New insights into the variability of upper airway morphology in modern humans.

The biological adaptation of the human lineage to its environment is a recurring question in paleoanthropology. Particularly, how eco-geographic factors (e.g., environmental temperature and humidity) have shaped upper airway morphology in hominins have been subject to continuing debate. Nasal shape is the result of many intertwined factors that include, but are not limited to, genetic drift, sexual selection, or adaptation to climate. A quantification of nasal airway (NA) morphological variation in modern human populations is crucial to better understand these multiple factors. In the present research, we study 195 in vivo CT scans of adult individuals collected in five different geographic areas (Chile, France, Cambodia, Russia, and South Africa). After segmentation of the nasal airway, we reconstruct 3D meshes that are analyzed with a landmark-free geometric morphometrics method based on surface deformation. Our results highlight subtle but statistically significant morphological differences between our five samples. The two morphologically closest groups are France and Russia, whose NAs are longer and narrower, with an important protrusion of the supero-anterior part. The Cambodian sample is the most morphologically distinct and clustered sample, with a mean NA that is wider and shorter. On the contrary, the Chilean sample form the most scattered cluster with the greatest intra-population variation. The South African sample is morphologically close to the Cambodian sample, but also partially overlaps the French and Russian variation. Interestingly, we record no correlation between NA volume and geographic groups, which raises the question of climate-related metabolic demands for oxygen consumption. The other factors of variation (sex and age) have no influence on the NA shape in our samples. However, NA volume varies significantly according both to sex and age: it is higher in males than in females and tends to increase with age. In contrast, we observe no effect of temperature or humidity on NA volume. Finally, we highlight the important influence of asymmetries related to nasal septum deviations in NA shape variation.

Vitamin D and adolescent idiopathic scoliosis, should we stop the hype? A cross-sectional observational prospective study based on a geometric morphometrics approach.

PURPOSE: There is strong evidence supporting the presence of fluctuating asymmetry (FA) in Adolescents with Idiopathic Scoliosis (AIS). Additionally, recent research investigating the relationship between vitamin D and AIS found a relation between them. We hypothesize a negative correlation between FA and vitamin D. METHODS: We performed a surface scan of the torso of 53 AIS patients, a blood test to measure vitamin D and the radiographic Cobb angle. A correlation analysis between vitamin D and FA was carried out to test our hypothesis, and a regression of vitamin D on 3D shape was performed to observe shape differences between the vitamin D deficiency and insufficiency groups. RESULTS: There was no correlation between vitamin D and FA. We found a strong negative correlation between vitamin D and the Cobb angle only in the premenarche group (n = 7; r = - 0.92). Differences in shape were observed between the deficiency and insufficiency groups, and that differences were related to the width of the torso, but not the rotation or lateral flexion. CONCLUSIONS: Our results do not support the massive screening of vitamin D in AIS. Shape analysis revealed differences between the shape of the deficiency and insufficiency groups related to robustness. However, this finding had no relation with the scoliosis characteristics, it just reflected different body composition, and its importance should be explored in future.

Unilateral Hypofunction of the Masseter Leads to Molecular and 3D Morphometric Signs of Atrophy in Ipsilateral Agonist Masticatory Muscles in Adult Mice.

Mice are commonly used to study mandibular dynamics due to their similarity in chewing cycle patterns with humans. Adult mice treated unilaterally with botulinum toxin type A (BoNTA) in the masseter exhibit atrophy of this muscle characterized by an increase in the gene expression of atrophy-related molecular markers, and a reduction in both muscle fiber diameter and muscle mass at 14d. However, the impact of this muscle imbalance on the non-treated masticatory muscles remains unexplored. Here, we hypothesize that the unilateral masseter hypofunction leads to molecular and 3D morphometric signs of atrophy of the masseter and its agonist masticatory muscles in adult mice. Twenty-three 8-week-old male BALB/c mice received a single injection of BoNTA in the right masseter, whereas the left masseter received the same volume of saline solution (control side). Animals were euthanized at 2d, 7d, and 14d, and the masticatory muscles were analyzed for mRNA expression. Five heads were harvested at 14d, fixed, stained with a contrast-enhanced agent, and scanned using X-ray microtomography. The three-dimensional morphometric parameters (the volume and thickness) from muscles in situ were obtained. Atrogin-1/MAFbx, MuRF-1, and Myogenin mRNA gene expression were significantly increased at 2 and 7d for both the masseter and temporalis from the BoNTA side. For medial pterygoid, increased mRNA gene expression was found at 7d for Atrogin-1/MAFbx and at 2d-7d for Myogenin. Both the volume and thickness of the masseter, temporalis, and medial pterygoid muscles from the BoNTA side were significantly reduced at 14d. In contrast, the lateral pterygoid from the BoNTA side showed a significant increase in volume at 14d. Therefore, the unilateral hypofunction of the masseter leads to molecular and morphological signs of atrophy in both the BoNTA-injected muscle and its agonistic non-injected masticatory muscles. The generalized effect on the mouse masticatory apparatus when one of its components is intervened suggests the need for more clinical studies to determine the safety of BoNTA usage in clinical dentistry.

Efficacy of shear strain gradients as an osteogenic stimulus.

The question of which mechanical variables are responsible for inducing osteogenic activity is unresolved despite extensive experimental and theoretical investigation. Candidate variables include strain magnitude, loading frequency, the interaction of magnitude and frequency (strain rate), and strain gradients. An additional challenge is discerning the coordination of periosteal and endosteal expansion during growth, and whether this coordination (or lack thereof) is fully dependent or partially independent of the local mechanical environment. In this study, under the assumption that calculated stresses correspond to relative strain magnitudes, we specify alternative growth algorithms of bone cross-sectional size and geometry to explore skeletal growth under alternative scenarios of osteogenic activity that are tracking 1) an attractor stress, 2) local stress magnitude or 3) steepness of stress gradients. These developmental simulations are initiated from two initial geometries (symmetrical and asymmetrical ellipses) under a time-varying torsional load whose magnitude is proportional to body size growth in a model primate. In addition, we model endosteal expansion under three conditions hypothesized in the literature, in which endosteal expansion is 1) independent of the mechanical milieu, 2) completely dependent on the mechanical milieu, and 3) a "hybrid" model in which intrinsic biological (independent) growth is operative early but gives way to mechanically-sensitive (dependent) growth at later ages. Three variables were recorded over each growth simulation: the safety factor (ratio of yield stress to actual stress), an efficiency ratio (invested bone area per unit of stress), and proximity to an isostress condition (an optimal design criterion in which stress is invariant throughout the structure). The attractor stress algorithm produces the most "adapted" bones in terms of mechanical competence and economy of material. Localized osteogenic activity that is guided in direct proportion to stress magnitude produces competent bones but with variable adult geometries depending on conditions of endosteal expansion. Stress gradients also produce functional but relatively inefficient bones, with widely variable safety factors during growth and heterogeneous stress fields. If, in fact, the osteocyte network monitors strain gradients to generate osteogenic signals, the resulting morphology is competent but falls well short of an optimal mechanical solution.

Morphometric Characterization of Asymmetric Mandibles Due to Condylar Hyperactivity.

PURPOSE: Mandibular asymmetry related to condylar hyperactivity (CH) presents a complex set of morphologic features that pose challenges for its correction. Using state-of-the-art morphometric techniques, this report provides a detailed and hierarchical description of the features present in CH-related asymmetric mandibles and offers new knowledge for the surgical treatment of CH. MATERIALS AND METHODS: Sixty patients were included in the sample. Thirty had CH-related asymmetric mandibles and the other 30 had clinically symmetric mandibles. Twenty-eight 3-dimensional landmarks were placed on computed tomographically based reconstructions of each participant's mandible and analyzed using geometric morphometric analysis for the quantitative and qualitative comparison of their morphologic features. RESULTS: All 60 participants exhibited asymmetry. However, those with CH exhibited a broad range of shapes and even shared several morphologic features with the controls. Mainly the ramus and then the body were the main contributors of the differences between groups. CONCLUSIONS: There is considerable overlap of anatomic features characterizing symmetric and asymmetric mandibles; based on shape alone, the 2 groups can be easily misclassified. The ramus and body of the affected side in CH-related asymmetric mandibles were the main contributors to asymmetry of the structure. The chin, a usual diagnostic structure, did not greatly contribute to the structural asymmetry of the mandible.

Dental malocclusions are not just about small and weak bones: assessing the morphology of the mandible with cross-section analysis and geometric morphometrics.

OBJECTIVES: Dental malocclusions in modern populations would be the result of small and weak jaws developing under low masticatory loads. We assess the validity of this by characterising the external and internal morphology of mandibles affected by class II and III malocclusions and comparing them with those from individuals with different masticatory load patterns. MATERIALS AND METHODS: CTs from up to 118 individuals exerting intensive, medium and low masticatory loads with harmonic occlusion, and from class II and III individuals, were used to compare their external shape using geometric morphometrics, as well as their internal amount and distribution of cortical bone. RESULTS: The low-load groups (harmonic, class II and III occlusion) are externally more gracile than the intense and medium load groups. But more relevant in shape variation is a marked allometric pattern, which differentiates class II (small) and III (large) mandibles. Despite gracility, the relative amount of cortical bone in the low-load groups is larger than in the remaining groups. CONCLUSIONS: There is no evidence that the modern mandible, including class II and III individuals, is intrinsically small and weak. Instead, there is a rather large degree of morphological variation, which could be linked to a lack of constraints derived from low masticatory loads. Thus, the effect of other factors such as genetics, but also basal metabolism, should be looked in more depth. CLINICAL RELEVANCE: Dental malocclusions are a common disorder whose aetiology has not been unravelled, and several to be considered in the prevention and therapy of malocclusion.

Masseter muscle atrophy impairs bone quality of the mandibular condyle but not the alveolar process early after induction.

BACKGROUND: Masseter muscle function influences mandibular bone homeostasis. As previously reported, bone resorption markers increased in the mouse mandibular condyle two days after masseter paralysis induced with botulinum toxin type A (BoNTA), followed by local bone loss. OBJECTIVE: This study aimed to evaluate the bone quality of both the mandibular condyle and alveolar process in the mandible of adult mice during the early stage of a BoNTA-induced masseter muscle atrophy, using a combined 3D histomorphometrics and shape analysis approach. METHODS: Adult BALB/c mice were divided into an untreated control group and an experimental group; the latter received one single BoNTA injection in the right masseter (BoNTA-right) and saline in the left masseter (Saline-left). 3D bone microstructural changes in the mandibular condyle and alveolar process were determined with high-resolution microtomography. Additionally, landmark-based geometric morphometrics was implemented to assess external shape changes. RESULTS: After 2 weeks, masseter mass was significantly reduced (P-value <0.001). When compared to Saline-left and untreated condyles, BoNTA-right condyles showed significant bone loss (P-value <0.001) and shape changes. No significant bone loss was observed in the alveolar processes of any of the groups (P-value >0.05). CONCLUSION: Condyle bone quality deteriorates at an early stage of BoNTA-induced masseter muscle atrophy, and before the alveolar process is affected. Since the observed bone microstructural changes resemble those in human temporomandibular joint degenerative disorders, the clinical safety of BoNTA intervention in the masticatory apparatus remains to be clarified.

The biting performance of Homo sapiens and Homo heidelbergensis.

Modern humans have smaller faces relative to Middle and Late Pleistocene members of the genus Homo. While facial reduction and differences in shape have been shown to increase biting efficiency in Homo sapiens relative to these hominins, facial size reduction has also been said to decrease our ability to resist masticatory loads. This study compares crania of Homo heidelbergensis and H. sapiens with respect to mechanical advantages of masticatory muscles, force production efficiency, strains experienced by the cranium and modes of deformation during simulated biting. Analyses utilize X-ray computed tomography (CT) scan-based 3D models of a recent modern human and two H. heidelbergensis. While having muscles of similar cross-sectional area to H. heidelbergensis, our results confirm that the modern human masticatory system is more efficient at converting muscle forces into bite forces. Thus, it can produce higher bite forces than Broken Hill for equal muscle input forces. This difference is the result of alterations in relative in and out-lever arm lengths associated with well-known differences in midfacial prognathism. Apparently at odds with this increased efficiency is the finding that the modern human cranium deforms more, resulting in greater strain magnitudes than Broken Hill when biting at the equivalent tooth. Hence, the facial reduction that characterizes modern humans may not have evolved as a result of selection for force production efficiency. These findings provide further evidence for a degree of uncoupling between form and function in the masticatory system of modern humans. This may reflect the impact of food preparation technologies. These data also support previous suggestions that differences in bite force production efficiency can be considered a spandrel, primarily driven by the midfacial reduction in H. sapiens that occurred for other reasons. Midfacial reduction plausibly resulted in a number of other significant changes in morphology, such as the development of a chin, which has itself been the subject of debate as to whether or not it represents a mechanical adaptation or a spandrel.

Congenital muscle dystrophy and diet consistency affect mouse skull shape differently.

The bones of the mammalian skull respond plastically to changes in masticatory function. However, the extent to which muscle function affects the growth and development of the skull, whose regions have different maturity patterns, remains unclear. Using muscle dissection and 3D landmark-based geometric morphometrics we investigated the effect of changes in muscle function established either before or after weaning, on skull shape and muscle mass in adult mice. We compared temporalis and masseter mass and skull shape in mice with a congenital muscle dystrophy (mdx) and wild type (wt) mice fed on either a hard or a soft diet. We found that dystrophy and diet have distinct effects on the morphology of the skull and the masticatory muscles. Mdx mice show a flattened neurocranium with a more dorsally displaced foramen magnum and an anteriorly placed mandibular condyle compared with wt mice. Compared with hard diet mice, soft diet mice had lower masseter mass and a face with more gracile features as well as labially inclined incisors, suggesting reduced bite strength. Thus, while the early-maturing neurocranium and the posterior portion of the mandible are affected by the congenital dystrophy, the late-maturing face including the anterior part of the mandible responds to dietary differences irrespective of the mdx mutation. Our study confirms a hierarchical, tripartite organisation of the skull (comprising neurocranium, face and mandible) with a modular division based on development and function. Moreover, we provide further experimental evidence that masticatory loading is one of the main environmental stimuli that generate craniofacial variation.

Validity and sensitivity of a human cranial finite element model: implications for comparative studies of biting performance.

Finite element analysis (FEA) is a modelling technique increasingly used in anatomical studies investigating skeletal form and function. In the case of the cranium this approach has been applied to both living and fossil taxa to (for example) investigate how form relates to function or infer diet or behaviour. However, FE models of complex musculoskeletal structures always rely on simplified representations because it is impossible completely to image and represent every detail of skeletal morphology, variations in material properties and the complexities of loading at all spatial and temporal scales. The effects of necessary simplifications merit investigation. To this end, this study focuses on one aspect, model geometry, which is particularly pertinent to fossil material where taphonomic processes often destroy the finer details of anatomy or in models built from clinical CTs where the resolution is limited and anatomical details are lost. We manipulated the details of a finite element (FE) model of an adult human male cranium and examined the impact on model performance. First, using digital speckle interferometry, we directly measured strains from the infraorbital region and frontal process of the maxilla of the physical cranium under simplified loading conditions, simulating incisor biting. These measured strains were then compared with predicted values from FE models with simplified geometries that included modifications to model resolution, and how cancellous bone and the thin bones of the circum-nasal and maxillary regions were represented. Distributions of regions of relatively high and low principal strains and principal strain vector magnitudes and directions, predicted by the most detailed FE model, are generally similar to those achieved in vitro. Representing cancellous bone as solid cortical bone lowers strain magnitudes substantially but the mode of deformation of the FE model is relatively constant. In contrast, omitting thin plates of bone in the circum-nasal region affects both mode and magnitude of deformation. Our findings provide a useful frame of reference with regard to the effects of simplifications on the performance of FE models of the cranium and call for caution in the interpretation and comparison of FEA results.

Morphologic variability of nonsyndromic operated patients affected by cleft lip and palate: a geometric morphometric study.

INTRODUCTION: In this study, we compared patterns of morphologic variations of the craniofacial skeleton between patients affected by clefts who were operated on and unaffected subjects, aiming to discuss possible morpho-functional consequences of treatment in craniofacial development. METHODS: The lateral cephalograms of 76 subjects, comprising patients with operated unilateral cleft lip and palate (OpC) and a group matched for sex and age without cleft, were used. Thirteen landmarks were used as variables in geometric morphometric tests quantifying and describing overall shape variation, differences between group means, allometry, and upper-lower face covariation. RESULTS: The OpC group showed broader shape variations including noncleft group characteristics, but mainly a retrognathic maxilla, a vertically elongated face, a more open mandibular angle, and a more closed basicranial angle. Group means differed mainly in the maxillomandibular relationships. Allometry differed between groups, with the smallest OpC patients showing the most altered morphology. Upper and lower face covariation was stronger in the OpC group, showing mainly vertical changes in the anterior face. CONCLUSIONS: Operated patients affected by clefts achieve a broad range of morphologies; the most altered were found in those with skeletal Class III and small size. Furthermore, their strongest upper and lower face shape covariation suggests that a harmonic dental occlusion could be a key factor in achieving "normal" craniofacial morphology.

Beyond skeletal studies: A computational analysis of nasal airway function in climate adaptation.

OBJECTIVES: Ecogeographic variation in human nasal anatomy has historically been analyzed on skeletal morphology and interpreted in the context of climatic adaptations to respiratory air-conditioning. Only a few studies have analyzed nasal soft tissue morphology, actively involved in air-conditioning physiology. MATERIALS AND METHODS: We used in vivo computer tomographic scans of (N = 146) adult individuals from Cambodia, Chile, Russia, and Spain. We conducted (N = 438) airflow simulations during inspiration using computational fluid dynamics to analyze the air-conditioning capacities of the nasal soft tissue in the inflow, functional, and outflow tract, under three different environmental conditions: cold-dry; hot-dry; and hot-humid. We performed statistical comparisons between populations and sexes. RESULTS: Subjects from hot-humid regions showed significantly lower air-conditioning capacities than subjects from colder regions in all the three conditions, specifically within the isthmus region in the inflow tract, and the anterior part of the internal functional tract. Posterior to the functional tract, no differences were detected. No differences between sexes were found in any of the tracts and under any of the conditions. DISCUSSION: Our statistical analyses support models of climatic adaptations of anterior nasal soft tissue morphology that fit with, and complement, previous research on dry skulls. However, our results challenge a morpho-functional model that attributes air-conditioning capacities exclusively to the functional tract located within the nasal cavity. Instead, our findings support studies that have suggested that both, the external nose and the intra-facial soft tissue airways contribute to efficiently warming and humidifying air during inspiration. This supports functional interpretations in modern midfacial variation and evolution.

Muscle-Bone Crosstalk in the Masticatory System: From Biomechanical to Molecular Interactions.

The masticatory system is a complex and highly organized group of structures, including craniofacial bones (maxillae and mandible), muscles, teeth, joints, and neurovascular elements. While the musculoskeletal structures of the head and neck are known to have a different embryonic origin, morphology, biomechanical demands, and biochemical characteristics than the trunk and limbs, their particular molecular basis and cell biology have been much less explored. In the last decade, the concept of muscle-bone crosstalk has emerged, comprising both the loads generated during muscle contraction and a biochemical component through soluble molecules. Bone cells embedded in the mineralized tissue respond to the biomechanical input by releasing molecular factors that impact the homeostasis of the attaching skeletal muscle. In the same way, muscle-derived factors act as soluble signals that modulate the remodeling process of the underlying bones. This concept of muscle-bone crosstalk at a molecular level is particularly interesting in the mandible, due to its tight anatomical relationship with one of the biggest and strongest masticatory muscles, the masseter. However, despite the close physical and physiological interaction of both tissues for proper functioning, this topic has been poorly addressed. Here we present one of the most detailed reviews of the literature to date regarding the biomechanical and biochemical interaction between muscles and bones of the masticatory system, both during development and in physiological or pathological remodeling processes. Evidence related to how masticatory function shapes the craniofacial bones is discussed, and a proposal presented that the masticatory muscles and craniofacial bones serve as secretory tissues. We furthermore discuss our current findings of myokines-release from masseter muscle in physiological conditions, during functional adaptation or pathology, and their putative role as bone-modulators in the craniofacial system. Finally, we address the physiological implications of the crosstalk between muscles and bones in the masticatory system, analyzing pathologies or clinical procedures in which the alteration of one of them affects the homeostasis of the other. Unveiling the mechanisms of muscle-bone crosstalk in the masticatory system opens broad possibilities for understanding and treating temporomandibular disorders, which severely impair the quality of life, with a high cost for diagnosis and management.

Mandibular Bone Loss after Masticatory Muscles Intervention with Botulinum Toxin: An Approach from Basic Research to Clinical Findings.

The injection of botulinum toxin type A (BoNT/A) in the masticatory muscles, to cause its temporary paralysis, is a widely used intervention for clinical disorders such as oromandibular dystonia, sleep bruxism, and aesthetics (i.e., masseteric hypertrophy). Considering that muscle contraction is required for mechano-transduction to maintain bone homeostasis, it is relevant to address the bone adverse effects associated with muscle condition after this intervention. Our aim is to condense the current and relevant literature about mandibular bone loss in fully mature mammals after BoNT/A intervention in the masticatory muscles. Here, we compile evidence from animal models (mice, rats, and rabbits) to clinical studies, demonstrating that BoNT/A-induced masticatory muscle atrophy promotes mandibular bone loss. Mandibular bone-related adverse effects involve cellular and metabolic changes, microstructure degradation, and morphological alterations. While bone loss has been detected at the mandibular condyle or alveolar bone, cellular and molecular mechanisms involved in this process must still be elucidated. Further basic research could provide evidence for designing strategies to control the undesired effects on bone during the therapeutic use of BoNT/A. However, in the meantime, we consider it essential that patients treated with BoNT/A in the masticatory muscles be warned about a putative collateral mandibular bone damage.

Normal and altered masticatory load impact on the range of craniofacial shape variation: An analysis of pre-Hispanic and modern populations of the American Southern Cone.

The reduction of masticatory load intensity resulting from dietary changes in human evolution has been proposed as an important factor that alters craniofacial shape in past and current populations. However, its impact on craniofacial variation and on the perceived differences among populations is unclear. The maxillomandibular relationship, which alters masticatory force direction, is a factor often neglected but it can contribute to variation in craniofacial morphology, particularly among modern/urban populations where the prevalence of dental malocclusions is greater than in prehistoric populations. This study investigates the influence of masticatory load intensity and maxillomandibular relationship as a proxy for force direction on the human craniofacial skeleton. By using 3D imaging and geometric morphometrics, we analyzed craniofacial shape variation among 186 individuals from pre-Hispanic and modern Chilean and Argentinean populations that differ in diet consistency (a proxy for masticatory load intensity) and maxillomandibular relationship. We predicted that masticatory load would have a subtle effect on the upper craniofacial bones and that this would be more marked in the maxilla. Our results showed no clear influence of masticatory load on craniofacial shape, particularly in modern/urban populations. Allometry, on the contrary, shows a stronger effect. The degree of integration between the upper craniofacial bones and the load-bearing maxilla depends on masticatory load intensity, decreasing from high to low but showing a conservative pattern of covariation among the groups. The degree of variation in the shape of the maxilla is greater than the upper craniofacial bones. These results suggest that masticatory load has a limited effect in determining differences in craniofacial morphology among populations. This effect is slightly greater for the maxillary region of the face. We propose that the reduction of functional constraints is key to greater shape variation found in modern/urban populations.

On the relationship between maxillary molar root shape and jaw kinematics in Australopithecus africanus and Paranthropus robustus.

Plio-Pleistocene hominins from South Africa remain poorly understood. Here, we focus on how Australopithecus africanus and Paranthropus robustus exploited and-in part-partitioned their environment. Specifically, we explore the extent to which first maxillary molar roots (M1) are oriented and thus, by proxy, estimate the direction of loads habitually exerted on the chewing surface. Landmark-based shape analysis of M1 root reconstructions of 26 South African hominins and three East African Paranthropus boisei suggest that A. africanus may have been able to dissipate the widest range of laterally directed loads. Paranthropus robustus and P. boisei, despite having overlapping morphologies, differ in aspects of root shape/size, dento-cranial morphologies, microwear textures and C4 food consumption. Hence, while Paranthropus monophyly cannot be excluded, equivalence of dietary niche can. The South African hominins occupied distinct ecological niches, whereby P. robustus appears uniquely adapted to dissipate antero-posteriorly directed loads.

Early molecular response and microanatomical changes in the masseter muscle and mandibular head after botulinum toxin intervention in adult mice.

BACKGROUND: Masseter muscle paralysis induced by botulinum toxin type A (BoNTA) evokes subchondral bone loss in mandibular heads of adult rats and growing mice after 4 weeks. However, the primary cellular and molecular events leading to altered bone remodeling remain unexplored. Thus, the aim of the current work has been to assess the molecular response that precedes the early microanatomical changes in the masseter muscle and subchondral bone of the mandibular head in adult mice after BoNTA intervention. METHODS: A pre-clinical in vivo study was performed by a single intramuscular injection of 0.2 U BoNTA in the right masseter (experimental) of adult BALB/c mice. The contralateral masseter was injected with vehicle (control). Changes in mRNA levels of molecular markers of bone loss or muscle atrophy/regeneration were addressed by qPCR at day 2 or 7, respectively. mRNA levels of receptor activator of nuclear factor-κB ligand (RANKL) was assessed in mandibular heads, whilst mRNA levels of Atrogin-1/MAFbx, MuRF-1 and Myogenin were addressed in masseter muscles. In order to identify the early microanatomical changes at day 14, fiber diameters in transversal sections of masseter muscles were quantified, and histomorphometric analysis was used to determine the bone per tissue area and the trabecular thickness of subchondral bone of the mandibular heads. RESULTS: An increase of up to 4-fold in RANKL mRNA levels were detected in mandibular heads of the BoNTA-injected sides as early as 2 days after intervention. Moreover, a 4-6 fold increase in Atrogin-1/MAFbx and MuRF-1 and an up to 25 fold increase in Myogenin mRNA level were detected in masseter muscles 7 days after BoNTA injections. Masseter muscle mass, as well as individual muscle fiber diameter, were significantly reduced in BoNTA-injected side after 14 days post-intervention. At the same time, in the mandibular heads from the treated side, the subchondral bone loss was evinced by a significant reduction in bone per tissue area (-40%) and trabecular thickness (-55%). CONCLUSIONS: Our results show that masseter muscle paralysis induced by BoNTA leads to significant microanatomical changes by day 14, preceded by molecular changes as early as 2 days in bone, and 7 days in muscle. Therefore, masseter muscle atrophy and subchondral bone loss detected at 14 days are preceded by molecular responses that occur during the first week after BoNTA intervention.

The Effect of Varying Jaw-elevator Muscle Forces on a Finite Element Model of a Human Cranium.

Finite element analyses simulating masticatory system loading are increasingly undertaken in primates, hominin fossils and modern humans. Simplifications of models and loadcases are often required given the limits of data and technology. One such area of uncertainty concerns the forces applied to cranial models and their sensitivity to variations in these forces. We assessed the effect of varying force magnitudes among jaw-elevator muscles applied to a finite element model of a human cranium. The model was loaded to simulate incisor and molar bites using different combinations of muscle forces. Symmetric, asymmetric, homogeneous, and heterogeneous muscle activations were simulated by scaling maximal forces. The effects were compared with respect to strain distribution (i.e., modes of deformation) and magnitudes; bite forces and temporomandibular joint (TMJ) reaction forces. Predicted modes of deformation, strain magnitudes and bite forces were directly proportional to total applied muscle force and relatively insensitive to the degree of heterogeneity of muscle activation. However, TMJ reaction forces and mandibular fossa strains decrease and increase on the balancing and working sides according to the degree of asymmetry of loading. These results indicate that when modes, rather than magnitudes, of facial deformation are of interest, errors in applied muscle forces have limited effects. However the degree of asymmetric loading does impact on TMJ reaction forces and mandibular fossa strains. These findings are of particular interest in relation to studies of skeletal and fossil material, where muscle data are not available and estimation of muscle forces from skeletal proxies is prone to error. Anat Rec, 299:828-839, 2016. © 2016 Wiley Periodicals, Inc.

The relationship between skull morphology, masticatory muscle force and cranial skeletal deformation during biting.

The human skull is gracile when compared to many Middle Pleistocene hominins. It has been argued that it is less able to generate and withstand high masticatory forces, and that the morphology of the lower portion of the modern human face correlates most strongly with dietary characteristics. This study uses geometric morphometrics and finite element analysis (FEA) to assess the relationship between skull morphology, muscle force and cranial deformations arising from biting, which is relevant in understanding how skull morphology relates to mastication. The three-dimensional skull anatomies of 20 individuals were reconstructed from medical computed tomograms. Maximal contractile muscle forces were estimated from muscular anatomical cross-sectional areas (CSAs). Fifty-nine landmarks were used to represent skull morphology. A partial least squares analysis was performed to assess the association between skull shape and muscle force, and FEA was used to compare the deformation (strains) generated during incisor and molar bites in two individuals representing extremes of morphological variation in the sample. The results showed that only the proportion of total muscle CSA accounted for by the temporalis appears associated with skull morphology, albeit weekly. However, individuals with a large temporalis tend to possess a relatively wider face, a narrower, more vertically oriented maxilla and a lower positioning of the coronoid process. The FEAs showed that, despite differences in morphology, biting results in similar modes of deformation for both crania, but with localised lower magnitudes of strains arising in the individual with the narrowest, most vertically oriented maxilla. Our results suggest that the morphology of the maxilla modulates the transmission of forces generated during mastication to the rest of the cranium by deforming less in individuals with the ability to generate proportionately larger temporalis muscle forces.

The Predictability from Skull Morphology of Temporalis and Masseter Muscle Cross-Sectional Areas in Humans.

To carry out functional simulations of the masticatory system that aim to predict strain magnitudes it is important to apply appropriate jaw-elevator muscle forces. Force magnitude estimation from directly measured muscle physiological cross-sectional area or anatomical cross-sectional area (CSA) is not possible for fossils and skeletal material from museum collections. In these cases, muscle CSAs are often estimated from bony features. This approach has been shown to be inaccurate in a prior study based on direct measurements from cadavers. Postmortem alterations as well as age changes in muscle form might explain this discrepancy. As such, the present study uses CT images from 20 living individuals to directly measure temporalis and masseter muscle CSAs and estimated cross-sectional areas (ECSAs) from bony features. The relationships between CSAs and ECSAs were assessed by comparing mean values and by examining correlations. ECSAs are up to 100% greater than CSA and the means of these variables for each muscle differ significantly. Further, ECSA is significantly correlated with CSA for temporalis but not masseter. Cranial centroid size is only significantly associated with CSA for temporalis. These findings indicate that ECSAs should be employed with caution in simulations of human masticatory system functioning; they do not reflect CSAs and it is plausible that this also applies to studies of closely related living and fossil taxa. When ECSAs are used, sensitivity analyses are required to determine the impact of potential errors.