Bioceramics. Proceedings of the 4th International Symposium on Ceramics in Medicine London, UK, September 1991

339 Pages·1991·15.097 MB·English

by  W. Bonfield, G. W. Hastings and K.E. Tanner (Eds.)

Bioceramics
Volume 4
Proceedings of the 4th International Symposium on
Ceramics in Medicine
London, UK, September 1991
Editedby
W.Bonfield
G.W.Hastings
K.E.Tanner
U T T E R W O R TH
I N E M A N N
Butterworth-Heinemann Ltd
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First published 1991
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Cover picture
Squared pedestal shape bulb pot, solid blue Jasper with
white bas-relief of 'Sacrifice to Aesculapius' modelled by
William Hackwood. Date circa 1790.
By courtesy of the Trustees of the Wedgwood Museum,
Barlaston, Staffordshire, UK.
Printed in Great Britain at the University Press, Cambridge
Preface
Bioceramics is a major underpinning technology in the development of second generation
implants and prostheses with an extended lifetime in patients. The advantage of traditional
ceramics, such as alumina, which provide a wear-resistant, bioinert surface in replacement
articulating joints, has been well demonstrated. In addition the recent application of novel
bioactive ceramics, glasses and composites, to secure osseointegration, has opened exciting
clinical opportunities for stable implant fixation, with hydroxyapatite surface coatings already
being utilised in total hip arthroplasty.
The leading edge of international research in bioceramics is reported in this volume, which
constitutes the proceedings of the Fourth International Symposium on Ceramics in Medicine
(Bioceramics 4) held in London on September 11-13, 1991. This symposium follows on
from the foundation meeting in Kyoto (1988), with the momentum of an international forum
being maintained in the following meetings in Heidelberg (1989) and Terre Haute (1990). It
is apparent that rapid progress is being made towards bioceramics applications in a range of
orthopaedic, cardiovascular, dental, urological, oncological and opthalmic devices. The
continuing trend to bioactive ceramics, glasses and composites is well demonstrated in the
collected papers, together with an emphasis on the optimisation o fthe implant-tissue interface
as evaluated in either in vitro or in vivo studies. We hope that the Proceedings will prove of
interest to clinicians, scientists, engineers, industrialists and government officials requiring
both a perceptive overview and an in-depth appreciation of the current world-wide activity in
Ceramics in Medicine.
W. Bonfield G.W. Hastings K.E. Tanner
Chairman Vice-Chairman Secretary General
Queen Mary and Westfield College, London, U.K.
Organising Committee
President Dr H. Oonoshi, Osaka, Japan
Chairman Professor W. Bonfield, London, U.K.
Vice Chairman Professor G.W. Hastings, London, U.K.
Secretary General Dr K.E. Tanner, London, U.K.
Member Dr S. Best, Oxford, U.K.
Scientific Committee
Professor M. Anseau, Mons, Belgium
Professor H. Aoki, Tokyo, Japan
Dr P. Christel, Paris, France
Professor D. Dowson, Leeds, U.K.
Professor P. Ducheyne, Philadelphia, PA, U.S.A.
Professor K. de Groot, Leiden, Netherlands
Professor G. Heimke, Clemson, SC, U.S.A.
Professor L.L. Hench, Gainsville, FL, U.S.A.
Professor S.F. Hulbert, Terre Haute, IN, U.S.A.
Professor T. Kokubo, Kyoto, Japan
Professor Sir Ronald Mason, Stoke-on-Trent, U.K.
Professor K. Ono, Osaka, Japan
Professor R. Pilliar, Toronto, Canada
Dr A. Ravaglioli, Faenza, Italy
Professor C. Rey, Toulouse, France
Dr Lek Uttamasil, Bangkok, Thailand
Professor T. Yamamuro, Kyoto, Japan
Organising Committee
President Dr H. Oonoshi, Osaka, Japan
Chairman Professor W. Bonfield, London, U.K.
Vice Chairman Professor G.W. Hastings, London, U.K.
Secretary General Dr K.E. Tanner, London, U.K.
Member Dr S. Best, Oxford, U.K.
Scientific Committee
Professor M. Anseau, Mons, Belgium
Professor H. Aoki, Tokyo, Japan
Dr P. Christel, Paris, France
Professor D. Dowson, Leeds, U.K.
Professor P. Ducheyne, Philadelphia, PA, U.S.A.
Professor K. de Groot, Leiden, Netherlands
Professor G. Heimke, Clemson, SC, U.S.A.
Professor L.L. Hench, Gainsville, FL, U.S.A.
Professor S.F. Hulbert, Terre Haute, IN, U.S.A.
Professor T. Kokubo, Kyoto, Japan
Professor Sir Ronald Mason, Stoke-on-Trent, U.K.
Professor K. Ono, Osaka, Japan
Professor R. Pilliar, Toronto, Canada
Dr A. Ravaglioli, Faenza, Italy
Professor C. Rey, Toulouse, France
Dr Lek Uttamasil, Bangkok, Thailand
Professor T. Yamamuro, Kyoto, Japan
Bioceramics, Volume 4 Edited by W. Bonfield, G. W. Hastings and K. E. Tanner
{Proceedings of the 4th International Symposium on Ceramics in Medicine,
London, UK, September 1991)
© 1991 Butterworth-Heinemann Ltd
Osteogenic Response of Rat Bone Marrow Cells in Porous Alumina,
Hydroxyapatite and Kiel Bone
M. Okumura*, C.A. van Blitterswijk, H.K. Koerten and H. Ohgushi*
Department of Orthopaedics, Nara Medical University, Kashinara, Nara 634, JAPAN and
*Biomaterials Research Group, ENT Department, Building 25, University of Leiden,
Rijnsburgerweg 10, 2333 AA Leiden, THE NETHERLANDS.
ABSTRACT
Porous alumina, hydroxyapatite(HA) and Kiel bone combined with rat marrow cells were
implanted subcutaneously in the back of syngeneic Fischer rats. The implants were
harvested 8 weeks after implantation. Undecalcified sections of the implants were examined
by light microscopy or fluoromicroscopy and the de novo bone/implants interfacial areas
were observed by scanning electron microscopy. All marrow cell loaded alumina and HA
ceramics showed newly bone formation. However, Kiel bone combined with marrow cells did
not show consistent osteogenesis. Also intervening fibrous tissue was observed between Kiel
bone and de novo bone(distance osteogenesis). Fluorochrome study revealed that the
osteogenesis formed in alumina was first seen away from the surface of the ceramics in the
porous regions and proceeded in a centrifugal direction, resulting in a contact with alumina
ceramics(contact osteogenesis). In contrast, the osteogenesis in HA proceeded in a centripetal
direction towards the center of the pores(bonding osteogenesis).
INTRODUCTION
Hydroxyapatite ceramics having bioactive nature have been used widely for bone substitute.
Alumina ceramics are one of the major types of bioinert materials(l), and have been utilized
in artificial prosthesis surgery because of their good biocompatibility and high mechanical
strength(2-9). However, when alumina ceramics are implanted in osseous defects, a layer of
connective tissue is initially observed at the interface between the ceramic and bone tissue.
Therefore, the prosthesis required a mechanical interlocking for stronger fixation of the
alumina/bone interfaced, 10). This connective tissue interposition could be caused by motion
between implanted ceramics and host bone. In other words, precisely fitted implant in bony
defect may not show the initial connective tissue interposition, thus experimental condition
may affect the bone formation process. In the present work, cell mediated bone formation in
porous materials was studied. When porous hydroxyapatite ceramic was used, interposed
connective tissue could not be detected between the de novo bone and hydroxyapatite(l 1-15).
Therefore, this experimental approach is very reliable to analyze the bone dynamics and
bone/biomaterial interface. By using this method, we analyzed the bone formation process and
bone/material interface in porous alumina, HA and Kiel bone.
IMPLANT MATERIALS
Polycrystaline alumina ceramics (Kyocera, Kyoto, Japan), coralline hydroxyapatite ceramics
(Interpore International, California, USA) and Kiel bone (Braun Melsungen, Germany) were
used in this experiment. The alumina has a mean void volume of 70% and fully
interconnected pores, measuring 100-600μπι in diameter. The average pore size of
hydroxyapatite implants is 200μπι in diameter, the void volume averages 66%. Implants with
the standard size of 5 x 5 x 5 mm were used.
4 Bioceramics Volume 4
METHOD
Marrow Cells Preparation and Surgical Procedure
The detailed procedures were previously reported(l 1-15). Briefly, femora and tibiae of
Fischer strain 344 rats (7 week-old males) were removed and placed in saline. The marrow
plug from the diaphyseal portion was then hydrostatically forced into a test tube containing
heparinized phosphate buffered saline(PBS). The marrow in PBS was disaggregated by
sequential passage through needles to obtain a cell suspension. The cell suspension was
centrifuged at 250G for 5 minutes and the cell pellet was resuspended in 200 μΐ of the
supernatant by vortex mixer. To make the composite graft, the porous implants were soaked
in this disaggregated marrow cell suspension.
Syngeneic Fischer rats were anesthetized by intraperitoneal injection of Nembutal.
Subcutaneous pouches were created by blunt dissection in the back of the rat. Implants of
alumina, HA and Kiel bone alone and combined with marrow cells were implanted in these
subcutaneous pouches. The implants were harvested at 8 weeks after implantation. During the
postoperative time, multiple fluorochrome labelings were performed to observe the newly
formed bone dynamics in the porous region. The rats were given one dose each of tetracycline
(50 mg/kg, subcutaneously) 5 weeks, calcein (15 mg/kg, intravenously) 6 weeks, and xylenol
orange (90 mg/kg, intravenously) 7 weeks after implantation respectively.
Histological Evaluation
Each implant was fixed in 70% ethyl alcohol, stained with Villanueva bone stain, dehydrated
in graded series of ethyl alcohol and acetone and then embedded in methyl methacrylate. After
cutting sections (200 μπι in thickness) by using a milling machine, they were ground on the
grinding machine with a diamond lap disc (Speed Lap, Maruto Ltd, Japan) to a thickness of
7 to 10 μπι. These specimens were examined by light microscopy or fluoromicroscopy. The
surface of the implant was coated with a thin layer of carbon, and the bone/materials
interface was observed by using a scanning electron microscope (SEM525M, Philips,
Netherlands). The implant was first observed in the backscattered electron imaging mode.
The bone/ceramic interface was then visualized by secondary electron imaging.
RESULTS
The histological section of all control implants without marrow cells showed fibro-vascular
tissue invasion into the porous regions. However, there was no bone formation in the porous
regions of each implant. On the other hand, all alumina (6 out of 6), and HA (6 out of 6)
combined with marrow cells showed osteogenesis in the porous regions. Four out of 6 Kiel
bone with marrow cells also showed small amount of newly formed bone. However, connective
tissue interposition was observed between Kiel bone and newly formed bone together with
moderately infiltrated small round cells and giant cells. The fibrous tissue intervention was
also observed at the interface between de novo bone and alumina surface in the porous
regions(Fig.la). In some interfacial areas, alumina ceramics was in contact with bone tissue
without fibrous tissue intervention(Fig.lb). Fluorochrome study showed that tetracycline
administered 5 weeks after implantation was seen close to the center of the pore and calcein
and xylenol orange (administered 6 and 7 weeks after implantation) were seen near the surface
of the alumina ceramics(Fig.2). This finding indicates that the bone formation began in the
pore area away from the ceramic surface in the porous region and proceeded bi-directionally
both to the surface of the alumina and to the center of the pores. In contrast, the osteogenesis
formed in HA proceeded in a centripetal direction toward the center of the pores.
Osteogenic Response of Rat Bone Marrow Cells: M. Okumura et al 5
Figure 1. Scanning electron micrograph of A1203 with marrow cells 8 weeks after implantation
Upper(a) shows an area where connective tissue (arrows) intervention between bone(B) and
alumina(A) is visible. Lower(b) shows an area where connective tissue is not observed at the
interface. Bar; 10 iim
6 Bioceramics Volume 4
Figure 2. Eight weeks after subcutaneous implantation of A1203 with marrow cells.
Upper(a) is the undecalcified section under light microscopy (Villanueva bone stain,original
magnification x200). The newly formed bone (B) seemed to be contact with alumina(A) in the
porous region. Lower(b) shows the same section under fluoromicroscopy. Tetracycline(T),
calcein(C) and xylenol orange(X) were administrated 5, 6 and 7 weeks respectively after
implantation. Tetracycline was seen close to the center of the pore and calcein and xylenol
orange were seen near the surface of the alumina ceramics. This indicates that osteogenesis
began away from the surface of the alumina ceramics and proceeded in a centrifugal direction.
Osteogenic Response of Rat Bone Marrow Cells: M. Okumura et al l
DISCUSSION AND CONCLUSIONS
Osborn and Newesely(16,17) classified the different patterns of osteogenesis as distance,
contact and bonding osteogenesis in terms of their biodynamics. They described that most
biotolerant metals are incorporated in bone by distance osteogenesis, bioinert materials by
contact osteogenesis, and bioactive implants by bonding osteogenesis. Our previous
reports(l 1-15) and the present work clearly showed these different types of osteogenesis in
the ectopic sites. Bioactive materials such as HA and TCP with marrow cells induced
consistent bone formation 3 weeks after subcutaneous implantation, and the bone formation
started directly on the surface of the materials and proceeded to the center of the pores
(bonding osteogenesis). On the other hand, traditional porous bone graft substitute of Kiel
bone made of bovine cancerous bone when combined with marrow cells also showed de novo
bone formation in the pore area(14). However, the bone formation was inconsistent and there
was no contact between the de novo bone and Kiel bone (distance osteogenesis). Alumina
ceramics combined with marrow cells also showed this pattern of bone formation.
Fluorochrome labeling demonstrated that the bone formation starts not from the ceramic
surface but in the pore regions away from the surface and progressed in a centrifugal manner
as described by Köster et al(19). Although the direction of the bone formation in both Kiel
bone and alumina ceramics is the same, microscopically, some area of bone/alumina ceramics
interface has no intervening fibrous tissue which always existed in bone/Kiel bone interface.
In the present report, data on the bone forming capacity (amount of the bone) between HA
and alumina ceramics are not given because of the differences in structure (porosity and pore
size). We believe that the most important factor in bone bonding is the interaction between the
material surface and osteogenic cells. As clearly demonstrated in the present study, this
bioinert alumina ceramics lacks the interaction resulted in "contact osteogenesis". In contrast,
bioactive materials such as HA shows this interaction as evidenced by the finding that bone
formation starts from the material surface by osteogenic cells and results in "bonding
osteogenesis"(12-18). These results show that ectopic bone formation induced by bone marrow
cells is a useful model to study the bone/biomaterial interaction.
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