challenges with current delivery systems. Current research in nonviral gene delivery has focused on improving efficiency, through development of new delivery vehicles and delivery mechanisms, but has paid little attention to factors that control cell responsiveness to gene transfer, which in the context of many biotechnological applications, may be controlled by interactions between a biomaterial and cell. These interactions, which typically involve cell attachment to proteins adsorbed at biomaterial surfaces have been shown to control cellular behaviors such as cell adhesion, spreading and proliferation, which in turn may dictate the efficiency of nonviral gene delivery to these cells. Self-assembled monolayers (SAMs) of alkanethiols on gold were used as model biomaterials with controllable surface chemistry to investigate the effect of the cell-biomaterial interactions on nonviral gene delivery. SAMs presenting different surface functional groups were used to modulate the amount, conformation, and activity of extracellular matrix (ECM) molecules known to adsorb on biomaterial surfaces, which in turn may modulate cellular behaviors critical to the efficacy of nonviral gene delivery. As an example ECM protein, fibronectin was adsorbed to both hydrophobic and hydrophilic (charged and uncharged) SAMs and mouse fibroblasts (NIH/3T3) were then seeded onto the SAMs with immobilized fibronectin. DNA complexes (formed with plasmid encoding firefly luciferase complexed with polyethylenimine or Lipofectamine 2000) were subsequently delivered by bolus methods and transfection was assayed at 48 hours with standard luciferase assays. Transfection on ECM-immobilized SAMs was compared to transfection in cells adhered to SAMs without immobilized ECM, as well as traditional polystyrene surfaces. Fibronectin immobilized on hydrophobic SAMs resulted in increased transfection levels relative to hydrophilic SAMs or control polystyrene surfaces, which suggests that changes in adsorbed fibronectin orientation may result in altered integrin binding profiles, which in turn modulate gene transfer. Furthermore, transfection was shown to increase as the concentration of fibronectin on each SAMs was increased, which suggests that changes in cell-biomaterial interactions that affect focal adhesion strength may also enhance the ability of the cells to respond to gene delivery. An increased understanding of the cell-biomaterial interface, including how the architecture of ECM proteins controlled by substrate binding and the resulting cellular behaviors translated from the extracellular environment through integrin receptors and focal adhesions, can be used to design tissue engineering scaffolds that promote nonviral gene delivery.