Alveolar ridge augmentation in rats by combined hydroxylapatite and osteoinductive material ELSE MARIE PTNHOLT''-\ GISLE BANG^ AND HANS REIDAR HAANAES-' ^ Institute for Surgical Research, Rikshospilalet, Urtiver.sity of Oslo, 'Department of Oral Pathology and Forensic Dentistry, University of Bergen, ''Department of Oral Surgery and Oral Medicine, Dental Faculty, University of Osto, Oslo, Norway

Pinholt EM, Bang G, Haanaes HR: Alveolar ridge augmentalion in rats by combined hydroxylapatite and o.stcoinduciivc mtiterial. Scand J Dent Res 1991; 99: 64- 74. Abstract The purpo.se of ihis study vi'as to examine if increased bony ingrowth developed when hydroxylapalite was implanted together with an osteoinductive substrate. Dense hydroxylapatite grantilcs (HA) (Caleitile, Caleitek, San Diego, GA, USA) were mixed with equal volumes of allogenif, demineralized, Iyophilized dentin or bone and implanted subperiosteally for alveolar ridge atigmciitatioii purposes and heterotopically in the abdominal muscles in rats. Light microseopic evaluation revealed tliat HA was surrounded by fibrous connective tissue containing foreign body giant cells and il had neither an osteoinductive nor an osteoconductive effect. The nevvh' lormed bone induced from the implanted allogenic, demineralized, Iyophilized dentin or bone was never fotmd in close contad with the H.A, and did not incorporate the implant. Key words: alveolar ridge augmentation: hydroxylapalite; jaws; osteoinductive material. I'^. M. Pinholt. Instilute for Stirgical Research, Rikshospitalet, University of Oslo, N-0027 Oslo I, Norway. .Aceeijted for ptiblieatioti 10 Jttne 1990.

CalcititTt ph(),s|5halc implant materials arc al present the nio.st biocompatible s\'nthetie bone stibstituies a\ailable (1), although tlieir bony incorporation lor alveolar ridge augmentation purposes i.s incomplete (2-9). Both tricaleiittn pho.sphate (TCP) Ca,, (PO.|).,, and hydroxylapatite (HA) Cam (PO,|),;(OH).j have demonstrated histoeompatibility and osteoconductive properties (10-16) but otily H.\ has a calcium to phosphate ratio (1.67) identical with that of hu-

man vertebral bone and tootli mineral and thi.s seem.s to be of importanee for its osteotropic beliavior (17). Mi.xing HA with aulogenou.s bone ha.s been reported to increase bony ingrowth (18). It is, however, desirable to avoid this additional harvesting procedure. Therefore calcium phosphate implants have been mixed with osteoconductive and osteoinductive substrates. It has been shown that allogenic, demineralized bone and dcntin are osteoinductive (19-21). Bone mor-

ALVEOLAR RIDGE AUGMENTATION phogenetic protein (BMP) has been determined as the local stimulating factor initiating the cascade of events during bone induction (22). Mesenchymal cells are transformed into cartilage and/or bone-forming celis by the BMP (22), which is concentrated especially in the cortical layers of bone and in dentin (22). Osteoconduction is a healing process where the implanted material acts as a scaffold which is gradually resorbed during creeping substitution and the healing proceeds with bone ingrowth from the adjacent living bone (13). Good results were obtained when apatite was mixed with purified collagen (23, 24) and when the osteoconductive TCP was substituted with osteoinductive material (25, 26). Demineralized, allogenic bone was reported to increase new bone formation in combination with different implants (27, 28) including dense HA (29), Osteoinductive materials therefore seem useful in replacing autogenous bone to enhance bony ingrowth. The aim of the present study was to evaluate if enhanced bony ingrowth develops around dense hydroxylapatite (HA) granules when mixed with allogenic, demineralized, lyophiiized dentin or bone.

Material and methods Wistar male rats weighing 310 g (SD 10) were used for the experimental (Group 1 = 10 animals), control (Group 11= 10 animals), and donor (eight animals) groups. They were fed a standard diet (Standard diet lor rats, Rikshospitalet, The National Hospital, Oslo, Norway) and given water ad libitum. Incisors and long bones from aJlogenic rats were harvested, cleaned, demineraiized in 0.2 N HCI at 4°G for 48 h, lyopbilized and sterilized in cthylene oxide gas for 3 h as described previously (30, 3]). Hydroxylapatite implants were dense granules, size 20/40 mesh, Calcitite (Calcitek Company, San Diego, CA, USA), In Group I, implantations were performed in the maxilla and in the abdominal muscles. The surgery was done under general anesthesia.

65

Hypnorm/Dormicum 0.15 ml/100 g intramuscularly (hHypnorm +l:Stcrile water mixed with l:Dormicum + I:Sterile water). On the premaxilla, where the horizontal plate of the hard palate eontinues into the vertical lateral plate between the incisor and the first molar, an incision was made to bony contact. Subperioslea! dissection was carried out to create a small pocket on the lateral aspect of the premaxilla. Alveolar ridge augmentation was performed with a syringe implanting equal volumes of dense HA granules and granules of allogenic, demineralized, lyophiiized bone on tbe right side. The surgical site was closed with Prolene monofi! suture. The same procedure was performed on the left side where equal volumes of dense HA granules and granules of allogenie, demineralized, ]yophiH7,ed dentin were deposited, A midline incision was carried out to the rectus abdominus muscle and blunt dissection aJong the musele sheet was performed on each side of the midline, A small poeket in the rectus abdominus muscle was created andon the right side equal volumes of dense HA granules and granules of allogenic, demineralized, lyophiiized bone were placed. On the left side the same procedure was performed and dense HA granules mixed with equal volumes of ailogenic, demineralized, and lyophiiized dentin were implanted. The pockets were closed with Prolene monoill suture. An additional intramuscular pocket was created on the right side and granules of dense HA only were implanted. The midline ineision was elosed with 2-0 Dexon. In Group II the same surgieal procedures were carried out as sham operations without any implants. To observe proper wound healing, clinical evaluations were performed daily thefirstthree postoperative days and thereafter on a weekly basis to evaluate development of infections or weight loss. After 4 wk the animals were killed. The premaxilla and the abdominal muscles were immediately dissected from the animal, tagged for orientation purposes and the specimens fixed in 4% neutral formalin. The specimens were radiographed for orientation of possible hard tissue formation and then demineralized in 17% formic acid, dehydrated, and embedded in paraffin wax. Serial sections were eut at 5 nm and stained with Harris hematoxyiin and Results For unknown reasons one rat in Group I lost 100 g in weight the first postoperative week

66

PINHOLT ET AL.

F/^'. /. A, ral alveolar ridge (AR) augmentation by allogenie, demineralized bone (DB) and dense hydroxylapatite granules (HA) (revealed as empty spaees due to demineralization). Most of the implanted bone is resorbed and has induced a eonsiderable amount of new bone formation, both in eontinuity wilh host bone (A) and separately in soft tissue (•). HA granules are surrounded by fibrous tissue with foreign body giant eells and have no osteoinduedve or osteoeonductive effeet. H & E, x 65. B, Etilarged part from A showing implanted allogenic, demineralized bone (DB) indueing new bone formation, •^. The HA granules are surrounded by fibrous tissue and have no osteoinductive or osteoeonduetive effect. H & E, x 165.

ALVEOLAR RIDGE AUGMENTATION and was subsequently killed. In Group II one rat died immediately postoperatively and therefore each of the two groups comprised nine animals. These animals gained weight during the 4 wk of observation. No clinical infections were observed at any time. In Group I al! implants were recovered. Hydroxylapatite did not induce bone formation either on the maxilla or in the abdominal muscles (Figs. 1-5). No osteoconduction was seen in connection with the HA on the maxilla (Figs. 1, 4). Both on the maxilla and in the abdominal muscle fibrous connective tissue with foreign body giant cells separated the HA granules from the implanted demincraiized bone and dentin including the induced newly formed bone (Figs, i—5). Both allogenic, demineralized bone and dentin induced new bone formation and exhibited bone conduction in a high percentage of the eases (Table 1, Figs. 1,

2, 4, 5). No inflammatory or foreign body reaction was observed around the demineralized bone or dentin. The sham operations in Group II induced no new bone formation in any ofthe animals either in the maxilla or in the abdomen. Discussion In the present study bone incorporation around dense HA granules was not enhanced by simultaneous implantation of osteoinductive material in adult rats. The method of healing of calcium phosphates has been discussed widely in the past and the reported results are quite divergent. Where there is no demand for mechanical strength, porous HA can offer a good functional structure with integration of implant and host bone {32-35). In the alveolar ridge ofthe maxillae stress-

Fig. 2, RcmnanLs of aliogenic, demineralized bone (DB) and hydroxyiapatite granules (HA) in rat abdominal muscle (M). Non-vital, cell-free bone implant has induced hcterolopic osteogenesis (NB) while hydroxyiapatite granules have not. The laLter are surrounded by afibi^ouscapsule containing foreign body giant cells. H & E, x 100.

68

PINHOLT ET AL.

Fig. 3. Dense hydroxylapatite granules (HA) implanted in rat abdominal muscle (M). HA granules are surrounded by fibrous lissue containing foreign body cells. No heterotopic new bone formation is seen. H & E, X 70.

es are mainly compressive and dense calcium phosphates appear to be better suited as an implant material (1, 36, 37). These products are synthetic anorganic materials which are expected to cause tissue reactions similar to previously tested anorganic implants (9, 38). Anorganic material is knovyn to resorb slowly and incompletely and heal by fibrous encap.sulation (38). Similar findings were observed in our study where the HA was surrounded by fibrous connective tissue with multinucleated giant cells. The newly formed bone induced from the implanted allogenic, demineralized dentin or bone was never found in close contact with the HA. Positive results of increased incorporation of dense HA granules have been obtained in investigations using autogenous bone (2, 18). It is, however, desirable to avoid the additional harvesting

procedure of bone, and both within orthopedic and maxillofacial surgery searches have been performed to obtain a substitute for that purpose. Purified bovine collagen was used (23, 24, 39) in combination with apatite crystals and the mixture, called Collapat, was claimed to be osteoinductive. The substrate was tested in animal experiments (39) and was later used in patients (24, 39). The main indication for its use was deep bony cavities following tumor resection and defects after autogenous bone harvesting. When used for spondyloidesis, fractures, or pseudarthrosis it was necessary to add autogeneous bone to the implant. No foreign body reaction to the hydroxylapatite was reported and osteoinduction was recognized from collagen as well as from apatite (39). Since Collapat has no mechanical stability it was not recom-

ALVEOLAR RIDGE AUGMENTATION

69

fj^. 4. A, rat alveolar ridge (AR) augmcnUlion by allogenic, demineralized dentin (DD) and dense hydroxylapatite granules (HA). E, gingival epithelium. Implanted dentJn has induced new bone Ibrmation, [A.) both in continuity with host alveolar ridge (AR), in excavation chambers in the implani, • ) and separately at a distance from ho,st bone (•), HA granules are sun'ounded by fibrous tissue with foreign body giant cells and have no osteoinductive or osteoconductive effect. H & E, x 30, B, enlarged pan from A showing implanted aliogenic, demineraiized dentin (DD) inducing new bone formation (A) both in continuity with host alveolar ridge (AR), in excavation chambers in implant ( • ) , and separately at a distanee from host bone (•). HA granules are surrounded by fibrous tissue containing foreign body giant cells and have no osteoinduetive or osteoconductive effect. H & E, x 70.

70

PINHOLT ET AL.

Fig. 5. Remnants of allogenic, demineralized dentin (DD) and hydroxyiapatite granules (HA) in rat abdominal muscle (M). Implanted dentin has induced heterotopic new bone formation (NB) and cartilage (C). HA granule.s are surrounded by fibrous tissue containing foreign body giant cells, and have not resulted in any osteoinductive activity. H & E, x 70.

mended for u.se in maxillofacial surgery (23). It is debatable wiietiier collagen stimulates bone growth. In vitro investigations (40) revealed that eollagen direetly stimulates eell growth of fibroblasts. In the same study it was shown that soluble unsintered HA had a cytotoxie effeet on the fibroblasts (40). Ani-

Table 1 .Nttmber of implattls ittdttcing new bone formation

Type and place of implant HA, jaw HA, adb. muscle Bone, jaw Bone, abd. muscle Dentin, ]aw Dentin, abd. muscle Sham operation, jaw Sham oper., abd. muscle

No. inducing bone formation (%) 0/9 (0%) 0/9 (0%) 8/9 (90%) 9/9 (100%) 8/9 (90%) 8/9 (90%) 0/18 (0%) 0/18 (0%)

mal experiments on rats (41, 42) showed that neither collagen nor collagen combined with tricalcium phosphate or hydroxyiapatite resulted in heterotopic new bone formation (41). Foreign body reaction was observed around Collapat (42) and SCHWARZ (42) concluded that Collapat was osteoconductive. This was in contrast to the conclusions by MiTTELMEiER & KATTHAGEN (23, 24) and KATTHAGEN & MiTTELMEiER (39). In oral and maxillofacial surgery purified bovine collagen has been used in combination with particulate hydroxyiapatite (43). In this investigation collagen mainly had a carrier function to facilitate handling ofthe implant. The compaction of the implant in this clinical study was confirmed from animal studies (44) to be due to replacement ofthe collagen component by dense fibrous connective tissue. In search of other materials suitable for

ALVEOLAR RIDGE AUGMENTATION enhancement of bone incorporation of alioplastic substrates, the osteoinductive principle has been investigated (25, 26, 42, 46). Bone morphogenetic protein (BMP) wa.s bound to calcium phosphate ceramics implanted in calvarial defects in dogs (25, 26), and tricaicium phosphate (TCP) was used alone and in combination with BMP. The investigators found that TCP combined with BMP promoted bony ingrowth and host bed regeneration of the skull defects more completely than TCP aionc (25, 26). The TCP, an osteoconductive substrate, acted as a carrier for the osteoinductive BMP. Unfortunately the TCP was absorbed incompletely during the observation period. These fmdings were confirmed by the studies of KAWAMURA el al. (45). Bone matrix morphogenetic proteins were bound to hydroxyiapatite (BMP-HA) in in vitro cxperimcnls of rat mcsenchymal muscle cell explants and in in vivo studies in muscle pouches of mice and bone marrow cavities of rabbit femora and tibial condyies. Undifferentiated mesenchymal ceils migrated poorly into the pores of the BMP-HA. Foreign body reaetion was observed inside the pores. Therefore it was concluded that the environment in the apatite internal pores was not suitable for the interaction between eells and BMP. Other osteoinduetive substrates have been used together with different implants to enhance bony ingrowth (27, 28). Polysulfone implants were impregnated with demineralized allogenic bone and implanted into the abdominal muscles of rats (27). In spite of foreign body reaction around the aHoplastic material new bone formation around the allogenie material was not inhibited. These results are comparable with the ones from the present study. KOEHLER & KREICBERGS (28) reimplanted bone supplemented with allogenic demineralized bone matrix in ulnar defects in rabbits. Better incorporation witii demineralized allogenie bone was observed than without the combination.

Corraline implants have been implanted in combination with freeze-dried bone (46) in alveolar intrabony defects of dogs. Periodontal ligament formation was observed from the corraline implant (46) with new bone formation and fsbrovaseular tissue penetration into the porous implant. Around the freeze-dried bone, whieh was only superficially demineralized, a connective tissue encapsulation was recognized. This is in accordance with the results of the studies by HOLMES & HAGLER (35). He showed that corraiine HA without bone substitutes was invaded by bone. Up to 61% of the surface of the implant was infiltrated by bone as a result of osteoconduction. Allogenic, demineralized bone was interpreted to induee new bone when implanted in combination with corraline hydroxylapatite (47). In this study it was concluded that the aHoplastic implant acted as a scaffold for heterotopie bone formation in the thoracic muscles of primates. This finding is in contrast to the findings of EIAWAMURA ^t al. (45). AspENBERG et al. (48) observed failure of bone induction by bone matrix in primates. The allogenic bone matrix in the monkeys was treated in exactly the same way as allogenic bone matrix of rats. He therefore concluded that the monkey was not a suitable laboratory' animal for osteoinductive purposes. Ailogenic, demineralized bone matrix mixed with dense HA was implanted in dogs to enhance the bony incorporation (29). The bony ingrowth was delayed compared to that obtained from autogenous bone mixed with dense HA in the same dogs. The difference was not statistically significant, possibly due to too small an animal sample. Intereonnecting bone between demineraiizcd, allogenic bone and HA granules was observed. This finding is in contrast to the findings in our study since new bone induced from aliogenie bone or dentin did not connect to HA. The allopiastie material in our investigation was surrounded by fibrous eonnective tissue sepa-

72

PINHOLT ET AL.

rating the HA froni the newly formed bone. Implantation of calcium phosphates is implantation of mineral-containing substrates. BANG (49) found that the bone induction process is retarded when dentin is implanted undemineralized and REDDI & HUGGINS (50) concluded that fibroblasts will not transform when mineral is present. Therefore osteoinduetion is not expected from HA. Bone ingrowth around this material is formed by osteointegration by the host bone and only where the bone is able to grow (9). HA is classified as a bioactive material (13, 51) and physicochemical characterization of the new material associated with HA ceramics implanted in nonosseous sites (52) showed an organic and inorganic phase close to that of bone. However, histologic and morphologic characterizations did not fit with that of true bone. It was concluded from the present investigation that dense granules of HA do not show osteoinductive or osteoconductive properties and that dense granules of HA are not incorporated into new hone formed by osteoinduction. References I.JARCI-IO M. Calcium phosphate ceramics as hard tissue prosthetics. Clin Orlliop 1981; 157: 259-78. 2. KENI JN, QLIINNJH, ZIDE MF, GUERRA LR,

responses to hydroxyiapatite particles of different shapes. J Oral Maxillofac Surg 1984; 42: 150-60. . DROBECK HP, ROTHSTEIN SS, GUMAER KI, SHERER AD, SLIGHTER RG. Histologic obser-

vation of soft tissue responses to implanted, multifaceted particles and discs of hydroxyiapatite. J Oral Maxillofac Surg 1984; 42: 143-9. . PiNHOLT E M . Effects of ionizing radiation on interfaces of hydroxyiapatite implanted in canine man-

dibles. Thesis, University of Minnesota, Minneapolis, 1988; 1-47. . DoNATH K. Histopathologische Befunde von Hydroxylapatitkeramiken im Kieferbereich. In: WATZEK G V, MATEJKA M , eds. Der zahnlose Unterkiefer. Seine chirurgisch-prothetische Re-

habilitation. Vienna: Springer, 1988; 17785. . KALLENBERGER A. Die Wirkung von Biokeramik (Kalziumphosphat keramik) auf kultivierte Kaninchenfibroblasten. S S 0 1978; 88; 90 9. . KoESTER K, KARBE E, KRAMER H , HEIDE

H, KoENiG R. Experimenteller Knochenersatz durch resorbierbare Calciumphosphat-Keramik. Langenbecks Arch Chir 1976; 341: 7786. 12 . KLEIN CPAT, GROOT K DE, DRIESSEN AA, LuBBE HBM VAN DER. Interaction of biodegradable beta-whitlockite ceramics with bone tissue: an in vivo study. Biomaterial.s 1985: 6: 189-92. BucHoi.z RW, CARLION A, HOLMES RE. Hy-

droxyapatite and tricalcium phosphate bone graft substitutes. Orlliop Clin North Am 1987; 18: 323-34.

BoYNE PJ. Alveolar ridge augmentation using 14 DENISSEN HW, GROOT K D, MAKKES PC, nonresorbable hydroxyiapatite with or withHooFF A V D, KLOPPER PJ. Tissue response out autogenous cancellous bone. J Oral Maxilto dense apatite implants in rats. J Biomed lofac Sttrg 1983; 41: 629 42. Mater Res 1980, 14: 713 21. 3. BLOCK MS, KENTJN. Long-term radiograph15 K.A.TO K , AoKi H , ENGt-{ D, T A B ATA T, O G O S I ic evaluation of hydroxylapatite-augmented M. Biocompatibility of apatite ceramics in mandibular alveolar ridges. J Oral Ma.xi.llofac mandibles. Biomater Med Devices Arlif Organs Surg 1984; 42: 793 6. 1979; 7: 291-7. 4. RoTiisTEiN SS, PARIS D. SAGE B. L'se ofdura-

16 JARCHO M , KAY JF, GUMAER KI, DOREMUS

patite for the rehabilitation of resorbed alveoRH, DROBECK HP. Tissue, cellular and subcellar ridges. J Am Denl .A.ssoc 1984; 109: 571 4. lular events at a bone-ceramic hydroxyiapatite 5. CHANG CS, MATUKAS VJ, LEMONS JE. Histointerlace. J Bioeiig 1977; 1: 79 92. logic study ol hydroxyiapatite as an implant 17. OsBORN JF, NEWESELY H . The material scimaterial for mandibular augmentation. J Oral ence of calcium phosphate ceramics. BiomateriMaxilltfac Surg 1983; 41: 729-37. als 1980; 1: 108 11. 6. MisiEK DJ, KENT JN, CARR RF. Soft tissue 18. BLOCK MS, K E N T J N . Healing of mandibular

ALVEOLAR RIDGE AUGMENTATION ridge augmentations using hydroxylapatite with and without autogenous bone in dogs. J

73

model in guinea pigs. Scand J Dent Res 1973; 81: 240-50.

31. PiNHOLT EM, BANG G , HAANAES HR. AlveoOral Maxiltofac Surg 1985; 43: 3-7. lar ridge augmentation by osteoinduction in 19. URIST MR. Bone histogenesis and morphorats. Scand J Dent Res: in press. genesis in implants of demineralized enamel 32. WHITE E , SHORS EG. Biomaterial aspects of and dentin. J Oral Surg 1971; 29: 88-102. Interpore-200 porous hydroxyapatite. Dent 20. PuTTE K VAN DE, URIST MR. Osteogenesis Clin .North Am 1986; 30: 49-67. in the interior of intramuscular implants of decalcified bone matrix. Clin Orthop 1965; 43: 33. HENCH L L , WILSON J . Surface-active biomaterials. Science 1984; 226: 630-6. 257-'7O. 34. KLAWITTER JJ, HuLBERT SF. Application of 21. BANG G , URIST MR. Bone induction in excaporous ceramics for the attachment of load vation chambers in matrix of decalcified denbearing orthopedie applications. J Biomed Matin. Arch Surg 1967; 94: 781-9. ler Res 1971; 2: 161 (only). 22. URIST MR, NILSSON OS, HUDAK R , et al. Im-

munologie evidence of a bone morphogenetic protein in the milieu interieur. Ann Biol Clin (Pans) 1985; 43: 755-66. 23. MiTTELMEiER H, KATTHAGEN BD. Klinische Erfarungen mit Gollagen-Apatit-Implantation zur lokalen Knochenregeneration. Z OrIhop 1983; 121: 115 23. 24. MiTTELMEiER H, KATTHAGEN BD. Neue Wege des Knochencrsalzes. Orthop Praxis 1984; 5: 389 98. 25.

26.

27.

28.

35. HOLMES RE, HAGLER H K . Porous hydroxyl-

apatite as a bone graft substitute in mandibular contour augmentation: a histiometric study. ,7 Oral Maxillofac Surg 1987, 45: 421~ 9. 36. JARGHO M . Biomaterial aspects of calcium phosphates. Properties and applications. Dent Clin .North Am 1986; 30: 25-47. 37. JASTY V, JARGHO M , GUMAER KI, SAUER-

SCHEI.L R, DROBECK HP. Bone tissue response to dense hydroxylapatite disc implants in URIST MR, LIETZE A, DAWSON E . Beta-tricalmongrel dogs: a light and eleetron tnicroscopic cium phosphate delivery system for bone morstudy, vol II. Ninth International Congress on phogenetic protein. Clin Orlhop 1984; 187: Electron Microscopic Study, Toronto, 277 80. 1978. URIST MR, NILSSON O , RASMUSSEN J, et al. 38. HJ6RTING-H.\NSEN E . Studies on implantation of Bone regeneration under the influence of a anorganic bone. Thesis, University of Copenhabone morphogenetic protein (BMP) beta trigen, 1970; 1- 198. calcium phosphate (TCP) composite in skull 39. KATTHAGEN BD, MITTELMEIER H . Experimentrephine defects in dogs. Clin Orlhop 1987; 214: tal animal investigation of bone regeneration 295-304. with collagen-apatite. Arch Orthop Trauma Surg V.\N DER STEENHOVEN JJ, SPECTOR M . O S 1984; 103: 291 302. teoinduction within porous polysulfone im40. JOOS U V, MiTTERMAYER C, SCHILLI VV, K A plants at extraosseous sites using demineralDEN A. Der Einfluss von Kollagen, Hydroxylaized allogeneic bone matrix. J Biomed Mater patit und Gelastypt S auf die Zellkultur. Dtsch Res 1983; 17: 793-806. Zahnaerzll Z 1982; 37: 22 4. KoEHLER P, KREICBERGS A. Incorporation of 41. R U E G E R J M , SIEBERT HR, WAGNER K J , PANautoclaved autogeneic bone supplemented NiKE A. Synthetischc und biologische Knowith allogeneic demineralized bone matrix. chenersatzmittel. Tierexperimentelle UntersuAn exprimeiUal study in the rabbit. Clin Orthop chungen der osteoinduktiven Eigen.schall. CItir 1987; 218: 247 58. Forum 1984; 223 7.

29. Bi.oGK MS, KENT JN, ARDOIN RC, DAVEN-

POR'i' VV. Mandibular augmentation in dogs with hydroxylapatite combined wilh detnincralized bone. J Oral Maxillofac Surg 1987; 45: 414--20. lO. BANG G . Induction ol hctcrotopic bone (brmation by demineralized dentin: an experimental

42. ScHWARz N, REDL H , SGHLAG G , et al. Experi-

mental osteoinduction in rats. Collagen-apatile versus osteogenin-containing gelatine. Arch Orthop Trauma Surg 1987; 106: 113 9. 43. MEHLISH D R , TAYLOR TD, LEIBOLD DG, et

at. Evaluation of collagen/hydroxylapalite for augmenting deficient alveolar ridges: a pre-

74

PINHOLT ET AL, liminary report, J Oral Maxillofac Surg 1987, 45: 408-13,

44, HATCHER JE,

LEMONS JE,

MATUKAS

VJ,

CoMPTON RC, Laboratory animal studies of apatite and collagen biomaterials, J Denl Res 1986; 65: 347 (Abstract), 45, KAWAMURA M , IWATA H , SATO K , MIURA T,

Chondroosteogenetic response to crude bone matrix proteins bound to hydroxyapatite, Clin Orlhop 1987; 217: 281-92, 46, WEST TL, BRUSTEIN DD, Freezc-dried bone and corraline implants compared in the dog, J Periodonlol 1985; 56: 348 51, 47, EL DEEB MN,

HOSNY M , SHARAWY M ,

Os-

teogenesis in composite grafts of allogenic demineralized bone powder and porous hydroxylapatite, ,7 Oral Maxillofac Surg 1989; 47: 50-6, 48, ASPENBERG P, LoHMANDER L S , T H O R N G R E N

KG, Failure of bone induction by bone matrix in adult monkeys, J Bone Joint Surg (Br) 1988, 70-B: 625-7, 49, BANG G, Induction of heterotopic bone formation by demineralized dentin in rats and guinea pigs, Scand J Dent Res 1973; 81: 230-9, 50, REDDI A H , HUGGINS CB, Influence of geometry of transplanted tooth and bone on transformation of fibroblasts, Proc Soc Exp Biol Med 1973; 143: 634-7, 51, GROOT K DE, EfTect of porosity and physicochemical properties on the stability, resorption, and strength of calcium phosphate ceramics, Ann .A'' 1^ Acad Sci 1988; 523: 227-33, 52, HEUGHEBAERT M , LEGEROS R Z , GINESTE M ,

GuiLHEM A, BoNEL G, Physicochemical characterization of deposits associated with hydroxylapatite ceramics implanted in nonosseous sites, J Biomed Maler Res 1988; 22: 257-68,