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Gene delivery to primary human cells is a technology of critical interest to both life science research and therapeutic applications. However, poor efficiencies in gene transfer and undesirable safety profiles remain key limitations in advancing this technology. Here, we describe a materials-based approach whereby application of a bioresorbable mineral coating improves microparticle-based transfection of plasmid DNA lipoplexes in several primary human cell types. In the presence of these mineral-coated microparticles (MCMs), we observed up to 4-fold increases in transfection efficiency with simultaneous reductions in cytotoxicity. We identified mechanisms by which MCMs improve transfection, as well as coating compositions that improve transfection in three-dimensional cell constructs. The approach afforded efficient transfection in primary human fibroblasts as well as mesenchymal and embryonic stem cells for both two- and three-dimensional transfection strategies. This MCM-based transfection is an advancement in gene delivery technology, as it represents a non-viral approach that enables highly efficient, localized transfection and allows for transfection of three-dimensional cell constructs.

Vascularendothelial growth factor (VEGF) activity is highly regulatedvia sequestering within the ECM and cell-demanded proteolysis to releasethe sequestered VEGF. Numerous studies have demonstrated that VEGFactivity mediates cellular events leading to angiogenesis and capillaryformation in vivo. This has motivated the study of biomaterials tosustain VEGF release, and in many cases, the materials are inspiredby the structure and function of the native ECM. However, there remainsa need for materials that can bind to VEGF with high specificity,as the in vivo environment is rich in a variety of growth factors(GFs) and GF-binding moieties. Here we describe a strategy to controlVEGF release using hydrogel microspheres with tethered peptides derivedfrom VEGF receptor 2 (VEGFR2). Using biomaterials covalently modifiedwith varying concentrations of two distinct VEGFR2-derived peptideswith varying serum stability, we analyzed both biomaterial and environmentalvariables that influence VEGF release and activity. The presence oftethered VEGF-binding peptides (VBPs) resulted in significantly extendedVEGF release relative to control conditions, and the resulting releasedVEGF significantly increased the expansion of human umbilical veinendothelial cells in culture. VEGF release rates were also stronglyinfluenced by the concentration of serum. The presence of Feline McDonoughSarcoma-like tyrosine kinase 1 (sFlt-1), a serum-borne receptor fragmentderived from VEGF receptor 1, increased VEGF release rates, althoughsFlt-1 was not sufficient to recapitulate the release profile of VEGFin serum. Further, the influence of serum on VEGF release was notdue to protease activity or nonspecific VEGF interactions in the presenceof serum-borne heparin. VEGF release kinetics correlated well witha generalizable mathematical model describing affinity-mediated releaseof VEGF from hydrogel microspheres in defined conditions. Modelingresults suggest a potential mechanism whereby competition betweenVEGF and multiple VEGF-binding serum proteins including sFlt-1, solublekinase insert domain receptor (sKDR), and α2-macroglobulin (α2-M)likely influenced VEGF release from microspheres. The materials andmathematical model described in this approach may be useful in a rangeof applications in which sustained, biologically active GF releaseof a specific GF is desirable. [ABSTRACT FROM AUTHOR]