Growth potential in onlay bone grafts: a comparison of vascularized and free calvarial bone and suture bone grafts

Vascularized bone grafts are characterized by a viable cell population with osteogenic potential. These features suggest that continued growth can be anticipated following vascularized membranous bone transfer in a growing craniofacial skeleton. The present paper compares the potential for appositional bone growth in vascularized and free calvarial onlay bone grafts. In seven 8-week-old beagles, growth was assessed by direct caliper measurements of graft dimensions intraoperatively and 16 weeks postoperatively. Vascularized grafts demonstrated a 50 to 60 percent increase in size in all dimensions compared to 10 to 20 percent growth in free grafts (p less than 0.01). Microradiography revealed preservation of calvarial bony architecture and minimal resorption in vascularized grafts, while triple-fluorochrome labeling confirmed subperiosteal appositional bone formation. Free grafts were characterized by significant resorption and a delay in subperiosteal bone formation.     

Fate of donor cells in vascularized bone grafts: identification of systemic chimerism by the polymerase chain reaction

Systemic chimerism, or the movement of cells from a transplanted tissue into host organs, is a phenomenon known to occur in association with development of immunological tolerance in allotransplantation. However, little is known about the fate and movement of cells into or out of autogenous free tissue transfers, including vascularized bone grafts. The purpose of this study was to identify systemic chimerism in vascularized bone grafts by transplantation of a vascularized tibiofibular graft from isogenous (inbred) male Lewis rats to female recipients. Donor (male) cells could be identified in the recipient (female) tissues by semiquantitative polymerase chain reaction analysis for a Y chromosome-specific DNA sequence. Chimerism was assessed at 1, 12, 18, and 24 weeks after transplantation. Competitive polymerase chain reaction study using the specific primers for a Y-chromosome marker ( gene) and an autosomal gene (GAPDH) allowed detection of small amounts of male cells in a large pool of female cells and measurement of their relative proportions as a function of time. Of 19 nonimmunosuppressed recipients, nine animals (47 percent) showed low-level chimerism (<0.1 percent) in the peripheral blood. Nine (47 percent), three (16 percent), and two (11 percent) recipients showed high-level chimerism (>1 percent) in the spleen, liver, and thymus, respectively, at final assessment. Donor cells were detected in all bone grafts and in six contralateral tibial bones (i.e., 67 percent of sampled contralateral tibial bones) at 18 and 24 weeks after transplantation. Twenty-four recipients were immunosuppressed with FK506 (tacrolimus) to suppress reaction to a minor histocompatibility barrier present on the Y chromosome. In this group, 14 animals (58 percent) showed low-level chimerism in peripheral blood and 12 (50 percent), eight (33 percent), and one (4 percent) recipients showed high-level chimerism in the spleen, thymus, and liver, respectively. Transplanted cells were detected in nine contralateral tibial bones (i.e., 60 percent of sampled contralateral tibial bones) at 12 and 18 weeks after surgery. The results indicate that polymerase chain reaction for the Y chromosome is a useful tool for differentiating between donor and recipient cell populations experimentally using sex-mismatched tissues in a rat model. This study demonstrated that systemic chimerism occurs after successful vascularized bone transplantation. Transplanted cells not only survive in the graft but also gradually migrate into the recipient's body.     

Reconstruction of the eyebrow in the pediatric burn patient

We have reviewed our experience with reconstruction of eyebrow alopecia secondary to thermal injury in the pediatric patient. Reconstruction was performed with free composite strip grafts or vascularized island pedicle flaps. The complication rates for eyebrows reconstructed with vascularized island pedicles with respect to loss of a significant portion of the flaps (30.8 percent) and malalignment of the grafts (23.1 percent) were significantly greater (p less than 0.001) than the significant tissue loss (10.6 percent) or graft malalignment (7.9 percent) observed for free composite grafts. Hair density was more predictably restored with the free composite graft technique (p = 0.0004). The patients reconstructed with composite grafts had 89.4 percent acceptable results in contrast to 38.5 percent acceptable results obtained with the island pedicle technique. Based on these findings, we reserve the use of the vascularized island pedicle technique for male patients with unilateral alopecia and heavy hair density in the remaining eyebrow and in cases where free composite grafts have failed. The remaining patients are initially treated with free composite grafts with acceptable results in the overwhelming majority of cases.     

A torsional strength comparison of vascularized and nonvascularized bone grafts

Comparisons of torsional strength are made on the ulnae from the forelegs of short haired hounds where a nonvascularized graft was performed on one leg and a vascularized graft performed on the other. By using the forelegs of a dog as the experimental model and microsurgical techniques, a vascularized bone segment was used to graft a five centimeter nonunion in one leg and at the same time a conventional bone graft was performed on a similar nonunion in the other leg. Similar segments of rib bone were used for each graft. Torsional strength data are shown for nine experimental animals. A successful method for mounting the bones for testing of torsional strength in a torsion machine is given. In each case for which the bones healed properly, the vascularized bone graft proved to be significantly stronger in torsion.     

The early revascularization of membranous bone

The experimental finding that membranous onlay bone grafts maintain volume and viability to a greater extent than do endochondral grafts may be related to the more rapid vascularization of membranous bone. Microangiographic techniques were used to study the rates of vascularization of membranous and endochondral bone grafts in adult white New Zealand rabbits at 1, 3, 7, 14, and 21 days after bone grafting. Vascularization patterns were quantified microscopically using a modified point-counting technique. At 3 days, membranous bone grafts demonstrated vessel ingrowth from both soft tissue and host bone. Little ingrowth was seen in endochondral grafts. By day 7, 2.5 vessels per square were identified entering membranous grafts, while an average of 0.6 vessels per square were counted for endochondral bone grafts. At day 14, there was an average of greater than 20 vessels per square for membranous grafts versus 1.8 for their endochondral counterparts. At 21 days, the endochondral grafts demonstrated persistent avascular central areas not seen in membranous grafts. Membranous onlay bone grafts in the rabbit are more rapidly vascularized than endochondral grafts. This factor may affect the greater volume maintenance seen in experimental membranous grafts.     

Growth in revascularized bone grafts in young puppies.

The results of microvascular transfers of growing ulnas in puppy forelegs have been studied. These transfers were carried out both heterotopically and orthotopically. The growth in the revascularized bone grafts has been compared to that in heterotopic, nonvascularized ulna transfers and to normal ulnar growth. The growth in the vascularized bone grafts was significantly greater than in the nonvascularized grafts, but significantly less than in normal ulnar growth. A metaphyseal contribution to the blood supply of the growing portion of long bones is suggested.     

Differential patterns of incorporation and remodeling among various types of bone grafts

The early (3 months) and later (6 months) patterns of incorporation and bone formation have been evaluated histomorphometrically for different types of bone grafts; that is, vascularized and nonvascularized autografts with and without ciclosporin, and vascularized and nonvascularized dog leukocyte antigen (DLA)-mismatched allografts with and without ciclosporin. The vascularized bones were superior to the nonvascularized ones in healing and remodeling their grafted segments. In the autograft bones, ciclosporin did not alter the incorporation process 3 months after transplantation but delayed and increased the remodeling activities in the long run (6 months). Nonvascularized allografts underwent vigorous resorption, and were markedly porotic. Ciclosporin administration significantly reduced resorption and enhanced remodeling in nonvascularized allografts. The remodeling of allografts was similar to that of autografts in the presence of ciclosporin, but stopped soon after the administration of ciclosporin ceased.     

Fixation effects on membranous and endochondral onlay bone graft revascularization and bone deposition.

This paper investigates the relationship between bone resorption, the process of bone revascularization, and graft fixation. Vital staining techniques and microangiography were used to study the extent of graft revascularization of fixed and nonfixed endochondral (rib) and membranous (skull) onlay bone grafts in 20 adult sheep mandibles bilaterally. This assessment was carried out at 2 and 20 weeks postoperatively. Sequential fluorochrome staining was performed to examine the pattern of new bone deposition. Fixation was achieved using the lagscrew technique. At 2 weeks, membranous bone demonstrated a greater area of graft revascularization if fixed than if the graft was not fixed. The opposite result was seen for endochondral grafts, where nonfixed grafts showed a greater area of revascularization than fixed grafts. At 20 weeks, all bone that was present was fully vascularized. The inconsistencies in the results on the relationship between fixation and revascularization for membranous and endochondral grafts in the early stages of healing (2 weeks) suggest that although revascularization is a necessary precondition for bone resorption and deposition, biomechanical and structural factors may be a more satisfactory explanation for the differences observed in the maintenance of bony volume.     

Growth and survival of vascularized and nonvascularized membranous bone: an experimental study

Although nonvascularized membranous bone grafts to the craniofacial skeleton demonstrate improved survival over similar grafts of endochondral origin, the comparative fate of vascularized membranous grafts is unknown. It is also unknown whether onlay membranous bone grafts in immature animals have the ability to grow. To examine these questions, a model was developed in New Zealand white rabbits in which a segment of the zygomatic arch was transferred to the subjacent mandible as either a vascularized or nonvascularized transfer. At harvest 16 weeks later, residual graft volume and bone architecture were analyzed. Results demonstrate no improved survival for vascularized membranous grafts in adult animals (n = 7), while in the immature animals (n = 6), growth of the vascularized bone transfers was documented. We conclude that in the majority of instances in craniofacial reconstruction, nonvascularized onlay membranous grafts are to be preferred. Specific instances for the use of vascularized transfers will be discussed.     

Bone reconstruction of the lower extremity: complications and outcomes

A study was performed to analyze the results and final outcomes of bone reconstruction of the lower extremity. Twenty-six patients presented with type IIIB open fractures, nine with type IIIC open fractures, and 15 with chronic osteomyelitis. Seven patients underwent primary amputation, and reconstruction was attempted for 43 patients. The mean bone defect size was 7.7 cm (range, 3 to 20 cm). Bone reconstruction was achieved with conventional bone grafts in 16 cases, in association with either local (13 cases) or free (three cases) flaps. Vascularized bone transfer was performed in 24 cases, with either osteocutaneous groin flaps (10 cases), soleus-fibula flaps (12 cases), or osteocutaneous lateral arm flaps (two cases). For three patients, bone reconstruction was performed with a technique that combines the induction of a membrane around a cement spacer with the use of an autologous cancellous bone graft. Infections were observed to be responsible for prolonged hospital stays and treatment failures. The cumulative rates of sepsis were 4.6 percent at 1 week after injury and 62.8 percent at 2 months. Vascular complications were also related to infections and were responsible for four secondary amputations. One patient asked for secondary amputation because of a painful nonfunctional lower limb. Bone healing occurred in 37 of 43 cases, and the average time to union was 9.5 months, with an average of 8.7 procedures. The mean lengths of stay were 49 days for conventional bone grafts and 62 days for vascularized bone grafts. All of the 50 patients were able to walk, with an average time of 14 months. All of the patients with amputations underwent prosthetic rehabilitation. Patients mostly complained about the reconstructed limb (62.8 percent). Joint stiffness was present in 40 percent of the cases. Other long-term complications were pain (nine cases), lack of sensation (five cases), infection (five cases), and pseudarthrosis (one case). However, all of the patients with successful reconstructions preferred their salvaged leg to an amputation. Of 41 patients who were working before the injury, 26 returned to work.     

Long-term remodeling of vascularized and nonvascularized onlay bone grafts: a macroscopic and microscopic analysis

The present study was performed to compare vascularized and nonvascularized onlay bone grafts to investigate the potential effect of graft-to-recipient bed orientation on long-term bone remodeling and changes in thickness and microarchitectural patterns of remodeling within the bone grafts. In two groups of 10 rabbits each, bone grafts were raised bilaterally from the supraorbital processes and placed subperiosteally on the zygomatic arch. The bone grafts were oriented parallel to the zygomatic arch on one side and perpendicular to the arch on the contralateral side. In the first group, vascularized bone grafts were transferred based on the auricularis anterior muscle, and in the second group nonvascularized bone grafts were transferred. Fluorochrome markers were injected during the last 3 months of animal survival, and animals were killed either 6 or 12 months postoperatively. The nonvascularized augmented zygoma showed no significant change in thickness 6 months after bone graft placement and a significant decrease in thickness 1 year after graft placement (p < 0.01). The vascularized augmented zygoma showed a slight but statistically significant decrease in thickness 6 months after graft placement (p < 0.003), with no significant difference relative to its initial thickness 1 year after graft placement. In animals killed 6 months after bone graft placement, both the rate of remodeling and the bone deposition rate measured during the last 3 months of survival were significantly higher in the vascularized bone grafts compared with their nonvascularized counterparts (p < 0.02). By 1 year postoperatively, there were no significant differences in thickness, mineral apposition rate, or osteon density between bone grafts oriented perpendicular and parallel to the zygomatic arch. These findings indicate that the vascularity of a bone graft has a significant effect on long-term thickness and histomorphometric parameters of bone remodeling, whereas the direction of placement of a subperiosteal graft relative to the recipient bed has minimal effect on these parameters. In vascularized bone grafts, both bone remodeling and deposition are accelerated during the initial period following graft placement. Continued bone deposition renders vascularized grafts better suited for the long-term maintenance of thickness and contour relative to nonvascularized grafts.     

Bone regeneration within a coralline hydroxyapatite implant.

The hypothesis that incomplete resorption of osteons in an autogenous cortical bone graft may limit its replacement by new bone regeneration was explored by implanting a hydroxyapatite replica of a coral skeletal structure into bone gaps. This implant contained channels and interconnections similar to those in osteon-evacuated bone grafts. In 6 implanted mandibular defects in dogs, two of which were examined at two, 4, and 6 months, 11 percent, 46 percent, and 88 percent of the implant areas were filled with regenerated bone. The regenerated bone was a woven type at two months, but changed to a lamellar type by 6 months. In two implanted defects examined at 12 months, biodegradation of 29 percent of the implant had occurred. The bone regeneration was physiological, the implant was biocompatible, and the biodegradation began after the bone had regenerated.     

The comparative analysis of homologous fresh frozen bone and autogenous bone graft,associated or not with autogenous bone marrow,in rabbit calvaria: a clinical and histomorphometric study

The aim of this study was to evaluate the potential of fresh frozen homologous and autogenous grafts, associated or not with autogenous bone marrow, to form bone. Sixty titanium cylinders were used, and were fixed to the skulls of 30 rabbits. These cylinders were filled with (A) autogenous bone (AM) autogenous bone associated with the bone marrow (H) fresh frozen homologous bone (HM) fresh frozen homologous bone associated with the bone marrow (M) pure autogenous bone marrow and (C) blood clot. The animals were sacrificed after 02 and 03 months. After clinical evaluation, the samples were stained with hematoxylin, eosin and Mallory Trichrome dyes for optical microscopy analysis and histomorphometric analysis. Experimental groups that received mineralized materials (A, AM, H, HM) showed the best bone formation results, presenting no statistical difference between them (P > 0.05). Groups that did not receive mineralized materials (M and C) showed the worst results (P < 0.05), but the M group showed better results than the C group. Most of the autogenous and homologous bone particles were resorbed and there was a larger amount of residual particles in the homologous graft (H, HM) when compared with the autogenous graft (A, AM; P < 0.05). These findings suggest that fresh frozen homologous grafts produced similar amounts of new bone when compared with the autogenous grafts. However, the amount of residual bone particles was larger in the homogenous groups, which may indicate a slower remodeling process. The homologous fresh frozen bone seems to be a good osteoconductive material. The use of only autogenous bone marrow showed better results when compared to the bood clot. However, this research indicates that association with mineralized materials is required.     

The effects of vascularity and cyclosporin A on the mechanical properties of canine fibular autografts.

We studied the biomechanical behavior of orthotopic canine autografts as influenced by vascularized supply and the administration of cyclosporin A at three months and six months post-surgery. The model was the proximal 8 cm of the fibula in young adult dogs. In vascularized grafts, blood supply was re-established by microvascular re-anastomosis. Experimental controls were sham-operated and unoperated bones. Mid-graft test sections were subjected to loading-to-failure in torsion to determine the strength and stiffness. In both three- and six-month groups, vascularized grafts were significantly stronger and stiffer than contralateral nonvascularized grafts. Vascularized grafts were not significantly different from sham-operated bones. A 30-day regimen of cyclosporin A was found to have no measurable effect on mechanical properties for any individual treatment group. The results indicate that re-established blood supply can be a major factor in maintaining the mechanical integrity in large-segment cortical autografts.     

Fixation effects on membranous and endochondral onlay bone-graft resorption

Difficulties arise in the prediction of maintenance of graft volume over time when bone grafts are used for facial contour reconstruction. We hypothesize that graft fixation will decrease movement and lead to decreased resorption. Fixed and nonfixed endochondral (rib) and membranous (skull) onlay bone grafts measuring 30 X 10 X 4 mm were grafted to the mandible bilaterally in 10 adult sheep. Fixation was achieved using the lag-screw technique. Volume measurements using caliper technique were made 20 weeks postoperatively. The volume of graft present at 20 weeks was significantly greater for the fixed bone grafts (p less than 0.001): fixed membranous, 85.9 percent; fixed endochondral, 76.2 percent; nonfixed membranous, 55 percent; and nonfixed endochondral, 16.6 percent. The results are explained using biomechanical theories related to the effects of strain. At present, it is suggested by this study that when onlay bone grafts are stabilized, improved results with respect to graft resorption can be expected.     

Vascularized bone allograft transplantation in a genetically defined rat model

A heterotopic subcutaneous model for experimental vascularized bone allograft transplantation has been presented. This model uses genetically defined rats and allows serial assessment of graft viability. The reliability of this model has been proven by successful isograft transplantation. This model was used to study the effect of matching at the major histocompatibility complex on vascularized bone allograft survival. Whereas grafts transplanted across a minor histocompatibility barrier survived until sacrifice, grafts transplanted across a major histocompatibility barrier were victims of an acute rejection process. This study, therefore, showed genetic disparity to be a critical determinant of vascularized bone allograft survival. It indicates that primary vascularized bone allografts are as susceptible to rejection as heart and kidney allografts. For these reasons, it can be anticipated that genetic matching will be important in clinical vascularized bone allograft transplantation. The model used in this study should be useful for obtaining further fundamental immunologic information concerning vascularized bone allograft transplantation.     

One-stage reconstruction of composite bone and soft-tissue defects in traumatic lower extremities

Management of bone loss that occurs after severe trauma of open lower extremity fractures continues to challenge reconstructive surgeons. Sixty-one patients who had 62 traumatic open lower extremity fractures and combined bone and composite soft-tissue defects were treated with the following protocol: extensive debridement of necrotic tissues, eradication of infection, and vascularization of osteocutaneous tissue for one-stage bone and soft-tissue coverage reconstruction. The mechanism of injury included 49 motorcycle accidents (80.3 percent), five falls (8.2 percent), three crush injuries (4.9 percent), two pedestrian-automobile accidents (3.3 percent), and two motor vehicle accidents (3.3 percent). The bone defects were located in the tibia in 49 patients (79 percent; one patient had bilateral open tibial fractures), in the femur in seven patients (11.3 percent), in the calcaneus bone in four patients (6.5 percent), and in the metatarsal bones in two patients (3.2 percent). The size of soft-tissue defects ranged from 5 x 9 cm to 30 x 17 cm. The average length of the preoperative bony defect was 11.7 cm. The average duration from injury to one-stage reconstruction was 27.1 days, and the average number of previous extensive debridement procedures was 3.4. Fifty patients had vascularized fibula osteoseptocutaneous flaps, six had vascularized iliac osteocutaneous flaps, and five patients had seven combined vascularized rib transfers with serratus anterior muscle and/or latissimus dorsi muscle transfers. One patient received a second combined rib flap because the first combined rib flap failed. The rate of complete flap survival was 88.9 percent (56 of 63 flaps). Two combined vascularized rib transfers with serratus anterior muscle and latissimus dorsi muscle flaps were lost totally (3.2 percent) because of arterial thrombosis and deep infection, respectively. Partial skin flap losses were encountered in the five fibula osteoseptocutaneous flaps (7.9 percent). Postoperative infection for this one-stage reconstruction was 7.9 percent. Excluding the failed flap and the infected/amputated limb, the primary bony union rate after successful free vascularized bone grafting was 88.5 percent (54 of 61 transfers). The average primary union time was 6.9 months. The overall union rate was 96.7 percent (59 of 61 transfers). The average time to overall union was 8.5 months after surgery. Seven transferred vascularized bones had stress fractures, for a rate of 11.5 percent. Donor-site problems were noted in six fibular flaps, in two iliac flaps, and in one rib flap. The fibular donor-site problems were foot drop in one patient, superficial peroneal nerve palsy in one patient, contracture of the flexor hallucis longus muscle in two patients, and skin necrosis after split-thickness skin grafting in two patients. The iliac flap donor-site problems were temporary flank pain in one patient and lateral thigh numbness in the other. One rib flap transfer patient had pleural fibrosis. Transfer of the appropriate combination of vascularized bone and soft-tissue flap with a one-stage procedure provides complex lower extremity defects with successful functional results that are almost equal to the previously reported microsurgical staged procedures and conventional techniques.     

Cranial reconstruction for metastatic breast cancer

All women with advanced breast cancer who are medically stable despite their disease are candidates for tumor extirpation and reconstruction. Advanced breast cancer today is incurable, and many prognostic factors can be used to try to predict a clinical course and response to therapy; however, no guidelines are available. Our case report most likely represents a metastasis to the calvarium with intracranial extension, reported to occur in about 3 percent of primary breast cancer patients. As demonstrated here, tumor ablation with immediate, one-stage reconstruction of large scalp defects is possible without the need for free tissue transfer or a delay in adjuvant therapy. Local tissue rearrangement has been employed for coverage of defects up to 50 percent of the cranium. The resulting donor defects can be closed with split-thickness skin grafts over pericranium. Serial tissue expansion and rearrangement can be used secondarily to replace skin grafts with hair-bearing scalp. Bony defects can be managed with either autogenous or alloplastic materials. Split-calvarial bone grafts can be harvested from the same operative field and cover small to medium-sized defects. Other sources of autogenous grafts include split ribs and iliac bone. Metals, calcium ceramics, and polymers such as methylmethacrylate can be used to cover intracranial contents and restore calvarial contour when defects are large or when autogenous material is not available. Palliation from tumor burden, prevention of pathologic fracture and oncologic emergencies, controlling pain, and enhancing quality of life are the goals of the oncologic and reconstructive surgeons in cases of advanced breast cancer. These goals are becoming even more important as new forms and combinations of chemotherapy, radiation, and gene therapy are extending the life expectancy of women with breast carcinoma.     

Use of the AO plate for immediate mandibular reconstruction in cancer patients

Free vascularized bone grafts have revolutionized mandibular reconstruction, yet their use in all mandibulectomy patients is not always necessary. A recently developed alternative to bony reconstruction has been the use of the AO reconstruction plate. We compared the use of the AO reconstruction plate with immediate free bone graft mandibular reconstruction in 31 patients. Reconstruction plates were used in 20 and immediate free bone grafts were used in 11 patients. The overall success rate for use of the plate was 15 of 20 (75 percent). There were 6 anterior reconstructions, of which only 2 (33 percent) were successful. This is opposed to 13 of 14 (93 percent) lateral reconstructions that were successful in lateral plate placements. There were 11 immediate composite free flaps: 4 iliac crest, 4 scapula, 2 fibula, and 1 composite radial forearm flaps. Six repairs were for anterior defects, and there were 5 full-thickness defects, 3 of which were in the anterior position. All 11 flaps were successful. In conclusion, we believe the reconstruction plates are a useful adjunct for mandibular replacement in the head and neck cancer patient but should be reserved for lateral defects. For anterior reconstructions, even in patients with locally advanced disease, free-tissue transfer of composite osteocutaneous flaps is the reconstructive method of choice.