Intraoperative photograph of fronto-orbital craniotomy model. Note intact dura and that the posterior border of osteotomy is defined by the coronal suture. Exposed frontal sinus is located at the bottom left of the craniotomy.
Craniometric landmarks (Felis domestica) and craniometric measurements. OB indicates orbital breadth; OH, orbital height; B, basion OB; N, nasion (A to B); F, frontal midpoint (A to ECT); A, akrokranion OH; ENT, entorbitale (F to ECT); and ECT, ectorbitale (N to ENT).
Two specimens from group 3. Note reestablishment of the left coronal suture and smooth implant borders (left) and areas of new bone intersecting implant (right). The implant is indicated by arrowheads.
Undecalcified section of hydroxyapatite cement–reconstructed calvaria 5 months after implantation. New bone (green) is seen intersecting the original implant (brown). Note the direct apposition of bone to implant and the almost complete coverage of the implant by bone (mineralized bone stain, ×10).
Lykins CL, Friedman CD, Costantino PD, Horioglu R. Hydroxyapatite Cement in Craniofacial Skeletal Reconstruction and Its Effects on the Developing Craniofacial Skeleton. Arch Otolaryngol Head Neck Surg. 1998;124(2):153-159. doi:10.1001/archotol.124.2.153
To assess the effects of hydroxyapatite cement (HAC) on the developing feline craniofacial skeleton.
Fronto-orbital craniotomies were performed on 14 kittens and reconstructed by autograft or HAC. By design, animals in which the craniofacial skeleton was reconstructed with HAC also underwent obliteration of the left frontal sinus. After achievement of skeletal maturity, animals were sacrificed and compared by 11 standardized cranial measurements obtained by sliding caliper. Additional analyses included histological studies, histomorphometry, and computed tomography.
Twenty-one 12-week-old female cats were divided into 3 groups, composed of 7 specimens.
The control animals underwent periosteal elevation alone (group 1). The remaining animals underwent unilateral fronto-orbital craniotomy and subsequent reconstruction with orthotopic bone flap replacement (group 2) or HAC (group 3).
All animals survived the study with no evidence of wound infection or implant failure. Gross morphological studies demonstrated excellent contour reconstruction in both experimental groups. Craniometric analysis detected 1 intergroup difference that consisted of a wider skull in group 3 on the reconstructed side. An intragroup difference in orbital height was also seen in group 3. Computed tomography demonstrated a solid appearance of the implant with obliteration of the left frontal sinus in group 3. Histological studies showed that HAC was osseointegrated to native bone, with areas of new bone interspersed throughout the implants. No significant inflammatory response or fibrous encapsulation was noted. Histomorphometry demonstrated that implants were replaced by osseous tissue in 44% to 50% of the animals within 5 months.
Hydroxyapatite cement is safe and effective for craniofacial reconstruction in the developing feline and may be appropriate for similar applications in humans.
ALLOPLASTIC implants have a long history of use in the contour reconstruction of the craniofacial skeleton. Despite the variety of materials available to the surgeon today, none currently fits the criteria of an ideal implant as set by Scales.1 These criteria require a material to be chemically inert, incapable of inducing hypersensitivity or foreign body reaction, noncarcinogenic, and easily shaped. Another level of complexity is added when alloplastic implants are used in the developing craniofacial skeleton because studies on the most commonly implanted materials, titanium plates and screws, have pointed to local growth restriction and distal compensation as the neurocranium presumably attempts to maintain a normal volume in the presence of locally restricted bone growth.2- 9 As a result, autogenous bone has remained the standard material for the repair of bony defects in the developing craniofacial skeleton. A new class of alloplastic materials, represented by hydroxyapatite cement (HAC; OsteoGenics Inc, Richardson, Tex), may avoid some of the shortcomings of traditional materials and allow greater ease in the contour reconstruction of the developing craniofacial skeleton.
Hydroxyapatite cement differs from other hydroxyapatite preparations in that it is nonceramic and may be applied as a paste that can be contoured intraoperatively. Developed at the Paffenbarger Research Center of the American Dental Association Health Foundation, Gaithersburg, Md, HAC is composed of tetracalcium phosphate and dicalcium phosphate anhydrous. When placed in an aqueous environment, these compounds react isothermically to form hydroxyapatite. The cement sets in 15 minutes, with conversion to hydroxyapatite in 4 hours.10 Testing has shown the compressive strength of the material to be approximately 37 MPa, which, while less than ceramic hydroxyapatite, is sufficient for the reconstruction of non–load-bearing bone.11 Improved material formulation has increased this value to 60 MPa. Pilot studies have demonstrated histological behavior similar to ceramic hydroxyapatite, namely excellent biocompatibility with no evidence of toxic reaction, no foreign body giant cell response, and osseointegration of the implant to bone.12 Unlike ceramic hydroxyapatite, however, animal studies using HAC have found this material capable of osseoconversion. When used in cranioplasties in an adult feline model, HAC was progressively replaced by bone without significant loss of implant volume.13 Friedman et al13 found that after 6 months, new bone accounted for only 5% of the implant volume, with this value increasing to 63% at 18 months. In all of the subjects, fibrous tissue never represented more than 10% of new tissue. In addition, the study by Friedman et al13 demonstrated successful obliteration of the feline frontal sinus.
Twenty-one 12-week-old cats (Felis domestica) were used in the study. All guidelines set by the Yale University (New Haven, Conn) Animal Care and Use Committee for animal experimentation were followed. Surgery was performed under sterile conditions using endotracheal isoflurane anesthesia following induction with intramuscular ketamine hydrochloride (25 mg/kg) and atropine (0.04 mg/kg). A single midline scalp incision extending from the nasal dorsum to the inion was made, following which the overlying fascia, temporalis muscle, and periosteum were dissected to reveal the frontoparietal skull. Following the group-specific surgical intervention, the tissues were closed in layers using resorbable sutures.
Postoperatively, all animals received 0.1 mL of buprenorphine hydrochloride before extubation for pain control. Antibiotic coverage consisted of 250 mg of cefazolin given before and after the operation. In addition, 2 mg of dexamethasone was given before and after the operation to minimize postoperative edema. All care was overseen by the Yale University Veterinary Care Service to ensure proper and humane animal treatment.
Animals were randomly assigned to 1 of 3 groups, consisting of 7 cats each. Animals in group 1 served as controls and underwent periosteal elevation alone. In groups 2 and 3, a pneumatic side-cutting burr was used to complete a left fronto-orbital craniotomy similar to that used in the study by Lin et al.2 Unlike that study, however, the coronal suture served as the posterior margin of the osteotomy (Figure 1). During the bone resection, saline irrigation was applied to minimize thermal injury to the bone. The craniotomy was designed to overlie the expanding portion of the craniofacial skeleton associated with the developing eye, the pneumatizing frontal sinus, and the enlarging frontal lobe. The dura mater was carefully preserved intact during the craniotomy procedure. In 2 separate procedures, small dural perforations occurred, but no specific dural repair was performed because of the limited nature of the perforations. By design, the frontal sinus was disrupted by the osteotomy because the frontal sinus is commonly encountered and ablated in the surgical repair of plagiocephaly. Following exposure of the sinus, the entire mucoperiosteum was carefully removed. Next, the bone flap was returned to its orthotopic position in animals in group 2 and secured in place by reapproximation of the periosteum. The surgical defects in animals in group 3 were repaired using HAC. Hydroxyapatite cement was mixed immediately before implantation using 0.3 mL of sterile water per gram of cement, resulting in the creation of a dense paste that was then shaped to fill the defect, including the exposed left frontal sinus.14 Following contour reconstruction of the defect using the contralateral side as a model, the cement was allowed to dry for 15 minutes before a drain was placed and the tissue closed in layers. In most cases, only 1 g of cement was required to completely reconstruct the defect.
All animals were sacrificed 5 months after the operation, at the time coinciding with full skeletal maturity. Animals were sedated using intramuscular ketamine hydrochloride, after which a lethal intracardiac dose of pentobarbital sodium (14.3 mg/kg) was administered. After the animals were sacrificed, the overlying soft tissues were sharply dissected from the skull, and the specimens were preserved in 10% formalin. Before craniometric analysis, thin-section coronal computed tomography scans were obtained on all animals using a helical scanner (GE Helical Scanner, General Electric, Medical Imaging Group, Milwaukee, Wis). Craniometric analysis was performed using an SPI 150-mm polydial sliding caliper. To minimize error, all measurements were performed in triplicate, and an average was obtained. In total, 5 paired measurements and 2 midline measurements were obtained using standardized craniometric landmarks (Figure 2). To account for variations in skull size between individual animals, all measurements were normalized by dividing each individual measurement by the skull length, resulting in the creation of a unitless ratio. The full cranial module15 was not used because we believed that 3 measurements introduced more sources of potential error into each ratio. We decided, with concurrence of R. Sherwood, PhD (Yale University Department of Physical Anthropology, personal communication, September 1994), that 1 consistently reproducible measurement, in this case the length of the upper neurocranium defined as the distance from the nasion to the akrokranion, was the most accurate way to standardize each ratio. Ratios between the experimental groups were then compared using a multivariate analysis of variance (MANOVA). After the demonstration of any statistically significant differences among the 3 groups, follow-up univariate analyses of variance (ANOVAs) and Student-Newman-Keuls multiple comparison procedures were performed to compare individual measurements between groups and between sides within each group.
All histological studies and histomorphometry were performed at the Harrington Arthritis Research Center, Phoenix, Ariz. The operative site was first removed en bloc, after which specimens were dehydrated in 70% to 100% ethanol, cleared in xylene, and infiltrated and embedded in methyl methacrylate. Undecalcified specimens were cut with a microtome at 15- to 20-µm sections using a tungsten carbide–tipped knife. Specimens were stained using a Paragon stain (Ladd Multiple Stain, Burlington, Vt) or Villanueva mineralized bone stain (Harrington Arthritis Research Center) for delineation of osteoid, mineralized bone, and fibroconnective tissue. All histomorphometry was performed using a fluorescence microscope (Zeiss, Danbury, Conn) and a digitizing system using the Bioquant System IV program (R&M Biometrics Inc, Nashville, Tenn).
All animals survived the duration of the study. There were no wound infections, toxic reactions, or implant extrusions in any animals. Before the animals were sacrificed, no gross facial abnormalities or growth disturbances were detected among the groups. Our observations of the dissected whole calvarial gross specimens are noted in the following paragraphs.
In group 1, no gross asymmetries were noted. In group 2, the consistent changes noted in all animals included a taller orbit, an altered shape of the superior orbital rim, and a thinner lateral spine of the frontal bone on the side of the skull on which the operation was performed. Despite the involvement of the osteotomy of a large portion of the left coronal suture, 3 of the 7 animals were found to retain what appeared to be a coronal suture on the reconstructed side. In addition, 4 animals were noted to have slight asymmetries of the frontal bone, which we believed to be the result of the reconstruction failing to restore normal preoperative form.
In group 3, consistent morphological changes were similar to those noted for animals in group 2, namely, a taller orbit, asymmetry in the shape of the orbital rim, and a narrower, more elongated lateral spine of the frontal bone on the side on which the operation was performed. Again, despite the ablation of a large portion of the coronal suture in the osteotomy, 4 of the 7 animals had the formation of a new coronal suture. In 2 of the 4 cases, this appeared as a new strip of bone that divided the implant in half. Also, despite the possibility of surgical error in reconstruction of the frontal bone, only 4 of the animals appeared to have asymmetric heights of the frontal bone. In all cases, there appeared to be excellent integration of the implant, with significant osseoconversion to bone. Implants ranged in appearance from solid, light yellow, and triangular to multiple small circular islands of HAC with new bone interposed between the pieces of the implant (Figure 3). It is interesting that in all animals, the entirety of the orbital rim was composed of new bone despite the complete reconstruction with HAC of the posterior half of the superior orbital rim.
The initial MANOVA was performed to detect any differences in skull shape when skulls were compared as a whole on the side of the operation and the contralateral side. A statistically significant difference existed between groups (P<.05) and between the reconstructed and contralateral sides of the skulls (P<.01). The interactive effect between the surgical group and side of the skull was not significant. Follow-up ANOVAs were then run to further clarify these general findings.
A statistically significant difference in the measurement from the frontal midpoint to the ectorbitale was noted when the reconstructed side of the skull was compared between groups (P<.01). A Student-Newman-Keuls multiple comparison procedure demonstrated that this distance in group 3 was significantly greater than in group 1 or 2 (P<.01). This finding indicates that animals in which the operative defects were reconstructed with HAC had wider skulls in the area adjacent to the site of reconstruction. No significant differences in any measurements were noted when the unoperated sides of the skull were compared between groups.
A significant difference in the orbital heights of animals in group 3 was noted when sides were compared within that group. No significant intragroup differences were noted in group 1 or 2. A second ANOVA was computed on the single midline measurement of akrokranion to basion, which detected no statistically significant difference between the 3 groups.
It is interesting that the differences in orbital height approached significance (P=.08) when the side of the skull on which the operation was performed was compared between groups. Study of the normalized craniometric measurements showed that the average orbital height of group 2 (0.409) was greater than that in the control group or the HAC group (Table 1). In addition, a trend (P=.09) was also seen when orbital heights were compared within group 2.
All specimens reconstructed with HAC had radiographic evidence of excellent contour reconstruction. Implants had a solid appearance on computed tomography, with a signal density similar to that of cortical bone. There was no evidence of defect formation within the substance of the implants. All specimens had complete obliteration of the left frontal sinus with no cystic areas that suggest mucocele formation.
Consistent with the findings of previous studies, we noted no significant inflammatory response or fibrous encapsulation of the implant.12- 14,16 In all cases, implants were found to be in direct contact with bone, with areas of new bone interspersed throughout the implant (Figure 4). New tissue formation was predominantly new bone with little fibroconnective tissue. Histomorphometry was performed on 3 specimens in group 3 (Table 2). During 5 months, approximately 50% of the implant volume was converted to new cortical and trabecular bone. Of this trabecular bone, fibrovascular tissue accounted for 6.8% to 18.4% of the total volume, with the remainder consisting of new bone.
In the present study, the effects of HAC on the developing craniofacial skeleton were assessed when used in the contour reconstruction of a large cranial defect. In addition to its excellent biocompatibility, HAC is replaced by new bone over time, with maintenance of shape and volume. These properties would seem to make HAC an ideal material for use in pediatric craniofacial surgery, which often requires extensive osteotomies and autologous nonvascularized bone grafts. The cement properties of HAC would obviate the need for rigid fixation, which may avoid some of the growth restrictions observed in previous developmental studies.2- 9 We hope also that the ability of HAC to be osseoconverted to new bone will allow the implant to permit growth within the remainder of the craniofacial skeleton. Because HAC is not approved for use in the pediatric population, this study was intended to serve as a pilot study for the eventual use of this material in children.
Cats were chosen as the model for this experiment because of previous baseline investigations that used cats and to avoid the costs and limited animal availability associated with the use of a primate. Cats are a phylogenetically closer model to humans than are rabbits and are considered a "short-faced" species, like humans and unlike the dog. Studies have suggested that the domestic cat may be an economical alternative animal model to the use of nonhuman primates, at least for the study of human midfacial growth.17,18 A large fronto-orbital craniotomy was used in an effort to simulate the osteotomy used in the surgical repair of unilateral coronal synostosis (plagiocephaly). The coronal suture was largely extirpated on the reconstructed side in an effort to create the greatest growth disturbance possible.
This study was patterned partially on a study by Lin et al,2 a developmental study that used cats as a model. We believed that this would in part allow comparison of the effects of HAC to rigid fixation. Unfortunately, 2 of the measurements used in the study by Lin et al2 (ie, height of the frontal bone and frontal sinus volume) could not be used, preventing direct comparison. We believed that changes detected in the height of the frontal bone would reflect the accuracy of reconstruction and that changes caused by growth disturbance alone would be impossible to isolate. We believe the measurements used in the present study and the study by Lin et al2 consistently used identifiable landmarks that are well established in the archeological literature.19
Consistent with the findings of previous studies, we found that HAC was well tolerated, with no evidence of toxic reaction, implant failure, or wound infection. This last point is particularly remarkable because 3 animals in this group experienced small dural tears during the osteotomy, with direct exposure of the intracranial contents to the frontal sinus. While there are no reported studies on the effects of HAC when directly applied to neural tissue, previous studies have demonstrated that HAC does not induce scar formation, inflammation, or changes in cortical architecture when applied directly to the dura in adult cats. If direct neural tissue contact were to occur, we would anticipate no adverse wound healing effects, and the expected histological response (although unknown and not the focus of the present study) would be the same as direct surgical manipulation or incision of neural tissue. Supporting the findings of a previous study by Friedman et al,13 we found HAC to be effective in obliteration of the frontal sinus. In conjunction with clinical evidence, computed tomography demonstrated complete obliteration of the left frontal sinus, with no evidence of cystic structures that would suggest mucocele formation.
In all cases, implants were in direct contact with bone, with no evidence of fibrous encapsulation. In addition, histomorphometry revealed rapid replacement of the implant by bone. In comparison with the results of the study by Friedman et al13 of adult cats, which showed 5% bony replacement of HAC in 6 months, our study produced 44% to 50% bony replacement of HAC in 5 months when kittens were used. This finding demonstrates that HAC implants are able, at least in part, to respond to different physiologic wound environments with different performance in the form of more accelerated conversion to bone when placed in the developing skeleton. The ratio of soft tissue to bone in the new tissue also compares favorably with previous studies of hydroxyapatite that have claimed percentages of soft tissue from 31% to 42% when implants were explanted at 12 months.20,21 It is worth noting that the study by Holmes and Hagler21 found that the new tissue found in split-rib autografts consisted of 57% soft tissue and 43% bone when implants were retrieved after 6 months in an adult canine model. It would thus seem that the fibrovascular tissue component of HAC implants falls well within normal limits when compared with nonvascularized autogenous bone grafts.
Prior to sacrifice, all subjects were indistinguishable from one another morphologically. After craniometric analysis, animals that had undergone reconstruction with HAC seemed similar to both of the other experimental groups, differing in only 2 measurements despite a fairly radical reconstruction with an alloplastic material. The widening of the skull at the area of implantation is consistent with the study by Wong et al3 that showed an increase in skull width at the area of the plated coronal suture. This finding is not surprising because the suture responsible in part for the anteroposterior growth of the calvaria was disrupted. When bony growth is impeded in the area of a suture, growth tends to occur in a direction parallel to the involved suture.22 In addition, this measurement lies in an area that was partly reconstructed with HAC, so the surgical technique may contribute to this difference as well as to any growth disturbance potentially caused by this material.
Direct comparisons with the effects of rigid fixation in the study by Lin et al2 could not be made because the plated group differed on the side of operation from control subjects in 2 measurements that were not used in our study because of design differences. However, HAC-like rigid fixation did not cause widespread growth disturbances when implanted into an identical model for the same period. In addition, only 1 difference was noted between the sides within the HAC group (orbital height) compared with the osteotomy and plate fixation group in the study by Lin et al,2 which differed in 3 of the 7 measurements (ie, height of frontal bone, frontal sinus volume, and akrokranion-ectorbitale). It is interesting that the orbital height in group 3 was not significantly different from the orbital height in the other 2 groups when the reconstructed or contralateral sides were compared for intergroup differences. Rather, the intragroup difference in orbital height seen in group 3 seemed to be caused by a "taller" orbit on the reconstructed side and a "shorter" orbit on the contralateral side. Despite a more extensive craniotomy involving the surgical extirpation of a suture, almost normal growth was allowed to occur on both sides in the animals that underwent reconstruction with HAC. Again, surgical technique and accuracy of reconstruction could have contributed to the differences noted in orbital height.
It is interesting that the gross differences observed in orbital shape on the side of operation of dissected specimens in groups 2 and 3 were not significantly different from controls when compared quantitatively. This may point to the limitation of caliper measurements in detecting complex changes in shape. It is likely that 2 measurements to assess orbital shape are not enough to detect complex differences in a circular shape. This may point to the necessity of using more complex methods of measurement, such as euclidean distance matrix analysis,23- 25 or a fully triangulated surface area comparison. However, the clinical significance and validity of these analysis methods remains to be determined.
While these results are encouraging, there are shortcomings in the experimental design that prevent direct comparisons with humans. The surgical procedure in this study was performed in an animal model that had attained 70% of its maximum adult size. Because surgery often occurs at a much earlier stage of development in human craniofacial surgery, it is not possible to say whether the implantation of HAC at an earlier stage would have a more significant effect on subsequent growth. Specifically, the human brain doubles in size during the first 6 months of life and does not reach 80% of adult size until 2 to 3 years of age. This rapid period of growth has thus already occurred in this feline model, and the developmental effects of implanted materials may be underestimated. In addition, it is likely that the developmental pattern of the growing feline skull is not identical to that of the developing human. Because the cat possesses an incomplete orbit, it is difficult to speculate how the orbital disturbances observed in this study would manifest themselves in species with complete orbits, such as humans.
While osteotomies were performed in an effort to duplicate a clinical situation often encountered in pediatric craniofacial surgery, the osteotomies were performed on healthy rather than diseased bone, and in group 2, the bone was replaced in its original position without reshaping or fixation. While this does not correlate directly with the clinical situation, the absence of any statistical difference from controls indicates that osteotomy alone produces only mild growth disturbance in cats. It is unknown what effects HAC would have had if the coronal suture had not been partially ablated.
Despite these issues, the preliminary results of this study indicate that HAC can safely be used for craniofacial reconstruction in the developing feline and may be appropriate for similar applications in humans. Hydroxyapatite cement could therefore be a safe alternative to free bone grafts in the contour reconstruction of the developing craniofacial skeleton. In all instances, HAC seemed to be as well tolerated in the developing skeleton as in the adult model. Despite extensive reconstruction with an alloplastic material, HAC did not produce global growth disturbances, affecting only orbital height and parietal bone width. In addition, HAC, when used in kittens, permitted new bone formation at a rate much faster than that seen when used in adult cats. As with previous developmental studies, timing of surgical intervention may be the paramount factor in the extent of growth disturbance. While no statistically significant differences in growth were detected in group 2, the production of grossly altered orbital morphological features and a statistical trend toward larger orbits indicate that osteotomy alone results in a slight craniofacial asymmetry. Whether these changes are the result of surgical technique or are secondary to growth disturbance alone cannot be determined. As with all surgery, clinical judgment must be used. The benefits of immediate contour reconstruction must be weighed against the possibility of growth disturbance and the production of new dysmorphologic features with the passage of time.
Accepted for publication September 12, 1997.
Presented at the 1996 Fall Meeting of the American Academy of Facial Plastic and Reconstructive Surgery, September 28, 1996.
Reprints: Craig D. Friedman, MD, Craniofacial Tissue Engineering, 4 Greystone Farm Ln, Westport, CT 06880 (e-mail: C_Friedman@FCCC.edu).