Radiation Attenuated Mandibular Distraction Osteogenesis Healing Is Quantified By The Percent And Degree Of Bone Regenerate Quality Using Micro-CT (µCT) Algorithms

Daniel A. Schwarz, MD; Daniel A. Zeldes, BS; Mehreen Kakwan; Krikor Arman, BA; Ameen Jamali and Steven R. Buchman, MD

Introduction: Determining the degree by which therapeutic radiation (XRT) attenuates bone healing is integral before attempting to consider the use of mandibular distraction osteogenesis (DO) in head and neck cancer reconstruction. While prior animal studies have shown results which vary from non-union to purported success with lower therapeutic doses (1, 2), no documented metrics have outlined the extent of either XRT attenuated bone healing or the impact of radiation on the quality of DO regenerate. Our goal was to utilize non-destructive imaging measures to characterize the precise and initial deficits within DO after XRT in the lower therapeutic ranges. Once these baseline parameters are established, we can gradually increase the XRT to determine the order and extent of bone healing variables that are attenuated in a dose-response curve. Bone Mineral Density (BMD) and Bone Volume Fraction (BVF) are two key µCT measurements that quantify the degree of new bone formation, along with the percentage of bone within an established region, respectively. A recently validated, novel μCT algorithm, μ-Densitometry, will allow us to determine the degree of mineralization within the regenerate volume. Based upon this, we hypothesized that lower therapeutic XRT given pre-operatively, will decrease the amount of bone mineral density and impede the maturation of normal bone healing, assessed by the degree of mineralization.

Methods: Male Sprague-Dawley rats (~325-400g) underwent 36Gy pre-operative external beam irradiation (X-Ray), fractionated at 3.75Gy/day x 10 days to the left hemi-mandible followed by 2 weeks of recovery before surgery. A titanium frame distractor was secured bilaterally onto the mandible and a vertical osteotomy was created posterior to the 3rd molar on the left hemi-mandible (Figure 1). Our established DO protocol was performed: 4d latency, 0.3mm distraction every 12hrs x 8.5 days to a total of 5.1mm distraction. After 28 days of consolidation, animals were euthanized, mandibles harvested, and dissection of the surgical hemi-mandible was performed. Our un-operated controls (uC-XRT) followed the same XRT and 2 wk recovery protocol.

µCT: Mandibles were scanned at a 45μm resolution then reconstructed. Established region of interest (ROI) templates were placed by three independent reviewers (Figure 2)., using anatomical and surgical landmarks, centered to encompass the gap and matched between the DO-XRT and uC-XRT groups Bone mineral content (BMC), bone mineral density (BMD) and bone volume fraction (BVF), along with micro-densitometric mineralization assessment, in Hounsfield units, was calculated (3).

Data Analysis: We used data for DO and uC, not exposed to XRT, obtained from the preceding study using the exact protocols as described above. Groups: I) DO, n=24; II) DO-XRT, n=6; III) uC, n=18 and IV) uC-XRT, n=8. Data was recorded as mean +/- SD. We used ANOVA followed by appropriate post-hoc comparisons for statistical analysis, with a p-value of < 0.05 considered significant.

Results: While BMD was lower in the DO-XRT group compared to the DO regenerate group (p=0.06), no statistically significant differences were found when comparing BMD or BVF in the uC vs. uC-XRT. However, using μ-densitometry, there was a significantly greater amount of low and medium mineralized regenerate bone in the DO-XRT samples compared with standard, or non-radiated DO regenerate. Similarly, the percentage of highly mineralized bone was significantly lower in the DO-XRT specimens vs. DO group (Figure 3).

Figure 3. μCT novel algorithm of the percentage mineralization within the regenerate DO vs. DO-XRT

Conclusions: This study utilized novel, non-destructive µCT imaging techniques and algorithms to delineate which particular facets of bone healing are diminished in DO after XRT. We found a significant decrease in the percentage of highly mineralized, mature bone, along with a significantly greater percent of low and medium mineralized, immature bone in the DO-XRT group when compared to the DO regenerate in the non-irradiated group. We also found a substantial trend toward lower overall BMD in DO-XRT, approaching statistical significance.

The data generated in this study quantified and outlined the extent of XRT attenuated healing and the reduced quality of regenerate in mandibular DO after irradiation utilizing established metrics that we have developed in our lab. This allowed us to determine the initial quantifiable affects on attenuation, using lower therapeutic doses of XRT, to determine what aspect of regenerate healing is being affected first, and to what degree. Our findings, and continued studies should help direct therapeutic intervention aimed at mitigating the specific detrimental effects of XRT on DO. Establishing these parameters as quantitative outcomes, should thereby allow us to facilitate the use of this technique as a predictable and durable means of post-oncologic head and neck reconstruction.

References:

1.  Gantous A, et al. Distraction osteogenesis in the irradiated canine mandible. Plast Reconstr Surg, 93(1):164-168, 1994

2.  Girod A, et al. Experimental study of mineralization in mandibular bone distraction with irradiation during the consolidation phase. J Craniomaxillofac Surg. 33(6):386-394, 2005

3.  Schwarz DA, et al. Dual Energy X-Ray Absorptiometry vs. μCT Analysis of BMD in Mandibular Distraction Osteogenesis: Plast Reconstr Surg. 116(3) Supplement: 37-38, 2005.