Project Report

With age, location, injury, and disease progression, hydroxyapatite (HA, Ca10(PO4)6(OH)2), the hard mineral component of bone can have different materials properties. These differences include amount of carbonate substitution, size, crystallinity and more. Bone remodeling cells such as osteoblasts which form new bone may be sensitive to these changes in the bone microenvironment and respond differently. Designing platforms with variation in bone-like mineral properties to control bone-matrix formation on surfaces may provide insight to how osteoblasts are affected by properties of existing bone matrix to form, remodel, and heal bone and provide new materials for bone repair. In order to investigate how differences in hydroxyapatite change the microenvironment surrounding osteoblasts and how this in turn affects osteoblast activity, model surfaces were developed to emulate such environments. Hydroxyapatite nanoparticles were synthesized through a wet precipitation reaction and obtained commercially. These particles were dry annealed for 0, 1, and 3 days at 200 ºC in order to change crystallinity while maintaining the morphology of the particles. The particles were then mixed with polylactide (co-glycolide) (PLG) in tetrahydrofuran (THF) and spin-coated onto glass substrates. Surfaces without particles were used as controls. Surfaces were examined via scanning electron microscopy to determine surface coverage of particles, Fourier Transform Infra-red spectroscopy to determine crystallinity of the HA-surface, and Alizarin Red staining to determine the amount o HA exposed to the surface for cellular interactions. Primary human osteoblasts obtained from the tibial plateau of patients going through knee-replacement surgery. These cells were seeded onto the surfaces and examined for proliferation rate, metabolic activity, protein production, and matrix production. It was determined that osteoblasts seeded on non-HA-containing control surfaces spread further and were larger than osteoblasts seeded onto HA-containing surfaces, however the metabolic activity was higher for HA-containing surfaces with increased activity for commercial HA surfaces observed by day 8 of culture and increase for precipitated HA surfaces by day 14 of culture through an Alamar blue assay. Additionally, osteoblasts seeded onto commercial HA surfaces had a faster proliferation rate than surfaces containing precipitated HA particles. On examination of the protein and matrix production of the osteoblasts via confocal imagining of surfaces stained immunoflurescently for fibronectin, surfaces containing particles with longer heat treatment appeared to have produced greater amounts of fibronectin. Further analysis of matrix content is currently being conducted. This project aims to contribute towards understanding how osteoblasts react to different biologically relevant environments by designing tunable platforms to examine cell-surface interaction as a result of differences in a simulated bone microenvironment. Studying the cell-environment relation may lead better incorporation of materials into hard tissues as well as provide platforms to better understand the mineralization and healing process of bone. Improving knowledge of osteoblastic functions in bone development and healing will contribute to the development and design of improved healthcare solutions to damaged and diseased bone and improve the quality of life for patients suffering from damage or disease affecting the skeleton. This work was done through a collaboration between Cornell University and the Queensland University of Technology with support from the National Science Foundation East Asia and Pacific Summer Institute Program and the Australian Academy of Sciences.

National Science Foundation (NSF)
Office of International and Integrative Activities (IIA)
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Carter Kimsey
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Lin Deng W
United States
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