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

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.Ph.D.Chemical EngineeringUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/78906/1/mcuddihy_1.pd

Poly(lactic-co-glycolic acid) Bone Scaffolds with Inverted Colloidal Crystal Geometry

Abstract Controllability of scaffold architecture is essential to meet specific criteria for bone tissue engineering implants, including adequate porosity, interconnectivity, and mechanical properties to promote bone growth. Many current scaffold manufacturing techniques induce random porosity in bulk materials, requiring high porosities (>95%) to guarantee complete interconnectivity, but the high porosity sacrifices mechanical properties. Additionally, the stochastic arrangement of pores causes scaffold-to-scaffold variation. Here, we introduce a biodegradable poly(lactic-co-glycolic acid) (PLGA) scaffold with an inverted colloidal crystal (ICC) structure that provides a highly ordered arrangement of identical spherical cavities. Colloidal crystals (CCs) were constructed with soda lime beads of 100-, 200-, and 330-μm diameters. After the CCs were annealed, they were infiltrated with 85:15 PLGA. The method of construction and highly ordered structure allowed for ease of control over cavity and interconnecting channel diameters and for full interconnectivity at lower porosities. The scaffolds demonstrated high mechanical properties for PLGA alone (>50 MPa), in vitro biocompatibility, and maintenance of osteoblast phenotype, making them promising for a highly controllable bone tissue engineering scaffold.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/63333/1/ten.tea.2007.0142.pd

Three-Dimensional Cell Culture Matrices: State of the Art

Traditional methods of cell growth and manipulation on 2-dimensional (2D) surfaces have been shown to be insufficient for new challenges of cell biology and biochemistry, as well as in pharmaceutical assays. Advances in materials chemistry, materials fabrication and processing technologies, and developmental biology have led to the design of 3D cell culture matrices that better represent the geometry, chemistry, and signaling environment of natural extracellular matrix. In this review, we present the status of state-of-the-art 3D cell-growth techniques and scaffolds and analyze them from the perspective of materials properties, manufacturing, and functionality. Particular emphasis was placed on tissue engineering and in vitro modeling of human organs, where we see exceptionally strong potential for 3D scaffolds and cell-growth methods. We also outline key challenges in this field and most likely directions for future development of 3D cell culture over the period of 5–10 years.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/63369/1/teb.2007.0150.pd