Barbieri Thesis - BioMedical Materials program (BMM)

Chapter 3 – Instructive composites: effect of filler content on osteoinduction
years into non–toxic components that can be cleared from the body. Polylactide is
widely used in the medical field (e.g. screws, sutures) and as scaffold for tissue
engineering and drug delivery applications. It has adequate mechanical properties for
certain applications (e.g. screws and plates), but it is too elastic for load–bearing bone
replacement purposes. [305] Thus, introducing calcium phosphates into polylactide was
an attempt to have osteoconductive materials with suitable mechanical properties. It
has been shown that adding nano–hydroxyapatite particulate or fibres into polylactide
materials improves the mechanical properties of the material, [239, 240] while composites
containing more than 40%wt. hydroxyapatite renders the material osteoconductive. [241]
Further, the presence of hydroxyapatite particles increased protein adsorption and
enhanced osteoblast adhesion. [241–243] So far, only one composite of polyester and
ceramic has been reported to be osteoinductive [235] and, to our best knowledge, no
literature about the effects of ceramic filler content on composite–related
osteoinduction is available. In the current study we present an approach to produce
instructive composites by introducing different amounts of nano–sized apatite into
poly(D,L–lactic acid). We evaluated the materials both in vitro and in vivo regarding
their chemistry, ion release rate, surface mineralisation and osteoinductivity and
hypothesize that adding increasing amounts of nano–apatite to a polymer will
generate a specific surface micro–structure that will result in an osteoinductive
composite. More apatite particles exposed at the surface will allow higher ion release,
protein adsorption and surface mineralization that should further enhance the bone
regenerative properties. [86, 228, 237, 263]
3.2. Materials and methods
3.2.1. Nano–apatite synthesis and physico–chemical characterization
Nano–apatite was prepared using a wet–precipitation reaction [306] where (NH4)2HPO4
(Fluka, Steinheim, Germany) aqueous solution (c=63.1 g L –1 ) was added to
Ca(NO3)2·4H2O (Fluka) aqueous solution (c=117.5 g L –1 ) with controlled speed (12.5
mL min –1 ) at 80±5ºC. The reaction pH was kept above 10 by using ammonia (Fluka)
as buffer. After precipitation, the resulting apatite powder was aged overnight,
washed with distilled water to fully remove ammonia and finally suspended in acetone
(Fluka) at a concentration of 0.1 g mL –1 . A small amount of the powder was then
sintered at 1100°C for 200 min (Nabertherm C19, Nabertherm, Lilienthal, Germany)
for chemical characterization. Both the unsintered and sintered apatite powders were
analysed by Fourier transform infrared spectrometer (FTIR, Perkin Elmer Spectrum
1000, Perkin Elmer, Waltham, MA, USA) and X–ray diffractometer (XRD, Rigaku
MiniFlex I, Rigaku, Tokyo, Japan). FTIR was run according a typical KBr pellet
protocol and spectra were collected in the range 400–4000 cm –1 and analysed with
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