4 Bones Substitues

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Tissue Reaction and Material Characteristics of

Four Bone Substitutes
Article in The International journal of oral & maxillofacial implants · January 1996
DOI: 10.7202/037238ar · Source: PubMed
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Tissue Reaction and Material Characteristics of Four

Bone Substitutes
Simon Storgård Jensen/Merete Aaboe, DDS/Else Marie Pinholt, DDS, MS, Dr
Odont/Erik Hjørting-Hansen, DDS, Prof Dr Odont/Flemming Melsen, MD, PhD,
Prof Dr Med/I. Eystein Ruyter, Dr Philos, Dr Rer Nat
The aim of the present study was to qualitatively and quantitatively compare
the tissue reactions around four different bone substitutes used in orthopedic
and craniofacial surgery. Cylinders of two bovine bone substitutes (Endobon
and Bio-Oss) and two coral-derived bone substitutes (Pro Osteon 500 and
Interpore 500 HA/CC) were implanted into 5-mm bur holes in rabbit tibiae.
There was no difference in the amount of newly formed bone around the four
biomaterials. Interpore 500 HA/CC resorbed completely, whereas the other
three biomaterials did not undergo any detectable biodegradation. Bio-Oss was
osseointegrated to a higher degree than the other biomaterials. Material
characteristics obtained by diffuse reflectance infrared Fourier transform
spectrometry analysis and energy-dispersive spectrometry did not explain the
differences in biologic behavior.
(INT J ORAL MAXILLOFAC IMPLANTS 1996;11:55–66)
Key words: bone substitute, bovine bone, calcium carbonate, coral, material
characteristics, material resorption, porous hydroxyapatite
Autogenic and allogenic bone grafts have been used with some success to improve
repair of bone defects. However, autogenic transplantation requires a second surgical
site, the surgeon has limited quantities available, and the autograft has a tendency to
resorb.1 Allografts possess the disadvantage of an uncertain immune response, the risk
of disease transmission, and a tendency to be resorbed.1 Therefore, development of
bone substitutes has been of major interest for a number of years.2
An ideal bone substitute should be biocompatible, and it should gradually be
replaced by newly formed bone and preferably possess osteoinductive or
osteoconductive properties. Osteoinduction is a healing process in which local
stimulating factors cause mesenchymal cells to disaggregate, migrate, reaggregate,
proliferate, and differentiate into chondroblasts or osteoblasts.3,4 In osteoconduction,
the implanted material serves as a scaffold for ingrowth of capillaries, perivascular
tissue, and osteoprogenitor cells from the recipient bed.1
In recent years, hydroxyapatite (HA) bone substitutes have been developed
synthetically,5,6 derived from corals or algae,7-9 or originated from natural bone
10
mineral.10 These bone substitutes are generally regarded as being biocompatible and
osteoconductive,7,11 although reports concerning their biodegradability have been
inconclusive.7,8,12-14
The present study qualitatively and quantitatively evaluated tissue reactions
around two bovine and two coral-derived bone substitutes when implanted into the
rabbit tibia, with special emphasis on differences in rate of resorption. Further, it
relates possible differences in biologic behavior of the four mentioned implant
materials compared to material characteristics obtained by energy-dispersive
spectrometry and infrared spectrometrical analysis of the materials.
Materials and Methods
A license to perform the study was obtained from the Danish National Experimental
Inspectorate. Twelve adult female rabbits (Oryctolatus cuniculus, Copenhagen
white) each weighing 3.75 kg (standard deviation [SD] ± 0.22) were used for this
study and were divided into an experimental group of eight animals and a control
group of four animals. They were fed a standard rabbit diet, given water ad libitum,
and kept separately in cages at the animal research facilities of the Panum Institute,
University of Copenhagen.
Biomaterials. Cylinders of the following materials, 5 mm in diameter and 10
mm in length, were used:
1. Endobon (Merck Biomaterialien, Darmstadt, Germany) is pyrolized (at
temperatures exceeding 1,000°C) ceramized cancellous bovine bone with an
interconnecting macropore system (porosity 30% to 80%, pore diameter 100 to
1,500 µm). From a block measuring 2 × 2 × 1 cm, two to four cylinders were
prepared with a trephine bur for the present study.
2. Bio-Oss (Geistlich Pharma, Wolhusen, Switzerland) is deproteinized sterilized
cancellous bovine bone with a porosity of 75% to 80%. The material is available
as cancellous granules, granules from cortical bone, and cancellous blocks
measuring 1 × 1 × 2 cm. For the present study, two to three cylinders were
prepared from each cancellous block with a trephine bur.
3. Pro Osteon 500 (Interpore International, Irvine, CA) is a coralline HA derived
from the goniopora coral by a hydrothermal exchange reaction with phosphate
(pore volume 65%, mean pore diameter 600 µm, and their interconnections 260
µm). The cylinders were prefabricated.
4. Interpore 500 HA/CC (Interpore International) is a composite bone substitute
derived from the goniopora coral and thus has the same pore configuration as
Pro Osteon 500. The outer 3 µm of the calcium carbonate (CC) is converted to
HA by the same exchange reaction as for Pro Osteon 500.15 The cylinders were
prefabricated.
Surgical Procedure. Surgery was performed under general anesthesia produced
by intramuscular injections of 0.5 mL of Hypnorm (Janssen Pharmaceutica, Beerse,

Belgium) per kg of body weight and 0.4 mL of Stesolid (Dumex, Copenhagen,
Denmark) per kg of body weight, supplemented by a submucosal injection of 1 mL
of 2% Xylocaine adrenalin (Astra Läkemedel, Södertälje, Sweden) in each field of
surgery for the purpose of local anesthesia.
An antibiotic, 1 mL of penicillin (Streptocillin vet 200.000 IE, Novo Industri
A/S, Copenhagen, Denmark) before surgery and 0.5 mL for each of the 3 following
days was injected intramuscularly. The surgical field of both tibiae was shaved and
washed with 0.2% chlorhexidine gluconate. Skin incision and subperiosteal
dissection was carried out on both tibiae. With a trephine (5-mm outer diameter),
four unicortical implant beds were prepared, two in each tibia. The implants were
placed at random order into the implant beds and in contact with the inside of the
opposing cortex. Thereafter, the implants were cut off to the level of the outer
cortex. For localization purposes, two gutta percha points were placed in the bone
between the implant beds. The periosteum was closed with nonresorbable suture
(Miralene 4-0, Braun Melsungen AG, Melsungen, Germany) and the skin with
resorbable suture (Vicryl 4-0, Ethicon, Norderstedt, Germany). Liquid plaster
(Nobecutan, Astra Tech A/S, Albertslund, Denmark) was applied to the wound.
After surgery, radiographs were taken of both tibiae, one in the axis of the implant
and one perpendicular to this axis (Kodak Ektaspeed occlusal film, 60 mA, 1.0
seconds).
For bone marking purposes, fluorochromes were injected intravenously at 4 and
3 days presurgery (20 mg of oxytetracycline [Terramycin vet, Pfizer, Copenhagen,
Denmark] per kg of body weight) and at 4 and 3 days before sacrifice (25 mg of
alizarin red [Alizarin Complexone, Sigma, St Louis, MO] per kg of body weight
dissolved in 10 mL of isotonic natrium bicarbonate [infusion liquid 14 g/L Sygehus
Apotekerne i Danmark, Copenhagen, Denmark]).16,17
Clinical and Histologic Evaluation. The control group was sacrificed 1 week
after surgery. The observation period of the experimental group was 8 weeks.
Observation for proper wound healing was performed daily the first postoperative
week and thereafter twice a week. The animals were sacrificed with an overdose of
20% weight per weight of Mebumalnatrium (Rigshospitalets Apotek, Copenhagen,
Denmark). The tibiae were dissected from the animals and fixated in 4% buffered
formaldehyde for 14 days, and then they were stored in 70% alcohol.
Soft tissue was removed from the specimens, and the tibiae were resected
approximately 5 mm from the implants. The anterior cortical layer was removed to
facilitate dehydration and infiltration of the embedding material, methyl
methacrylate resin. Undecalcified consecutive nonrandomized sections were sawn at
a thickness of 220 µm with an Exakt saw (blade thickness 230 µm) parallel to the
long axis of the bone and of the implant cylinders. The sections were stained with
modified Goldner trichrome stain for qualitative and quantitative histologic
evaluation. The section in the middle of each implant was used for quantitative
measurements. Point counting was used to estimate volume fractions and relative
surfaces. The volume fraction indicates the percentage of the reference space (cortex
or marrow) occupied by a given structure (bone or implant material), while the
relative surface indicates the percentage of the implant surface area covered with
bone in a given reference space (cortex or marrow). Thickness of cortical structures
was estimated by direct measurements on projected sections.
Diffuse Reflectance Infrared Fourier Transform Spectrometry (DRIFTS)
Analysis. Using an agate mortar, each sample was ground together with potassium
bromide (KBr) (Uvasol for spectrometry, Merck) to a fine powder for DRIFTS
analysis in the range 5,200 to 450 cm-1 (System 2000, Perkin-Elmer, USA). One
hundred scans per sample were performed with a resolution of 1.0 cm-1.
The spectra from the implant materials were compared with reference spectra
obtained by DRIFTS analysis of hydroxyapatite (Ca10[PO4]6[OH]2, Euro-Crystals,
The Netherlands), calcium carbonate (CaCO3, Pro analysi, Merck), and calcium
hydroxide (Ca[OH]2, Pro analysi, Merck).
Energy-Dispersive Spectrometry (EDS). The cylinders of the four
biomaterials were coated with carbon for conduction and were analyzed in an
energy-dispersive x-ray spectrometer (Edax DX-4i, Edax International, Mahwa, NJ)
combined with a scanning electron microscope (Philips XL30i, Philips Electronics
NW, Eindhoven, The Netherlands). Endobon, Bio-Oss, and Pro Osteon 500 were
analyzed in an area of 100 µm2 with the energy of the electron beam fixed at 10 keV.
Interpore 500 HA/CC was analyzed with increasing energy (5.3, 6, 7, 10, 15, 20, and
30 keV) to expose changes in the calcium-to-phosphate ratio from the surface to the
bulk of the material.
Statistical Analysis. For the statistical evaluation of material resorption,
Wilcoxon’s nonparametric rank sum test was used to compare the volume fractions
of implant material in the cortex and the marrow after 1 and 8 weeks of observation.
The Friedman two-way analysis of variance was used to compare the biomaterials
with respect to the amount of new bone in the cortex and the marrow, the relative
area of implant surface covered with bone in the cortex, and cortex thickness inside
and outside the former defect. Wilcoxon’s nonparametric signed rank test was used
to analyze the thickness of cortical bone inside and outside the former defect.
Results
Clinical Evaluation. All animals remained healthy during the observation period,
and all sites of implantation healed uneventfully. There were no signs of infection,
edema, or dehiscence.
Qualitative Histologic Evalation. A wide rim of woven bone with a primitive
trabecular pattern was often observed after 1 week. This bone extended from the
periosteal and endosteal surface of the cortical bone neighboring the defect toward
the implants (Fig 1). To a degree, this bone extended into the porous system of the
implants. Highly vascularized connective tissue was observed between the
trabeculae of the primitive bone. On the endosteal surface opposite the defect, a
similar phenomenon was noted.
After 8 weeks, osseous healing had taken place. In the cortical areas of the
experimental cavities, woven bone filled the varying residual porous system of the
implants; however, haversian systems were seen where an initiating remodeling of
the newly formed bone was observed. The primitive periosteal and endosteal bone,
seen after 1 week of observation, could no longer be observed after 8 weeks. In the
marrow space, newly formed bone was seen on the trabeculae of all the implant
materials.
Most of the cortical areas of the cavities filled with Endobon were occupied by
newly formed bone, generally separated from the implant material by a thin fibrous
connective tissue membrane (Fig 2). However, complete osseointegration with
intimate contact between bone and implant could be seen both on sharply
demarcated surfaces and on blurred surfaces. In the marrow space of the tibia, the
surface of the implant was covered with connective tissue, and only pointwise was a
sparse amount of newly formed bone observed.
In most of the cortical areas of the cavities filled with Bio-Oss, the implant
material was completely osseointegrated with intimate contact between the surfaces
of the porous system of Bio-Oss and the newly formed bone (Fig 3). Fibrous
connective tissue membranes were noted in small foci only and were in no way a
dominating feature. A small amount of connective tissue and newly formed bone
was seen in relation to the implant projecting into the marrow space; in comparison
with Endobon, however, more of the Bio-Oss implant surface was characterized by
the presence of newly formed bone (Fig 4). There was no sign of surface dissolution,
but a few resorption lacunae were present, especially in the marrow (Fig 5).
Most of the cortical areas of the cavities filled with Pro Osteon 500 were
occupied by newly formed bone. Bone was seen in intimate contact with both
sharply demarcated and blurred surfaces; however, most often a thin rim of fibrous
connective tissue separated the bone from the implant material (Fig 6). In the
marrow, a small amount of bone was seen on the surface of the implant material
with no connection to cortical bone.
Bone was woven closely into the few remnants of Interpore 500 HA/CC in the
cortical areas of the cavities (Fig 7). Nearly all of the implant material extending into
the marrow space was resorbed (Figs 8 and 9). Where fragments were present, bone
was often seen in close apposition, frequently with no connection to the cortical
bone.
Because of the section thickness, a detailed evaluation of cellular events and thus
the nature of decomposition, whether cellular or biochemical, of the biomaterials
could not be described. The histologic observations are summarized in Table 1.

Quantitative Histologic Evaluation. The quantitative histologic evaluation
showed a statistically significant decrease in volume fraction of the Interpore 500
HA/CC material from week 1 to week 8 (P < .005) (Tables 2 and 3). Although this
material almost disappeared, the other three materials did not demonstrate any
statistically significant change in volume fraction (P > .05). After 8 weeks of
observation, no statistically significant differences were observed in the amount of
newly formed bone in the defects filled with the four different biomaterials (Table 4
), whereas the relative area of bone-covered implant surface was significantly higher
on Bio-Oss than on Endobon and Pro Osteon 500 (P < .01) (Table 5); the residual
amount of Interpore 500 HA/CC was too small to estimate quantitatively.
Cortical bone thicknesses were measured corresponding to the former defects,
and comparison of these revealed no statistically significant difference between the
four biomaterials (P > .1) (Table 6). However, when cortical bone thickness outside
the former defect was used as a reference value, the defects filled with Interpore 500
HA/CC had become significantly thicker (P < .01) (Table 6). This was not observed
in the defects filled with Endobon, Bio-Oss, and Pro Osteon 500 (P > .1). The
volume fraction of the newly formed bone on the implant surfaces in the the marrow
space was negligible on all four biomaterials as seen in Table 7.
DRIFTS Analysis. Endobon. The absorption peaked at 3,573.2 and 632.9 cm-1,
representing monomeric OH- groups in the apatite structure.18,19 Monomeric OH-
groups of a silicate material containing OH- gave rise to the absorption that peaked at
3,697.6 and 3,644.7 cm-1.20 The absorption peaks at 2,148.7; 2,077.6; 2,047.8;
2,003.2; and 1,988.3 cm-1 were overtones of the antisymmetric stretching vibrations
of the PO43- group represented by the peaks at 1,088.7; 1,055.2; and 1,025.4 cm-1.
Symmetric stretching vibrations of the PO43- group gave rise to the absorption that
peaked at 962.8 cm-1.18,19 The two peaks at 602.3 and 574.3 cm-1 were the result of
antisymmetric bending motions of the PO43- group.18 Absorption peaked at 1,460.4;
1,414.2; and 874.7 cm-1, representing CO32- absorptions in the type B carbonated
apatite. The CO32- absorption of the type A carbonated apatite was vaguely seen as
the peak at 879.6 cm-1.21 The band around 2,350 to 2,300 cm-1 was noise from the
KBr background specter. It was concluded that Endobon is an HA containing
moderate amounts of type B carbonated apatite, small amounts of type A carbonated
apatite, and an OH--containing silicate material.
Bio-Oss. The absorption peak at 3,571.5 and the weak shoulder at 634.1 cm-1
represented monomeric OH- in the apatite structure. The absorption peaks at 2,139.2;
2,075.5; and 1,999.4 cm-1 were overtones of the antisymmetric stretching vibrations
of the PO43- group represented by the peak at 1,045.8 cm-1. Symmetric stretching
vibrations of the PO43- group caused the peak at 962.8 cm-1. Antisymmetric bending
motions of the phosphate group gave rise to the peaks at 605.8 and 567.7 cm-1.
Absorption peaks at 1,454.4; 1,418.2; and 873.6 cm-1 represented CO32- absorptions
of the type B carbonated apatite. The presence of type A carbonated apatite was
vaguely seen as the shoulders at 1,498.8 and 879.2 cm-1. The band around 2,350 to
2,300 cm-1 was noise from the KBr background specter. It was concluded that
Bio-Oss is an HA containing type B carbonated apatite and a minor portion of type
A carbonated apatite.
Pro Osteon 500. Absorption peaks at 3,571.8 and 633.2 cm-1 represented
monomeric OH- groups in the apatite structure. The absorption bands at 1,089.2 and
1,050.4 cm-1 were the result of stretching vibrations of the PO43- group. Symmetric
stretching vibrations of the PO43- group in HA gave rise to the absorption peak at
962.4 cm-1. The absorption bands at 2,144.7; 2,075.0; 2,047.9; 2,001.6; and 1,988.6
cm-1 were overtones of the PO43- antisymmetric stretching vibrations around 1,000
cm-1. Peaks at 603.8 and 572.2 cm-1 were caused by antisymmetric bending motions
of the PO43- group. Absorption peaks at 1,547.2; 1,457.6; and 880.0 cm-1 represented
CO32- absorptions of type A carbonated apatite, and the CO32- absorptions at 1,457.6;
1,411.0; and 873.5 cm-1 represented type B carbonated apatite. It was concluded that
Pro Osteon 500 is an HA containing both types A and B carbonated apatite.
Interpore 500 HA/CC. The absorption pattern of Interpore 500 HA/CC
generally corresponded to the CaCO3 reference spectrum. The diffuse shoulder at
3,576.1 cm-1 and the peak at 631.9 cm-1 might have been traces of the monomeric
OH- group of an HA coating. The peaks at 1,085.6 and 1,014.1 cm-1 could indicate
the presence of the PO43- group of an apatite structure. It was concluded that
Interpore 500 HA/CC is a calcium carbonate, and traces of HA are indicated.
Energy-Dispersive Spectrometry. The results of the semiquantitative chemical
analysis of the four biomaterials are listed in the Tables 8 and 9.
Discussion
In the present study, two bovine and two coral-derived bone substitutes were
implanted into rabbit tibiae for qualitative and quantitative histologic evaluation of
tissue reactions. Diffuse reflectance infrared Fourier transform spectrometry
(DRIFTS) analysis and the energy-dispersive analysis (EDS) were performed to
relate material characteristics of the four bone substitutes to differences in biologic
behavior. During the 8 weeks of implantation into rabbit tibiae, Interpore 500
HA/CC was almost totally resorbed; whereas, the three other biomaterials did not
undergo any statistically significant degradation. Bio-Oss was osseointegrated to a
higher extent than Endobon and Pro Osteon 500, but there was no statistically
significant difference in the amount of newly formed bone in the cortical part of the
cavities filled with the four tested biomaterials.
The DRIFTS analyses indicated Endobon, Bio-Oss, and Pro Osteon 500 to be
HAs containing both types A and B carbonated apatite. In addition, Endobon was
found to contain an OH--containing silicate material, and Interpore 500 HA/CC was
found to consist of calcium carbonate with a thin outer layer of HA. The EDS
analyses of Endobon, Bio-Oss, and Pro Osteon 500 (see Table 8) indicate the
presence of a calcium phosphate material. Increasing the penetration depths by
increasing the energy levels of the electron beam in the analysis of Interpore 500
HA/CC (see Table 9) reveals an increasing calcium-to-phosphate ratio, indicating the
presence of a thin surface layer of HA.
The infrared spectrometric analysis was performed to determine the identity and
location of the different functional groups within the materials, and the EDS analysis
was chosen to reveal the presence of any significant amounts of foreign substances
13,22 in the biomaterials. Such could not be observed. Bio-Oss and Interpore 200
have previously been subjected to infrared spectrometry,13,23 and the results from
these analyses correspond to our DRIFTS findings for Bio-Oss and Pro Osteon 500.
Thus, the material characteristics did not reveal any impurities in the four
biomaterials that would make them unsuitable for clinical use.
The defects created in this study were not meant to be critical size defects
(CSD),24-26 even though Schlickewei and Paul11 proved no spontaneous regeneration
of an almost similar defect. The purpose for chosing this experimental model was
that each rabbit should serve as its own control, so that any statistically significant
difference in the parameters measured could be interpreted as a difference between
the four biomaterials in biologic behavior in bone healing. Undecalcified sections
were chosen to avoid shrinkage of the specimens and thereby to provide more
reliable quantitative measurements.27
The interconnected porous systems of the four investigated materials are of
optimal size and configuration for vascular ingrowth, which is necessary for new
bone formation.28,29 Ripamonti and Reddi30 and van Eeden and Ripamonti31
described the critical role of the geometry of the implant material in the
differentiation of bone. They demonstrated osteoinduction when cylinders of
Interpore 200 and Pro Osteon 500 were implanted heterotopically into baboon
muscle, but not when granules of the same materials were used. For this reason,
implant blocks were used in the present study. This experiment was conducted in an
osteogenic environment and thus cannot reveal any osteoinduction. The minor
portion of newly formed bone seen in relation to all the implant materials in the
marrow cavity, and especially on the surface of Bio-Oss (see Fig 4), may be a result
of the seeding phenomenon described by Heughebaert et al32 and Hjørting-Hansen et
al.33
The primitive trabecular bone seen in the 1-week specimens of each of the four
materials (see Fig 1) has an appearance similar to that of callus formation observed
in a stable bone fracture.34,35 The fact that this newly formed bone was not observed
in any of the 8-week specimens also indicates that it is a reaction to the surgical
trauma, independent of the kind of biomaterial used.
Although Mandelkow et al,36 in a study on 6-mm bur holes in sheep tibiae,
reported less bone formation in the Bio-Oss cavity than in that of the control, several
experimental and clinical studies have shown Bio-Oss to be highly biocompatible
and osteoconductive.11,14,37-39 Disagreements prevail as to the biodegradability of
Bio-Oss. Klinge et al14 reported almost total resorption of Bio-Oss granules
implanted in 5-mm rabbit skull defects after 14 weeks, whereas Schlickewei and
Paul,11 Bereiter et al,37 and Schlickewei et al40 only saw biodegradation that could
be ascribed to physiologic remodeling when different configurations of Bio-Oss
were used in experimental animals and for different clinical indications. In the
present study, a few resorption lacunae (see Fig 5) might indicate resorptive activity
so low that a possible decrease in implant volume was quantitatively not detectable
after 8 weeks. The morphology of the adjacent cells was that of osteoclasts. This
could indicate that osteoclasts are able to resorb Bio-Oss directly, as earlier
indicated.23 However, a low-to-moderate rate of biodegradation could readily be
hidden by the large interimplant variation in volume fraction seen after both 1 and 8
weeks (see Tables 2 and 3). To our knowledge, the relative surface area of Bio-Oss
covered with bone (see Table 5) has not been compared to other biomaterials
previously. However, other studies have described the same intimate contact
between Bio-Oss and newly formed bone (see Fig 3).14,40
The results of the material analyses do not explain the significant difference
between the two bovine bone substitutes. Through transmission electron microscopy
and x-ray diffraction analysis, the author has shown Bio-Oss to have the crystalline
structure of natural bone (crystal dimension 400 × 100 Å), whereas in the
manufacturing procedure Endobon has been exposed to temperatures exceeding
1,000°C, resulting in a crystal growth, according to the manufacturer’s description.
This does not change the basic structure of the bovine spongiosa but may cause the
minor degree of osseointegration because of the altered surface characteristics.22,41
However, a high degree of osseointegration and good clinical results have been
reported for Endobon.42-45
In a study on rabbit skull defects, Fukuta et al38 measured the area occupied by
bone and by implant material (Bio-Oss and Interpore 200) after 1, 2, 4, and 8
months. As in the present study, they did not observe any significant difference in
the amount of new bone formed or any signs of degradation of the materials. Holmes
and coworkers have performed histomorphometric studies12,46,47 on Interpore 200
and Interpore 500 (now Pro Osteon 500) implanted into long bone defects and used
for alveolar ridge augmentation in dogs. Although studies in dogs and rabbits are not
directly comparable, mean volume fractions of 45.2% and 51.2% newly formed
bone12,47 seem to be in accordance with the 39% found in the present study (see
Table 4). Their finding, however, of 90% of the implant surface covered with bone
12,47 is in contrast to the 38% found in the present study (see Table 5). Fibrous
encapsulation of most of the Pro Osteon 500 implant material could not be a result of
micromovements of the implants,33 because the cylinders were press-fitted into the
cavities.
Reports concerning biodegradability of blocks and granules of the coralline

hydroxyapatites Pro Osteon 500 and Interpore 200 cover resorption rates from 2% to
29% a year (Hjørting-Hansen et al. Unpublished data).7,12,48-50 Granules are reported
to biodegrade faster than blocks.50 It is generally accepted that an optimal bone
substitute is one that maintains mechanical stability and volume during the initial
healing and then subsequently totally resorbs.51 Interpore 500 HA/CC may be such a
material.15 Total bone healing of the osseous defect and almost total resorption of
the material during the 8 weeks of observation were observed (see Figs 8 and 9).
However, the defect in this study is not a CSD and therefore, it might regenerate
spontaneously.24-26 The idea of having a bone substitute at disposal where the rate of
resorption can be controlled with reference to clinical indication choosing different
thicknesses of HA surface layer15 is intriguing. However, the big interindividual and
intraindividual differences concerning healing potential and rate of resorption make
it difficult to predict which surface layer is optimal in a given situation.
Conclusion
Although one must not draw conclusions from studies conducted in rodents directly
to clinical use in humans, this study reveals important differences between four bone
substitutes all used commonly in orthopedic and craniofacial surgery. The better
osseointegration of Bio-Oss and the almost complete biodegradation of Interpore
500 HA/CC suggest that future experiments concentrate on these two categories of
materials. It would be interesting to test both materials in a critical size defect model
in an experimental animal situated higher on the phylogenetic ladder with
observation periods of, for example, 1, 2, 4, 8, and 12 months. Such a study with an
evaluation of thin sections could uncover the cellular events and the dynamics of
material biodegradation, and probably reveal whether Bio-Oss actually is replaced
by creeping substitution or just takes part in the physiologic remodeling of the bone.
In addition, it could be relevant to test the use of the two materials as carriers of
bone morphogenetic proteins or growth factors.
Acknowledgments
This study was financially supported by the Danish Medical Research Council
(J.nr. 12-1718-1 kg/mp and 12-2023-1 KG/Lpa), FUT (Danish Dental Associations
Foundation for the Support of Scientific Investigation), Colgate Scientific
Foundation, and Director Ib Henriksen’s Foundation.
The biomaterials were kindly supplied by the individual manufacturers.
The authors wish to thank the Department of Biostatistics for help with the
statistical analysis.
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Fig. 1 Primitive trabecular bone extending from the

periosteal (P) and endosteal (E) surfaces of the Endobon implant material (I) after 1
week of observation. M = marrow space; G = gutta percha point. Original linear
magnification × 10. Modified Goldner trichrome stain.
Fig. 2 Trabeculae of the Endobon

implant material (I) separated from the newly formed bone (B) by a thin fibrous
connective tissue membrane (arrows) after 8 weeks of observation. Original linear
magnification × 60. Modified Goldner trichrome stain.
Fig. 3 Intimate contact between

the Bio-Oss implant material (i) and newly formed bone (b) in the cortical part of the
cavity after 8 weeks of observation. Original linear magnification × 20. Modified Goldner
trichrome stain.
Fig. 4 Newly formed bone (arrow)

on the surface of the Bio-Oss implant material (i) in the marrow space (m) after 8 weeks
of observation. Original linear magnification × 60. Modified Goldner trichrome stain.
Fig. 5 Resorption lacuna (arrow)

on the surface of the Bio-Oss implant material (i) in the marrow space (m) after 8 weeks
of observation. Original linear magnification × 160. Modified Goldner trichrome stain.
Fig. 6 Trabeculae of the Pro

Osteon 500 implant material (i) separated from the newly formed bone (b) by fibrous
connective tissue (arrows) in the cortical part of the cavity after 8 weeks of observation.
Original linear magnification × 60. Modified Goldner trichrome stain.
Fig. 7 Remnants of the Interpore

500 HA/CC implant material (arrows) woven closely into the newly formed bone (b) in
the cortical part of the cavity after 8 weeks of observation. Original linear magnification ×
40. Modified Goldner trichrome stain.
Fig. 8 Cylinder of the Interpore 500 HA/CC implant

material (i) after 1 week of observation. Note the endosteal callus formation (c). The
central fissure is an artifact. Original linear magnification × 10. Modified Goldner
trichrome stain.
Fig. 9 Remnants of the Interpore 500 HA/CC implant

material (arrows) after 8 weeks of observation. Original linear magnification × 10.
Modified Goldner trichrome stain.
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Tissue Reaction and Material Characteristics of

Simon Storgård Jensen Else Marie Pinholt

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Erik Hjørting-Hansen Eystein I Ruyter

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Tissue Reaction and Material Characteristics of Four

by intramuscular injections of 0.5 mL of Hypnorm (Janssen Pharmaceutica, Beerse,

could not be described. The histologic observations are summarized in Table 1.

Reports concerning biodegradability of blocks and granules of the coralline

1. Burchardt H. The biology of bone graft repair. Clin Orthop 1983;174:28–42.

hydroxyapatite/calcium carbonate implants in rabbits. Trans Ortho Res Soc

42. von Schaller P, Geldmacher J, Freiberger N, Flügel M. Die Auffütterung kleiner

Fig. 1 Primitive trabecular bone extending from the

Fig. 2 Trabeculae of the Endobon

Fig. 3 Intimate contact between

Fig. 4 Newly formed bone (arrow)

Fig. 5 Resorption lacuna (arrow)

Fig. 6 Trabeculae of the Pro

Fig. 7 Remnants of the Interpore

Fig. 8 Cylinder of the Interpore 500 HA/CC implant

Fig. 9 Remnants of the Interpore 500 HA/CC implant

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