Bone Grafts and Implants

Introduction

Application of a freshly harvested cancellous bone autograft is considered to be the “gold standard” treatment for promoting the healing of bony defects. In spite of this, cancellous bone grafting is one of the most underutilized procedures in veterinary orthopedics.

Cancellous bone grafts benefit healing by three different mechanisms: 1) the matrix of cancellous bone contains bone morphogenetic proteins that stimulate uncommitted mesenchymal cells to differentiate into bone forming cells at the injury site (osteoinduction); 2) the trabecular surfaces of the cancellous bone graft allow invasion of the graft by newly formed blood vessels and provide a surface or scaffold upon which new bone can be deposited (osteoconduction); and 3) bone forming cells in the graft which survive transplantation may participate directly in early formation of bony callus (osteogenesis). The delicate trabecular structure of cancellous bone does not enable it to bear weight or support fixation devices (structural support function).

Clinical Indications for Cancellous Bone Grafting

Specific clinical indications for cancellous bone grafting fall into two major categories: 1) conditions where bone healing requires enhancement (nonunions, potential nonunions, highly comminuted fractures, and osteotomy procedures that depend upon rapid new bone formation for a successful outcome – e.g. triple pelvic osteotomy); and 2) treatment of bone loss secondary to trauma (high energy open fractures), disease (osteomyelitis and bone cysts), or surgical resection (arthrodeses, empty holes in bone after bone plate removal, and limb salvage procedures in patients with bone neoplasia).

Nonunions

Most nonunion fractures seen in dogs and cats are hypertrophic and are attributable to inadequate stabilization. Application of appropriate internal or external fixation improves the mechanical environment such that many of these fractures will heal successfully without bone grafting. Avascular nonunions, however, do require bone grafting. The sclerotic ends of such fractures are debrided back to bleeding bone using either a bone curette or rongeurs, and a Steinmenn pin, K-wire, or drill bit is used to reopen the medullary cavity. Fibrous tissue at the fracture site should also be resected. A liberal amount of freshly harvested cancellous autograft should be packed into the resulting defect to enhance bone healing.

Potential Nonunions

Decreased vascular density in the distal radius of small breed dogs has been implicated in the higher frequency of nonunion seen in these patients. Treatment usually involves open reduction, fixation with a bone plate and screws, and application of canellous bone autograft to the fracture region.

Comminuted Fractures

Highly comminuted fractures have avascular fragments that have a tendency to be resorbed. Additionally, fixation devices applied to such fractures must function in buttress mode, thus provision of optimal stability can be challenging. Vigorous early production of bridging callus stimulated by liberal application of cancellous autograft over the fracture region is often critical to a successful outcome in these cases.

Osteotomy Procedures

Rapid formation of bridging callus is critical to a successful outcome in canine patients that have undergone triple pelvic osteotomy (TPO). As most TPO patients are young dogs with relatively soft bone, pullout of fixation screws and loss of reduction can occur with repetitive loading. Liberal application of cancellous bone graft at the ilial osteotomy site facilitates rapid production of bridging callus thus promoting successful healing before implant loosening can occur. Corticocancellous bone removed from the pubis and the triangular piece of bone removed from the dorsal aspect of the caudal ilial bone segment can be cut into small pieces and mixed with the graft as a cancellous bone extender.

High Energy Open Fractures

Bone loss is a common feature of high energy open fractures, especially those accompanied by significant soft tissue shearing injuries. Initial treatment includes wound debridement and stabilization of the fracture. Cancellous bone grafting is often delayed for about 14 days to allow the surrounding soft tissue to recover sufficient vascularity to be supportive of the bone graft. If the graft cannot be covered with skin or muscle, it can be protected with a sterile nonadherent dressing followed by a bandage. If the cancellous bone graft fails, it is generally resorbed or expelled through the wound. Cancellous bone is the only type of bone graft that can be safely applied to contaminated or infected fractures, as grafts containing cortical elements will generally sequestrate.

Osteomyelitis

Chronic bone infection is characterized by large areas of avascular bone and dense scar tissue, which can be impenetrable barriers to parenterally administered antibiotics. When avascular bone and scar tissue are removed by debridement, this results in a large bony defect that is slow to heal and is prone to reinfection. Packing such a defect with freshly harvested cancellous bone is helpful in resolving infection and promoting bone healing. Rigid stabilization and appropriate antibiotic therapy are required in addition to the graft in order to obtain a successful outcome.

When cancellous autograft is applied to a contaminated or infected host bed, care must be taken to avoid contamination of the donor site. In this situation, the following procurement and application procedure is recommended: 1) perform the host site surgical procedure and any necessary debridement, lavage, obtain a sample for culture, and then cover the repair site with moist sponges; 2) change gloves and use a separate set of surgical instruments to harvest the cancellous bone graft; 3) close the donor site; 4) apply the cancellous graft to the recipient site; and 5) close the repair site.

Bone Cysts

Bone cysts are benign fluid-filled lesions of unknown etiology that may be monostotic (involving a single bone) or polyostotic (involving more than one bone). Clinical signs include pain and swelling, but cystic bone lesions can be asymptomatic until they reach a fairly large size or until a pathologic fracture occurs. Treatment involves curettage of the walls of the cyst, filling of the resulting defect with cancellous bone graft, and stabilization of the bone until healing occurs.

Arthrodesis

When arthrodesis is performed, stable bony union of multiple, often complex joint surfaces must be achieved as quickly as possible. A successful outcome depends upon adherence to the following principles: 1) removal of all cartilage from the surfaces that must undergo bony healing; 2) liberal application of cancellous bone graft to fill defects and to promote early callus formation; 3) rigid fixation; and 4) healing of the joint in a functional anatomic position.

Bone Plate Removal

Implant removal is performed in some patients that have undergone longbone fracture repairs with bone plates and screws. Implant removal results in a variable number of empty holes that can act as stress concentration points until healing occurs. Some surgeons advocate packing the empty bone holes with cancellous autograft bone to speed the healing of these defects.

Limb Salvage

Animals with a neoplastic lesion involving the distal radius are the best candidates for limb salvage. Wide resection of the neoplastic portion of the bone produces a defect that is too massive to be filled with cancellous bone alone. Typically, an allogeneic cortical bone segment is cut to fit the defect. Rigid stabilization is accomplished with bone plate and screw fixation that extends from the proximal radial host segment to the distal portion of the 3rd metacarpal bone. To promote rapid healing at the alloimplant-host bone junctions, freshly harvested cancellous autograft is packed into the medullary cavity at the proximal and distal ends of the alloimplant (composite grafting). Cancellous bone is also liberally applied over the proximal and distal interfaces of the alloimplant segment. Further discussion of this technique can be found in the section “Harvesting, Storage, and Application of Cortical Allografts” in a later section of this chapter. Distraction osteogenesis provides an alternative solution for dealing with large bony defects in limb salvage patients and this technique is covered in the last section of this Chapter.

Donor Sites for Cancellous Bone Grafts

In young adult animals, the metaphyseal regions of most major longbones can provide graft material with high osteogenic potential. With progressing age, bone marrow at some of these sites undergoes a slow transformation from hematopoietic marrow to fatty marrow. Cancellous bone obtained from areas where the bone marrow is still hematopoietic provides the highest level of osteogenic function. In this regard, the best sites for obtaining cancellous autografts in mature dogs are the proximal humerus, proximal femur, distal femur, and the wing of the ilium. Cancellous bone taken from these sites has a rich, deep reddish brown appearance. In contrast, cancellous bone taken from the proximal metaphysis of the tibia, a site where marrow becomes fatty, has more of a yellowish or tan appearance (Figure 54-1).

The 3 most commonly used sites for obtaining cancellous bone grafts in dogs are the craniolateral aspect of the proximal metaphysis of the humerus, the dorsal aspect the wing of the ilium, and the caudomedial aspect of the proximal metaphysis of the tibia. In terms of both the quality and the quantity of graft material that can be obtained, the proximal humerus is the best of these three alternatives. The proximal humerus is also an easily accessible site. The quality of cancellous bone from the wing of the ilium is quite good, but the volume of material that can be obtained is significantly less than for the humerus, and it is not as easily accessible. Although the proximal tibia is easily accessible, cancellous autograft from this site tends to provide less volume and lower quality material compared to the other two sites. If additional graft material is needed from the same donor site at a later date, restoration of cancellous bone is more rapid and complete in the proximal humerus compared to the proximal tibia. The recommended waiting time before returning to a site for a second graft harvest is 2 months.
 

The femur offers two additional donor sites for obtaining cancellous bone. A greater quantity can be obtained from the condylar region of the distal femur compared to the greater trochanteric region of the proximal femur. Graft quality is good at both locations. Access to these sites requires more dissection than is needed for access to the proximal humerus and proximal tibia. The distal femur offers a convenient location for harvesting cancellous bone graft to be used for a triple pelvic osteotomy procedure.

Obtaining a large amount of cancellous bone graft material is problematic in cats. The proximal metaphyseal region of the humerus is the preferred donor site, similar to dogs. Rib grafts can provide a larger volume of material and these can be harvested and cut into small chips with rongeurs. This corticocancellous graft material can be mixed with cancellous bone to fill large bony defects in feline patients.

Surgical Approaches to Donor Sites

Proximal Humerus

A 2 to 3 cm long skin incision is made over the craniolateral aspect of the greater tubercle, just cranial to the palpable acromial head of the deltoideus muscle. Subcutaneous tissue is separated by sharp dissection to reveal the periosteal surface of the bone. Exposure is maintained by insertion of a small Gelpi self-retaining retractor. An access hole is made with a Steinmann pin or drill bit through the thin outer layer of cortical bone in the proximal metaphyseal region of the humerus (Figure 54-2). It is important to maintain a safe distance from the growth plate in skeletally immature animals. It is also important to make the access hole in the metaphysis rather than in the hard cortical bone of the diaphysis. An access hole in this later location increases the risk of a postoperative iatrogenic fracture of the donor bone.
 

Proximal Femur

A 2 to 3 cm long skin incision is made directly over the lateral aspect of the greater trochanter. The subcutaneous tissues and the superficial gluteal muscle are sharply incised to reveal the surface of the bone. Exposure is maintained with a Gelpi retractor. An access hole is made with a Steinmann pin or drill bit.

Distal Femur

A 2 cm long incision is made over the bone halfway between the fabella and the proximal patella, parallel to the margin of the patella. The incision is made from the skin to the bone on either the medial or the lateral aspect of the femoral condyle. A Gelpi retractor is applied to maintain exposure, which reveals the stifle at the caudal margin of the reflection of the joint capsule. An access hole is made through the cortex of the condyle with a Steinmann pin or drill bit at the location shown in Figure 54-3.
 

Proximal Tibia

A medial skin incision 2 to 3 cm in length is made starting approximately 2 cm distal to the tibial plateau, midway between the tibial tubercle and the medial collateral ligament. Subcutaneous tissues and underlying muscle (insertions of sartorius and gracilis muscles) are separated with sharp and blunt dissection to reveal the proximal tibial metaphysis. The cross-sectional shape of the proximal tibia is triangular, with the base of the triangle located caudally. In light of this, the access hole in the metaphysis should be made at a caudomedial location as shown in Figure 54-4.
 

Iliac Crest

A 4 to 8 cm long skin incision is made directly over the dorsal aspect of the iliac crest. Deep fascia is incised along the entire length of the incision. The middle gluteal muscle is sharply incised from its attachment to the dorsal aspect of the iliac crest and then is subperiosteally elevated from the wing of the ilium to reveal the bone. The dorsomedial aspect of the ilium is exposed by sharp incision of the insertion of the sacrospinalis muscle. Exposure is maintained with Gelpi retractors. An access hole can be made in the dorsal surface of the ilium, or an osteotome or saw can be used to remove a cap of bone from the craniodorsal aspect of the iliac crest as shown in Figure 54-5. If a large quantity of bone graft is needed, this cap can be cut into multiple pieces with a pair of rongeurs to serve as a cancellous bone extender. The corticocancellous bone chips are then mixed with cancellous bone and applied to the recipient site. The iliac crest may be preferable to the previously mentioned longbone metaphyseal donor sites in young animals with open growth plates because there is less risk of clinically significant growth disturbance as a complication of graft procurement.
 

Instrumentation and Graft Harvesting Procedure

Minimal instrumentation is required for harvesting cancellous bone grafts from the humerus, femur or tibia. Placement of a small Gelpi retractor is useful to maintain exposure at the donor site. A Steinmann pin or drill bit is used to penetrate the thin cortex of the metaphysis. The access hole should not be made through the thicker cortical bone of the diaphysis. In young patients with open physes, it is important to locate the access hole a safe distance away from the growth plate. Growth deformities have been reported secondary to graft harvest in these patients. In very small patients with open physes, cancellous bone graft harvest from the wing of the ilium is a safer procedure than graft procurement from a longbone donor site. An osteotome or a bone saw are helpful for removing the craniodorsal portion of the wing of the ilium for improved access to the cancellous bone available for harvest.

A bone curette is inserted through the access hole to remove cancellous bone. The size of the bone curette can be varied according to the size of the patient, but a 5 mm curette works well in most cases. A rotational scooping movement of the curette is effective for harvesting cancellous chips. It is important to avoid penetration of the far cortex during graft procurement. A large amount of cancellous bone can be harvested through a single access hole, although the volume available is frequently underestimated (See Figure 54-6). If a larger opening is needed, the hole should be lengthened along the longitudinal axis of the bone, keeping the corners rounded. Square corners and extension of the access hole perpendicular to the long axis of the bone cause significant mechanical weakening that may predispose inatrogenic fracture of the bone through the graft site.
 

Graft Application Techniques

Cancellous bone graft material should be protected from dehydration in order to achieve optimal effect. Individual chips can be immersed in patient blood that has been aspirated from the access hole and placed in a small stainless steel cup. Alternatively, graft chips can be placed in a blood soaked sponge until they are applied to the recipient site. Graft chips should never be immersed in saline or disinfectant solutions.

Graft application should be the last thing done prior to soft tissue closure over the repair site. Adequate preparation of the recipient site is very important. The orthopedic repair and all debridement and lavage should be completed prior to graft harvesting and placement. Adequate nutrition to sustain the cancellous graft chips until they are revascularized is most likely to occur when they are applied to viable bone surfaces and immediately covered with viable soft tissues during surgical closure of the wound. When severe soft tissue trauma is present (i.e. shearing wounds), it may be prudent to delay cancellous bone grafting until sufficient wound healing has occurred to provide a supportive environment for the graft.

Graft chips that are about the size of a match head (2 to 3 mm in diameter) provide an ideal surface to volume ratio, thus facilitating nutrition of the graft by diffusion until revascularization takes place. Cancellous bone chips should not be densely packed into a defect as this may impair diffusion. Appropriately applied cancellous autograft chips have been shown to be extensively revascularized by 1 week after implantation.

“Closed” application of cancellous bone graft may be useful when highly comminuted shaft fractures are treated using a non-invasive repair technique such as closed application of an external skeletal fixator. “Closed” cancellous bone grafting involves making a 1 cm long access incision over the middle portion of the fracture region. This will accommodate a modified 3 cc syringe which is used to inject the bone graft material over the area of comminution. The tip of the syringe is cut off, the plunger is pulled back, and chips of cancellous bone graft are loaded into the empty cylinder. The loaded syringe is then inserted through the access incision down to the level of the fracture and graft material is injected. This process can be repeated at different angles to deposit 3 cc aliquots of cancellous bone throughout the area of comminution.

Avoiding Possible Complications of Cancellous Bone Grafting

Complications following cancellous bone grafting in dogs and cats are uncommon. Formation of a seroma or hematoma at the donor site is perhaps the most frequently encountered problem. Both of these events are easily preventable. If persistent hemorrhage is encountered from the access hole, it can be plugged with a piece of absorbable gelatin sponge (Gelfoam). Careful closure of overlying soft tissue layers (especially the deepest layer immediately over the access hole) to obliterate dead space, and proper attention to hemostasis will prevent postoperative seroma formation.

Iatrogenic fracture through the access hole has been reported after cancellous bone graft harvest from the proximal humerus and proximal tibia. Guidelines for avoiding this complication are as follows: 1) Make sure to locate the access hole in the metaphysis rather than in the diaphysis; 2) Make sure to direct the drill bit perpendicular to the cortex rather than obliquely when drilling, so that the access hole will be circular; and 3) If the access hole needs to be enlarged, an increase in its length is less detrimental to bone strength than an increase in width.

Premature closure of open physes and resultant growth deformities have been reported secondary to the harvesting of cancellous bone graft material from longbone metaphyses. In skeletally immature animals (< 13 months old), cancellous bone should be obtained from the wing of the ilium (instead of from the humerus, tibia, or femur) because there is little chance that disturbed growth will result in a major clinical problem at this site.

Infection at the donor site is a potential complication when the surgeon is confronted with an open, contaminated or infected fracture. In this situation, a separate set of sterile, uncontaminated instruments and a new pair of surgical gloves must be used for harvesting the cancellous bone graft. The surgeon must not go back and forth between the donor site and the repair site. The graft material is stored in a cup filled with blood obtained from the access hole or in a blood soaked sponge while the surgeon closes soft tissues over the donor site. The graft is then applied to the fracture region and the repair site is closed.

With proper attention to detail, all of the complications of cancellous bone grafting previously discussed are easily avoidable. In any case in which the use of a cancellous bone graft is anticipated, the surgeon must remember to clip and prepare an appropriate donor site to enable the use of this simple and an extremely valuable technique when needed.

Suggested Readings

Ferguson JF: Fracture of the humerus after cancellous bone graft harvesting in a dog. J Sm Anim Pract 37:232, 1996.

Johnson KA: Cancellous bone graft collection from the tibia in dogs. Vet Surg 15:334, 1986.

McLaughlin RM, Roush JK: Autogenous cancellous and cortico-cancellous bone grafting. Vet Medicine 93:1071, 1998.

Palmisano MP, Schrader SC: Premature closure of the proximal physis of the humerus in a dog as a result of harvesting a cancellous bone graft. J Am Vet Med Assoc 215:1460, 1999.

Penwick RC, Mosier DA, Clark DM: Healing of canine autogenous cancellous bone graft donor sites. Vet Surg 20:229, 1991.

Slocum B, Slocum TD: Bone graft harvest: Distal femoral condyles. In Bojrab MJ, ed: Current Techniques in Small Animal Surgery, 4th ed. Philadelphia: Lippincott Williams & Wilkins, 1998, p. 909.

Stallings JT, Parker RB, Lewis DD, et al: A comparison of autogenous cortico-cancellous bone graft obtained from the wing of the ilium with an acetabular reamer to autogenous cancellous bone graft obtained from the proximal humerus in dogs. Vet Comp Orthop Traumatol 10:79, 1997.

Trevor PB, et al: Evaluation of the proximal portion of the femur as an autogenous cancellous bone donor site in dogs. Am J Vet Res 53:1599, 1992.

Wilson JW, Rhinelander FW, Stewart CL: Vascularization of cancellous bone chip grafts. Am J Vet Res 46:1691, 1985.
 

Introduction

A large volume of corticocancellous bone graft may be readily obtained from the wing of the ilium using a powered acetabular reamer. Although graft incorporation is slower and less uniform when compared to similar volumes of cancellous bone graft, the corticocancellous bone grafting technique is advantageous because it yields a greater volume of graft, offers a more proximate location of the harvest site when performing procedures involving the hind limb, and produces graft with a consistency that is favorable for packing into bone defects, resulting in an intimate association with the recipient bed.

Surgical Procedure

The patient is placed in lateral recumbency. An area extending approximately 5 cm cranial to the wing of the ilium to 3 cm caudal to the greater trochanter, and from dorsal to the wing of the contralateral ilium to 5 cm ventral to the ilial body is clipped, aseptically prepared and draped for surgery.

The skin incision begins craniodorsal to the iliac spine and is continued caudally, paramidline to the level of the middle of the body of the ilium. Subcutaneous tissues and the deep gluteal fascia are incised along the same line as the skin incision, exposing the tuber sacrale. The origin of the middle gluteal muscle on the dorsal ilium is incised, allowing subperiosteal elevation of the middle gluteal muscle. Elevation is continued caudally to the level of the caudal dorsal iliac spine, but should not be continued beyond this point in order to preserve the cranial gluteal vein, artery, and nerve. Elevating the middle gluteal muscle off of the cranial aspect of the wing of the ilium improves exposure. Exposure is maintained by placing one or two Hohmann retractors from dorsal to ventral to expose the wing of the ilium. Gelpi retractors can also be placed to facilitate esposure.

An acetabular reamer (20 or 23 mm for small dogs, 26 or 29 mm for medium-sized dogs, and 29 or 32 mm for large dogs) attached to a low speed, high torque drill is used for harvesting the graft. Reaming is initiated on the lateral ilium immediately caudal to the iliac crest. It is continued caudally, removing the lateral cortex and cancellous bone while leaving the medial cortex and dorsal edge of the ilium intact. Reaming is carried as far caudally as is feasible, creating an oval-shaped defect (Figure 54-7). When the cup of the reamer is full, it is detached from the extension and the graft is removed to be stored in a sterile receptacle until required. Switching to a smaller diameter reamer generally allows the surgeon to extend reaming down the body of the ilium. Care must be taken to avoid penetrating the medial cortex of the ilium with the reamer. When the reaming is completed, additional exposed cancellous bone along the periphery of the defect can be harvested with a bone curette.

The donor site is thoroughly lavaged with sterile saline. A splash block of local anesthetic may be administered prior to closure to decrease postoperative discomfort. Closure is performed in multiple layers to decrease the risk of postoperative seroma formation. The superficial fascia of the middle gluteal muscle is apposed to its periosteal insertion or to the superficial fasia of the sacrospinalis muscle with a series of horizontal mattress sutures.
 

The gluteal fascia, subcutaneous tissues, and skin are closed routinely.

The harvested graft has a paste-like consistency, which facilitates packing the graft into bone defects and results in intimate contact with the recipient bed.

Postoperative Considerations

The cortiocancellous graft appears more radiodense than a cancellous graft on immediate postoperative radiographs because of the graft’s cortical component. Morbidity associated with ilial corticocancellous graft harvest is minimal; however, transient, self-limiting hind limb lameness and seroma formation may occur. Restricted postoperative activity is therefore recommended.

Suggested Readings

Culvenor JA, Parker RB: Collection of cortico-cancellous bone graft from the ilium of the dog using an acteabular reamer. J Small Anim Pract 37:513, 1996.

Piermattei DL, Johnson KA: An atlas of surgical approaches to the bones and joints of the dog and cat, 4th ed. Philadelphia: W.B. Saunders, 2004, 278.

Stallings JT, Parker RB, Lewis DD, et al.: A comparison of autogenous cortico-cancellous bone graft obtained from the wing of the ilium with an acetabular reamer to autogenous cancellous bone graft obtained from the proximal humerus in dogs. Vet Comp Orthop Traumatol 10:79, 1997.
 

Introduction

Cortical allografts have been used to enhance repair of long bone fractures in veterinary surgery for several decades.^1-4 An allograft is bone transferred from one individual to another individual of the same species. This type of graft elicits an immune response because of foreign cellular antigens of the allograft and the reaction of the host immune system. Fresh-frozen processed cortical allografts are the most commonly used cortical grafts in veterinary orthopedic surgery. Allografts are also considered alloimplants because they are a nonviable material (dead bone), and by definition, the term implant refers to any nonviable material placed in the body.

An autogenous allograft (autograft) is bone transferred from a donor site to a recipient site in the same individual. There are definite disadvantages to this type of graft. Sufficient bone is often not available, morbidity at the donor site is a concern, increased anesthetic and surgery time, and increased risk of infection. Fresh frozen allograft is preferred for convenience of storage, and reduction of disease and immunogenicity.^5-9

Frozen allografts provide structural (mechanical), osteoconductive, and osteoinductive support to fracture repair.¹⁰ Other methods of processing bone allografts include cryopreservation, freeze-drying (lypholized) and demineralized preparation. The processing of bone grafts by these methods is more involved technically and are not practical for the veterinary surgeon in practice. Inconvience of allograft harvesting, processing, storage, and quality assurance have limited their use.¹¹

Harvesting of Allografts

Harvesting of allografts is practical for the veterinary surgeon and requires adherence to strict asepsis, preparation and time. The procurement of cortical allografts begins with proper donor selection. Donors should be mature, healthy animals, preferably between the 1.5 to 8 years of age, with no preexisting neoplastic, metabolic, bacterial, or viral diseases. A complete physical examination and review of history are essential. Current vaccinations and blood screening for transmissible diseases should be performed. Immature donors have bones that may be brittle and less developed than older donors, and this factor may cause problems during implantation with stability (screw purchase).

Allografts can be harvested from dead donors and then sterilized with ethylene oxide. Although ethylene oxide is considered a superior sterilizing agent for surface contamination, but low residual levels may be toxic to recipient tissue and could interfere with healing. It may also affect the mechanical strength and incorporation of cortical allografts.^12,13 Freshly harvested cortical allografts are preferred.

Absolute aseptic surgical technique is required. All donors should be prepared as for any standard surgical orthopedic procedure, with proper aseptic scrubbing and draping. Donors are placed under general anesthesia, and standard approaches to the long bones are used. The bone should be exposed from metaphysis to metaphysis by removing as much soft tissues (muscle and periosteum) as possible. An oscillating bone saw is used to cut the bone. This saw should be cooled with liquid during cutting. After the bone is removed, it is placed in a solution of lactated Ringer’s or saline. This is temporary before final preparation of the graft. Once all the donor graft has been harvested, euthanasia is performed on the donor.

The grafts are then stripped of all remaining soft tissue attachments, and the medullary contents are removed. A sharp periosteal elevator or scalpel blade works best for stripping, whereas a bone curette works best for removal of medullary contents. The medullary cavity should be flushed out with sterile lactated Ringer’s or saline solution. Once the graft is clean, it can be cut into proximal, middle, and distal thirds, halved or maintained in its full length. The graft’s medullary cavity is cultured for aerobic and anaerobic organisms. The graft is placed in a suitable glass jar that has been previously autoclaved. Each jar with the graft should be marked, indicating left or right, with the name of the bone, segment of bone, date of harvesting, and donor identification. The jar and graft are then immediately placed in a household freezer at a temperature of -20° C. Any temperature warmer than this leads to improper freezing and possible autolysis. The American Association of Tissue Banks allows 6 months storage at -20° C and recommends -40° C for longer-term storage (up to 5 years); -70° C is preferred.¹¹ The colder temperatures inhibit molecular translations that result in degradation. The author has not had any problems safely storing bone grafts at -20° C for 1 year. Only grafts that culture negative are placed in the bone bank.

The most common indication for use of cortical allografts is replacement of bone in patients with highly comminuted fractures. Other indications are correction of nonunions, delayed unions, and mal-unions with or without bone loss, bone lengthening, limb-sparing procedures for bone tumors, and, in selected cases, osteomyelitis with bone loss due to sequestrum formation.¹⁴ This last indication should be considered a salvage procedure if amputation is not an option. In preparation for surgical implantation of an allograft, radiographs of the opposite limb should be made, and bone length measured. An estimate of the graft length needed is made by comparing the intact cortical segments on the lateral projection of the affected limb and subtracting this from the total length of the normal bone. (Figure 54-8). A graft is then selected based on this estimate as well as by visually comparing the width of the host and graft bone. Usually, the femur is used to replace a segment of femur, however the use of other long bones should not be discouraged because the width of other bones may be adequate if a near perfect match cannot be made with similar bones.
 

Surgical Application

Prior to surgery a proper allograft is selected based on previous radiographic planning, visual observation and comparison of available allografts. It is prudent to select an allograft that is slightly longer then is required should adjustment be needed during surgery. Most importantly, the diameter of the allograft bone should be as close as possible to the host bone. Prophylactic antibiotics are administered at the time of anesthesia, during the operation and postoperatively. An appropriate cancellous bone graft site is prepared. The addition of cancellous bone at the host-graft site increases the success of the procedure.⁷ Before the surgical procedure, the cortical allograft is allowed to thaw in a sterile bowl of lactated Ringer’s solution or normal saline solution. The patient is aseptically prepped, draped and strict surgical technique followed. If a large number of bone segments are to be removed, the surgeon must have a point of reference proximally and distally to maintain proper alignment with respect to rotation, varus and valgus. This is best done with small Kirshner wires placed parallel to each other, one in the proximal fracture segment and one in the distal segment. The fracture is exposed, and the comminuted fragments are removed. The fractured bone ends, proximally and distally are cut with a bone saw perpendicular to the long axis of the bone in preparation for the cortical allograft. The allograft is cut to the proper size; the surgeon must ensure that it is perpendicular to the long axis of the graft. This cut should allow 360° of cortical contact ideally, and not less than 50% contact at the host-graft interface. In some cases of delayed union, malunion, and nonunion, the callous formation may be larger than the allograft or cut surface. This is not of concern and can serve as a ridge for autogenous cancellous bone to be placed on to augment grafting. A dynamic compression plate is selected to allow for a minimum of five cortices (three screws) above and below the allograft. Standard ASIF plating technique is used. The plate is contoured to both the host bone and allograft. An alternate technique is to contour the plate preoperatively from the radiograph of the normal intact bone and make adjustments at the time of surgery. The plate is first applied to the allograft with a minimum of two screws (four cortices) in a neutral position. The allograft is aligned to the host bone to ensure as close to 360° of cortical contact as possible. This is not always possible, but the closer to 360° the better the stability. Care should be taken to test the reduction at both ends of the allograft before completing screw fixation of the plate. Alignment should be observed and rotation, varus and valgus corrected. The preplaced Kirschner wires and temporary cerclage wires or bone clamps aid in proper positioning. If any correction is needed the plate can be removed and the allograft cut for correction. If the correction causes the total bone length to be shorter, a new allograft should be used. Depending on the bone, most patients can tolerate shortening of 2 to 3 cm in the limb without an impact on function. The screw holes above and below the allograft are placed in the loaded position. This maneuver results in compression at the host-graft interfaces. The remaining screws are placed in a neutral position. The entire surgical site is flushed with lactated Ringer’s solution before placing an autogenous cancellous bone graft around the host-graft interfaces. Commercially prepared cancellous bone chips or cancellous bone chips and demineralized powder can also be used (Veterinary Transplant Services, Seattle, WA). The surgical site is cultured for aerobic and anaerobic organisms before routine closure.

Postoperative Care

Postoperative care consists of an appropriate coaptation with a modified Robert Jones dressing or padded bandage, depending on the long bone repaired, for 2 to 3 weeks. Activity should be restricted to leash only walks and cage or kennel confinement during this time period. Antibiotics are administered for 2 weeks postoperatively and are adjusted or discontinued based on the culture results. Radiographs are taken at 3 to 4 week intervals, to follow healing and implant stability for the first 3 months followed by radiographic exam every 6 to 12 months thereafter. Plate removal should only be considered in young patients or in those patients with allografts less than 3cm and only after 2 years post surgery. Plate removal is “staged” with 3 to 6 months between surgeries.

Bone Healing

Cortical allografts heal by proceeding through phases of inflammation, revascularization, osteoinduction, osteoconduction and remodeling. This process takes much longer due to the dense structure of cortical bone.¹⁵ Cortical allograft incorporation differs from autogenous cancellous bone in that initial repair is due to osteoclastic rather than osteoblastic activity.¹⁶ Resorption occurs rapidly shortly after transplantation and gradually declines to normal levels within a year.¹⁶ Resorption of the graft and replacement by host bone begin at the host-graft interface and move toward the center of the graft, with marked proliferation of periosteal and endosteal bone covering the graft surfaces.⁷ This process can take years depending on the length of the graft. Biopsy specimens taken at various levels of long allografts at 45.5 months⁷ and 92 months¹⁴ after implantation showed graft bone still present. As this process continues, mechanical strength is added to the graft. Predominately at the center portion of the allograft with live bone present in the external and internal circumferential lamella.¹⁴ The presence of dead bone matrix from the graft interspersed with interstitial lamellar and host osteons affords strength to the bone.¹⁶

Cortical allografts provide osteoconductive, osteoinductive and mechanical support in the repair of long bone fractures. Success has been reported to be over 80%.⁷ Outcome is dependent on case selection, degree of trauma, soft tissue damage, coexisting injuries and adherence to strict surgical asepsis and technique. Decreased surgical time, stability of the fracture repair, and rapid return to function are definite benefits. Harvesting of bone, aseptic technique, and bone plating principles may be a limitation, depending on training and surgical experience. The added cost of proper surgical equipment and time spent setting up the bone bank are also possible limitations. As a general rule, infected or open fractures and metaphyseal fractures that do not allow proper screw purchase are not indications for cortical allografts.

References

1. Fox S: Cancellous bone grafting in the dog: An overview. J Am Anim Hosp Assoc 20:840, 1984.
2. Hulse D: Pathophysiology of autogenous cancellous bone grafts. Compendium Continuing Education Pract Vet 2(2): 136, 1980.
3. Johnson A: Principles of bone grafting. Seminars Vet Med Surg (Small Animal) 6(1): 90, 1991.
4. Olds R, Sinibaldi K, DeAngelis M, et al: Autogenous cancellous bone grafting in small animals. JAAHA 9:454, 1973.
5. Johnson AL: Principles and practical applications of cortical bone grafting techniques. Compendium Contin. Educ Pract Vet 10(8): 906,1988.
6. Schena C, McCurnin D: The use of fresh cortical and cancellous allografts in the repair of a fractured femur in a dog: A case report. J Am Anim Hosp Assoc 19:352,1983.
7. Sinibaldi K: Evaluation of full cortical allografts in 25 dogs. J Am Vet Med Assoc 194(11):1570, 1989.
8. Henry W, Wadsworth P: Diaphyseal allografts in the repair of long bone fractures. J Am Anim Hosp Assoc 17:535, 1981.
9. Aaron A, Wiedel J: Allograft use in orthopedic surgery. Orthopedics 17(1):41, 1994.
10. Burchardt H: The biology of bone graft repair. Clinical Orthopedics 174:28,1983.
11. Fitch R, Kerwin S, Newman-Gage H, Sinibaldi K: Bone autografts and allografts in dogs. Compendium Continu Educ Pract Vet 19(5)558,1997.
12. Arizono T, Iwanoto Y, Okuyama K, Sugioka Y: Ethylene oxide sterilization of bone grafts: Residual gas concentration and fibroblast toxicity. Acta Orthop Scand 65(6):640,1994.
13. Wagner S, Manley P, et al: Failure of ethylene oxide-sterilized cortical allografts in two dogs. J Am Anim Hosp Assoc 30:181, 1994.
14. Sinibaldi KR, unpublished data.
15. Burchardt H, Enneking WF: Transplantation of bone. Surg Clin North Am. 58:403, 1978.
16. Enneking WF, et al. Physical and biological aspects of repair in dog cortical bone transplants. J Bone Joint Surg (Am) 57:237, 1975.
     

Introduction

Distraction osteogenesis is a technique capable of generating large amounts of bone by gradual distraction of osteotomized bone ends. This method is now widely accepted for the treatment of shortened limbs, bony defects from tumor or trauma and angular limb deformity.¹⁸ The technique is most commonly performed using circular external fixator systems and tensioned fine wires, a method introduced and refined by Ilizarov.¹⁸

Historical Perspectives

Gavril A. Ilizarov was a physician with no formal orthopedic training who practiced in a small industrial town in Western Siberia after World War II. Antibiotics were scarce and chronic osteomyelitis and non-unions were common post-war injuries among the population of patients he cared for. As a result, he found himself practicing orthopedics in his general practice in an isolated area of the world without access to any of the technological and medical advances that took place during the post-World War II era. He devised an innovative external fixator system comprised of modular rings and trans-osseous wires attached to the rings under tension to stabilize bone fragments. The phenomenon of distraction osteogenesis was discovered incidentally, when Ilizarov applied a fixator designed to create gradual compression of the fracture ends to a patient with an infected non-union. He instructed the patient to adjust specialized nuts on the frame several times daily in order to achieve compression at the fracture site. Instead, the patient mistakenly turned the nuts in the wrong direction, thereby lengthening the frame and creating distraction at the fracture site (Figure 54-9). Ilizarov observed significant new bone formation in the distraction gap and simultaneous resolution of the infection. He applied this technique successfully to some of the most challenging conditions in orthopedic surgery.^16,17 The reconstruction of bones affected by post-traumatic conditions, such as intercalary defects, shortening and deformity was the most common application of his method. In 1984, an Italian veterinarian by the name of Dr. Antionio Ferretti, began using the Ilizarov methods in veterinary patients. The use of circular fixation and distraction osteogenesis began to appear in North American veterinary literature in the early 1990s.²⁴ Currently, IMEX Veterinary (Longview, TX) manufactures a circular external fixator system that has lightweight design elements suitable for veterinary patients (Figure 54-10). Other circular external fixation systems are also available in North America and Europe.
 

Bone transport osteogenesis is a modification of the original distraction osteogenesis technique, involving the transport of a bone fragment across a bony defect with distraction osteogenesis occurring in the trailing pathway of movement (Figure 54-11). The bone fragment eventually contacts the opposite end of the defect, and is compressed to the adjacent bone in its new position, resulting in union between the bone fragment and the parent bone. The new bone that forms in the distraction pathway rapidly remodels into lamellar bone, thereby filling in the segmental defect. This method was used by Ilizarov to salvage many limbs that otherwise would have been amputated because of non-union, osteomyelitis or extensive segmental bone loss.¹⁸ Bone transport osteogenesis is also used in veterinary patients for limb salvage following segmental bone loss due to trauma or tumor excision.^11,12
 

Histomorphology of Distraction Osteogenesis

Distraction osteogenesis requires prolonged and gradual distraction of two freshly osteotomized bone ends (See Figure 54-9). The new bone by distraction osteogenesis or bone transport osteogenesis is termed regenerate bone. The process of new bone formation is often called osteoneogenesis. The biology of distraction osteogenesis has been extensively studied.^{1,4,6-8,10,21} The results of these investigations have greatly expanded the understanding of the histological, biochemical, vascular, radiographic, and mechanical properties of regenerate bone formation. Ilizarov mistakenly assumed that distraction osteogenesis recapitulated endochondral bone formation. This belief was generated by the radiographic observation that a radiolucent zone consistently occurred in the center of the regenerate bone (radiolucent central zone) until distraction was completed, similar to a growth plate which remains radiolucent until growth is completed. More recent studies have shown that bone formation during distraction osteogenesis results from both intramembranous and endochondral ossification, with intramembranous bone formation predominating at a ratio of 5:1.¹⁴ The radiolucent central zone is comprised of Type I collagen columns adjacent to a zone of newly formed vessels. This vasculature delivers proliferating and differentiating osteoblasts which migrate along the collagen columns and deposit osteoid. These collagen columns are formed in parallel and along the lines of distraction tension. Each of these osteoid-covered, longitudinal columns of collagen begins to mineralize starting from either end of the gap and progressing toward the central radiolucent zone. The mineralizing new bone columns resemble stalagmites and stalactites projecting from each osteotomy surface on radiographs. Each bone column expands transversely as more collagen fibers are incorporated circumferentially and mineralized until they reach a maximum diameter of 150 to 200 microns. The space between the bone columns consists of large, thin-walled vessels.^2,3,9,10,14 Once distraction is completed, the bone columns begin to bridge the peripheral aspect of the radiolucent central zone. Columns of mineralizing new bone then rapidly bridge the entire central radiolucent zone and are eventually interconnected transversely by woven bone plates forming a honeycomb-like pattern. Once bridging occurs, rapid secondary remodeling of the cortices ensues and the Haversian system is re-established. This remodeling process occurs much more rapidly than with classical fracture healing; partly because the collagen fibers are more orderly and aligned at the start of mineralization and therefore tend to remodel in a manner parallel with the long axis of the bone. In addition, the mechanical strain environment created in the distraction gap seems to promote robust angiogenesis, massive osteoblast recruitment and rapid production of osteoid.^13,14

Technique

Clinical Factors Influencing Distraction Osteogenesis

The environment created during distraction osteogenesis is not identical to the environment seen during fracture healing. The optimal mechanical environment in which bone formation occurs clinically has not been fully determined. Several unique factors are known to influence regenerate bone formation. These include: frame type, osteotomy technique, delay interval between surgery and distraction (latency period), the total distance moved per day (distraction rate), and the number of increments used to achieve the total distance moved per day (rhythm).

Frame Type

Although Ilizarov attributed special biological effects to the use of ring fixators, distraction osteogenesis can also be achieved using linear fixators or hybrid frames. The two major advantages of using circular external fixation systems are 1) the axial micromotion that occurs under compressive loads with fine wire fixation and 2) the versatility of fixator components and spatial configurations possible with circular frames that allow precise movement of bone fragments while not compromising overall frame stability. Fine wire fixation exhibits nonlinear biomechanical behavior. Specifically, controlled micromotion of the bone segments occurs during weight-bearing yet stiffness in bending and torsion similar is maintained in a manner similar or superior to conventional linear fixators. Controlled axial micromotion is thought to be beneficial to bone formation.^5,21,22

Ring diameter, wire tension, bone position within the frame and number of rings and wires per bone segment affect overall frame stability. Clinicians should be familiar with the biomechanical characteristics of circular fixator frames and the principles of Ilizarov to achieve ideal stability when designing and positioning the frame for a particular patient.^19,20,23

Osteotomy Technique

Ilizarov considered the preservation of the meduallary vascular system and periosteum to be essential for bone distraction osteogenesis. His original technique involved carefully cutting the cortex with an osteotome while preserving the periosteal sleeve, a procedure he termed corticotomy.^18,23 Since that time however, results of animal studies have shown that the quality and quantity of bone formed during distraction osteogenesis following an osteotomy created with an oscillating saw, osteotome or corticotomy is similar.¹⁵ No advantage has been seen with the corticotomy technique over the more standard osteotomy techniques. An important point to remember, however, is to avoid thermal damage to the bone. If an oscillating saw is used to create the osteotomy prior to distraction or bone transport osteogenesis, copious lavage with cool saline is required.

Latency Period

Latency refers to the amount of time between creation of the osteotomy and commencement of distraction. This period of time is important for the formation of a soft callous. Several factors influence the choice of latency period including the degree of trauma to the soft tissue envelope, age of the patient, location of the osteotomy (metaphyseal versus diaphyseal), and patient-related co-morbidity issues. In dogs, suggested latency periods range from 2 to 7 days.²³ In a healthy patient, the author typically uses a 3 day latency period unless there are significant co-morbidity factors. Co-morbidity factors include advanced patient age, soft tissue trauma or loss, or other conditions that would delay healing such as concurrent use of chemotherapy or diabetes, etc. When these conditions are present, the latency period may be lengthened. Too long a latency period will result in premature healing requiring re-fracture.

Distraction Rate

A rate of 1mm of distraction per day is the most common distraction rate used in veterinary medicine for linear distraction. Choice of distraction rate depends upon many of the same factors that influence latency period. Distraction can be performed more rapidly in young animals, sometimes up to 4 mm per day. In dogs, mineralized bone is usually visible within the distraction gap on radiographs by day 14 to 21 of distraction. It is important to monitor the appearance of the regenerate bone on radiographs during distraction because the distraction rate may need to be adjusted during distraction. Radiographic evaluation of the regenerate bone is recommended every 7 to 10 days. If the regenerate bone begins to take on a thinning, ductile shape resembling an hourglass or the radiolucent central zone begins to progressively widen, the distraction rate may need to be decreased. Alternatively, if the wires nearest the distraction gap begin to bend toward the distraction gap and the central radiolucent zone begins to disappear, the distraction rate may need to be increased to avoid premature consolidation.¹¹

Distraction Rhythm

The rhythm of distraction refers to the number of incremental lengthenings performed per 24 hour period to achieve the desired rate of distraction. The recommended rhythm for dogs ranges from 2 to 4. This means that the total amount of distraction achieved during any given 24 hour time period should be divided into 2 to 4 increments. The author recommends a distraction rhythm of 4.¹¹ This is easy for clients to do because the nuts used to perform distraction on the IMEX veterinary circular fixator system have four faces. The owner can be instructed to turn the nuts one face four times daily. The pitch of the all-thread rods used in the same fixator system is 1 mm. Therefore, one complete revolution of the distraction nuts will move the distraction wires 1 mm. Studies performed in goats have shown that increasing the number of increments up to 270 per day using an automated distractor system did not seem to have an advantage.²¹ In the author’s experience, diminishing the distraction rhythm to less than 3 or 4 tends to be associated with more soft tissue complications such as inflammation and tendon contracture. It is likely that the ideal distraction rhythm varies from patient to patient and is probably influenced by similar factors as rate and latency.

Consolidation Period

Once distraction is discontinued, the consolidation period begins. Consolidation involves rapid mineralization of the radiolucent central zone and remodeling of the regenerate bone. The new cortices become organized and marrow elements begin to reform. Consolidation will be delayed if the patient is not weight-bearing. There are likely many other factors that influence consolidation such as biomechanical properties of the fixator, anatomic location etc., but these factors are less well understood. Clinicians must use radiographic evaluation to decide when to remove the fixator. If mineralization and cross-sectional area of the regenerate are similar to the parent bone, it is typically safe to remove the fixator. Certain frame designs allow for progressive destabilization which may allow for a more rapid gain in stiffness. There is no exact formula for deciding when the fixator should be removed, but the more bone created by distraction osteogenesis, the longer consolidation will take to complete. A very rough rule of thumb is that the fixator should remain in place for 50% to 100% of the time needed to achieve the desired amount of new bone.

Regenerate Bone as a Bone Graft Alternative

The use of regenerate bone as a bone grafting alternative is less familiar to many surgeons and initially more technically complex. Distraction osteogenesis may not be the first option of choice in patients where standard autogeneous or allogeneic graft material is suitable. However, there are unique advantages to regenerate bone as a means to fill a defect or create a bony union, particularly in situations where infection is established or likely. Because regenerate is autogenous and its formation is associated with an immediate, robust blood supply, it can be used in situations when cortical allograft would be contraindicated. In human trauma, distraction osteogenesis is most commonly used for severe soft tissue and bone loss following extremity shear injuries, such as those sustained in motorcycle accidents. Because this method of reconstruction does not require internal fixation, such as with cortical allografts, surgeons can begin bony repair prior to establishment of a healthy soft tissue envelope. Other unique uses include chronic osteomyelitic non-unions. In these patients, distraction osteogenesis is used to achieve union, but also as a means to resolve the infection and replace resorbed bone. Ilizarov was the first to note the remarkable ability of distraction osteogenesis to treat osteomylelitis without the aid of antibiotics in his patients. His phrase for this observation was that distraction osteogenesis “burned the infection in the flame of the regenerate”, referring to the effect of angiogenesis and subsequent arrival of immune cells that successfully eradicated infection.

Theoretically, distraction osteogenesis can create limitless quantities of bone. This is in contrast to large-segment cadaveric bone allografts, where supply is often limited and procurement and storage is expensive; and to cortical autograft, where donor site morbidity limits the anatomic location and amount of bone available. In addition, the use of autogeneous tissue eliminates the risk of disease transmission from donor to recipient, a significant concern in human medicine.

Future Directions

To date, the major limitations to the use of distraction osteogenesis as an alternative to bone grafting in veterinary medicine have been the relatively lengthy period of time required to reconstruct large defects and the small number of veterinary surgeons comfortable with the technique. Circular and hybrid fixators have recently become more “main stream” as veterinarians become familiar with their versatility. Research is ongoing to understand more about the biology of osteogenesis, thereby allowing clinicians to manipulate the distraction osteogenesis process using novel growth factors and gene therapies to create bone more rapidly. Double level distraction osteogenesis has been described in veterinary patients to diminish the time needed to reconstruct large diaphyseal defects. Newer hybrid fixator designs and components allow distraction in more than one anatomic plane at a time and, as clinical experience accumulates, surgeons will become more comfortable with case selection and management. Clinical applications for distraction osteogenesis are likely to expand, but will probably be used as a solution in the more challenging orthopedic situations, rather than in cases where simple grafting is routinely successful.

References

1. Aronson, J. Experimental and clinical experience with distraction osteogenesis. Cleft Palate Craniofac. J. 31: 473-481, 1994.
2. Aronson, J. Temporal and spatial increases in blood flow during distraction osteogenesis. Clin. Orthop. Relat Res. 124-131, 1994.
3. Aronson, J., Good, B., Stewart, C., Harrison, B., Harp, J. Preliminary studies of mineralization during distraction osteogenesis. Clin. Orthop. Relat Res. 43-49, 1990.
4. Aronson, J., Harp, J. H. Mechanical forces as predictors of healing during tibial lengthening by distraction osteogenesis. Clin. Orthop. Relat Res. 73-79, 1994.
5. Aronson, J., Harp, J. H., Jr. Factors influencing the choice of external fixation for distraction osteogenesis. Instr. Course Lect. 39: 175-183, 1990.
6. Aronson, J., Harrison, B. H., Stewart, C. L., Harp, J. H., Jr. The histology of distraction osteogenesis using different external fixators. Clin. Orthop. Relat Res. 106-116, 1989.
7. Aronson, J., Johnson, E., Harp, J. H. Local bone transportation for treatment of intercalary defects by the Ilizarov technique. Biomechanical and clinical considerations. Clin. Orthop. Relat Res. 71-79, 1989.
8. Aronson, J., Shen, X. Experimental healing of distraction osteogenesis comparing metaphyseal with diaphyseal sites. Clin. Orthop. Relat Res. 25-30, 1994.
9. Aronson, J., Shen, X. C., Gao, G. G. et al. Sustained proliferation accompanies distraction osteogenesis in the rat. J. Orthop. Res. 15: 563-569, 1997.
10. Aronson, J., Shen, X. C., Skinner, R. A., Hogue, W. R., Badger, T. M., Lumpkin, C. K., Jr. Rat model of distraction osteogenesis. J. Orthop. Res. 15: 221-226, 1997.
11. Ehrhart, N. Longitudinal bone transport for treatment of primary bone tumors in dogs: technique description and outcome in 9 dogs. Vet. Surg. 34: 24-34, 2005.
12. Ehrhart, N., Eurell, J. A., Tommasini, M., Constable, P. D., Johnson, A. L., Feretti, A. Effect of cisplatin on bone transport osteogenesis in dogs. Am. J. Vet. Res. 63: 703-711, 2002.
13. Fink, B., Krieger, M., Strauss, J. M. et al. Osteoneogenesis and its influencing factors during treatment with the Ilizarov method. Clin. Orthop. Relat Res. 261-272, 1996.
14. Fink, B., Pollnau, C., Vogel, M., Skripitz, R., Enderle, A. Histomorphometry of distraction osteogenesis during experimental tibial lengthening. J. Orthop. Trauma 17: 113-118, 2003.
15. Frierson, M., Ibrahim, K., Boles, M., Bote, H., Ganey, T. Distraction osteogenesis. A comparison of corticotomy techniques. Clin. Orthop. Relat Res. 19-24, 1994.
16. Ilizarov, G. A. The tension-stress effect on the genesis and growth of tissues. Part I. The influence of stability of fixation and soft-tissue preservation. Clin. Orthop. 249-281, 1989.
17. Ilizarov, G. A. The tension-stress effect on the genesis and growth of tissues: Part II. The influence of the rate and frequency of distraction. Clin. Orthop. 263-285, 1989.
18. Ilizarov, G. A. The principles of the Ilizarov method. 1988. Bull. Hosp. Jt. Dis. 56: 49-53, 1997.
19. Lewis, D. D., Bronson, D. G., Cross, A. R., Welch, R. D., Kubilis, P. S. Axial characteristics of circular external skeletal fixator single ring constructs. Vet. Surg. 30: 386-394, 2001.
20. Lewis, D. D., Cross, A. R., Carmichael, S., Anderson, M. A. Recent advances in external skeletal fixation. J. Small Anim Pract. 42: 103-112, 2001.
21. Welch, R. D., Birch, J. G., Makarov, M. R., Samchukov, M. L. Histomorphometry of distraction osteogenesis in a caprine tibial lengthening model. J. Bone Miner. Res. 13: 1-9, 1998.
22. Welch, R. D., Lewis, D. D. Distraction osteogenesis. Vet. Clin. North Am. Small Anim Pract. 29: 1187-viii, 1999.
23. Welch, R. D., Lewis, D. D. Distraction osteogenesis. Vet. Clin. North Am. Small Anim Pract. 29: 1187-viii, 1999.
24. Yanoff, S. R., Hulse, D. A., Palmer, R. H., Herron, M. R. Distraction osteogenesis using modified external fixation devices in five dogs. Vet. Surg. 21: 480-487, 1992.