The mechanism of osteoinduction by nacre: Effects of soluble proteins and insoluble matrix

Nacre, the inner layer of mollucan shell, has attracted increasing attention over recent years as a potential material for bone and/or tooth implants. As shown in previous studies, nacre has osteoinductive properties causing it induce bone formation. But how and why nacre can induce bone formation remains unclear. The objective of this work is to determine the mechanism of nacre's osteoinudctive properties. Two alternative mechanisms have already been proposed to explain the osteogenic properties of nacre: (1) either nacre binds to bone via a hydroxyapatite layer or (2) the presence of soluble nacre proteins stimulates osteoblast differentiation.; In order to test those postulated mechanisms, nacre was first separated from abalone shell and soaked in a phosphate buffer solution to determine if nacre has the ability to surface-transform to hydroxyapatite (HAP). HAP formation is desirable for a bone/tooth implant because it can form strong bonds with natural bone. The mechanism of this surface transformation can be explained as a dissolution-precipitation mechanism.; Second, soluble nacre proteins were isolated and separated from nacre by acetic acid treatment. Those proteins were analyzed by amino acid analysis and matrix-assisted laser desorption/ionization mass spectrometry. The effects of the resulting proteins on osteoblast differentiation were also studied. Soluble nacre protein increased alkaline phosphatase (ALP) activity and osteopontin (OPN) expression in cultures of osteoblasts. Since both ALP and OPN are markers of osteoblast differentiation, our results support the hypothesis that nacre protein promotes the differentiation of osteoblast in vitro.; A material model that mimics nacre was also developed via a stepwise formation approach. The thickness of organic layer was about 10 nm and that of inorganic layer was on the scale of microns. Secondary ion mass spectrometry data were analyzed to reveal each layer of this multilayered material. The osteoblasts' response to this material model was also evaluated by cell adhesion and proliferation. It showed that the number of cells on this material model was comparable to nacre after 14-day culture.