S-O3-6
Plasma Processing of Artificial Bones and Bone Regenerative Medicine
人工骨プラズマ処理と骨再生医療
Akira Myoui1,2, Satoshi Hamaguchi3 and Hideki Yoshikawa2
名井 陽, 浜口智志，吉川秀樹
1The Medical Center for Translational Research, Osaka University Hospital
2-15 Yamadaoka, Suita, Osaka 565-0871, Japan大阪大学医学部附属病院未来医療センター 〒565-0871 吹田市山田丘2-15
2Department of Orthopaedics, Graduate School Medicine, Osaka University
2-2 Yamadaoka, Suita, Osaka 565-0871, Japan
大阪大学大学院医学系研究科器官制御外科学 〒565-0871 吹田市山田丘2-2
3
Center for Atomic and Molecular Technologies, Graduate School of Engineering, Osaka University
2-1 Yamadaoka, Suita, Osaka 565-0871, Japan
大阪大学大学院工学研究科原子分子イオン制御理工学センター 〒565-0871 吹田市山田丘2-1

In orthopedic surgeries, autologous bone grafting has been the golden standard for the treatment of bone
defects. However, recent advances in synthetic calcium phosphate ceramics have altered the picture andceramic bone substitutes have become as a standard along with autografts. Interconnected porous ceramics
is also useful as a scaffold for bone tissue engineering. However, surface property of hydroxyapatite
ceramics is hydrophilic. Recently, we found that plasma treatment was able to improve the hydrophilicity.
Our preliminary data suggest that plasma treatment may facilitate bone regeneration in the pores andimprove the integration of ceramics to host bone.

1. Bone Defect Treatment and Ceramics Bone
Substitutes
In orthopedic surgeries, autologous bone grafting
has been the golden standard for the treatment ofbone defects. However, it has several disadvantages,
such as the high incidence of complications related
to the harvesting procedures. To avoid thesedisadvantages, various bone substitutes have beendeveloped in Japan, and among them, porous
ceramics made of calcium phosphate,hydroxyapatite or beta tri-calcium phosphate havebeen considered to be the most promisingbiomaterials.
Together with Covalent Material Co., andNational Institute for Materials Science, wedeveloped a fully interconnected porous HA
ceramics, IP-CHA or “NEOBONE”[1]. This
material has an excellent macro-porous structurewith highly interconnected spherical pores(porosity: 75%) and appropriate compression
strength (about 10 MPa) for surgical handling.
Biocompatibility of IP-CHA as a bone substituteevaluated in a rabbit model of bone defect repair
was much better than early porous ceramics
products with less porosity and lessinterconnections suggesting that the interconnected
porous structure facilitated the bone tissue ingrowthin the material and improved the integration to hostbone. Until now, several products of interconnected

porous ceramics including IP-CHA are clinicallyavailable as bone substitutes and they are found to
be clinically very useful[2].
2. Bone Tissue Engineering Using Porous
Ceramics as a Scaffold
Using these bone substitutes as a scaffold, we
have been working on bone regeneration study. The
main principle of current tissue engineering strategy
for bone is a combination of biocompatible porous
scaffold and bone-inducing molecules such as bonemorphogenetic protein (BMP) or bone producing
cells such as osteoblasts. First, we combined
osteoblastic cells differentiated from bone marrowmesenchymal cells with IP-CHA discs as scaffold
and found vigorous bone forming activity in rat
subcutaneous ectopic bone formation model andrabbit forearm bone defect model[3]. We also tested
BMP/IP-CHA composite for bone formation inseveral in vivo models including canine and theresults were satisfactory. Furthermore, we recentlyinvestigated the safety and usefulness ofosteoblastic cell/IP-CHA composite as a clinicalresearch for the treatment of bone defects after theremoval of bone tumors and found good results.
3. Plasma Surface Treatment Improved
Hydrophilicity of HA
As discussed previously, interconnected porousHA ceramics are useful material as a bone
substitute and also as a scaffold material for bone
tissue engineering. However, the surface propertiesof HA ceramics is basically hydrophobic and may
not be an ideal surface for cell/tissue attachment orfor supporting bone regeneration as a scaffold. Ourrecent study revealed that treatment of HA byplasmas significantly improved its hydrophilicity[4].
IP-CHA discs (φ5mm x h2mm) were placed in thedischarge chamber and plasma treatment at a
sub-atmospheric pressure was applied. As shown in
Fig. 1, after plasma treatment, water infiltrated
easily into the pores, indicating plasma treatment
improved the hydrophilicity of inner pore surface.

Fig. 2. Rat calvaria defect model. Full thickness
calvaria bone defects were created by trephine andIP-CHA discs were placed in the defect. Micro-CT
analyses were performed at 4 and 8 weeks after
implantation. Plasma treatment significantly
improved the bone volume in the IP-CHA pores at4weeks.

Although our data are preliminary, they suggestthat plasma treatment improves the hydrophilicityof porous HA ceramics and in consequence, mayenhance the bone regeneration in the pores andIP-CHA integration to host bone. Surface property
alteration may also affect the performance of HA
ceramics as a scaffold for bone tissue engineering
by various ways, for instance by changing theattachment of bone forming cells or by modifying
the cytokine release kinetics in cytokine/IP-CHA
composite system.

Fig. 1. Gross appearance (top) and micro-CT
images (bottom) of IP-CHA discs treated withplasma for 3 minutes and 10 minutes, respectively.
A drop of water containing an X-ray contrast agent
was placed on the top surface of each disc andsubjected to micro-CT analysis. The white color
indicates the location of hydroxyapatite (HA),
gray the location of water (i.e., water filled pores),and dark the location of empty space (unfilledpores). It is seen that, with plasma treatment, mostpores are filled with added water, indicating theincrease of hydrophilicity of inner pore surfaces.

4. Plasma Surface Treatment Enhanced theIntegration of HA Bone Substitute in vivo
We have also examined the effect of plasma
surface treatment on the biocompatibility ofIP-CHA in two different in vivo bone defect models,
rabbit femoral condyle intramedullary bone defectmodel and rat calvaria full thickness bone defectmodel. In both in vivo models, we found a tendencyof better biocompatibility in plasma-treated group,with a significant difference in newly formed bonevolume in the pores analyzed by micro-CT at 4
weeks after implantation (Fig. 2).

References[1] N. Tamai, A. Myoui, T. Tomita, T. Nakase, J. Tanaka,
T. Ochi and H. Yoshikawa, J Biomed Mater Res 59(2002) 110.
[2] N. Tamai, A. Myoui, I. Kudawara, T. Ueda and H.
Yoshikawa. J Orthop Sci. 15 (2010) 560.
[3] M. Nishikawa, H. Ohgushi, N. Tamai, K. Osuga, M.
Uemura, H. Yoshikawa and A. Myoui. Cell Transplant.
14(2005) 829.
[4] D.-S. Lee, Y. Moriguchi, K. Okada, A. Myoui, H.
Yoshikawa, and S. Hamaguchi, IEEE Trans. PlasmaSci.(2011) in press.