Growth of DLD-1 Colon Cancer Cells on Variotis™ Scaffolds of Controlled Porosity: A Preliminary Study

Abstract

Tissue engineering will play an increasingly vital role in cancer research. Provision of biomimetic microenvironment systems for in vitro cancer models can be addressed in part by utilizing thick 3D scaffolds with high interconnective porosity . This approach gives rise to new analytical challenges and opportunities. In this preliminary study, Variotis™ synthetic scaffolds of high interconnected porosity and hierarchical structure were used. An effective macroscopic porosity of 94.3 ±1.74 vol% was attained by using microCT and finite element methods. The actual porosity was determined to be 94.6±0.29 vol%. Scaffolds were compressed in a customized jig to thicknesses of 99.5 mm, 74.6 mm, 46.3 mm (±0.5% tolerance) and then annealed to set respective porosities of 94.3 vol%, 93.2 vol%, 89.5 vol% (±1.5% tolerance). Scaffolds were then sectioned to 2mm thickness. DLD-1 colon cancer cells were grown on 3D scaffolds of three specified porosities for varying periods of time then imaged using confocal and scanning electron microscopy methods. Hoechst staining resulted with minimal scaffold autofluoresence while autofluoresence exceeded useful limits when used in conjunction with Alexa488-phalloidin under argon laser excitation in confocal microscopy. Using Hoechst staining, DLD-1 cells (nuclei) were observed to readily attach and proliferate on Variotis™ scaffolds. Normal DLD-1 cell morphologies were evident using scanning electron microscopy. The high interconnected porosity of the scaffolds allowed cells to be observed deep within scaffolds. Scaffolds remained structurally stable and unified throughout all culture experiments and provided ease of handling during cell culture and microscopy.