Bone health in children with inflammatory bowel disease : adjusting for bone age

OBJECTIVES: Clinical results of bone mineral density for children with inflammatory bowel disease are commonly reported using reference data for chronological age. It is known that these children, particularly those with Crohn disease, experience delayed growth and maturation. Therefore, it is more appropriate to compare clinical results with bone age rather than chronological age. MATERIALS AND METHODS: Areal bone mineral density (aBMD) was measured using dual energy x-ray absorptiometry, and bone age was assessed using the Tanner-Whitehouse 3 method from a standard hand/wrist radiograph. Results were available for 44 children ages 7.99 to 16.89 years. Areal bone mineral density measurements were converted to z scores using both chronological and bone ages for each subject. RESULTS: Areal bone mineral density z scores calculated using bone age, as opposed to chronological age, were significantly improved for both the total body and lumbar spine regions of interest. When subjects were grouped according to diagnosis, bone age generated z scores remained significantly improved for those with Crohn disease but not for those diagnosed with ulcerative colitis. Grouping of children with Crohn disease into younger and older ages produced significantly higher z scores using bone age compared with chronological for the older age group, but not the younger age group. CONCLUSIONS: Our findings, in accordance with those presented in the literature, suggest that aBMD results in children with Crohn disease should include the consideration of bone age, rather than merely chronological age. Bone size, although not as easily available, would also be an important consideration for interpreting results in paediatric populations. © 2009 by European Society for Pediatric Gastroenterology, Hepatology, and Nutrition and North American Society for Pediatric Gastroenterology, Hepatology, and Nutrition.

Influences of age and mechanical stability on volume, microstructure, and mineralization of the fracture callus during bone healing : Is osteoclast activity the key to age-related impaired healing?

Earlier studies have shown that the influence of fixation stability on bone healing diminishes with advanced age. The goal of this study was to unravel the relationship between mechanical stimulus and age on callus competence at a tissue level. Using 3D in vitro micro-computed tomography derived metrics, 2D in vivo radiography, and histology, we investigated the influences of age and varying fixation stability on callus size, geometry, microstructure, composition, remodeling, and vascularity. Compared were four groups with a 1.5-mm osteotomy gap in the femora of Sprague–Dawley rats: Young rigid (YR), Young semirigid (YSR), Old rigid (OR), Old semirigid (OSR). Hypothesis was that calcified callus microstructure and composition is impaired due to the influence of advanced age, and these individuals would show a reduced response to fixation stabilities. Semirigid fixations resulted in a larger ΔCSA (Callus cross-sectional area) compared to rigid groups. In vitro μCT analysis at 6 weeks postmortem showed callus bridging scores in younger animals to be superior than their older counterparts (pb0.01). Younger animals showed (i) larger callus strut thickness (pb0.001), (ii) lower perforation in struts (pb0.01), and (iii) higher mineralization of callus struts (pb0.001). Callus mineralization was reduced in young animals with semirigid fracture fixation but remained unaffected in the aged group. While stability had an influence, age showed none on callus size and geometry of callus. With no differences observed in relative osteoid areas in the callus ROI, old as well as semirigid fixated animals showed a higher osteoclast count (pb0.05). Blood vessel density was reduced in animals with semirigid fixation (pb0.05). In conclusion, in vivo monitoring indicated delayed callus maturation in aged individuals. Callus bridging and callus competence (microstructure and mineralization) were impaired in individuals with an advanced age. This matched with increased bone resorption due to higher osteoclast numbers. Varying fixator configurations in older individuals did not alter the dominant effect of advanced age on callus tissue mineralization, unlike in their younger counterparts. Age-associated influences appeared independent from stability. This study illustrates the dominating role of osteoclastic activity in age-related impaired healing, while demonstrating the optimization of fixation parameters such as stiffness appeared to be less effective in influencing healing in aged individuals.

Lateral bone density variations in the scoliotic spine

Adolescent Idiopathic Scoliosis (AIS) is the most common deformity of the spine, affecting 2-4% of the population. Previous studies have shown that the vertebrae in scoliotic spines undergo abnormal shape changes, however there has been little exploration of how scoliosis affects bone density distribution within the vertebrae. In this study, existing CT scans of 53 female idiopathic scoliosis patients with right-sided main thoracic curves were used to measure the lateral (right to left) bone density profile at mid-height through each vertebral body. Five key bone density profile measures were identified from each normalised bone density distribution, and multiple regression analysis was performed to explore the relationship between bone density distribution and patient demographics (age, height, weight, body mass index (BMI), skeletal maturity, time since Menarche, vertebral level, and scoliosis curve severity). Results showed a marked convex/concave asymmetry in bone density for vertebral levels at or near the apex of the scoliotic curve. At the apical vertebra, mean bone density at the left side (concave) cortical shell was 23.5% higher than for the right (convex) cortical shell, and cancellous bone density along the central 60% of the lateral path from convex to concave increased by 13.8%. The centre of mass of the bone density profile at the thoracic curve apex was located 53.8% of the distance along the lateral path, indicating a shift of nearly 4% toward the concavity of the deformity. These lateral bone density gradients tapered off when moving away from the apical vertebra. Multi-linear regressions showed that the right cortical shell peak bone density is significantly correlated with skeletal maturity, with each Risser increment corresponding to an increase in mineral equivalent bone density of 4-5%. There were also statistically significant relationships between patient height, weight and BMI, and the gradient of cancellous bone density along the central 60% of the lateral path. Bone density gradient is positively correlated with weight, and negatively correlated with height and BMI, such that at the apical vertebra, a unit decrease in BMI corresponds to an almost 100% increase in bone density gradient.

Runx2 overexpression in bone marrow stromal cells accelerates bone formation in critically-sized femoral defects

The repair of large non-unions in long bones remains a significant clinical problem due to high failure rates and limited tissue availability for auto- and allografts. Many cell-based strategies for healing bone defects deliver bone marrow stromal cells to the defect site to take advantage of the inherent osteogenic capacity of this cell type. However, many factors, including donor age and ex vivo expansion of the cells, cause bone marrow stromal cells to lose their differentiation ability. To overcome these limitations, we have genetically engineered bone marrow stromal cells to constitutively overexpress the osteoblast specific transcription factor Runx2. In the present study, we examined Runx2-modified bone marrow stromal cells, delivered via poly(caprolactone) scaffolds loaded with type I collagen meshes, in critically-sized segmental defects in rats compared to unmodified cells, cell-free scaffolds and empty defects. Runx2 expression in bone marrow stromal cells accelerated healing of critically-sized defects compared to unmodified bone marrow stromal cells and defects receiving cell-free treatments. These findings provide an accelerated method for healing large bone defects which may reduce recovery time and the need for external fixation of critically-sized defects.

Small animal bone healing models : standards, tips, and pitfalls results of a consensus meeting

Small animal fracture models have gained increasing interest in fracture healing studies. To achieve standardized and defined study conditions, various variables must be carefully controlled when designing fracture healing experiments in mice or rats. The strain, age and sex of the animals may influence the process of fracture healing. Furthermore, the choice of the fracture fixation technique depends on the questions addressed, whereby intra- and extramedullary implants as well as open and closed surgical approaches may be considered. During the last few years, a variety of different, highly sophisticated implants for fracture fixation in small animals have been developed. Rigid fixation with locking plates or external fixators results in predominantly intramembranous healing in both mice and rats. Locking plates, external fixators, intramedullary screws, the locking nail and the pin-clip device allow different degrees of stability resulting in various amounts of endochondral and intramembranous healing. The use of common pins that do not provide rotational and axial stability during fracture stabilization should be discouraged in the future. Analyses should include at least biomechanical and histological evaluations, even if the focus of the study is directed towards the elucidation of molecular mechanisms of fracture healing using the largely available spectrum of antibodies and gene-targeted animals to study molecular mechanisms of fracture healing. This review discusses distinct requirements for the experimental setups as well as the advantages and pitfalls of the different fixation techniques in rats and mice.

Error and uncertainty of adult age estimation of the pubic symphysis in an Australian sub-population using computed tomography

After attending this presentation, attendees will gain awareness of: (1) the error and uncertainty associated with the application of the Suchey-Brooks (S-B) method of age estimation of the pubic symphysis to a contemporary Australian population; (2) the implications of sexual dimorphism and bilateral asymmetry of the pubic symphysis through preliminary geometric morphometric assessment; and (3) the value of three-dimensional (3D) autopsy data acquisition for creating forensic anthropological standards. This presentation will impact the forensic science community by demonstrating that, in the absence of demographically sound skeletal collections, post-mortem autopsy data provides an exciting platform for the construction of large contemporary ‘virtual osteological libraries’ for which forensic anthropological research can be conducted on Australian individuals. More specifically, this study assesses the applicability and accuracy of the S-B method to a contemporary adult population in Queensland, Australia, and using a geometric morphometric approach, provides an insight to the age-related degeneration of the pubic symphysis. Despite the prominent use of the Suchey-Brooks (1990) method of age estimation in forensic anthropological practice, it is subject to intrinsic limitations, with reports of differential inter-population error rates between geographical locations1-4. Australian forensic anthropology is constrained by a paucity of population specific standards due to a lack of repositories of documented skeletons. Consequently, in Australian casework proceedings, standards constructed from predominately American reference samples are applied to establish a biological profile. In the global era of terrorism and natural disasters, more specific population standards are required to improve the efficiency of medico-legal death investigation in Queensland. The sample comprises multi-slice computed tomography (MSCT) scans of the pubic symphysis (slice thickness: 0.5mm, overlap: 0.1mm) on 195 individuals of caucasian ethnicity aged 15-70 years. Volume rendering reconstruction of the symphyseal surface was conducted in Amira® (v.4.1) and quantitative analyses in Rapidform® XOS. The sample was divided into ten-year age sub-sets (eg. 15-24) with a final sub-set of 65-70 years. Error with respect to the method’s assigned means were analysed on the basis of bias (directionality of error), inaccuracy (magnitude of error) and percentage correct classification of left and right symphyseal surfaces. Morphometric variables including surface area, circumference, maximum height and width of the symphyseal surface and micro-architectural assessment of cortical and trabecular bone composition were quantified using novel automated engineering software capabilities. The results of this study demonstrated correct age classification utilizing the mean and standard deviations of each phase of the S-B method of 80.02% and 86.18% in Australian males and females, respectively. Application of the S-B method resulted in positive biases and mean inaccuracies of 7.24 (±6.56) years for individuals less than 55 years of age, compared to negative biases and mean inaccuracies of 5.89 (±3.90) years for individuals greater than 55 years of age. Statistically significant differences between chronological and S-B mean age were demonstrated in 83.33% and 50% of the six age subsets in males and females, respectively. Asymmetry of the pubic symphysis was a frequent phenomenon with 53.33% of the Queensland population exhibiting statistically significant (χ2 - p<0.01) differential phase classification of left and right surfaces of the same individual. Directionality was found in bilateral asymmetry, with the right symphyseal faces being slightly older on average and providing more accurate estimates using the S-B method5. Morphometric analysis verified these findings, with the left surface exhibiting significantly greater circumference and surface area than the right (p<0.05). Morphometric analysis demonstrated an increase in maximum height and width of the surface with age, with most significant changes (p<0.05) occurring between the 25-34 and 55-64 year age subsets. These differences may be attributed to hormonal components linked to menopause in females and a reduction in testosterone in males. Micro-architectural analysis demonstrated degradation of cortical composition with age, with differential bone resorption between the medial, ventral and dorsal surfaces of the pubic symphysis. This study recommends that the S-B method be applied with caution in medico-legal death investigations of unknown skeletal remains in Queensland. Age estimation will always be accompanied by error; therefore this study demonstrates the potential for quantitative morphometric modelling of age related changes of the pubic symphysis as a tool for methodological refinement, providing a rigor and robust assessment to remove the subjectivity associated with current pelvic aging methods.

Quantitative morphometric examination of the human pubic symphyseal surface of Australian individuals with age using computed tomography

Current forensic practice in age estimation relies on the application of morphological standards as a means to characterize complex threedimensional skeletal surfaces. Research in our laboratory has demonstrated that the application of the morphologically based Suchey-Brooks method to a contemporary Queensland, Australian population demonstrated significant inaccuracy in age-estimation. Consequently, this study presents preliminary results to quantify age-related skeletal changes of the pubic symphysis in Queensland individuals using novel geometric and micro-architectural protocols that have the potential of improving age estimation in the forensic context. Computed tomography scans of the right and left pubis were obtained from Caucasian individuals aged 15–70 years (n=195) from the Queensland Health Forensic and Scientific Services. Morphometric variables including surface area, circumference, maximum height and width of the symphyseal surface, and micro-architectural assessment of cortical and trabecular bone structure were conducted in Rapidform XOS and Osteomeasure, respectively. Morphometric analysis demonstrated increases in maximum height and width of the surface with age independent of gender, with most significant (P<0.05) changes between the 25–34 and 55–64 year subsets. Sexual dimorphism and bilateral asymmetry were prominent features in the Queensland population. Micro-architectural analysis demonstrated degradation of cortical composition with age, with differential bone resorption between the medial, ventral and dorsal aspects of the symphysis. The ability to quantitatively model age-related changes to the pubic symphysis provides potential for future methodological refinement, where rigor and robust geometric assessment of the surface may remove the subjectivity associated with aging the pubic symphysis.

Thorough assessment of DNA preservation from fossil bone and sediments excavated from a late Pleistocene–Holocene cave deposit on Kangaroo Island, South Australia

Fossils and sediments preserved in caves are an excellent source of information for investigating impacts of past environmental changes on biodiversity. Until recently studies have relied on morphology-based palaeontological approaches, but recent advances in molecular analytical methods offer excellent potential for extracting a greater array of biological information from these sites. This study presents a thorough assessment of DNA preservation from late Pleistocene–Holocene vertebrate fossils and sediments from Kelly Hill Cave Kangaroo Island, South Australia. Using a combination of extraction techniques and sequencing technologies, ancient DNA was characterised from over 70 bones and 20 sediment samples from 15 stratigraphic layers ranging in age from >20 ka to ∼6.8 ka. A combination of primers targeting marsupial and placental mammals, reptiles and two universal plant primers were used to reveal genetic biodiversity for comparison with the mainland and with the morphological fossil record for Kelly Hill Cave. We demonstrate that Kelly Hill Cave has excellent long-term DNA preservation, back to at least 20 ka. This contrasts with the majority of Australian cave sites thus far explored for ancient DNA preservation, and highlights the great promise Kangaroo Island caves hold for yielding the hitherto-elusive DNA of extinct Australian Pleistocene species.

Results using Trabecular Metal™ augments in combination with acetabular impaction bone grafting in deficient acetabula

We examined whether the use of trabecular metal wedges to fill segmental defects is an effective method of socket reconstruction when used in combination with impaction grafting and implantation of a cemented socket. Fifteen hips in 14 patients underwent impaction grafting in combination with a TM wedge with a minimum of 2 years follow-up. All patients had their defects assessed using the Paprosky classification. Patients were reviewed with x-rays and migration of the implant was measured. Outcome scores were also collected. Mean follow-up was 39 months (25-83). The mean age at surgery was 67.8 (49-85) years. Seven of the patients had previously undergone impaction grafting with the use of a stainless steel rim mesh to constrain the graft. None of the patients had failed either clinically or radiologically.

Sex-specific compromised bone healing in female rats might be associated with a decrease in mesenchymal stem cell quantity

Introduction The clinically known importance of patient sex as a major risk factor for compromised bone healing is poorly reflected in animal models. Consequently, the underlying cellular mechanisms remain elusive. Because mesenchymal stem cells (MSCs) are postulated to regulate tissue regeneration and give rise to essential differentiated cell types, they may contribute to sex-specific differences in bone healing outcomes. Methods We investigated sex-specific variations in bone healing and associated differences in MSC populations. A 1.5 mm osteotomy gap in the femora of 8 male and 8 female 12-month-old Sprague-Dawley rats was stabilized by an external fixator. Healing was analyzed in terms of biomechanical testing, bridging and callus size over time (radiography at 2, 4, and 6 weeks after surgery), and callus volume and geometry by μCT at final follow-up. MSCs were obtained from bone marrow samples of an age-matched group of 12 animals (6 per gender) and analyzed for numbers of colony-forming units (CFUs) and their capacity to differentiate and proliferate. The proportion of senescent cells was determined by β-galactosidase staining. Results Sex-specific differences were indicated by a compromised mechanical competence of the callus in females compared with males (maximum torque at failure, p = 0.028). Throughout the follow-up, the cross-sectional area of callus relative to bone was reduced in females (p ≤ 0.01), and the bridging of callus was delayed (p 2weeks = 0.041). μCT revealed a reduced callus size (p = 0.003), mineralization (p = 0.003) and polar moment of inertia (p = 0.003) in female animals. The female bone marrow contained significantly fewer MSCs, represented by low CFU numbers in both femora and tibiae (p femur = 0.017, p tibia = 0.010). Functional characteristics of male and female MSCs were similar. Conclusion Biomechanically compromised and radiographically delayed bone formation were distinctive in female rats. These differences were concomitant with a reduced number of MSCs, which may be causative for the suboptimal bone healing.

Mice lacking β-Adrenergic receptors have increased bone mass but are not protected from deleterious skeletal effects of ovariectomy

Activation of β2-adrenergic receptors inhibits osteoblastic bone formation and enhances osteoclastic bone resorption. Whether β-blockers inhibit ovariectomy-induced bone loss and decrease fracture risk remains controversial. To further explore the role of β-adrenergic signaling in skeletal acquisition and response to estrogen deficiency, we evaluated mice lacking the three known β-adrenergic receptors (β-less). Body weight, percent fat, and bone mineral density were significantly higher in male β-less than wild-type (WT) mice, more so with increasing age. Consistent with their greater fat mass, serum leptin was significantly higher in β-less than WT mice. Mid-femoral cross-sectional area and cortical thickness were significantly higher in adult β-less than WT mice, as were femoral biomechanical properties (+28 to +49%, P < 0.01). Young male β-less had higher vertebral (1.3-fold) and distal femoral (3.5-fold) trabecular bone volume than WT (P < 0.001 for both) and lower osteoclast surface. With aging, these differences lessened, with histological evidence of increased osteoclast surface and decreased bone formation rate at the distal femur in β-less vs. WT mice. Serum tartrate-resistance alkaline phosphatase-5B was elevated in β-less compared with WT mice from 8–16 wk of age (P < 0.01). Ovariectomy inhibited bone mass gain and decreased trabecular bone volume/total volume similarly in β-less and WT mice. Altogether, these data indicate that absence of β-adrenergic signaling results in obesity and increased cortical bone mass in males but does not prevent deleterious effects of estrogen deficiency on trabecular bone microarchitecture. Our findings also suggest direct positive effects of weight and/or leptin on bone turnover and cortical bone structure, independent of adrenergic signaling.

Temporal Characterization of Ossification of the Crania in Australian Subadults: New standards for Age Estimation using Computed Tomography

After attending this presentation, attendees will gain awareness of the ontogeny of cranial maturation, specifically: (1) the fusion timings of primary ossification centers in the basicranium; and (2) the temporal pattern of closure of the anterior fontanelle, to develop new population-specific age standards for medicolegal death investigation of Australian subadults. This presentation will impact the forensic science community by demonstrating the potential of a contemporary forensic subadult Computed Tomography (CT) database of cranial scans and population data, to recalibrate existing standards for age estimation and quantify growth and development of Australian children. This research welcomes a study design applicable to all countries faced with paucity in skeletal repositories. Accurate assessment of age-at-death of skeletal remains represents a key element in forensic anthropology methodology. In Australian casework, age standards derived from American reference samples are applied in light of scarcity in documented Australian skeletal collections. Currently practitioners rely on antiquated standards, such as the Scheuer and Black1 compilation for age estimation, despite implications of secular trends and population variation. Skeletal maturation standards are population specific and should not be extrapolated from one population to another, while secular changes in skeletal dimensions and accelerated maturation underscore the importance of establishing modern standards to estimate age in modern subadults. Despite CT imaging becoming the gold standard for skeletal analysis in Australia, practitioners caution the application of forensic age standards derived from macroscopic inspection to a CT medium, suggesting a need for revised methodologies. Multi-slice CT scans of subadult crania and cervical vertebrae 1 and 2 were acquired from 350 Australian individuals (males: n=193, females: n=157) aged birth to 12 years. The CT database, projected at 920 individuals upon completion (January 2014), comprises thin-slice DICOM data (resolution: 0.5/0.3mm) of patients scanned since 2010 at major Brisbane Childrens Hospitals. DICOM datasets were subject to manual segmentation, followed by the construction of multi-planar and volume rendering cranial models, for subsequent scoring. The union of primary ossification centers of the occipital bone were scored as open, partially closed or completely closed; while the fontanelles, and vertebrae were scored in accordance with two stages. Transition analysis was applied to elucidate age at transition between union states for each center, and robust age parameters established using Bayesian statistics. In comparison to reported literature, closure of the fontanelles and contiguous sutures in Australian infants occur earlier than reported, with the anterior fontanelle transitioning from open to closed at 16.7±1.1 months. The metopic suture is closed prior to 10 weeks post-partum and completely obliterated by 6 months of age, independent of sex. Utilizing reverse engineering capabilities, an alternate method for infant age estimation based on quantification of fontanelle area and non-linear regression with variance component modeling will be presented. Closure models indicate that the greatest rate of change in anterior fontanelle area occurs prior to 5 months of age. This study complements the work of Scheuer and Black1, providing more specific age intervals for union and temporal maturity of each primary ossification center of the occipital bone. For example, dominant fusion of the sutura intra-occipitalis posterior occurs before 9 months of age, followed by persistence of a hyaline cartilage tongue posterior to the foramen magnum until 2.5 years; with obliteration at 2.9±0.1 years. Recalibrated age parameters for the atlas and axis are presented, with the anterior arch of the atlas appearing at 2.9 months in females and 6.3 months in males; while dentoneural, dentocentral and neurocentral junctions of the axis transitioned from non-union to union at 2.1±0.1 years in females and 3.7±0.1 years in males. These results are an exemplar of significant sexual dimorphism in maturation (p<0.05), with girls exhibiting union earlier than boys, justifying the need for segregated sex standards for age estimation. Studies such as this are imperative for providing updated standards for Australian forensic and pediatric practice and provide an insight into skeletal development of this population. During this presentation, the utility of novel regression models for age estimation of infants will be discussed, with emphasis on three-dimensional modeling capabilities of complex structures such as fontanelles, for the development of new age estimation methods.

The global burden attributable to low bone mineral density

Introduction The Global Burden of Disease Study 2010 estimated the worldwide health burden of 291 diseases and injuries and 67 risk factors by calculating disability-adjusted life years (DALYs). Osteoporosis was not considered as a disease, and bone mineral density (BMD) was analysed as a risk factor for fractures, which formed part of the health burden due to falls. Objectives To calculate (1) the global distribution of BMD, (2) its population attributable fraction (PAF) for fractures and subsequently for falls, and (3) the number of DALYs due to BMD. Methods A systematic review was performed seeking population-based studies in which BMD was measured by dual-energy X-ray absorptiometry at the femoral neck in people aged 50 years and over. Age- and sex-specific mean ± SD BMD values (g/cm2) were extracted from eligible studies. Comparative risk assessment methodology was used to calculate PAFs of BMD for fractures. The theoretical minimum risk exposure distribution was estimated as the age- and sex-specific 90th centile from the Third National Health and Nutrition Examination Survey (NHANES III). Relative risks of fractures were obtained from a previous meta-analysis. Hospital data were used to calculate the fraction of the health burden of falls that was due to fractures. Results Global deaths and DALYs attributable to low BMD increased from 103 000 and 3 125 000 in 1990 to 188 000 and 5 216 000 in 2010, respectively. The percentage of low BMD in the total global burden almost doubled from 1990 (0.12%) to 2010 (0.21%). Around one-third of falls-related deaths were attributable to low BMD. Conclusions Low BMD is responsible for a growing global health burden, only partially representative of the real burden of osteoporosis.

Changes in cervical spine bone mineral density in response to flight training

BACKGROUND High magnitude loads and unusual loading regimes are two important determinants for increasing bone mass. Past research demonstrated that positive Gz-induced loading, providing high loads in an unaccustomed manner, had an osteogenic effect on bone. Another determinant of bone mass is that the bone response to loading is site specific. This study sought to further investigate the site specific bone response to loading, examining the cervical spine response, the site suspected of experiencing the greatest loading, to high performance flight. METHODS Bone mineral density (BMD) and bone mineral content (BMC) was monitored in 9 RAAF trainee fighter pilots completing an 8-mo flight training course on a PC-9 and 10 age-height-weight-matched controls. RESULTS At completion of the course, the pilots had a significant increase in cervical spine BMD and total body BMC. No significant changes were found for the control group. CONCLUSIONS This study demonstrated that the physical environment associated with flight training may have contributed to a significant increase in cervical spine bone mass in the trainee PC-9 pilots. The increase in bone mass was possibly a response to the strain generated by the daily wearing of helmet and mask assembly under the influence of positive sustained accelerative forces.