Functionalization of microparticles with mineral coatings enhances non-viral transfection of primary human cells

Andrew S Khalil,Gianluca Fontana,Todd C Mcdevitt,Hunter J Johnson,Angela W Xie,Jennifer M Umhoefer,Xiaohua Yu,Tracy A Hookway,William L Murphy

doi:10.1038/s41598-017-14153-x

Abstract

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.

Highlights

Advancements in gene delivery technology are of great interest for both clinical and basic biomedical research applications[1,2,3,4]
mineral-coated microparticles (MCMs) reduced cytotoxic effects commonly associated with chemical transfection reagents, and improved transfection efficiency for several primary human cell types including dermal fibroblasts, embryonic stem cells, and mesenchymal stromal cells
Incubation of microparticles in specified mSBF solutions resulted in mineral coatings with distinct nano-structure and stability characteristics

Summary

Introduction

Advancements in gene delivery technology are of great interest for both clinical and basic biomedical research applications[1,2,3,4]. Physical methods include the use of ballistics[8], electric fields[9], osmotic pressure, or physical injection[10] to disrupt the cell membrane and deliver nucleic acids directly to the cytoplasm[5] Some of these physical methods have been refined to achieve high efficiencies relative to viral delivery with low toxicity in vitro, but have limited clinical promise in large animals or humans[5]. Upon incubation of microparticles in a simulated body fluid containing the ion species and concentrations of human blood plasma, modified to contain 2X calcium (mSBF), a mineral coating forms on the microparticle surface via a nucleation and growth mechanism These coatings are biocompatible, bioresorbable, charged, and have a high degree of nanometer-scale porosity, allowing for efficient delivery for a range of different biomolecules[20,22,23,24,25,26] including DNA complexes for chemical transfection. We showed that improved transfection can be achieved with a variety of microparticle core materials, and demonstrated efficient localized transfection via MCMs in both two-dimensional (2-D) and 3-D cell culture formats

Methods

Results

Discussion

Conclusion