Annually, 3 million musculoskeletal and orthopedic procedures are performed in US, including those for fractures (15%), joint problems (22%), and spinal disorders (12%). The musculoskeletal diseases, disabilities and trauma necessitating these procedures cost approximately $215 billion in health care costs and loss of economic productivity. The field of bone tissue engineering has been developed in response to limitations in contemporary therapeutic strategies for these musculoskeletal and orthopedic defects.

A biomimetic approach involving the self-assembly of mineral within the pores of 3-dimensional porous polymer scaffolds is a promising strategy to integrate biological advantages (e.g. bioactivity and osteoconductivity) of an inorganic phase with desirable material functions (e.g. biodegradability) of an organic phase into a single material for mineralized tissue engineering. As cells attach and grow on the surface of this hybrid material, biological functions of the cells could be regulated by the mineral surface, which may also undergo partial dissolution affecting cell function as well. Therefore, we hypothesized that proliferation and differentiation of multipotent mesenchymal stem cells are regulated by a tandem of solution and surface-mediated signals from the biomimetically synthesized apatite materials.

To test the hypothesis, we first demonstrated that bone-like carbonated apatites were self-assembled within the pores of 3-D porous PLGA scaffolds via a biomimetic process using a simulated body fluid (SBF). By adjusting the ionic activity product (IP) of the SBF, carbonate content (7.23% to 5.42%), Ca/P molar ratio (1.63 ± 0.005 to 1.51 ± 0.002) and crystallinity (FWHM at (112):​ 0.147 to 1.035) were controlled in a predictable manner. The crystallinity of apatite is one of main factors determining its resorbability, and the variances in chemical composition of the apatites, along with their dissolution products, could differentially influence cell function. Therefore, we also tested the dissolution behavior of the apatites and both their solution and surface-mediated effects on cell function.

The dissolution behavior of the apatites was characterized quantitatively by measuring the chemical composition of the dissolution products and qualitatively by changes in the structure and morphology of the apatite. Lower crystalline carbonated apatites with low Ca/P ratios were more resorbable and underwent bulky erosion on the mineral surface in both PBS and serum-supplemented MEMa media, whereas higher crystalline carbonated apatites with high Ca/P ratios were less resorbable and underwent surface erosion. When immersed in PBS, only dissolution occurred and crystallinity of the apatites increased over time. Both adsorption and dissolution of Ca and P were observed in serum-supplemented MEMa and crystallinity of the apatites maintained over time.

Observing that the mineralized scaffolds significantly adsorbed Ca and P from serum-supplemented MEMa media, we next tested effects of extracellular soluble Ca and P on cell functions, and concluded that proliferation and osteogenic differentiation of mouse BMSCs were inhibited by Ca,P-deficiency. This finding was generalized, when soluble Ca,P-deficiency also inhibited functions of cells seeded on surfaces of the carbonated apatites and PLGA. However, under conditions of normal soluble Ca and P in the media, mineralized surfaces exhibited significantly enhanced osteogenic differentiation and cell-mediated mineralization relative to non-mineralized surfaces. Among groups of the biomimetic apatites, the more-resorbable carbonated apatite, whose Ca/P ratio and crystallinity were closer to those of natural bone mineral apatite, had a stimulatory effect on osteogenic differentiation compared to the less-resorbable carbonated apatite, whose Ca/P ratio and crystallinity were close to those of hydroxyapatite.

From the experiments of this thesis, we conclude that mouse BMSCs proliferate and differentiate into osteogenic phenotypes in response to combined stimuli from two extracellular environments;​ solution-mediated effects and surface-mediated effects of calcium phosphate biomaterials. By uncoupling mechanisms of soluble and surface-mediated signals from carbonated apatite self-assembled on a polymer, it was elucidated that, regardless of the substrate that the cells are attached to and grow on, appropriate levels of extracellular soluble Ca and P are essential for proliferation and osteogenic differentiation of the mouse BMSCs. Polymer scaffolds coated with bone-like carbonated apatites can stimulate osteogenic differentiation to a greater extent than noncoated scaffolds when normal levels of extracellular soluble Ca and P are maintained. The work of this thesis also demonstrates that signals from both the solution and the surface of biomaterials should be taken into account when trying to optimize biological performance of a material. This combination of design criteria may also advance for the development of new materials for bone tissue engineering.