Inkjet Printing of Nanohydroxyapatite Gradients on Fibrous Scaffold for Bone–Ligament Enthesis

Abstract

The mineralized gradient found over an ACL–subchondral bone interface was accomplished by inkjet-printing, by creating a pattern of nanoparticulate hydroxyapatite (HANP) on the surface of a highly-aligned and chemically modified poly(lactic acid) (PLA) nanofiber scaffold. The printed HANP remained relatively stable on the surface of the PLA, with the gradient pattern clearly visible following a 72-h aqueous incubation, as confirmed by fluorescence microscopy. High-magnification SEM images displayed HANP distribution on and between fibers. Following immobilization of fibrin on the surface of the samples, the proliferation and differentiation response of human mesenchymal stem cells in vitro was quantitatively assessed. The onset of osteogenic differentiation appeared to have been greatly enhanced by the combined addition of fibrin and HANP, as indicated by enhanced ALP activity. Furthermore, the inclusion of fibrin enhanced osteogenic differentiation to a greater degree than the inclusion of HANP. The cell morphology appeared more cuboidal in shape over the high-HANP density regions compared to a more spindle-like shape in the low-HANP density regions, depending on HANP presence/density over the gradient region (1250 µm). This study illustrates the efficacy of producing an inkjet-printed pattern of HANP biomolecules capable of inducing a cellular response for ligament-to-bone enthesis.