Poster 125: Mandibular Lengthening by Distraction Osteogenesis (DO) in a Porcine Model: Changes in Anterior Digastric Muscle

Abstract

The effects of mandibular lengthening by distraction osteogenesis on the muscles of mastication are not well understood. The purpose of this investigation was to document the changes, over time, in the digastric muscle in response to unilateral mandibular lengthening. In this experiment, 17 mixed dentition Yucatan minipigs were used: Osteotomy and unilateral (right side) mandibular distraction osteogenesis using a protocol of 0-day latency, 1 mm/day rate of distraction for 12 days, followed by 24 days of fixation (n=14 minipigs); osteotomy and acute lengthening of 12 mm (n=2); and sham control with dissection and 0 mm advancement (n=1). Anterior digastric muscles were harvested from the distraction and contralateral sides at mid-DO, end-DO, mid-fixation, and end-fixation. The same muscle samples were harvested from the acute lengthening and sham control groups 24 days after operation. All muscle samples were fixed, embedded in paraffin, and sectioned transversely and longitudinally. Specimens were stained with Hematoxylin & Eosin (H&E) or immunostained against PCNA. To evaluate histologic changes, a blinded pathologist examined all the H&E slides under light microscopy for the presence of inflammation, hematoma, necrosis, fibrosis, edema and for the size of muscle fibers. Histologic images (400X) from H&E slides were used for computer histomorphometric analysis. The cross-sectional surface area of the muscle fibers was measured by histomorphometric software (Image J, NIH) and the data were analyzed for statistical significance (two sample T-test). Proliferation activity was evaluated by calculating the percentage of PCNA positive nuclei in six randomly chosen fields in each sample. Statistical significance of the means was evaluated using the Wilcoxon’s matched pairs test. In two of 4 pigs, at mid-DO, there was a mild and scattered inflammatory infiltrate in the experimental side muscle. There were no inflammatory changes observed in specimens at end-DO (n=4), mid-fixation (n=4) and end-fixation (n=2). Muscles from the sham control group also demonstrated no inflammatory changes. In the acute lengthening group (n=2), inflammatory infiltrate, fibrosis, edema, and hematoma were present in the digastric on the experimental side. Histomorphometric analysis showed that cross-sectional surface area of the anterior digastric myocyte fibers on the distracted side were significantly (10%) larger than controls at end-fixation. In the acute lengthening group there was a 23% reduction in myocyte fiber cross-sectional surface area 24 days postoperative. The size of myocyte fiber nuclei was increased on the distracted side at all time points. Data from PCNA immunohistochemistry revealed that the anterior digastric muscle, which is parallel to the distraction vector, had a 3.6-fold (mid-DO group), 7.1-fold (end-DO group), 10.4-fold (mid-fixation group), 10.3-fold (end-fixation group) increase in PCNA-positive myocytes compared with the contralateral samples. Anterior digastric muscle in the acute lengthening group had a 2.9-fold increase in PCNA-positive myocytes when compared to the contralateral muscle. The results of this study indicate that the anterior digastric muscle, whose fibers are parallel to the distraction vector, undergoes hypertrophy and proliferation in response to DO.