SUPORT CURS MASTERAT SUBSTITUENTI OSOSI

Dr. SIRBU PAUL

PROF. DR. PAUL BOTEZ
Bone defects of different etiology trauma, osteoporosis, tumors or metabolic
diseases represent an important medical issue with socio-economical involvement, due
mainly to the lack of spontaneous healing or to the treatment problems and long lasting
healing.
Approximately 10% of the bone surgery requires bone grafts or bone substitutes
usage ((Katsuko 2004)). Autografts are still considered the golden standard in treatment of
bone defects, even if the literature reports difficulties in graft harvesting and graft availability.
The allograft usage involves the infection risk (viral or bacterial) and high costs, while it
requires a bone bank with all facilities.
For over a century, scientists are trying to discover or synthesize inorganic products
that could be used as bone substitutes; these products must provide long lasting bone
biocompatibility and must provide bone healing abilities. These bone substitutes must solve
the autograft and allograft disadvantages.
The last decade literature is plenty of works regarding bone substitutes applications in
vivo and in vitro; the researchers are looking for the crystallographic, physical and
chemical and biomechanical properties of the bone substitutes and also for the biological
behavior during implantation in the living tissues. (Ribeiro 2006, Gisep 2004, Verne
2002).
The promising results of these studies have issued a classification for the bone
substitutes as follows: phospohocalcic compounds (hydroxiapatite and calcium
triphosphate), bone ceramics and various growth factors (bone morphogenetic proteins).
The usage of these bone substitutes for bone healing is based on inducing bone
apposition (e.g. osteoinductive products as hydroxiapatite or calcium triphosphate) or
bone neoformation from the precursor mesenchimal cells. (e.g. osteoinductive products
as bone morphogenetic proteins) (Yamamoto 2006, Hidaka 2006, El-Ghannam 2005,
Grauer J.N. 2005.).
Nowadays, there are described or synthesized many bone substitutes with different
osteointegrative properties; the multitude of products has imposed the generation and usage
of the experimental models and the evaluation methods required for an accurate comparison.
In most cases, in vivo studies for these compounds requires the usage of an
epiphyseal (trabecular) or diaphyseal bone defect, in which the bone healing process is
investigated in detail, by radiological, histological, morphometric and biomechanical means.
(Schmidhammer 2006, Misch 2006, Akca 2006, Li 2006.).
The evaluation and research of bone substitutes in bone defect treatment is a
complex process whose goal is the clinical usage. The last step represents a decisive and a
maximum interest subject for the practitioner and the patient.
References
1
Akca K, Cehreli MC., Biomechanical consequences of progressive marginal bone loss around oral
implants: a finite element stress analysis. Med Biol Eng Comput. 2006 Jul;44(7):527-535. Epub 2006 Jun 10.
PMID: 16937188
2
El-Ghannam A., Bone reconstruction: from bioceramics to tissue engineering. Expert Rev Med Devices.
2005 Jan;2(1):87-101. Review. PMID: 16293032
3
Gisep A, Kugler S, Wahl D, Rahn B., Mechanical characterisation of a bone defect model filled with
ceramic cements. J Mater Sci Mater Med. 2004 Oct;15(10):1065-71. PMID: 15516866
4
Grauer JN, Beiner JM, Kwon B, Vaccaro AR., The evolution of allograft bone for spinal applications.
Orthopedics. 2005 Jun;28(6):573-7; quiz 578-9. Review. PMID: 16138470
5
Hidaka C, Cunningham ME, Rodeo SA, Maher SA, Zhu W., Modern biologics used in orthopaedic
surgery. Curr Opin Rheumatol. 2006 Jan;18(1):74-9. Review. PMID: 16344622
6
Katsuko S. Furukawa, Shunsuke Miyauchi, Daisuke Suzuki, Yoshikazu Umezu, Tsuneo Shinjo, Takashi
Ushida, Miki Eguchi, Tetsuya Tateishi, Bone tissue engineering based on beadcell sheets composed of calcium
phosphate beads and bone marrow cells, Materials Science and Engineering C 24 (2004) 437440
7
Li ZH, Liao W, Liu SQ, Zhang YF, Wang CY, Zhao Q., The study of tissue-engineering bone for repair of

segmental bone defects, Zhonghua Zheng Xing Wai Ke Za Zhi. 2006 Jan;22(1):55-9. PMID: 16573169
8
Misch KA, Yi ES, Sarment DP., Accuracy of cone beam computed tomography for periodontal defect
measurements. J Periodontol. 2006 Jul;77(7):1261-6. PMID: 16805691
9
Ribeiro C.C., Barrias C.C., Barbosa M.A., Preparation and characterisation of calcium-phosphate porous
microspheres with a uniform size for biomedical applications. J Mater Sci Mater Med. 2006 May; 17 (5): 455-63.
PMID: 16688586.

10. Schmidhammer R, Zandieh S, Mittermayr R, Pelinka LE, Leixnering M, Hopf R,

Kroepfl A, Redl H., Assessment of bone union/nonunion in an experimental model
using microcomputed technology. J Trauma. 2006 Jul;61(1):199-205.
PMID: 16832271
11. Verne E, Bosetti M, Brovarone CV, Moisescu C, Lupo F, Spriano S, Cannas M., Fluoroapatite glass-ceramic
coatings on alumina: structural, mechanical and biological characterisation. Biomaterials. 2002
Aug;23(16):3395-403. PMID: 12099282
12. Yamamoto M, Takahashi Y, Tabata Y., Enhanced bone regeneration at a segmental bone defect by
controlled release of bone morphogenetic protein-2 from a biodegradable hydrogel. Tissue Eng.
2006 May;12(5):1305-11.

PMID: 16771643
The importance of bone substitutes
Autografts, allografts and bone synthetic substitutes plays an important role in
orthopaedic and reconstructive surgery while the understanding of their biological effects is
required for directed applications.
Bone grafting materials are those implants that promotes bone healing by one of the
following actions: osteogenesis, osteoinduction and osteoconduction.
Osteoconductive materials became more important, especially in bone pathology
while they are used as bone substitutes. These substances have a composition similar to the
bone mineral matrix and are biocompatible. Their main function is of bone tissue support,
allowing bone apposition on their surface; thus, they are used mainly for treating the bone
defects. More recently, they are used as a vehicle for osteoinductive substances, augmenting
bone formation.
While initially, only coral hydroxiapatite, calcium phosphate (Plaster-of-Paris) and then
bioactive glasses (bioviotroceramics) were used as bone substitutes, nowadays we are using
osteoconductive ceramic materials and phosphocalcic cements.
From the ceramic materials used in medical applications, we mention here the
calcium triphosphate (-TCP), hydroxyapatite (HA) (Feifel 1995, Mihil 2005, Ohura 1999)
and calcium biphosphate (or biphasic calcium phosphate or BCP or -TCP-HA).
Phosphocalcic cements (CPC) consist in one or more calcium phosphates (CaP)
soluble in aqueous solutions. Many experimental and clinical studies have used
phosphocalcic cements (Constantz 1998, Frankenburg 1998, Mihil 2005).
The proposed subject underlines the practical application by a histologic, histomorphometric and biomechanical investigation between organic bone substitutes (e.g.
phosphorilated cellulose) and import phosphocalcic compounds, whose prices are still
prohibitive, even if they are biocompatible and osteoinductive. This study will represent also
the first step for the homologation process for the Romanian products.
To reach this goal, we will use fundamental research methods, as in vivo research
on animal model followed by a histologic evaluation on calcified and decalcified bone and
electronic methods for biomechanical histomorphometric evaluation.
Up to now, the phosphocalcic cements were successfully used in maxillofacial surgery,
in radius lower extremity fractures or calcaneal fractures, trochanteric fractures, tibial
plate fractures, and also as filler for the osteoporosis in vertebrae bone. Another
application is represented by osteosynthesis augmentation by sponge screws or
pedicular screws in spine surgery.
In this domain, our proposed subject shows an important practical application, by
providing useful information in practical orthopaedics for phosphocalcic cement usage in
order to improve screws osteosynthesis for osteoporotic bones

The goal of this study is based on fundamental research methods in the

biomechanical field, as torsion and bending tests.
The usage of the bone morphogenetic proteins (BMP), alone or associated with
osteoconductive materials is a very actual subject regarding the bone healing
subject. Our proposed subject is important because it proposes to find an optimal
combination between the osteoconductive and osteoinductive material in order to
promote bone healing.
Bibliografie selectiv
1. Constantz BR, Barr BM, Ison IC, Fulmer MT, Baker J, McKinney L, Goodman SB, Gunasekaren S, Delaney
DC, Ross J, Poser RD. Histological, chemical, and crystallographic analysis of four calcium phosphate cements in
different rabbit osseous sites. J Biomed Mater Res. 1998 Winter;43(4):451-61. PMID: 9855204
2. Feifel H, Gerner A, Schmidt KH, Wimmer F, Schmitz HJ. Die Beeinflussung der Knochenregeneration in
phykogener Hydroxylapatitkeramik durch einen osteoinduktiven Proteinkomplex. Dtsch Z Mund Kiefer
GesichtsChir 19, 25-27 (1995).
3. Frankenburg EP, Goldstein SA, Bauer TW, Harris SA, Poser RD. Biomechanical and histological evaluation of a
calcium phosphate cement. J Bone Joint Surg Am. 1998 Aug;80(8):1112-24. PMID: 9730120
4. Mihil RI, Thurnher M, Kropik K, Moser Doris, Redl H, Tratamentul cu Algipore al defectelor osoase
segmentare
diafizare la iepure, Revista de Ortopedie i Traumatologie (Bucureti), 2005, vol. 15, nr. 3-4, p. 279-284

5. Ohura K, Hamanishi C, Tanaka S, Matsuda N. Healing of segmental bone defects in rats

induced by a beta-TCP
MCPM cement combined with rhBMP-2. J Biomed Mater Res 1999 Feb;44(2):16875. PMID: 10397918 Popescu Negreanu T. Utilizarea biovitroceramicilor i materialelor

compozit pe baz de biovitroceramici colagen n ortopedie, Revista de Ortopedie i

Traumatologie (Bucureti), 2001, vol. 11, nr. 1-2, 3-13

ACTUAL KNOWLEDE STAGE IN THE AREA RELATED TO THE SUBJECT PROPOSED

Autografts, allografts and synthetic bone substitutes are playing an important role in
reconstructive bone surgery and the understanding of their biological effects is requireed for
their coorect usage. Muschler and Lane are proposing the name of bone graft propun
denumirea de material de grefare for any implant that promotes the bone healing by one or
more of the following activities: osteogenesis, osteinduction and osteoconduction.
1. International research goals
1.1. The first research goal is represented by the discovery and usage of bone
subtitutes with osteoconductive and osteoinductive properties
In 1965 Urist reported that the demineralized bone matrix (DBM) induces bone formation
when implanted in the loose extraskeletal tissues; then he isolated from the demineralized
bone matrix a protein called bone morphogenetic protein (BMP) (fig.1).

Fig 1. Molecular structure of the human BMP2. Protein Databank www.rcsb.org

growth factor supergamily called TGF- (transforming growth factors). The most used BMP-s
are BMP-2 (Yang 2006, Patel 2006, Sciadini 2000, Singh 2006) and BMP-7 or the
osteogenic protein (OP-1) (Tsiridis 2006). An important study in the BMP study is
represented by the vehicle through which these proteins are adminstered locaaly. First, a
powder demineralized bone matrix was used. The mainly contains a type I collagenand
represented the golden standard for the comparison with later synthetised materials (Maeda
2004, Kim 2002). Many other biocompatible materials were evaluated as BMP carriers:
-Extracellular matrix components: various collaen types, fibrin, fibronectin, hialuronic acid,
glycosaminoglycans
-Ceramic materials: hydroxiapatitis, calcium triphosphate -Synthetic polymers:
polylactic and polyglycolic acid -Bone grafts: autogenous and allogenous
1.2. The second research direction is represented by the discovery, synthesis and
usage of the osteoconductive bone subtitutes.
Osteoconductive materials gain an increased importance in the biomaterial field, especially
for bone pathology while serving as bone substitutes. These substances has a composition
similar to the bone mineralized matrix and are also biocompatible. Their main function is of
bone support, allowing aposition of the new bone on their surface for this reason they are
used especially in the treatment of bone defects. Recently, they were used as vehicle for the
osteoinductive substances, augmenting the bone forming.

1.2.1. Ceramic materials (phosphocalcic products)

In the past 80 years the calcium phosphates were intensively investigated and used in
bone repair. The most important property of the phosphocalcic compunds (Bohner
2000) is the water solubility, so as a compound is more resorbable as it is water
soluble (e.g. -TCP) when a compound is less soluble in water and in the bone matrix,
it will be less or hard to be resorbed (e.g. HA). The most used compounds in the
medical field are represented by the calcium triphosphates (-TCP), hydroxiapatitis
(HA) (Mihil 2006, Ohura 1996) and calcium biphosphate (BCP or -TCP-HA).
1.2.2. Phosphocalcic cements (CPC)
They were discovered by Brown and Chow in the '80. CPC are composed by one or
more calcium phosphates (CaP) that are solved and precipitated in an aqueous solution.
During precipitation, the calcium phosphate crystals are growing and interweaving so as they
confer mechanical rigidity to the newly formed cement. Many experimental and clinical
studies have used the phosphocalcic cements (Constantz 1998, Frankenburg 1998,

Mihil 2006). By difference to the polymetilmetacrilate (PMMA), that is formed by a

polymerization reaction, the phosphocalcic cements are formed by a less exothermal
reaction while the cement volume remains almost constant during formation Regardless
their type, the phosphocalcic cements shows two big disadvantages by comparison with
porous phosphocalcic ceramics:
-They are fragile materials, and can be used alone (in low mechanical stress regions)
either
combined with osteosynthesis materials (in loading regions, with high mechanical stress)
-The pore dimension is around 1 micron that makes the bone growth difficult inside the
pores
while degradation is proceeding step by step. (outside-in).
2. National research
In Romania, Dr. Popescu-Negreanu and the research group led by Prof. Dr. D. Antonescu in
UMF C. Davila Bucharest have realized a biovitroceramic, commercially called PAW1,
homologated in 1994 and used in many national clinics (Antonescu 1997). Bioactivity tests
were performed by PAW1 implantation, as granules or adzed pieces in the rabbit tibia or the
dog femur. Microscopy studies at the bone implant interface showed a good
osteoconductivity and osseointegration for PAW1. Experimental results were confirmed by
the good clinical results of the PAW 1 in treatment of the tibial plate fractures, plumbing of the
bone defects after benign tumor resection or cyphosis treatment by anterior arthrodesis.
PAW1 showed good results in revision osteosynthesis after hip prostesis and was also
combined with antibiotics to treat osteitis with various localizations: humerus, femur, tibia.
The disadvantages of the auto-, allo- and xenografts used in bone substitution favorized the
development of synthetic bone substitutes. Between these substitutes, the bi- or triphasic
phosphocalcic ceramics plays themain roles. The efficiency of the bone substitutes characterized by
the bone material interaction s bound to the main physical properties and especially to the material
macroporosity (Botez 2002).(Fig.2)
Fig. 2 Material macroporosity and radiologic result of the material application in the spine
For limited indications on small and medium bone defects, if there is a good contact of the
receptor bone and a stable mechanic assembly, the phosphocalcic based and macroporous
bone substitut, with or without embedded antibiotic, represents a viable alternative to auto
and allografts.(Botez 2002)

3. Actual directions

In the past years, the literature is mentioning more and more the guided tissue regeneration
term. This treatment method was successfully used in stomatology, maxillofacial surgery and
experimental repair of the peripheral nerves. It consists in building a physical barrier around
the diseased organ and to isolate it from the surrounding parts, facilitating the healing process.
Using a tubular membrane around a diaphyseal bone defect has the following advantages
(Belloti 2006): 1. Confers a space for bone regeneration, 2. Build a bridge over the defect 3.
Impede the loose tissues to colonize the defect; 4. represent a scaffold for periosteal
regeneration, 5. creates an open space with an increased density of proosteogenic factors that
cannot diffuse in the surrounding tissues. Various membrane types have been used:
biodegradable polylactic (Belloti 2006, Simamora 2006), biodegradable polyurethanic (Chia

2006), expanded polytetrafluoroethylenic (Gore-Tex, non-biodegradable). Actual research is

directed toward the production of macroporous phosphocalcic cements in order to increase the
osteoconductivity and biodegradability without altering the biomechanical properties.
4. Objectives (trends)
Bone defects treatment remains a difficult problem and a challenge for the orthopedic
surgeon. Minor epiphyseal or diaphyseal defects, Defectele minore epifizo-metafizare sau
diafizare i pn la marile pierderi de substan osoas, precum cele rezultate din rezeciile
tumorale, se ntlnesc frecvent n practica medical. Modalitile lor de rezolvare sunt
multiple, variate i relativ complicate. Gama terapeutic se extinde de la clasica autogref
capabil numai de rezolvri limitate , la alogrefe legate i de existena unei costisitoare bnci
de esut osos i pn la heterogrefe (xenogrefe), cu dificulti de preparare, cu riscul unor
transmisii virale sau septice precum i cu pierderea specificitii antigenice. Dei diferitele
cercetri i experiene tiinifice din acest domeniu abund, martor fiind i bibliografia
conex, aceti nlocuitori ai grefelor nu au intrat nc, larg, n practica curent. Scepticismul,
parial ndreptit, este n raport de tolerabilitatea lor, de posibilitatea de producere a unor
relaii chimice cu esutul osos, de depunere de os sau chiar de a realiza o osteoinducie.
Dac pentru tratamentul defectelor osoase mici, situate n zone neportante este suficient
utilizarea numai a substituenilor de os (ceramici sau cimenturi), pentru tratamentul
defectelor osoase mari mai ales situate n zone portante este necesar asocierea unei
osteosinteze solide.
Bibliografie selectiv
1
Antonescu D M, Popescu Negreanu T, Popescu M, Stoica C, Cristea S T, Ursu T. Bioglassceramics as a
substitute for bone grafting, Acta Orthopaedica Helenica, vol. 48 (1997):169-177
2
Bauer T W, Muschler G F. Bone graft materials- An overview of the basic science, Clinical Orthopaedics
and Related Research 371 (2000):10-27 Indexat ISI ISSN: 0009-921X
3
Beloti MM, de Oliveira PT, Gimenes R, Zaghete MA, Bertolini MJ, Rosa AL. In vitro biocompatibility of a
novel membrane of the composite poly(vinylidene-trifluoroethylene)/barium titanate. J. Biomed Mater Res A.
2006,
Indexat ISI ISSN: 1549-3296
1
Bohner M. Calcium orthophosphates in medicine: from ceramics to calcium phosphatecements. Injury
2000 Dec; 31 Suppl 4:37-47. Indexat ISI ISSN: 0020-1383
2
Botez P, Petcu I. Analyse des applications chez lhomme dun substitut osseux a base de ceramique
biphasee (CERAFORM) - Etude prospective preliminaire. 8e Congres de LAOLF - Bucharest, 2002
6. Chia SL, Gorna K, Gogolewski S, Alini M. Biodegradable elastomeric polyurethane membranes as

chondrocyte carriers for cartilage repair. Tissue Eng. 2006 Jul;12(7):1945-53. Indexat ISI ISSN:
1076-3279
1

Constantz BR, Barr BM, Ison IC, Fulmer MT, Baker J, McKinney L, Goodman SB, Gunasekaren S,
Delaney DC, Ross J, Poser RD. Histological, chemical, and crystallographic analysis of four calcium phosphate
cements in different rabbit osseous sites. J Biomed Mater Res. 1998 Winter;43(4):451-61. Indexat ISI ISSN:

1549-3296
2
Feifel H, Gerner A, Schmidt KH, Wimmer F, Schmitz HJ. Die Beeinflussung der Knochenregeneration in
phykogener Hydroxylapatitkeramik durch einen osteoinduktiven Proteinkomplex. Dtsch Z Mund Kiefer
GesichtsChir 19, 25-27 (1995).
3
Frankenburg EP, Goldstein SA, Bauer TW, Harris SA, Poser RD. Biomechanical and histological
evaluation of a calcium phosphate cement. J Bone Joint Surg Am. 1998 Aug;80(8):1112-24. Indexat ISI ISSN:

0301-620X
4
Giardino R, Fini M, Aldini NN, Giavaresi G, Rocca M, Martini L, Zaffe D, Cane V. A reabsorbable tubular
chamber for the treatment of large diaphyseal bone defects. Experimental study in rabbits. Int J Artif Organs 1998
Aug;21(8):473-82. Indexat ISI ISSN: 0391-3988
5
Kim HD, Valentini RF., Retention and activity of BMP-2 in hyaluronic acid-based scaffolds in vitro. J
Biomed Mater Res. 2002 Mar 5;59(3):573-84. Indexat ISI ISSN: 1549-3296
6
Maeda H, Sano A, Fujioka K., Controlled release of rhBMP-2 from collagen minipellet and the
relationship between release profile and ectopic bone formation. Int J Pharm. 2004 May 4; 275(1-2):109-22.
Indexat ISI

ISSN: 0961-7671
1.

Mihil RI, Thurnher M, Schwarz N, Redl H. Studiul histoligic comparative ntre Norian i Biobon n
Substituenii de os n tratamentul defectelor osoase, Mihil RI, Redl H, Antonescu D, Schwarz N, Srbu PD,
Casa de Editur Venus, Iai, 2006 Editura Indexata CNCSIS

2.

3.
4.

Mihil RI, Thurnher M, Kropik K, Moser Doris, Spassova Else, Redl H. Tratamentul cu Algipore al
defectelor osoase segmentare diafizare la iepure n Substituenii de os n tratamentul defectelor osoase,
Mihil RI, Redl H, Antonescu D, Schwarz N, Srbu PD, Casa de Editur Venus, Iai, 2006, Editura
Indexata CNCSIS
Ohura K, Bohner M, Hardouin P, Lemaitre J, Pasquier G, Flautre B. Resorption of, and bone formation from,
new beta-tricalcium phosphate-monocalcium phosphate cements: an in vivo study. J Biomed Mater Res 1996
Feb;30(2):193-200. Indexat ISI ISSN: 1549-3296
Ohura K, Hamanishi C, Tanaka S, Matsuda N. Healing of segmental bone defects in rats induced by a betaTCPMCPM cement combined with rhBMP-2. J Biomed Mater Res 1999 Feb;44(2):168-75. Indexat ISI

ISSN: 15493296
5.
6.
7.

Patel AD. Bone morphogenetic proteins in orthopaedic trauma: recent clinical findings with human bone
morphogenetic protein-2 (rhBMP-2).Curr Med Res Opin. 2006;22 Suppl 1:S1-5. Indexat ISI ISSN: 03007995
Popescu Negreanu T. Utilizarea biovitroceramicilor i materialelor compozit pe baz de biovitroceramici
colagen n ortopedie, Revista de Ortopedie i Traumatologie (Bucureti), 2001, vol. 11, nr. 1-2, 3-13, Indexat
CNCSIS
Sciadini MF, Johnson KD. Evaluation of recombinant human bone morphogenetic protein-2 as a bone-graft
substitute in a canine segmental defect model. J Orthop Res 2000 Mar; 18(2):289-302. Indexat ISI ISSN:

07360266
8.

Singh K, Smucker JD, Boden SD., Use of recombinant human bone morphogenetic protein-2 as an adjunct in
posterolateral lumbar spine fusion: a prospective CT-scan analysis at one and two years. J Spinal Disord
Tech.2006 Aug;19(6):416-23.PMID: 16891977 Indexat ISI ISSN: 1536-0652