Cyclic loading failed to promote growth in a pig model of midfacial hypoplasia.

Yucatan miniature pigs, often used as large animal models in clinical research, are distinguished by a breed-specific midfacial hypoplasia with anterior crossbite. Although this deformity can be corrected by distraction osteogenesis, a less invasive method is desirable. We chose a mechanical cyclic stimulation protocol that has been successful in enhancing sutural growth in small animals and in a pilot study on standard pigs. Yucatan minipigs (n = 14) were obtained in pairs, with one of each pair randomly assigned to sham or loaded groups. All animals had loading implants installed on the right nasal and frontal bones and received labels for cell proliferation and mineral apposition. After a week of healing and under anesthesia, experimental animals received cyclic tensile loads (2.5 Hz, 30 min) delivered to the right nasofrontal suture daily for 5 days. Sutural strains were recorded at the final session for experimental animals. Sham animals received the same treatment except without loading or strain gauge placement. In contrast to pilot results on standard pigs, the treatment did not produce the expected sutural widening and increased growth. Although sutures were not fused and strains were in the normal range, the targeted right nasofrontal suture was narrowed rather than widened, with no statistically significant changes in sutural cell proliferation, mineral apposition, or vascularity. In general, Yucatan minipig sutures were more vascular than those of standard pigs and also tended to have more proliferating cells. In conclusion, either because the sutures themselves are abnormal or because of growth restrictions elsewhere in the skull, this cyclic loading protocol was unable to produce the desired response of sutural widening and growth. This treatment, effective in normal animals, did not improve naturally occurring midfacial hypoplasia in Yucatan minipigs.

Mechanobiology of bone and suture - Results from a pig model.

OBJECTIVE: To compare the morphology and mechanical function of sutures in normal pigs and minipigs to those of Yucatan minipigs, a natural model for midfacial hypoplasia. SETTING AND SAMPLE POPULATION: Research took place at the Department of Orthodontics at the University of Washington and used varying sample sizes of normal-snouted pigs and Yucatan minipigs. MATERIAL AND METHODS: Skulls and heads were examined for morphology of the nasofrontal suture using computed tomography and histology. Strain gauge recordings were made of sutural strain during mastication and during cyclic tensile loading of the nasofrontal suture. RESULTS: Sutures in Yucatans had narrower gaps than same-age normal pigs. The nasofrontal suture was simpler in construction and had more active osteoblasts on the bone fronts in Yucatans. The sutural ligament was less well organized, and based on a small sample, masticatory strain appeared to be lower than in normal minipigs. However, sutures were not fused and showed similar strains in response to the cyclic loading procedure. CONCLUSION: Midfacial hypoplasia in Yucatan pigs has the likely proximate cause of hyperossification. Yet prior to fusion, the sutures appear to be amenable to treatment that would promote their growth rate.

Respiratory internal kinematics of the tongue base and soft palate in obese minipigs with obstructive sleep apnea.

It is largely unknown how the tongue base and soft palate deform to alter the configuration of the oropharyngeal airway during respiration. This study is to address this important gap. After live sleep monitoring of five Yucatan and two Panepinto minipigs to verify obstructive sleep apnea (OSA), eight and four ultrasonic crystals were implanted into the tongue base and soft palate to circumscribe a cubic and square region, respectively. The 3D and 2D deformational changes of the circumscribed regions were measured simultaneously with electromyographic activity of the oropharyngeal muscles during spontaneous respiration under sedated sleep. The results indicated that both obese Yucatan and Panepinto minipigs presented spontaneous OSA, but not in three nonobese Yucatan minipigs. During inspiration, the tongue base showed elongation in both dorsal and ventral regions but thinning and thickening in the anterior and posterior regions, respectively. The widths showed opposite directions, widening in the dorsal but narrowing in the ventral regions. The soft palate expanded in both length and width. Compared to normal controls, obese/OSA ones showed similar directions of deformational changes, but the magnitude of change was two times larger in the tongue base and soft palate, and obese/OSA Panepinto minipigs presented 10 times larger changes in all dimensions of both the tongue base and the soft palate. The distance changes between the dorsal surface of tongue base and soft palate during inspiration increased in normal but decreased in obese OSA minipigs.

The nasal septum and midfacial growth.

The nasal septum is the only element of the chondrocranium which never completely ossifies. The persistence of this nonarticular cartilage has given rise to a variety of theories concerning cranial mechanics and growth of the midface. Previously, using pigs, we demonstrated that the septum is not a strut supporting the snout and that septal growth seems capable of stretching the overlying nasofrontal suture, a major contributor to snout elongation. Here we investigate whether abnormalities of the septum are implicated in cases of midfacial hypoplasia, in which growth of the midface is inadequate. Mild midfacial hypoplasia is common in domestic pig breeds and often severe in the Yucatan minipig, a popular laboratory breed. Normal-snouted and midfacial hypoplastic heads of standard (farm mixed breed) and minipigs ranging in age from perinatal to 12 months were dissected, imaged by CT, and/or prepared for histology. Even at birth, Yucatan minipigs with midfacial hypoplasia exhibited greater caudal ossification than normal; the ventral cartilaginous sphenoidal "tail" was diminished or missing. In addition, cells that morphologically appeared to have divided recently were less numerous than in newborn standard pigs. Juvenile Yucatan minipigs lacked caudal cartilaginous growth zones almost completely. In standard newborns, the ventral caudal septum was more replicative than the dorsal, but this trend was not seen in Yucatan newborns. In conclusion, accelerated maturation of the caudal septum was associated with midfacial hypoplasia, a further indication that the septum, particularly its ventral portion, is important for midfacial elongation.

Repeated botulinum treatment of rabbit masseter causes cumulative tissue damage.

OBJECTIVE: Botulinum neurotoxins (BoNT) are used in masticatory muscles for pain relief, unloading of the mandible, and cosmetic facial contouring. Treatment is often repeated every few months as function returns. This study assessed masticatory function and musculoskeletal structure after multiple BoNT treatment of the rabbit masseter. DESIGN: Female rabbits received 3 injections of BoNT (n = 13) or saline (n = 5) into one masseter muscle at intervals of 12 weeks. The contralateral side served as control. Periodic measurements of masticatory electromyography (EMG) and stimulated anterior bite force were made. After the final 12-week recovery interval, neuromuscular connection was investigated by stimulating the masseteric nerve to elicit an evoked EMG response. Mandibular specimens were collected for microCT analysis, and masseters were collected for histomorphometry and counts of replicating cells. RESULTS: Control and saline-injected muscles maintained consistent masticatory EMG and anterior bite force throughout the study. BoNT-injected masseters showed strong declines after each injection; during the 12-week recovery period, masticatory EMG and anterior bite force improved, although only electrical activity reached normal levels. Multiple injection resulted in persistently atrophied muscle fibers with fibrosis, and notable loss of bone from the mandibular body and condyle. The uninjected masseters of the BoNT group also showed evidence of mild toxin-related changes. CONCLUSIONS: Although muscle function is mostly regained after each injection, masseters receiving multiple doses of BoNT show extensive damage. In addition, mandibular bone density is decreased on the injected side.

Botulinum toxin in the masseter muscle: Lingering effects of denervation.

Botulinum neurotoxins (BoNTs) are paralytic agents used to treat a variety of conditions in jaw muscles. Although their effect is considered temporary, there are reports of persistent functional changes. Using rabbits that received BoNT injection in one masseter muscle, the recovery of neuromuscular connection was investigated using nerve stimulation to evoke an electromyographic (EMG) response, and the recovery of muscle fibers was investigated using histological morphometry and bromodeoxyuridine (BrdU) immunohistochemistry. One month after treatment, evoked EMG was greatly reduced in both amplitude and duration, indicating that little reinnervation had taken place. Muscle fibers were atrophied and collagenous tissue was increased. Three months after treatment, evoked EMG duration was normal, indicating that at least some neuromuscular junctions were functional. Histologically, some muscle fibers were hypertrophied, some were still atrophied, and some appeared to have died. Fibrosis was still apparent amid slight increases in dividing cells and regenerating fibers. The histological effects of BoNT were evident although attenuated at a distance of about 1 cm from the injection level, but no regional differences could be discerned for the evoked EMGs. In conclusion, there were persistent muscular deficits seen 3 months after BoNT treatment that may have been caused by the failure of some affected muscle fibers to become reinnervated.

Obstructive sleep apnea in obese minipigs.

OBJECTIVE: Obesity has reached epidemic proportions and is a strong risk factor for obstructive sleep apnea (OSA). However, the underlying mechanisms are poorly understood and current treatment strategies for OSA and obesity have critical limitations. Thus, establishment of an obesity-related large animal model with spontaneous OSA is imperative. MATERIALS AND METHODS: Natural and sedated sleep were monitored and characterized in 4 obese (body mass index - BMI>48) and 3 non-obese (BMI<40) minipigs. These minipigs were instrumented with the BioRadio system under sedation for the wireless recording of respiratory airflow, snoring, abdominal and chest respiratory movements, electroencephalogram, electrooclulogram, electromyogram, and oxygen saturation. After instrumentation, the minipigs were placed in a dark room with a remote night-vision camera for monitoring all behaviors. Wakefulness and different sleep stages were classified, and episodes of apneas and/or hypopneas were identified during natural and/or sedated sleep. RESULTS: No hypopnea episodes were observed in two of the non-obese minipigs, but one non-obese minipig had 5 hypopnea events. Heavy snoring and 27-58 apnea and/or hypopnea episodes were identified in all 4 obese minipigs. Most of these episodes occurred in the rapid eye movement stage during natural sleep and/or sedated sleep in Yucatan minipigs. CONCLUSIONS: Obese minipigs can experience naturally occurring OSA, thus are an ideal large animal model for obese-related OSA studies.