Barbieri Thesis - BioMedical Materials program (BMM)

Chapter 3 – Instructive composites: effect of filler content on osteoinduction
mineralization generates a suitable environment for inducible cells to form bone [217] and
allows osteoinduction occur. [248, 249]
Besides in vivo surface mineralization, protein adsorption may also play a role in
triggering ectopic bone formation. It has been reported that a bioinert Al2O3 ceramic
adsorbs proteins and induces bone formation in muscle of dogs while it was not shown
to surface calcify in vitro or in vivo. [251] It has also been shown that calcium phosphate
materials have high affinity for proteins (e.g. bone morphogenetic proteins, BMPs) [228, 249,
326] and that composites with high content of HA adsorb more proteins. [242] Based on
these assumptions, it may well be that 40% CaP composites could have adsorbed
higher amount of proteins (including BMPs) from the surrounding body fluids. Such
proteins may have then contributed to instruct (stem) cells to osteogenically differentiate
and trigger inductive bone formation. Conversely, composites with lower apatite content
would not adsorb enough proteins to start osteoinduction. However, the proteins that are
adsorbed and play a potential role in 40% CaP osteoinduction need to be evaluated in
further studies.
Another aspect regarding inductive bone formation in 40% CaP may be ion release
(especially calcium) from the composites. Calcium is believed to be an extra–cellular
signaling molecule in bone [321–323] and it has been shown that ions released from calcium
phosphate materials do not only enhance the bioactivity of materials [227, 228, 324] but also
the proliferation, differentiation and mineralization of osteogenic cells. [321–323] We
observed higher amounts of calcium ions released in vitro from 40% CaP composites
compared to the others and, as shown in this study, only 40% CaP gave rise to inductive
bone formation, indicating a possible role of the amount of calcium ion released in
osteoinduction. The higher release of calcium ions from 40% CaP, as compared to the
other three considered materials, might be due to the fact that it exposes more apatite
on its surface. Moreover, at the contact with surrounding fluids, the exposed apatite may
have dissolved releasing calcium. Besides this, hydrolysis of the polymer matrix could
have occurred further exposing apatite from its bulk and enhancing ion release. Thus the
large amount of calcium released may have triggered 40% CaP composite–related
osteoinduction. [325] However, the importance that calcium ions may have in triggering
heterotopic bone formation still needs to be understood and demonstrated.
Finally, the surface micro–structural changes due to the increased apatite content in the
four composites could have played a role as well. Several studies have reported that
controlled micro– or nano–structures do not only have positive effects on the
differentiation of (mesenchymal) stem cells into osteogenic cells, [195] but also influence
protein adsorption and ion release. [217, 228, 326] As shown in Figure 4, the 40% CaP has a
rougher surface micro–structure as compared to those composites with lower apatite
contents. Previous studies have reported that specific surface micro–structure on a
chemically unaltered ceramic can render osteoinductive ceramics. [217, 228, 237] Further, we
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