Age-related changes in craniofacial morphology in GDF-8 (myostatin)-deficient mice.

It is well recognized that masticatory muscle function helps determine morphology, although the extent of function on final form is still debated. GDF-8 (myostatin), a transcription factor is a negative regulator of skeletal muscle growth. A recent study has shown that mice homozygous for the myostatin mutation had increased muscle mass and craniofacial dysmorphology in adulthood. However, it is unclear whether such dysmorphology is present at birth. This study examines the onset and relationship between hypermuscularity and craniofacial morphology in neonatal and adult mice with GDF-8 deficiency. Fifteen (8 wild-type and 7 GDF-8 -/-), 1-day-old and 16 (9 wt and 7 GDF-8 -/-), 180-day-old male CD-1 mice were used. Standardized radiographs were taken of each head, scanned, traced, and cephalometric landmarks identified. Significant mean differences were assessed using a group x age, two-way ANOVA. Myostatin-deficient mice had significantly (P < 0.01) smaller body and masseter muscle weights and craniofacial skeletons at 1 day of age and significantly greater body and masseter muscle weights at 180 days of age compared to controls. Myostatin-deficient mice showed significantly (P < 0.001) longer and "rocker-shaped" mandibles and shorter and wider crania compared to controls at 180 days. Significant correlations were noted between masseter muscle weight and all cephalometric measurements in 180-day-old Myostatin-deficient mice. Results suggest that in this mouse model, there may be both early systemic skeletal growth deficiencies and later compensatory changes from hypermuscularity. These findings reiterate the role that masticatory muscle function plays on the ontogeny of the cranial vault, base, and most notably the mandible.

Age-related peridural hyperemia in craniosynostotic rabbits.

OBJECTIVE: Craniosynostosis is the premature fusion of the calvarial sutures and is associated with aesthetic impairment and secondary damage to brain growth. Associated neurological injuries can result from increased intracranial pressure (ICP) and abnormal cerebral blood flow (CBF). Arterial spin-labeling (ASL) MRI was used to assess regional CBF in developing rabbits with early-onset coronal suture synostosis (EOCS) and age-matched wild-type controls (WT). METHODS: Rabbits were subjected to ASL MRI at or near 10, 25, or 42 days of age. Differences in regional CBF were assessed using one-way ANOVA. CONCLUSION: CBF was similar in WT and EOCS rabbits with the exception of the peridural surfaces in EOCS rabbits at 25 days of age. A twofold increase in peridural CBF at 25 days of age coincides with a transient increase in ICP. By 42 days of age, CBF in peridural surfaces had decreased.

Noggin inhibits postoperative resynostosis in craniosynostotic rabbits.

UNLABELLED: Inhibition of bone formation after surgery to correct craniosynostosis would alleviate the need for secondary surgeries and decrease morbidity and mortality. This study used a single dose of Noggin protein to prevent resynostosis and improve postoperative outcomes in a rabbit model of craniosynostosis. INTRODUCTION: Craniosynostosis is defined as the premature fusion of one or more of the cranial sutures, which causes secondary deformations of the cranial vault, cranial base, and brain. Current surgical intervention involves extirpation of the fused suture to allow unrestricted brain growth. However, resynostosis of the extirpated regions often occurs. Several bone morphogenetic proteins (BMPs), well-described inducers of ossification, are involved in bone healing. This study tested the hypothesis that a postoperative treatment with Noggin, an extracellular BMP inhibitor, can inhibit resynostosis in a rabbit model of human familial nonsyndromic craniosynostosis. MATERIALS AND METHODS: Thirty-one New Zealand white rabbits with bilateral coronal suture synostosis were divided into three groups: (1) suturectomy controls (n = 13); (2) suturectomy with BSA in a slow-resorbing collagen vehicle, (n = 8); and (3) suturectomy with Noggin in a slow-resorbing collagen vehicle (n = 10). At 10 days of age, a 3 x 15-mm coronal suturectomy was performed. The sites in groups 2 and 3 were immediately filled with BSA-loaded gel or Noggin-loaded gel, respectively. Serial 3D-CT scan reconstructions of the defects and standard radiographs were obtained at 10, 25, 42, and 84 days of age, and the sutures were harvested for histological analysis. RESULTS: Radiographic analysis revealed that Noggin-treated animals had significantly greater coronal suture marker separation by 25 days and significantly greater craniofacial length at 84 days of age compared with controls. 3D-CT analysis revealed that Noggin treatment led to significantly greater defect areas through 84 days and to increased intracranial volumes at 84 days of age compared with other groups. Histological analysis supported CT data, showing that the untreated and BSA-treated groups had significant healing of the suturectomy site, whereas the Noggin-treated group had incomplete wound healing. CONCLUSIONS: These data support our hypothesis that inhibition of BMP activity using Noggin may prevent postoperative resynostosis in this rabbit model. These findings also suggest that Noggin therapy may have potential clinical use to prevent postoperative resynostosis in infants with craniosynostosis.

Craniosacral therapy: the effects of cranial manipulation on intracranial pressure and cranial bone movement.

STUDY DESIGN: Quasi-experimental design. OBJECTIVES: To determine if physical manipulation of the cranial vault sutures will result in changes of the intracranial pressure (ICP) along with movement at the coronal suture. BACKGROUND: Craniosacral therapy is used to treat conditions ranging from headache pain to developmental disabilities. However, the biological premise for this technique has been theorized but not substantiated in the literature. METHODS: Thirteen adult New Zealand white rabbits (oryctolagus cuniculus) were anesthetized and microplates were attached on either side of the coronal suture. Epidural ICP measurements were made using a NeuroMonitor transducer. Distractive loads of 5, 10, 15, and 20 g (simulating a craniosacral frontal lift technique) were applied sequentially across the coronal suture. Baseline and distraction radiographs and ICP were obtained. One animal underwent additional distractive loads between 100 and 10,000 g. Plate separation was measured using a digital caliper from the radiographs. Two-way analysis of variance was used to assess significant differences in ICP and suture movement. RESULTS: No significant differences were noted between baseline and distraction suture separation (F = 0.045; P>.05) and between baseline and distraction ICP (F = 0.279; P>.05) at any load. In the single animal that underwent additional distractive forces, movement across the coronal suture was not seen until the 500-g force, which produced 0.30 mm of separation but no corresponding ICP changes. CONCLUSION: Low loads of force, similar to those used clinically when performing a craniosacral frontal lift technique, resulted in no significant changes in coronal suture movement or ICP in rabbits. These results suggest that a different biological basis for craniosacral therapy should be explored.

Testing causal mechanisms of nonsyndromic craniosynostosis using path analysis of cranial contents in rabbits with uncorrected craniosynostosis.

OBJECTIVE: Various causal mechanisms of familial nonsyndromic craniosynostosis have been presented. One hypothesis suggests that overproduction of bone at the suture is the primary origin of craniosynostosis, which affects brain and cranial growth secondarily through altered intracranial pressure (Primary Suture Fusion Model). Other hypotheses suggest that decreased cranial base growth or abnormal brain growth are the primary cause of craniosynostosis (Cranial Base, Brain Parenchyma Models, respectively). This study was designed to investigate which model best describes neurocranial changes associated with craniosynostosis in a rabbit model through multivariate path analysis. DESIGN: Serial magnetic resonance imaging scans and intracranial pressure measurements were obtained at 10, 25, and 42 days of age from 18 rabbits: six controls, six with delayed-onset synostosis, and six with early-onset synostosis. Five variables were collected from each rabbit: calvarial thickness at the affected suture, cranial base length, brain volume, cerebrospinal fluid volume, and intracranial pressure. This data set was used to test causal pathway relationships generated by the proposed models. Goodness of fit was measured by experimental group for each model. RESULTS: Primary Suture Fusion Model best explained the variables in both delayed-onset and early-onset synostotic rabbits (Goodness of fit = 93%, 97%, respectively). Cranial Base Model (Goodness of fit = 94%) best explained the data in control rabbits. CONCLUSION: Results suggest that the primary site of craniosynostosis in craniosynostotic rabbits is most likely the synostosed suture. Other cranial vault anomalies are most likely secondary compensatory changes. Results of the present study may provide insight regarding the causal pathway of craniosynostosis.

Intracranial volume changes in craniosynostotic rabbits: effects of age and surgical correction.

BACKGROUND: The premature fusion of one or more cranial sutures, termed craniosynostosis, alters normal brain growth patterns and results in compensatory changes in the cranial vault. The authors previously reported that bilateral coronal suture fusion resulted in a reduction in intracranial volume in a rabbit model of nonsyndromic, familial coronal suture synostosis. METHODS: The current follow-up study involved collecting cross-sectional three-dimensional computed tomographic head scans from 142 rabbits (70 normal, 44 with uncorrected synostosis, and 28 synostosed rabbits with coronal suturectomy) at 0, 10, 25, 42, 84, and 126 days of age. Intracranial contents were reconstructed, and indirect intracranial volume was calculated. RESULTS: Results revealed a significant (p < 0.05) postsynostotic reduction of intracranial volume (23 percent) by 25 days of age in rabbits with uncorrected craniosynostosis compared with normal controls, which continued through 84 days of age. Also, rabbits with surgically released synostosis, using a simple strip suturectomy, showed significantly (p < 0.05) greater intracranial volume at 25 days of age compared with unoperated synostosed rabbits. However, no changes in intracranial volume were noted between 42 and 84 days of age in rabbits with surgically released synostosis, at which point their intracranial volume was 30 percent less than that in normal control rabbits. CONCLUSIONS: These data suggest that in rabbits with uncorrected craniosynostosis, compensatory changes in the neurocranium were not capable of compensating for the loss of sutures as growth sites. The results also showed that that surgical release of the synostosed suture improved intracranial volume in the short term (25 to 42 days) but failed to change it in the long term (42 to 84 days), possibly because of rapid resynostosis of the suturectomy site. This study suggests that surgical release of the suture fusion site alone may not be adequate to allow for normal intracranial volume growth in synostotic rabbits. For this reason, it may be efficacious to design and develop adjunct protein and gene therapies to prevent resynostosis and improve postoperative intracranial volume in craniosynostotic individuals.

Age-related changes in lateral ventricle morphology in craniosynostotic rabbits using magnetic resonance imaging.

BACKGROUND AND AIMS: Craniosynostosis occurs in 300-500 per 1,000,000 live births and results in secondary craniofacial, ocular, and intracranial anomalies. Neurologic problems associated with craniosynostosis include changes in intracranial morphology such as dilation of the cerebral ventricles, however, clinical studies are confounded by small sample sizes, heterogenous samples, and lack of age-matched controls. The present study was designed to assess age-related changes in the lateral ventricle volume of the brain in normal rabbits and rabbits with naturally-occurring coronal suture synostosis using serial magnetic resonance imaging. METHODS: Eighteen rabbits (6 wild-type controls, 6 with early-onset [ approximately 21 days gestation], and 6 with delayed-onset [approximately 25 days post-gestation] coronal suture synostosis) had magnetic resonance imaging (MRI) at 10, 25, and 42 days of age. RESULTS: The results demonstrate that rabbits with early-onset synostosis had significantly (p<0.001) dilated and larger lateral ventricles (by 77% at 10 days of age) than wild-type and delayed-onset synostosis rabbits, which progressively worsened by day 42. CONCLUSION: This finding suggests that uncorrected coronal suture synostosis may have early effects on lateral ventricle volume hypertrophy, possibly through obstructed cerebrospinal fluid and/or venous drainage and circulation.

Early neuromotor behavior in craniosynostotic rabbits.

OBJECTIVE: Clinical studies have shown both abnormal and normal mental and psychomotor development in patients with craniosynostosis. However, a number of confounding variables make study comparisons difficult. For these reasons, the present study describes early neuromotor development in an homogeneous rabbit model of craniosynostosis. DESIGN: Fifty-three newborn New Zealand white rabbit kits were used: 13 were wild-type, normal control rabbits; 23 had delayed-onset coronal suture synostosis (onset is approximately 57 to 74 days post conception); and 17 had early-onset coronal suture synostosis (onset is approximately 21 to 25 days post conception). All rabbits were observed individually and blindly in an open field, daily for 2 minutes, from birth through the first 14 days of life. The first day of emergence of 10 different mature behaviors and developmental events (in developmental order of appearance: falling, righting, cliff avoidance, first sign of fur, body elevation, head elevation, circling, dragging, eye opening, and hopping) was recorded for each kit. Daily activity levels (grid crossing), and body weights were also recorded. RESULTS: Significant group (p <.05) differences were observed in 9 of 11 measures. Both synostosed groups had significantly (p <.05) accelerated onset of behavior in 8 of 9 measures, compared with wild-type controls. The early-onset synostosis group had significantly (p <.05) accelerated onset in five of eight measures, compared with wild-type controls, and three of eight measures, compared with the delayed-onset synostosis group. CONCLUSIONS: Synostotic rabbits showed precocious neuromotor development possibly through frontal lobe constrictions and altered brain activity from increased intracranial pressure, although primary genetic effects cannot be ruled out.