Bone grafting procedure involves placement of a bone graft in periodontal bone defect to achieve re-establishment of the lost bone volume. In orthopaedics bone grafts have been used for years. The first recorded bone implant was performed in 1668. It was Hegdus, who in 1923 attempted the use of bone grafts for the reconstruction of bone defects produced by periodontal disease 1. After this, many researchers used different forms of bone grafts in periodontal bone defects. A detailed description of historical aspect of bone grafts is available in “History of periodontal regenerative therapy”.
Before we read in detail about bone grafts, following terms should be clearly understood.
In this case, viable osteoblasts within the grafted material deposit new bone. It occurs in case of autogenous bone grafts.
Osteoconduction occurs when the bone graft material serves as a scaffold for new bone growth that is perpetuated by the native bone.
It is a process in which new bone is induced to form through the action of factors contained within the grafted bone, such as proteins or growth factors.
It occurs when the grafted material does not possess the osteoinductive properties but enhances osteoinduction by promoting bone formation. For example, it has been shown that enamel matrix derivative do not stimulate denovo bone growth alone, but when used with demineralized freeze dried bone allograft (DFDBA), it enhances the osteoinductive effect of DFDBA 2.
Bone fill is the presence of hard tissue in a periodontal osseous defect.
These are bone grafts harvested from patient’s own body. These are considered ideal because they possess osteoconductive and osteoinductive properties and also source of osteoprogenitor cells.
The isograft is material that is taken from one individual and transplanted into another genetically identical individual, such as an identical twin. In isograft cases donor and the recipient must have the same genotype.
Allografts are derived from other individuals of the same species. Examples include freeze-dried bone allografts (FDBA) and demineralized freeze-dried bone allograft (DFDBA). These are most commonly used grafting materials.
Xenografts are derived from nonhuman animal sources such as bovine. The anorganic bovine bone is chemically treated to remove its organic components,leavinga trabecular and porous architecture similar to human bone.
Alloplasts are synthetic bone substitutes. These are made from hydroxylapatite, a naturally occurring mineral that is also the main mineral component of bone. Bioactive glasses are commonly used alloplasts.
Ideal properties of a bone graft:
There are some basic requirements for a bone grafting material to be designated as ideal. Every grafting material has its own advantages and disadvantages. The ideal grafting material should possess the following properties 3,
- Should be biocompatible and non immunogenic
- Should have no risk of disease transmission
- Should be physiologically stable
- Should be osteoconductive
- Should be osteoinductive
- Should have the ability to osteointegrate i.e. the ability to chemically bond to the surface of bone without an intervening layer of fibrous tissue
- Should induce osteogenesis
- Should have minimal post operative sequelae
- Should be cost effective
- Should be acceptable by the patient
Autografts are considered to be closest to an ideal grafting material because they possess all the above stated properties. Allografts have osteointegrative and osteoconductive properties and may also exhibit osteoinductive potential, but they are not osteogenic because they do not contain viable cells. Alloplasts or synthetic graft materials possess only osteointegrative and osteoconductive properties. In the following diagram the essential elements for new bone formation have been elucidated,
Requirments for new bone formation
Classification of bone grafts:
Based on the source of procurement, grafts have been divided into following categories 4,
- Autogenous bone grafts
- Xenogenic bone grafts
- Synthetic/Alloplastic bone grafts
- Composite grafts
Following flow chart describes classification of various bone grafts on the basis of their sources of procurement,
Classification of bone grafts
Autogenous Bone Graft:
These bone grafts are obtained from the same individual on whom the bone grafting has to be performed. They may be obtained from a local remote location. As they possess most of the properties of an ideal graft material, they are considered the gold standard bone replacement graft. The graft obtained has viable cells, which participate in new bone formation hence has osteogenic properties. Major advantages of using autogenous grafts are: these grafts are osteogenic and no risk of disease transmission. Major disadvantages of autografts include procurement morbidity, limited availability in case of intraoral sites and high cost of procedure in case of extraoral graft harvesting.
Autografts are subdivided in three groups: cancellous, cortical and or corticocancellous autografts. The cancellous bone grafts is derived from cancellous part of bone. Because of the cancellous nature of the graft, most number of viable cells are available in this type of autograft. Although, after transplantation majority of cells present in the graft die as result of ischemia but mesenchymal stem cells present in the bone marrow which are resistant to ischemia, may survive the grafting procedure. On the other hand, cortical autografts act as osteoconductive graft providing support for bone formation. Because of presence of thick cortical bone, the revascularization and integration of cortical bone grafts is slow. The cortico-cancellous graft offers stability and osteogenic capacity. It can be used in weight-loaded areas. As in case of cortical grafts, vascularisation is slow in these grafts.
Intraoral autogenous bone grafts can be harvested from the edentulous alveolar areas, maxillary tuberosity, healing bony wound, extraction sites and mental and retromolar areas 5.
Cortical bone chips:
The use of cortical bone chips can be traced back to the work of Nabers and O’Leary (1965) 6. They used shavings of cortical bone removed by chisel during osteoplasty and ostectomy, in periodontal bone defects. This type of autogenous bone graft did not get much popularity and is rarely used today because of large size (1,559.6 × 183 μm) of the graft particles, which have more vulnerability for sequestration 7.
The rationale behind use of osseous coagulum and bone blend was that smaller the size of graft particles, more are the chances for its resorption and replacement with host bone 8. The bone particles were harvested by using round burs, and were then mixed with blood. The rationale for mixing blood with graft particles was to provide osteogenic progenitor cells and morphogens to the wound site and to promote a stable clot formation. Studies on monkey model demonstrated that a small particle size (100 μm) led to earlier and higher osteogenic activity than did the larger particles 9. The major disadvantage of this procedure is difficult graft collection procedure and inability to aspirate during graft collection. Another problem is unknown quantity of the bone fragments collected and material fluidity.
Bone blend procedure involves collection of cortical or cancellous bone with the help of trephine or rongeurs. Then it is put in an amalgam capsule and triturated to achieve the consistency of a slushy osseous mass. The resultant particle size is in the range of 210 x 105 μm 7. Research reports have shown clinically significant bone fill in areas treated with bone blend 8, 10.
Newer techniques for bone graft collection:
The graft material collected by osseous coagulum or bone blend techniques is sufficient for small regenerative procedures. A recent technique for collecting the bone graft by using bone collectors was introduced by Blay et al 11. The bone particles are collected with the help of bone filters. Bacterial contamination during graft collection is one major problem during this procedure. So, a stringent aspiration protocol, preoperative oral chlorhexidine rinse and antibiotic prophylaxis should be applied.
More recently, piezoelectric device (Piezosurgery) has been introduced for different bone augmentation procedures. This device used for ultrasonic cutting of bone create microvibrations that are caused by the piezoelectric effect which was first described by French physicists Jean and Marie Curie, in 1880. The frequency used to cut the bone is 25–29 kHz because the micromovements that are created at this frequency (ranging between 60 to 21 μm) cut only mineralised tissue. The neurovascular other soft tissues are cut at frequencies higher than 50 kHz 12-14. Irrespective of the method used, all the mechanical techniques have some amount of detrimental effects on viability of the cells. Keeping all the biological factors into consideration the bone harvesting procedure should be carefully performed.
Extraoral bone graft harvesting is a popular procedure, especially in cases where large amount of bone graft is required. Many authors believe that extraoral cancellous bone and marrow grafts have greatest potential for new bone formation 15-17. Most suitable site for extraoral bone graft harvesting is iliac crest. According to Rosen et al. (2000), autografts from iliac cancellous bone and marrow provide a great osteogenic potential because they have been shown to induce cementogenesis, bone regeneration and Sharpey’s fibers reattachment 18. Major disadvantages of extraoral iliac graft include: postoperative complications, expense, time, and extra surgical procedure involved in procuring these grafts. These all factors make autogenous iliac crest grafts a less than desirable alternative.
In majority of cases large amount of bone graft is not required, making intraoral bone graft harvesting more preferable.
An allograft is a graft obtained from genetically dissimilar members of the same species. Allografts are obtained under sterile conditions from fresh cadavers, usually within 24 hours of the death of the donor. Their major advantages are unlimited availability and osteoinductive potential comparable with autogenous bone. These are available primarily in two forms: freeze dried bone allograft (FDBA) and decalcified freeze dried bone allograft (DFDBA).
So, what is the need for decalcify / demineralize a cortical bone allograft ?
Demineralization of the cortical bone allograft improves the osteoinductive potential by exposing bone morphogenic proteins, and other inductive factors known to increase bone formation. This is why; FDBA provides an osteoconductive scaffold and elicits resorption when implanted in mesenchymal tissues whereas DFDBA also provides an osteoconductive surface along with osteoconductive scaffold. There are set standards for procuring, processing and sterilization of allografts (FDBA/DFDBA). Most of the bone banks adhere to the guidelines of the American Association of Tissue Banks (AATB) as described by Centers for Disease Control and Prevention 19. According to AATB, allografts should not be collected if,
- Donor is from high-risk groups, as determined by medical testing and behavioral risk assessments.
- Donors has been tested positive for HIV antibody by ELISA
- Autopsy of donor reveals occult disease
- Donor bone has been tested positive for bacterial contamination
- Donor and bone has been tested positive for hepatitis B surface antigen (HBsAG) or hepatitis C virus (HCV)
- Donor has been tested positive for syphilis
Processing of allografts:
Although allograft manufacturing companies do not disclose the exact method of bone processing they follow, however following is the description of basic technique of bone processing,
The first and foremost step is obtaining the bone from a suitable donor and reducing it to bone pieces of small size of approximately 5 mm. Second step is elimination of bone marrow and cellular debris. The elimination of bone marrow and cellular debris is achieved with fluids and detergents, which, by their clearing effect, improve the osteoconductive capacity of the bone. Pressurization allows full penetration of inactivating or eliminating agents into the bone. Various chemical solutions used in this procedure include saline, acetone, ethanol or hydrogen peroxide which remove bioburden and reduce antigenicity. Then bone particles are treated with antimicrobial, antimycotic and antifungal solutions. After this, bone particles are kept in liquid nitrogen at a temperature as low as −80°C. Then the bone particles are freeze dried. Freeze-drying has a logistical advantage in that it allows further storage of the tissue at room temperature. Bone particles are treated with repetitive solvent washes to eliminate moisture content. After this, bone particles are further reduced in size ranging between approximately 250 and 750 μm. The processing for freeze dried bone graft (FDBA) ends here and packaging of the graft in sterile containers follows. At the end, low-dose of γ radiation at low temperatures is applied on the graft to ensure sterility.
For preparing decalcified freeze dried bone graft (DFDBA), the decalcification process follows after final particle size of 250 to 750 μm is achieved. The bone particles are immersed in a hydrochloric acid bath at concentrations ranging from 0.5 to 0.6 N for various lengths of time. Then these acid treated particles are immersed in a buffering solution to remove residual acid. The demineralized allograft is further rinsed with various solutions (e.g., distilled water) to remove residual buffer solution. Then graft is packed in sterile containers followed by irradiation of low-dose of γ radiations at low temperatures to ensure sterility.
Bone graft processing results in exponential reduction in graft contamination and disease transfer, or both. Sterility assurance level (SAL) is a term used to describe the probability that an item will not be sterile after it has been subjected to a validated sterilization process 20. With proper processing, allografts for dental purposes routinely achieve sterility assurance level (SAL) of 10-6. In other words we can say that the odds of an organism’s surviving after allograft processing are lessthan one in 1 million 21.
After processing bone allograft has to undergo certain tests which include:
Visual inspection test:
Visual detection is done for problems such as gross graft contamination, packaging defects and product mislabeling.
Residual moisture test:
Testing of freeze-dried allografts is done to ensure residual moisture is 6 percent or less.
Residual calcium test:
Testing of demineralized freeze-dried bone allograft is done to ensure residual calcium content is 8% or less.
Commercially available allografts:
This is commercially available allograft obtained from human source. It is subjected to patented tutoplast process which gently removes the undesirable components of the bone including fats, cells, antigens, and inactivates pathogens, while preserving the valuable minerals and collagen matrix, leading to complete and rapid bone regeneration 22. Various chemicals are used to eliminate these unwanted components including acetone to remove lipids, H2O2 to oxidize remaining proteins, solvent dehydration with acetone to preserve the collagenous fiber structure, osmotic treatment and low dose gamma irradiation. It is claimed by the manufacturers that this procedure yields a bone graft with nicely preserved trabecular pattern of bone and mineral structure, as compared to freeze-drying process which results in a more osteoconductive material.
This allograft is derived from cadaver long bones. The procured bone is subjected to chemical processing under aseptic conditions to remove unwanted bone components such as lipid, blood, and cellular components before it is frozen. The resultant bone is subjected to reduction in particle size and stored in sterile containers. The graft is combined with a glycerol carrier to stabilize the proteins and improve the graft handling 23.
Grafton DBM (BioHorizons, Birmingham, Alabama)
As already stated, xenografts are grafts transferred between genetically dissimilar members of different species. These are derived from three sources: bovine bone, porcine bone and natural coral. These are osteoconductive, biocompatible and similar to human bone in structure 24, 25.
Bovine derived Xenografts:
Bovine derived Xenografts are processed by removing all the organic content of the bone to minimize any chances of graft rejection. Procedure involves application of various chemicals on the bone. The raw bone is subjected to boiling in alkalis (such as potassium hydroxide) followed by maceration in hydrogen peroxide and ethylenediamine. The resultant graft has a hydroxyappatite skeleton with porous internal structure with large surface area, which facilitates revascularization and integration into the host bone 26, 27. A porous structure also facilitates cell mediated resorption thereby its replacement with newly formed bone. As compared to synthetic bone substitutes, xenografts can also be used as structural component similar to human bone which makes them a better osteoconductive bone substitute.
Commercially available bovine derived xenografts:
This is a bovine derived bone grafting material. The unwanted organic material within the bone is removed by chemical processing at low temperature resulting in a porous mineral matrix. Processing at low temperature has an advantage of preservation of natural architecture of bone. This graft material is available in cancellous and cortical granules and blocks. The particle size of the graft is approximately 100 × 200 × 500 Å.
Bio-Oss ® (Osteohealth Co., Shirley, NY)
This graft has inorganic and organic components consisting of Bio-Oss Spongiosa granules (0.25–1 mm) and 10% highly purified porcine collagen. The addition of collagen in graft material improves handling characteristics of the graft without acting as a barrier. The graft particles are adhered to each other due to presence of collagen which facilitates graft placement without membrane. The collagen component is resorbed within 4-6 weeks.
OsteoGraf / N ® (CeraMed Dental, LLC, Lakewood, CO):
It is a natural anorganic bovine-derived microporous hydroxylapatite. It is the only xenograft that meets all ASTM (American Society for Testing and Materials) standards for “Composition of Anorganic Bone for Surgical Implants (F1581-95). It achieves a hydrophilic – cohesive consistency when hydrated. It remodels to vital bone at the same rate as host bone. It is manufactured as radiopaque, rounded particles and is available in two particle sizes:
- OsteoGraf / N-300 (250–420 μm) packaged in 1-g and 3-g vials
- OsteoGraf / N-700 (420–1,000 μm) packaged in 1-g and 3-g vials
OsteoGraf / N-300
PepGen P-15 ® (Dentsply Friadent, Mannheim, Germany):
It is a tissue-engineered bone replacement graft material which mimics the inorganic and organic components of autogenous bone. It consists of a uniquely designed P-15 peptide, a synthetic biomimetic of the 15 amino acid sequence of Type-I collagen, which is uniquely involved in the binding of cells, particularly fibroblasts and osteoblasts. It is available as granules with particle size range: PepGen P-15 particulate (250-420 microns) packaged in 1-gram and 3-gram vials. It is also available as flowable material PepGen P-15 FLOW which is PepGen P-15 particulate suspended in inert biocompatible hydrogel packaged in 0.5 cc and 1.0 cc syringes.
Porcine derived Xenografts:
Xenografts derived from porcine cortical and cancellous bone have also been developed to be used as bone substitutes.
It is commercially available xenograft of porcine origin. It is a heterologous cortico-cancellous collagenated bone mix. It must always be hydrated before use. It is thoroughly mixed with a few drops of sterile physiological solution to activate its collagen matrix and to enhance its adhesivity. It can also be mixed either with OsteoBiol Gel or with patient’s blood. A major advantage of this material is that it can act as carrier for various therapeutic agents and drugs. The collagen present in this bio-material facilitates blood clotting and the subsequent invasion of repairing and regenerative cells, thus favouring bone formation. It also provides a cohesive environment for graft particle. This material has been claimed to have high osteoconductive activity. The particle size of the graft is 250-1,000 μm.
Coralline calcium carbonate:
These are derived from natural coral. The structural features of commercially used coral, Porites, are similar to that of cancellous bone, which makes them a suitable bone substitute.
It is a commercially available grafting material derived from natural coral which has a calcium carbonate structure composed primarily of aragonite. It has a porous structure with pore size of 100 to 200 μm. This porous structure provides a large surface area helping in a better cellular resorption and replacement by natural bone. Its osteoconductive properties are comparable to other bone substitutes.
Various commercially available bone substitutes
Type of substitute
Commercial name and brand
|Allograft||Human bone||Puros (Zimmer Dental, Carlsbad, California)
Grafton DBM (BioHorizons, Birmingham, Alabama)
|Xenograft||Bovine derived||Bio-Oss ® (Osteohealth Co., Shirley, NY)
Bio-Oss Collagen ® (Osteohealth Co., Shirley, NY)
OsteoGraf/N ® (CeraMed Dental, LLC, Lakewood, CO)
PepGen P-15 ® (Dentsply Friadent, Mannheim, Germany)
|Porcine derived||OsteoBiol ® Gen-Os|
|Coralline calcium carbonate||Biocoral ® (Inoteb,Saint Gonnery, France)|
|Alloplast||Polycrystalline ceramic form of pure densely sintered hydroxylapatite||Calcitek, Inc., Carlbad, CA.
OsteoGraf / D300 and OsteoGraf / D700 (CeraMed Corp., Lakewood, CO)
|Coralline porous non-resorbable hydroxylapatite||Interpore 200 (Interpore International, Irvine, CA).
Pro-Osteon 500R (Interpore Cross International, Irvine, CA, USA).
|Resorbable nonceramic hydroxylapatite||OsteoGen ® (Impladent, Holliswood, NY).
OsteoGraf / LD-300 ® (CeraMed Corp., Lakewood, CO).
Cerabone ® (Coripharm GmbH & Co. KG, Dieburg, Germany).
|Nanocrystalline hydroxyapatite (NHA)||Ostim™(Heraeus Kulzer, Hanau, Germany).|
|Fluorohydroxyapatitic (FHA) biomaterials||FRIOS ® Algipore ® (Friadent GmbH, Mannheim, Germany)|
|β-Tricalcium Phosphate (TCP)||Bioresorb ® ( Sybron Implant Solutions GmbH Bremen, Germany).
Cerasorb ® (Curasan, Kleinostheim Germany).
Vitoss ® (Orthovita, Malvern, PA, USA).
|Biphasic alloplastic materials||Calcitec ® Inc. (Austin, TX).
Osteogen ® (Impladent Ltd, Holliswood, NY).
Tricos ® (Baxter, Bern, Switzerland).
Osteon TM (Genoss Co. Ltd., Suwon, Korea).
Bone Ceramic ® (Straumann, Basel, Switzerland).
Ceraform ® (Teknimed SA, Vic-en Bigorre, France).
MBCP+™(Biomatlante biologics solutions).
|Calcium Phosphate Cements (CPC)||Norian ® PDC™ (Shofu Inc., Kyoto, Japan).
Augmentech AT (Wetzlar, Germany).
Calcibon ® (Biomet).
|Bioactive Glasses||PerioGlas ® (Block Drug Co., NJ, USA).
PerioGlas ® Plus(Block Drug Co., NJ, USA).
Unigraft ® (Unicare Biomedical Inc., Laguna Hills, CA, USA).
Biogran™ (Orthovita Inc., Malvern, PA, USA).
|Biocompatible osteoconductive polymers||HTR TM Synthetic Bone (Bioplant, Norwalk, CT)|
|Composite Grafts||Ceramics and bioactive molecules||Healos ® (Orquest, Mountain View, CA).
Collagraft ® (Zimmer Corp, Warsaw, IN).
Tricos ® (Baxter BioSciences BioSurgery).
Alloplasts are synthetic bone substitutes that are readily available and also eliminate the need for a patient donor site. Like other bone grafting materials, the ideal properties of alloplastic materials include: it should be biocompatible with host tissues, non-antigenic, non-carcinogenic, and non-inflammatory. Along with this, the graft material,
- Should be sufficiently porous and should allow tissues to grow into and around the implant (osteoconduction),
- Should be able to stimulate bone induction, should be resorbable and replaceable by bone,
- Should be radio-opaque in order to be visualized radiographically,
- Should be able to withstand sterilization without losing favourable qualities,
- Should be stable in varying temperatures and humidity,
- Should have surface electrical activity (i.e., be charged negatively),
- Should be Hydrophilic,
- Should be easy to manipulate clinically
- Should have high compressive strength
- Should be inexpensive, and easily attainable.
Various synthetic graft materials include hydroxyapatite, tricalcium phosphate, calcium sulfate (plaster of Paris), bio-active glasses, and hard tissue replacement polymers. Following is a detailed description of these materials.
Hydroxyapatite (Ca4 (PO4)6(OH) 2):
Hydroxyapatite (HA) is basic component of inorganic components of bone. Hydroxyapatite ceramics have a stoichiometry similar to that of bone mineral 28, 29. It can be manufactured from natural reef building coral skeleton by a “hydrothermal exchange reaction”, where the trabecular, bone imitating structure of the coral remains unchanged and the calcium carbonate (CC) skeleton is converted to calcium phosphate, the main inorganic salt of bone 30. The main difference between calcium carbonate and hydroxyappatite is markedly slow resorption of hydroxyapatite as compared to calcium carbonate. Biodegradation should not occur before the graft implant has filled with bone. The slow resorption of hydroxyapatite allows formation of new bone in the graft due to its porosity. HA lacks capacity to induce bone growth.
Other methods of manufacturing porous HA include, homogenizing calcium phosphate powder with appropriately sized naphthalene particles, resulting in macroporous material after the naphtalen has been removed. The resultant structure is sintered at high temperature (1100-1300°C). Another method utilizes hydrogen peroxide to generate a pore-filled structure. Porous HA is brittle and can be used only in non-loading sites. Its compressive strength is enhanced by bone ingrowth but it is only comparable to that of cancellous bone 31.
Depending upon method of procurement and processing, HA grafts are available in following forms,
Polycrystalline ceramic form of pure densely sintered HA:
It is a polycrystalline form of HA prepared in relatively large particle size (18–40 mesh). Calcitite, a commercially available HA graft ((Calcitek, Inc., Carlbad, CA), has particle size of 420-840 μm. OsteoGraf / D300 and OsteoGraf / D700 ((CeraMed Corp., Lakewood, CO), have particle sizes 250–420 μm and 420–1,000 μm respectively.
Coralline porous non-resorbable hydroxylapatite:
As already stated they are derived from marine coral skeleton from which organic components have been removed. Interpore 200 (Interpore International, Irvine, CA) and Pro-Osteon 500R (Interpore Cross International, Irvine, CA, USA) are examples of commercially available coralline porous non-resorbable hydroxylapatite.
Resorbable nonceramic hydroxylapatite:
It is a resorbable form of HA which is highly microporous and non-sintered (nonceramic). Particle size varies from 300–400 μm. Commercially available examples of this type of HA grafts include OsteoGen ® (Impladent, Holliswood, NY) which is a low-temperature hydroxylapatite (HA) material. It is manufactured by processing at low temperature and is not sintered. These non sintered precipitated particles have size measuring from 300-400 μm. Other examples include OsteoGraf/LD-300 ® (particles are sized between 250 and 420 μm) (CeraMed Corp., Lakewood, CO) and Cerabone ® (Coripharm GmbH & Co. KG, Dieburg, Germany).
Nanocrystalline hydroxyapatite (NHA):
As the name indicates, nanocrystalline hydroxyapatite (NHA) bone graft has nano sized particles. The main advantage of this biomaterial is improved osteoconductive properties and complete resorption of the material within 12 weeks. Commercially available example of this graft is Ostim™ (Heraeus Kulzer, Hanau, Germany) (NHA), which is a synthetic nanocrystalline hydroxyapatite (NHA) paste containing 65% water and 35% nanostructured apatite particles.
Fluorohydroxyapatitic (FHA) biomaterials:
Historically, it was observed that some forms of calcified algae have internal structure similar to cancellous bone 32. So, its potential use as bone substitute was proposed.
It is commercially available fluorohydroxyapatitic (FHA) bone graft. It is derived from calcifying marine algae (Corallina officinalis). This material has got microperforations which are interconnected. The average diameter of the pores is around 10 μm and every pore is limited by one layer of small FHA crystallites with a size of 25–35 nm. The material has good osteoconductivity.
β-Tricalcium Phosphate (TCP):
Tricalcium phosphate is present in two forms: α and β. The α-tricalcium phosphate is monoclinic and consists of columns of cations, while the beta tricalcium phosphate has a rhombohedral structure. The physical properties of these two forms are different. α- TCP is less stable than β- TCP and when mixed with water it forms calcium-deficient hydroxyapatite. β- TCP has physical structure more close to bone but its compressive strength is almost 1/20th of cortical bone. The osteoconductive properties of this grafting material have been shown to be good 33, 34. After placement in the bone defect this material is slowly resorbed and replaced by natural bone which is facilitated by the porous structure of the material. The exact mechanism of bone graft resorption is still under investigation. Because of the absence of osteoclasts around the material in a rabbit experiment, it was proposed that the material is lost mainly by dissolution in biological liquids 35. Some other researchers propose a cell mediated bioresorption because of presence of osteoclast like giant cells 36, 37.
Commercially available β-Tricalcium Phosphate (TCP):
Various commercially available β-Tricalcium Phosphate (TCP) graft materials include:
It is a β-Tricalcium phosphate bone grafting material having a porous structure similar to human bone. The manufacturers claim a micropore sizes range from 0.5 – 10μm, and the macropore sizes are 50 – 700μm which provides a highly porous structure new bone formation and slow graft resorption. The major advantage of using pure β-Tricalcium Phosphate (TCP) is its high osteoconductivity but at the same time it not as stable as hydroxyapatite and is resorbed at a faster rate.
Bioresorb®( Sybron Implant Solutions GmbH Bremen, Germany)
This synthetic graft material has particle size of granules between 150-500 µm. It is a pure-phase β-TCP. It is completely resorbed in 4-12 months and its porous scaffold during that time helps the natural bone formation to occur. It does not contain HA, so its resorption occurs completely, uniformly and parallel to the formation of the surrounding bone. The granules are radio-opaque, resorbing gradually from the inside out, so clinicians can easily monitor the status of grafted site.
It is a β-tricalcium phosphate synthetic bone substitute. The material is is 3-dimensionally macroporous, containing spaces into which bone ingrowth takes place. It does not have significant compressive strength by itself.
Biphasic alloplastic materials:
Various bone grafts have been developed by combining of hydroxyapatite (HA) and tricalcium phosphate. This combination has been proposed to have better osteoconductive properties as compared to individual material. In these materials the quantity of HA is higher than TCP because it has been suggested that higher HA ratio shows accelerated new bone formation 38. Various commercially available biphasic alloplastic materials include,
Calcitec ® Inc. (Austin, TX):
This synthetic bone graft contains hydroxyapatite (HA) and tricalcium phosphate crystals and is claimed to have high osteoconductive properties as claimed by manufacturers.
It is a biphasic alloplast containing synthetic bioactive resorbable crystals and crystal clusters in the range of 300 to 400 μm which are osteoconductive, highly hydrophilic, and resorbable. The graft material has highly porous crystalline clusters which act as a slowly-resorbing matrix, permitting the infiltration of bone-forming cells and the subsequent deposition of host bone.
Tricos ® (Baxter, Bern, Switzerland):
This synthetic bone graft contains 60% HA and 40% β-TCP. The β-TCP is quickly resorbed and has a high osteogenetic potential and constitutes a source of P+ and Ca++ ions whereas hydroxyapatite is slowly resorbed and ensures the long-term stability of the material.
Osteon TM (Genoss Co. Ltd., Suwon, Korea):
It is composed of 70% HA and 30% β-tricalcium phosphate (β -TCP). Material porosity is in the range of 300–500 nm.
Bone Ceramic ® (Straumann, Basel, Switzerland):
It is composed of 60% hydroxyapatite (100% crystalline), and 40% β TCP (particulate form). The graft material porosity is in the range of 100–500 μm.
Ceraform ® (Teknimed SA, Vic-en Bigorre, France):
It is composed of 65% HA and 35% TCP. The material has a mean granular diameter is between 900 and 1,200 μm and is available in block or granular form.
MBCP+™ (Biomatlante biologics solutions):
MBCP is a Micro Macroporous Resorable Biphasic Calcium Phosphate bone substitute. It is a 3D interconnected scaffold of bone graft with 20% HA and 80% β-TCP.
Calcium Phosphate Cements (CPC):
Calcium phosphate cement (CPC) utilizes two phases of components, one in powder form and another in liquid form. Powder is usually a mix of different calcium phosphate salts where almost all products contain one or several of the components including: amorphous calcium, phosphate (ACP), dicalcium phosphate dihydrate (DCPD), dicalcium phosphate anhydrous (DCPA), α-tricalcium phosphate (α-TCP), dicalcium phosphate (DCP), tetra-calcium phosphate (TTCP), monocalcium phosphate monohydrate (MCPM) and calcium carbonate (CC). The liquid component usually contains sodium phosphate solution. When mixed, the material sets to form a solid mass. After mixing the material, a workable consistency of the material is achieved and the bone defect is filled and contoured. The porosity of the set mass can vary between 30% and 50%, depending on the processing conditions, e.g. liquid-to-powder ratio. The setting, rheological and mechanical properties of the CPC can be adjusted by modifying different factors, such as the chemical composition of reactants, particle size or presence of nucleating agents 39-42. Various studies have shown that CPC are highly biocompatible and osteoconductive materials, which can stimulate tissue regeneration 43-45. These cements can also be used as carriers for drugs and other mediators like growth factors which promote bone formation. A large-scale, prospective, blinded, and randomized controlled clinical trial study demonstrated that the use of rh PDGF-BB + β-TCP was safe and effective in the treatment of periodontal osseous defects 46.
The problem associated with these materials is prolonged setting time and the inability to set in the presence of blood. To overcome these problems new improved cements have been developed. Following are the commercially available calcium phosphate cements used in periodontal bone defects,
Norian ® PDC™ (Shofu Inc., Kyoto, Japan):
It is an injectable, moldable, fast setting, bioabsorbable calcium phosphate cement having high compressive strength (55 MPa). The powder of the cement consists of a-tricalcium phosphate (a Ca3[PO4]2), monocalcium phosphate monohydrate (Ca[H2PO4]2·H2O) and calcium carbonate (CaCO3). The liquid contains a solution of sodium phosphate. The pore diameter of the set mass is around 300 Å.
Augmentech AT (Wetzlar, Germany):
This is also a moldable, fast setting, bioabsorbable cement having high compressive strength. The powder of the mix contains tricalcium phosphate (TCP), magnesium phosphate, magnesium hydrogen phosphate and strontium carbonate. The liquid is a watery solution of diammonium hydrogen phosphate.
Calcibon ® (Biomet):
It is a synthetic, biodegradable, calcium phosphate based bone substitute. The powder of the mix contains Powder: α-TCP (61%), DCP (26%), CaCO3(10%), and PHA (3%). Solution contains H2O and Na2HPO4. The set material has got high compressive strength (up to 60 MPa). Compressive strength increases during setting of the material. After 6 hours it is comparable to cancellous bone. The final compressive strength is reached after 3 days.
These are silicate based alloplasts containing calcium and phosphate. Hench was the first one to develop bioactive glass that could bind to tissue 47. The formation of carbonated hydroxyapatite layer on its surface when placed in body fluids make this material bioactive. The FDA approved composition of bioactive glass is designated as 45S5. The 45S5 bioactive glass is composed of Si02 (46.1 mol%),CaO (26.9 mol%), Na20 (24.4 mol%) and P205 (2.6 mol%) 48. The high amounts of Na2O and CaO, as well as the relatively high CaO/P2O5 ratio make the glass surface highly reactive in physiological environments 49. 45S5 is able to form HCAP (hydroxycarbonated apatite) in less than 2 hours and binds to tissues.
The bioactive glasses are obtained basically by two processes: melting and sol-gel process. Sol-gel process is a preferred process because it requires lower temperature as compared to the conventional melting process. It has also been reported that glasses made from sol-gel technique have increased bioactivity 50.
There are two types of responses generated when alloplasts is implanted in body: biochemical and cellular. Biochemical response is generated due to change in environment around alloplast. When bioactive glasses are placed in vivo, a highly basic environment with pH around 10 is created, because of which a layer rich in silica gel is formed on the surface of glass particles. Because of interaction with surrounding fluid, a layer of calcium phosphate is formed which is made up of hydroxycarbonate apatite (HCA). Hydroxycarbonate apatite is chemically and structurally very similar to composition of bone. The cellular reactions include colonization, proliferation and differentiation of relevant (bone) cells 49, 51.
Flow chart showing various interfacial reactions involved in forming a bond between bone and a bioactive glass 52
With development in materials over years, new bioactive glass compositions have been introduced. These materials contain no sodium or have additional elements incorporated in the silicate network such as fluorine 53, magnesium 54, 55, strontium 56-58, iron 59, silver 60, 61, boron 62-64 , potassium 65 or zinc 66, 67.
Bioactive glass posses superior mechanical strength compared with calcium phosphate products, as a result of a strong graft-bone bonding 68.
Commercially available bioactive glass alloplasts:
PerioGlas ® (Block Drug Co., NJ, USA):
PerioGlas is a synthetic absorbable osteoconductive bone graft substitute composed of a calcium phospho-silicate bioactive glass, Bioglass. This graft material is available for use since 1995. The particle size of the graft particles varies from 90-710 μm. It is supplied sterile, packaged either in a Tyvek-sealed PET-G cup or in a filled syringe within a second sterile barrier package. While using, the graft is mixed with sterile water, saline, the patient’s own blood or marrow or with autogenous or allograft bone to form a wet sandy paste that is applied to the defect 69.
PerioGlas®(Block Drug Co., NJ, USA)
PerioGlas ® Plus(Block Drug Co., NJ, USA):
It is composed of a calcium phosphosilicate material and a calcium sulfate binder. The inorganic calcium and phosphorous components are thermally incorporated in a sodium silicate network (PerioGlas®) designed specifically for its absorbability and osteoconductive nature. The calcium sulphate incorporated in the graft material binds the bioactive glass particles and is absorbed with due course of time. The calcium sulfate in the PerioGlas® Plus is absorbed between 4 and 8 weeks after implantation, depending on the graft site, size and material used. The remaining PerioGlas® particle left behind are also absorbed and slowly replaced with host bone usually within six months 70.
Unigraft ® (Unicare Biomedical Inc., Laguna Hills, CA, USA):
Unigraft is made of synthetic bioactive glass material. The graft material is composed of fused oxides of calcium, phosphorus, silicon and sodium. Upon implantation, the material begins to dissolve by gradually releasing a steady stream of Ca and P ions, along with soluble silica into the bony defect. This increased concentration of local bone mineral ions has been demonstrated to enhance bone regeneration and exhibit an anti-bacterial effect. The particle size of the graft material varies from 200 μm to about 420 μm. It is supplied sterile in foil-sealed polyolefin vial. The graft material is mixed with sterile saline or with patient’s blood to form a sandy paste that is to be applied to the defect 71.
Biogran™ (Orthovita Inc., Malvern, PA, USA):
Biogran is a synthetic bone graft material consisting of 300-355 μm diameter bioactive glass granules. It is composed of bioactive salts of Si, Ca, Na, P. The graft material can be mixed with the patient’s blood or sterile saline within the disposable dappen dish cup and subsequently delivered to the defect site. With due course of time, it is slowly resorbed and replaced by host bone.
Biocompatible osteoconductive polymers:
These are nonresorbable, particulate of calcium layered with polymethylmethacrylate and hydroxyethylmethacrylate (PMMA-PHEMA) 72. The polymers used as bone substitutes can be classified as natural and synthetic polymers which can further be divided as degradable and nondegradable 73. Present biocompatible polymers contain calcium hydroxide and polymethylmethacrylate (PMMA) and polyhydroxylethylmethacrylate (PHEMA). Commercially available polymer grafts is HTR TM Synthetic Bone (Bioplant, Norwalk, CT). This composite is prepared from a core of PMMA and PHEMA with a coating of calcium hydroxide 74. The graft material is highly porous with 150–350 μm interled pore size. Being polymers, these grafts are hydrophobic with a negative surface charge (−8 to −10 mV), which is believed to impede development of infection 75. The hydrophobic nature of the material enhances clotting and negative particle surface charge aids in adherence to bone 25. Various studies have provided evidence of bone formation around graft particles 76-78.
These are newly emerging grafting materials. Composite grafts combine scaffolding properties with biological elements to stimulate cell proliferation and differentiation and eventually osteogenesis. These contain a synthetic osteoconductive matrix and osteogenic cells and growth factors which make the graft osteoinductive. Because this combination provides a scaffold as well as molecules which have osteogenic properties, these graft material become a close replacement of autogenous bone grafts. The osteoconductive matrix becomes a carrier for the bioactive molecules. The potential combinations of graft materials for making a composite graft include: bone marrow/synthetic composites, ultraporous β-TCP/BMA composite, osteoinductive growth factors and synthetic composites, BMP/polyglycolic acid polymer composites and BMA/BMP/poly-glycolic acid polymer composite 79.
Commercially available composite grafts:
Healos ® (Orquest, Mountain View, CA):
Healos® is a matrix of bovine fibrillar Type I collagen coated with hydroxyapatite mineral, which constitutes approximately 25% of the implant by weight. It can be mixed with bone marrow aspirate to provide osteogenic and osteoinductive potential. Another addition to this bone graft is MP52, which is a member of the BMP family. Addition of this protein further increases the osteogenic potential of this bone graft.
Collagraft ® (Zimmer Corp, Warsaw, IN):
Collagraft is a composite of suspended fibrillar collagen and a porous calcium phosphate ceramic, in a ratio of 1:1. The fibrillar collagen is highly purified collagen obtained from bovine dermis. Autologous bone marrow aspirate can be added to these materials or it can be mixed with autologous bone as a bone graft extender. It does not offer structural support by itself and its movement may be difficult to control 80, 81.
Tricos ® (Baxter BioSciences BioSurgery):
Tricos® is a combination of hydroxyapatite-coated beta tricalcium phosphate (HA/TCP) granules and a fibrin matrix. This bioactive material provides a three-dimensional osteoconductive environment for the formation of new bone.
Collagen based delivery systems in tissue engineering:
The biocompatible properties of collagen have resulted in the use of collagen as a matrix or scaffold for tissue regeneration. Collagen has been used as a carrier for various therapeutic agents and bioactive molecules. These include antibiotics, steroids, anticoagulants, antineoplastics, immunosuppressants, growth factors, cytokines and gene therapeutics (e.g., plasmid DNA) 82. The main aim of using a carrier is to deliver the drug or bioactive molecule in a sufficient concentration for appropriate time duration at site where its desired effect is required. Collagen matrices are available in the form of films, sheets, wafers, discs, gels, sponges, 3D scaffolds, and nanofibers (alone and in combination with a plethora of natural and synthetic fibers as well as ceramics).
Another major field of research is culturing specific cell types directly on the collagen-based matrix prior to in vivo application. Delivering these specific cells to the site promotes healing. Various aspects of tissue engineering have been discussed in “Tissue engineering in periodontics”.
Factors affecting success of bone grafts:
Defect size and topography:
It is a very important factor which affects the success of bone graft. The predictability of graft success generally increases as the number of remaining bony walls increases. In other words we can say the graft success is most predictable with three wall and least predictable with one wall defect. A deep narrow defect is more predictable for regeneration than a shallow and wide defect 83, 84.
Presence of infection:
Presence of infection at the grafting site is a major factor responsible for graft failure. Under low pH conditions, bone and graft material are rapidly absorbed through solution mediated resorption. Therefore, it is important to eliminate all reasons for inflammation before placement of bone graft.
The graft material should be stable at its position to facilitate a proper biological response during healing. An improperly placed graft material with vulnerability to movement is bound to fail.
The area in which regeneration has to take place requires space maintenance. If the graft material resorbs too quickly, there is no sufficient time for new bone formation and the defect gets filled with connective tissue rather than new bone 85, 86.
The healing period varies from defect to defect. For smaller three wall defects healing period may be smaller as compared to large defect in which large amount on bone graft is placed with less autogenous bone and fewer remaining walls. An adequate healing period must be given for regeneration to take place 85, 87.
Adequate blood supply:
For proper healing in area where bone graft has been placed, an adequate blood supply is of paramount importance. The blood supply in this area is derived from two sources: the cortical or cancellous bone and soft tissue covering the defect. The cortical bone has few arterioles as compared to cancellous bone 87, 88.
Primary closure of the soft tissue at the operated site is paramount for graft success. The opening of the incision line is one of the most common complications during post-operative healing. As a result of this, the graft material may be lost, contaminated and vascularisation is delayed causing graft failure.
Regional acceleratory phenomenon (RAP):
It is a local response to injury; in which healing takes place at a faster rate as compared to normal regeneration process 87. It is basically attributed to increase in the concentration of chemical mediators like growth factors, which may enhance the healing process to almost 10 times that normal 89. RAP begins few days after the injury, reaches its peak at one to two months, lasts four months in bone and totally subsides within 6-24 months.
Affect of growth factors:
Presence of growth factors is required for regeneration to take place. These growth factors are derived from various cellular sources like platelets, macrophages and other cells involved in local inflammatory response. various growth factors involves are: epidermal growth factor, fibroblast growth factor, insulin-like growth factor, keratinocyte growth factor, platelet derived growth factor, transforming growth factor, vascular endothelial growth factor etc. A detailed description of these factors is available in “Growth factors in periodontal regeneration”.
Particle size of graft material:
The acceptable size of graft particles ranges from 125-1000 μm. A minimum space of 100 μm is required between the graft particle to allow vascularisation and bone formation. A particle size less than 100 μm elicits macrophage resorption of the graft particles causing their early loss 7. Most commercially available grafts have particle size ranging from 250-750 μm.
Systemic conditions like diabetes mellitus, hyperparathyroidism, thyrotoxicosis, osteoporosis, Paget’s disease etc. have adverse affects on healing of the graft.
Smoking and alcohol also adversely affect the healing of the graft.
Bone substitutes and their application in periodontal regenerative therapy is presently a major field of research in periodontology. In the above discussion, we have discussed in detail various bone grafts and their sources in detail. Although autogenous bone graft is closest to ideal bone graft but it requires extra surgical site to harvest the graft. The allografts provide an alternative to autograft but are not as osteoinductive as autografts are. Synthetic bone substitutes such as various forms of hydroxylapatite, β-Tricalcium Phosphate (TCP), biphasic alloplastic materials, calcium phosphate cements (CPC) and bioactive glasses have provided us a good alternative to autogenous bone grafts. As these materials have a composition which is very similar to bone, they get resorbed and replaced by host bone in due course of time.
The future of bone graft materials are composite bone grafts, which have combined properties of scaffold and biologically active molecules such as bone morphogenetic proteins/growth factors. In other words we can say that the graft material acts as a carrier for biologically active molecules. Incorporation of collagen matrix into these biomaterials makes them ideal for carrying these biologically active molecules. Although the research on these materials is in its initial stage, they may be the closest replacement of autografts. The future directions in bone grafts and other aspects of periodontal regeneration have been discussed in “Tissue engineering in periodontics”.
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