Biomimetic Mineralization of Hydrogel Biomaterials for Bone Tissue Engineering - Biomimetics: Advancing Nanobiomaterials and Tissue Engineering - page 58

3.4 Enzymatically-Induced Mineralization Using

Alkaline Phosphatase (ALP)

The enzyme alkaline phosphatase (ALP) supports mineralization of bone
in vivo by cleavage of phosphate from organic phosphate [44]. The enzyme
acts as a catalyst for the hydrolysis of the organic phosphoesters, thereby
increasing the local concentration of inorganic phosphate groups, which
results in deposition of carbonated apatites. The use of ALP to induce homo-
geneous mineralization of hydrogels to increase their mechanical strength
or render them more suitable for bone replacement applications is an alter-
native to incorporation of CaP particles, which tend to aggregate [22, 45].
ALP crosslinked to dentine-derived collagen sheets has induced min-
eralization in vivo [46-48]. Regarding applications as a biomaterial com-
ponent to aid bone regeneration, ALP has been covalently linked to
bioactive glass scaffolds by Verné et al. to increase bioactivity in vitro [49].
Osasthanon et al. covalently grafted ALP to a porous fi brin scaffold to
induce mineral deposition in vitro and promote bone formation in vivo in
a mouse calvarial defect [50]. ALP has also been electosprayed to form a
coating on titanium implants [51, 52].
Mineralization of hydrogel materials has been achieved by incorpora-
tion of ALP followed by soaking in a solution containing calcium ions
and glycerophosphate as a substrate for ALP, which cleaves off phosphate
which is then free to react with calcium to form insoluble CaP within
the gel. ALP-mediated mineralization has induced formation of calcium
phosphate phases including apatite [47, 53, 54].
Studies on ALP incorporation into hydrogels to induce mineralization
can be divided into two categories. In the fi rst category, ALP-induced min-
eralization is an analytical tool to aid fundamental research into bone cell
behavior, principles of biomineralization, and mineralizability of hydrogel
materials. In the second category, ALP-induced mineralization is intended
to improve hydrogels' suitability as scaffold materials for bone regeneration.

3.4.1 ALP-Induced Hydrogel Mineralization for Fundamental

Research

In the fi rst category, Filmon et al. cultivated osteoblasts on PHEMA hydro-
gels containing ALP, as a result of which the hydrogel surface was covered
by CaP calcospherites found in natural bone [55]. Cells anchored to these
calcospherites showed an increased spreading area. The calcospherites
also promoted adsorption of the matrix proteins fi bronectin and bone sia-
loprotein (BSP). In other work by the same group, the ability of different
biphosphonates to inhibit ALP-induced mineralization of such hydrogels
was compared [56, 57].