The Use of 3-D Culture in Peptide Hydrogel for Analysis of Discoidin Domain Receptor 1-Collagen Interaction

Abstract

The aim of this study is to examine a novel drop culture model using a biologically inspired self-assembling peptide: hydrogel (RAD16-I, also called PuraMatrix), which produces a nanoscale environment similar to native extracellular matrix (ECM) for a cell line weakly adherent to a plastic surface during cell culture. Our work investigates quantitatively analyzing discoidin domain receptor (DDR) 1-mediated protein interactions between collagen type I and matrix metalloproteinase (MMP)-2 or -9, as well as cell invasion, using, as a scaffold, PuraMatrix, a novel peptide hydrogel. Results demonstrate that the dynamic cell culture technique produced a highly stable reharvesting of cells throughout the constructs with HP-75, human pituitary adenoma cell line when compared to the traditional seeding methods. Secretion of MMP via collagen type I was observed quantitatively in the supernatant (EC50; MMP-2, 50.4 ng/ml: MMP-9, 57.6 ng/ml). In PuraMatrix gel impregnated with 50 ng/ml of collagen type I, transfection of the vector encoding full-length DDR1 or siRNA targeting DDR1 up- or down-regulated respectively secretion of MMP-2 and -9, and cell invasion. Our results show that incorporation of this peptide with each ECM component provides a more permissive environment to elucidate ECM to cell signal interaction.