Page 1 - Dr. Louise Connell Chitosan-Silica-Bone-Regeneration-Poster
P. 1
Chitosan - Silica
Hybrid Scaffolds for Bone Regeneration
Louise S. Connell, Esther M. Valliant, Julian R. Jones Freeze-dried Scaffolds
Department of Materials Science, Faculty of Engineering, Imperial College London
l.connell10@imperial.ac.uk RGANIC CONTENTS GREATER THAN 40 wt% and polymer concentrations less than
Marta Suarez O17 mg/mL were required to produce mechanically stable scaffolds that did not disintegrate
Fundación ITMA, Parque Tecnológico de Asturias, Spain. to a powder on handling (Fig. 4). µ
Department of Nanostructured Materials, Universidad de Oveiedo, Spain 40 wt% organic 50 wt% organic 65 wt% organic
Introduction 37.5 mm 36.8 mm 36.6 mm
B IOACTIVE GLASSES FULFILL most of the requirements for use as bone regeneration
scaffolds but they are too brittle to be used in load sharing applications. A hybrid is a
material containing organic and inorganic components that interact at a molecular level so that Figure 4: Increased organic content increases the mechanical
the two components cannot be distinguished above the nanoscale (Fig. 1). Type II hybrids stability of the scaffold.
contain covalent bonding between the components, providing a strong interaction. This combines 65 wt% organic 50 wt% organic Figure 6: Mercury intrusion porosimtery (MIP) showing the effect of
the mechanical strength of glass with the toughness of polymers to produce improved scaffolds freezing temperature on the modal interconnect diameter.
1
for bone regeneration. These hybrids can be fabricated by introducing natural polymers, such
as gelatin and chitosan, and a covalent coupling molecule into the sol-gel process. a) 0.25
o
-20 C 0.2
Previously, chitosan-silica hybrid scaffolds 0.15
have been fabricated using a freeze-drying Yield Stress /MPa 0.1
2
method and gelatin-silica hybrid scaffolds Organic polymer - Chitosan 0.05
have been produced using a sol-gel foaming Inorganic network - Silica
0
3
‐80
method. Until now, chitosan has not been b) 3 ‐20 Freezing temperature /°C ‐196
incorporated into the foaming process and /MPa 2.5
the two methods have not been compared. o 2
-80 C modulus 1.5
The authors present chitosan-silica hybrid Young’s 1
scaffolds formed by combining the sol-gel 0.5
process with both freeze-drying and foaming Coupling molecule - GPTMS 0 ‐20 ‐80 ‐196
steps. Optimisation of the foaming process c) 20 Freezing temperature /°C
18
is discussed and the morphological and 16
14
mechanical properties of the resulting Yield Strain /% 12
10
o
materials are compared as potential bone -196 C 8
6
regeneration scaffolds. Figure 1: Representation of a chitosan-silica hybrid showing the 4
2
covalent coupling between the different components. 0
‐20 ‐80 ‐196
Freezing temperature /°C
Chitosan Requirements for tissue engineering scaffolds
Figure 5: SEM images showing the reduction in pore size as the Figure 7: The effect of freezing temperature on:
• Deacetylated derivative of chitin. • Biocompatible and bioactive. freezing temperature reduces. The pores, formed by ice a) compressive strength
• Sourced from crustacean shells • Pores with interconnect diameters of 100 µm for cell crystals, are angular and elongated. Microcracking can b) Young's modulus
o
o
and the cell walls of fungi. penetration and vascularisation. be observed in the -20 C and -80 C scaffolds whereas c) yield strain of 65 wt% organic scaffolds.
o
it is not observed in the -196 C scaffolds.
• Biocompatible and degradable • Resorbable with controlled degradation rate.
by lysozyme. • Mechanical properties close to cancellous bone Freeze-dried scaffolds show elongated and angular pores with interconnect diameters that
• Functional groups that can react. (compressive strength 2-12 MPa). reduce in size as the freezing temperature reduces (Fig. 5).
• Soluble in dilute acid. • Easy fabrication process.
Mechanical testing results show that the compressive strengths of the freeze-dried scaffolds
Figure 2: Chitosan monomer unit. are around 20 times lower than that of cancellous bone (Fig. 7a). Despite this, the yield strain
is around 10 % (Fig. 7c), confirming that the incorporation of polymers has reduced the brittle
nature of the silica sol-gel materials making them more suitable for load sharing applications.
Scaffold Fabrication
Mercury intrusion porosimetry (MIP) data (Fig. 6) shows that the modal interconnects of
o
o
o
µ
HITOSAN WAS FUNCTIONALISED with 3-glycidoxypropyl trimethoxysilane (GPTMS) in scaffolds frozen at -196 C are ~9 m and in the -20 C and -80 C scaffolds diameters are
o
Cdilute HCl (pH 5) for 24 h. In a separate container, tetraethyl orthosilicate (TEOS) was ~100-150 µm. The -196 C scaffolds show an improvement in compressive strength over
o
o
hydrolysed in a water: HCl vol ratio of 3:1 and a TEOS: water mol ratio of 1:4. those frozen at -20 C and -80 C, which is attributed to the reduction in microcracks that form
during processing visibility under SEM (Fig. 5) and by the reduction in noise observed in the
The hydrolysed TEOS was mixed with the polymer solution to produce a sol.
stress-strain curves. However, the modal interconnect diameter is far too small for the material
to be used as a tissue engineering scaffold.
a) b)
Foamed Scaffolds
TEOS Chitosan
H O/HCl GPTMS
2
CAFFOLDS WITH THE BEST pore morphology and interconnect diameters of 100 µm
pH 2 pH 5
Swere fabricated by combining an organic content of 30 wt% and a polymer concentration
o
of 50 mg/mL. SEM images (Fig. 9a-c) confirm that at a freezing temperature of -80 C the
dominant pore forming process is the foaming method.
65 wt% organic 50 wt% organic 40 wt% organic 30 wt% organic In contrast to the freeze-dried scaffolds,
an organic content of less than 40 wt% is
required to produce stable scaffolds.
3 days Age
Sol
At organic contents higher than this, the
concentration of silica precursors in the sol
Freeze-dried sample is too low to form a spanning network that
o
-20/-80/-196 C Figure 8: Increasing silica content leads
to stable foam formation. gels. The insert in (Fig. 8) shows the isolated
Polymer
At low silica contents, milky silica microspheres formed at low silica
Gelling agent Surfactant
Silica microspheres suspensions of silica precursor concentrations.
(2-5 µm diameter) microspheres are formed.
3 days Age
Increasing the polymer concentration, and
a) b)
hence increasing the concentration of silica
Foamed sample
o
-80 C
precursors, improves the pore morphology.
It is hypothesised that the higher silica
Figure 3: a) Fabrication process to produce freeze-dried and foamed scaffolds. content will provide improved compressive
b) Hypothesised reaction between chitosan and GPTMS during functionalisation.
strength over that of the freeze-dried scaffolds
bringing it closer to cancellous bone.
Freeze-dried scaffolds: c) d)
The sol was poured into moulds where it gelled to form monolith samples. These were aged Figure 9: SEM images showing improving pore morphology
o
o
o
o
for 3 days in sealed moulds at 40 C and frozen at -20 C, -80 C or -196 C. of 30 wt% organic scaffolds as the polymer
concentration increases from:
The samples were then freeze-dried for 5 days to produce scaffolds. a) 17 mg/mL to b) 30 mg/mL and c) 50 mg/mL.
d) MIP shows that at all concentrations, the
interconnect diameter is around 100 m.
µ
Foamed scaffolds:
Triton-X 100 surfactant was added to the mixed sol and was vigorously agitated to produce
a foam. The foam was stabilised by adding HF to accelerate gelling. Just prior to the gelling Conclusions
point, the foam was poured into moulds.
ŸChitosan-silica hybrid scaffolds have been fabricated using the sol-gel method and
o
o
The gelled foams were aged for 3 days at 40 C, frozen at -80 C and finally freeze-dried for 5 days. processed into scaffolds using both freeze-drying and foaming methods.
ŸThe freeze-drying process results in angular and elongated pores with interconnect
diameters that vary with freezing temperature while foaming gives spherical pores with
References interconnect diameters of around 100 µm.
1 Valliant, E.M. and J.R. Jones, Soft Matter, 7:5083-5095, 2011. ŸThe compressive strength of the freeze-dried scaffolds is well below that of cancellous
2 Shirosaki Y, et al., Chem Eng J, 137:122-128, 2008 bone but the materials show a reduction in brittle nature.
3 Mahony O, et al., Adv Funct Mater, 20: 3835–3845, 2010
ŸFoamed scaffolds can incorporate a higher inorganic content than freeze-dried scaffolds.
Acknowledgements It is hypothesised that this will lead to increased compressive strength and bring it closer
The authors would like to acknowledge EPSRC for funding especially EP/E057098 to that of cancellous bone.
100
95
75
25
5
0