1. IN VITRO ANALYSIS OF SURFACE MODIFIED STAIN-
ETCHED POROUS SILICON MICROPARTICLES
Nancy Wareing2, Yuan Tian1, Roberto Rodriguez1, Armando Loni3, Leigh T. Canham3, Giridhar Akkaraju2, and Jeffery L. Coffer1
1Department of Chemistry, Texas Christian University, Fort Worth, TX, 76129, USA;
2Department of Biology, Texas Christian University, Fort Worth, TX 76129, USA
3pSiMedica Ltd., Malvern Hills Science Park, Geraldine Road, Malvern, Worcestershire WR14 3SZ, UK
I. INTRODUCTION
Biomaterial applications of nanostructured porous silicon (pSi) exploit its ability to be resorbed in vitro / in vivo in a non-toxic manner, its diverse range of surface functionalities, and its high surface area. Due to the relative ease of processing, pSi derived from the stain-etch method is an appealing candidate for use in such applications. Here we show the biocompatibility and high-affinity membrane interaction of surface oxidized, metal-assisted stain-etched mesoporous silicon (MASE pSi) microparticles with human embryonic kidney (HEK293) cells, and nucleic acid electrostatic coupling capabilities following functionalization, suggesting the possibility of using such material for targeted transfection and drug delivery.
II. RESULTS
MASE pSi microparticles exhibit minimal cytotoxic effects on HEK293 cells at concentrations up to 5.0 mg/mL (Fig. 1).
Surface functionalization of MASE pSi microparticles can be demonstrated with APTES and FITC (Scheme 1).
Ultra-sonication of MASE pSi microparticles results in substantial reduction in mean diameter (Fig. 2).
HEK293 cells form high-affinity membrane associations with APTES-FITC MASE pSi microparticles following ultra-sonication (Fig. 3).
DNA-binding capability of APTES-MASE pSi microparticles was found to be dependent on silicon:DNA mass ratios (Fig. 4)
III. CONCLUSION
We demonstrate the ability of non-toxic, surface-functionalized MASE pSi microparticles to adsorb onto the membrane of HEK293 cells with high-affinity and bind pDNA. These findings suggest the possibility of employing MASE pSi microparticles for targeted drug delivery.
Fig. 2 | Size distribution of unsonicated and ultra-sonicated [3 min, 4-5W] APTES-MASE was analyzed by TEM. Diameter was taken from long axis of microparticles. Mean diameter of unsonicated microparticles [828 nm] was nearly double that of ultra-sonicated particles [422 nm]. a b
Fig. 3 | (a) Two dimensional confocal image of β-actin immunofluorescence stained HEK293 cells with FITC-conjugated MASE pSi microparticles (b) 3-D confocal Z-stack rendering shows adsorption of FITC-labeled microparticles onto the membrane of a β-actin-labeled HEK293 cell.
Fig. 1 | Cytotoxicity of MASE pSi microparticles [356m2/g BET surface area, 0.292ml/g pore volume, 3.5nm-4.7nm average pore diameter] on HEK293 at various concentrations [0.05, 0.5, and 5.0 mg/mL] was analyzed by Trypan blue exclusion assay.
Fig. 4 | APTES-MASE pSi microparticles were electrostatically coupled to the CMV-LacZ plasmid at varying silica:DNA mass ratios. The surfactant Triton X-100 was added to indicated samples. DNA-binding efficiency was analyzed via gel electrophoresis.
4 hrs.
then
wash
Visualization
DNA coupling
CMV-LacZ plasmid
Scheme 1 | Following ultrasonication, APTES-FITC surface functionalized microparticles were added to HEK293 cells following indirect immunofluorescence staining of cytoskeletal components.
We thank the TCU IS initiative (GA, JLC), the TCU Department of Biology (NW, GA), the Robert A. Welch Foundation (JLC), and the NIH (JLC) for their financial support of this research.