Utilizing Inverted Colloidal Crystal Scaffolds to Engineer In Vitro Bone and Bone Marrow.

Abstract

Numerous studies have shown that cells and tissues grown in 2D substrates behave dramatically differently than in the body, and that culturing them in three dimensional (3D) scaffolds can restore some of this lost functionality. 3D scaffolds can provide structural support to an injury site in the body, aiding the growth of healthy tissue. Outside of the body, scaffolds can be used to test pharmaceuticals, collecting data in an environment that represents the body better than traditional 2D cultures. This could potentially save millions of dollars in drug development costs.

For these reasons, this dissertation focuses on the utilization of inverted colloidal crystal (ICC) scaffolds for bone and bone marrow engineering. ICC scaffolds are matrices that have a highly ordered 3D structure of interconnected spherical cavities. This dissertation describes the first ICC scaffold composed of a biodegradable polymer, poly(lactic-co-glycolic acid) (PLGA). Within these scaffolds, the scaffold cavity sizes were controlled on the micro-scale by utilizing different beads sizes, 100, 200, and 330 μm, as the scaffold template. Additionally, the size of the channels that connect the cavities were controlled within the range of 660-710 ◦C by changing the annealing temperature. The compressive moduli of these scaffolds were in the range of 55-63 MPa. Lastly, biocompatibility with a human osteoblast cell line was demonstrated.

Next, the dissertation utilized polyacrylamide hydrogel ICC scaffolds to engineer a human hematopoietic stem cell (HSC) niche. ICC scaffolds demonstrated significantly greater HSC expansion than 2D cultures. This work was continued by comparing the ICC scaffolds to Matrigel and 2D cultures. Here, it was observed that ICC cultures demonstrated stable numbers of HSCs throughout 14 days. 2D cultures expanded the number of HSCs 6-8× over 14 days and Matrigel cultures expanded differentiated cells but few HSCs. These results indicated that physical cell-cell interactions cause quiescence or preservation of the HSC phenotype, and the absence of direct cell-cell interactions causes HSC differentiation. Lastly, preliminary data was collected in utilizing ICC scaffolds as a tool for drug testing.