Browsing by Author "Organ, Jason"
Now showing 1 - 13 of 13
- Results Per Page
- Sort Options
Item Absolute and Relative Morphometric Differences in the Craniofacial Skeleton of OIM-/- Mice and Wild-Type Littermates(2019-03-05) McBride, Alexandra H.; Organ, Jason; Menegaz, Rachel A.; Ladd, SummerPurpose: Osteogenesis Imperfecta (OI, or “Brittle Bone Disease”) is a disorder caused by genetic point mutations in COL1A1/COL1A2 which affect the synthesis of type I collagen (Col1). Humans with the severe type III OI exhibit increased susceptibility to skeletal fractures and shortened stature, as well as cranial dysmorphologies and dental malocclusions. Mouse models of Col1 defects report postcranial phenotypes similar to those seen in humans, with a limited number of studies reporting alterations to cranial and dental integrity. This project tests the hypothesis that the reduced craniofacial dimensions reported in both humans and mice with Col1 defects are linked to an overall reduction in body size. Methods: The homozygous OI murine (OIM-/-) is a mouse strain with a nonlethal recessively inherited mutation of the COL1A2 gene. Wild-type (WT) and OIM-/- littermates were weaned at 21d and raised until adult (16 weeks). 3D morphometric landmarks were collected from serial in-vivo µCT scans at 4, 10, and 16 weeks using etdips software. Past 2.17 software was used to Procrustes-transform (rotate and translate) the landmark data, and to calculate interlandmark distances (ILDs) and centroid sizes. ILDs were scaled against skull/mandible centroid size and skull/mandible length to account for the effect of size. Mann-Whitney U tests (α=0.05) were used to compare centroid sizes and both absolute and relative (scaled) ILDs between the genotypes. Results: When comparing absolute morphometric distances, adult OIM-/- mice have shorter skulls, basicrania, palates, mandibles, and toothrows. However, OIM-/-mice are smaller overall than their WT littermates as measured by both body mass and craniomandibular centroid sizes. When the effects of size are accounted for, the trend for interlandmark distances in WT mice to be greater than those in OIM-/- mice is significantly reduced or even reversed. For example, when scaled to centroid size, no significant difference exists between WT and OIM-/- mice in skull, basicranial, or mandibular length. OIM-/- mice have a relatively short midface, short nasal bones, tall mandibular corpora and long mandibular toothrows. Conclusions: These findings underscore the importance of size and scaling in morphometric analyses. The deleterious effect of Col1 mutations on global skeletal dimensions, in combination with localized morphometric changes, may underlie the facial phenotype seen in human patients with OI type III. Attempts to identify these localized changes should first account for the restricted growth and small body sizes present in individuals with OI.Item Contrast-enhanced micro-CT approaches for visualizing musculoskeletal development in neonatal mice(2023) Stalls, Javan; Miller, Courtney; Gonzales, Lauren; Lesciotto, Kate; Handler, Emma; Organ, Jason; Menegaz, Rachel A.Contrast-enhanced micro-CT approaches for visualizing musculoskeletal development in neonatal mice Javan A. Stalls, Courtney A. Miller, Jason M. Organ, Emma K. Handler, Lauren A. Gonzales, Kate M. Lesciotto, Rachel A. Menegaz Purpose: While there are many forms of radiological imaging that can be used to gather anatomical data from biological specimens, computed tomography (CT) imaging has been the gold standard for visualizing dense tissue, such as bone, with detailed resolution. However, this imaging modality is not well suited for soft tissues (muscle, brain, abdominal organs, cartilage, etc.) due to their decreased tissue density. The inability to distinguish between soft tissues in CT scans limits our ability to investigate the bone-muscle interactions known to stimulate and direct bone modeling during early postnatal development. The development of contrast-enhancing staining agents, capable of binding materials to increase their radiodensity, has allowed for more accurate and enhanced visualizations of less dense soft tissues, such as muscle and brain structures. Contrast agents such as iodine have differential affinities for the different soft tissues in the body allowing for easier visualization and segmentation of soft tissues in relation to the skeleton. Previous studies have used contrast-enhanced CT (CE-CT) scanning to analyze early development of mice from prenatal stages to postnatal day 7. However, additional CE-CT imaging during the first three postnatal weeks is needed to understand muscle-bone interactions during critical periods of behavioral development, such as suckling and weaning. The goal of this project is to develop a CE-CT protocol and corresponding anatomical atlas showing the development of skeletal and soft tissue structures in the crania of neonatal mice from birth to weaning. Methods: Neonatal and preweaning mice (B6C3Fe a/a-Col1a2OIM/J) were euthanized on day of birth (P0), postnatal day 7 (P7), and postnatal day 14 (P14). Ethanol-fixed tissues were submerged in 1.25% iodine in 70% ethanol (I2E) for 2-14 days, with the skin intact in order to preserve cutaneous musculature. Both pre-stained and post-stained tissues were scanned using a MRS CT-80 micro-CT machine (20 µm3 voxel resolution). Results: Preliminary CE-CT scans following 10 days in an iodine stain present improved visualization of soft tissue (brain structures, cranial muscles, salivary glands) when compared to the baseline bone CT scans. Conclusion: These scans will be used to develop 3D models of musculoskeletal ontogeny from birth-weaning, providing insights into this critical developmental period. The use of CT contrast agents such as iodine offers new opportunities to investigate the anatomical interactions of bone and muscle during early development, and can be applied to investigate models of both normal growth and pathological disorders affecting musculoskeletal growth.Item Cranial Bone Ossification Trajectories in a Mouse Model of Osteogenesis Imperfecta(2023) Miller, Courtney; Lugo, Laura; Husain, Tooba S.; Organ, Jason; Handler, Emma; Gonzales, Lauren; Menegaz, Rachel A.Purpose: Osteogenesis imperfecta (OI) is a genetic disorder that affects the production of type I collagen. Altered collagen production results in delayed or impaired skeletal formation and biomineralization. It also results in the defining characteristics of OI: brittle bones and high rates of fractures. Investigations of skeletal growth in OI have primarily focused on the postcranial skeleton, where interrupted, atypical, and disorganized ossification is seen at long bone growth plates. However, few studies have investigated changes in craniofacial growth in OI and there are currently few early interventions to improve growth trajectories in this region. The current medication prescribed for children with OI to improve skeletal growth, such as bisphosphonates, have major side effects and are not suitable for long-term use. A better understanding of craniofacial development in OI can help with targeting specific developmental stages when new treatments can be administered to provide the best results. The aim of this study is to examine cranial ossification from birth to weaning to determine where and when differences in growth occur in OI. We hypothesize that starting at birth mice with OI will have delayed craniofacial growth due to the poor collagen formation. Methods: To test our hypothesis, we collected cranial bone volumes from micro-CT scans of the homozygous recessive OI murine model (OIM or B6C3Fe a/a-Col1a2oim/oim) and compared them to their wild type (WT) littermates. The OIM model has a COL1A2 mutation that has been found to express a similar skeletal phenotype to the severe form (type III) of OI in humans. Bone volumes were collected from birth (P0) and weaning (P21) from the nasal, frontal, parietal, interparietal, and occipital bones (n=2/genotype/timepoint). Results: At birth, OIM and WT bone volumes were similar. By weaning, bone volume was lower in OIM mice compared to WT mice. Our results demonstrate that OIM mice have reduced rates of bone ossification between birth and weaning, and these differences are most profound in the facial and occipital regions. Additionally, OIM skulls are characterized by low bone volume and potential delays in the closure of cranial sutures and fontanelles. Conclusions: This study suggests that the divergence in cranial ossification rates related to COL1A2 mutations occurs postnatally. Interventions to recover craniofacial bone growth in this experimental model should focus on the critical growth period between birth and weaning. Results from this research have the potential to assist in developing treatments and highlight the importance of early life development of the craniofacial bones in human patients with OI.Item Craniofacial Bone Mineral Density During Growth in Mice with Osteogenesis Imperfecta (OI)(2021) Miller, Courtney; Wright, Tommy; McBride, Alexandra H.; Organ, Jason; Menegaz, Rachel A.Purpose: Osteogenesis imperfecta(OI) is a genetic connective tissue defect resulting in fragile bones due to mutations affecting formation of type I collagen. Low bone mineral density (BMD) in the post-cranial skeleton has been reported in human patients and murine models with OI, yet little is known about craniofacial biomineralization in the disorder. Typically, skeletal mineralization is responsive to the strain environment. The aim of this study is to investigate longitudinal changes in craniofacial BMD in a mouse model of OI type III (most severe form), and to quantify BMD in regions relative to feeding biomechanical forces. Methods: Homozygous recessive OI murine (OIM), a mouse strain with a COL1A2 mutation modeling OI type III, and unaffected wild-type (WT) littermates were micro-CT scanned at weeks 4, 10, and 16. BMD in eight regions was analyzed using Bruker CTAnalyzer software and Mann-Whitney U tests. Results: OIM mice had significantly (p< 0.05) lower BMD than WT mice in all eight regions during week 4, no significant differences in week 10, and significant differences at the parietal bone, mandibular symphysis, and maxillary incisor regions during week 16. Absolute BMD was higher within regions proximal to the bite point at skeletal maturity. Conclusions: These results support a trend that OIM mice have lower BMD in the craniofacial skeleton compared to WT mice throughout growth and BDM in all mice is affected by proximity to bite forces. Understanding craniofacial mineralization patterns in OI could assist in the implementation of pharmaceutical interventions to increase BMD.Item Craniofacial Bone Mineral Density in Mice with Osteogenesis Imperfecta (OI)(2019-03-05) Ladd, Summer; Organ, Jason; Menegaz, Rachel A.; McBride, Alexandra H.Purpose: Osteogenesis imperfecta (OI) is a rare genetic disorder characterized by the abnormal synthesis and assembly of type I collagen (Col1), a major organic component of bone. Clinical manifestations of the severe OI type III include small body size, limb deformities, and low bone mineral density (BMD) within the post-cranial skeleton. OI type III often co-occurs with craniofacial defects, such as dentinogenesis imperfecta (DI). The goals of this study are: (1) to examine whether Col1 defects, as seen in OI type III, affect BMD within the craniofacial skeleton; (2) to examine whether craniofacial BMD covaries with diet-related biomechanical loading. Methods: The homozygous recessive murine mouse (OIM-/-) is a model for OI Type III. Similar to human OI patients, OIM-/- mice exhibit low post-cranial BMD, smaller body size, and DI. OIM-/- mice and WT littermates were weaned at 21 days and raised on either hard (high loading) or soft (low loading) diets. This resulted in four genotype x diet treatment groups: OIM-hard (n=6), OIM-soft (n=3), WT-hard (n=9), and WT-soft (n=3). Micro-CT scans were collected at 16 weeks (skeletal maturity). BMD was measured using Bruker CTAnalyzer software for eight regions of interest (ROIs) within the mandible (TMJ, corpus at the second molar, and symphysis), facial skeleton (nasal bone, maxilla at the second molar, premaxilla at the incisor), and cranial vault (frontal and parietal bones). Pairwise Mann-Whitney U tests were used to statistically compare BMD between treatments (α = 0.05). Results: At all ROIs except for the frontal bone, WT-hard mice had significantly (p p = 0.052) with the current sample sizes. Similarly, at the mandibular and cranial vault ROIs, WT-soft mice tended to have higher BMD than OIM-hard and/or OIM-soft mice (p Conclusions: These results suggest that craniofacial BMD is generally lower in individuals with Col1 defects, consistent with the postcranial presentation. WT mice raised on a hard diet were observed to have the highest BMD measurements across the craniofacial skeleton, however no significant differences were observed between OIM-/- mice raised on hard versus soft diets. While diet-associated loading may influence craniofacial BMD, in this study Col1 status appears to be the primary determinant of BMD.Item Craniofacial Morphology of Juvenile Mice with Osteogenesis Imperfecta(2020) Menegaz, Rachel A.; Organ, Jason; Steele, Ashley T.Osteogenesis imperfecta (OI) type III is a severe genetic disorder of type I collagen (Col1) which results in bone fragility, reduced stature, and impaired craniofacial growth. To investigate the mechanisms by which Col1 mutations alter craniofacial growth, we used the homozygous recessive OI murine (OIM) mouse model, which is known to exhibit human-like adult phenotypes but for which the juvenile phenotype is unknown. Weaning OIM and wild type (WT) littermates were µCT scanned at 21 days. Craniofacial landmarks were collected using 3D Slicer software. Interlandmark distances (ILDs) and centroid sizes were calculated using Past 2.17 software. ILDs were scaled against skull/mandible centroid size to remove the effect of overall body size for shape analyses. Mann-Whitney U-tests were used to compare absolute and relative (scaled) ILDs between genotypes. Craniomandibular centroid sizes and absolute linear distances (skull, rostrum, palate, and mandible lengths) demonstrate that OIM mice are smaller overall compared to WT littermates. When scaled to centroid size, juvenile OIM mice have a decrease in midface height, nasal and mandibular diastema length but increased hemimandible length compared to WT mice. For a given skull length, OIM mice have shorter faces in both the anteroposterior and dorsoventral dimensions. The morphometric changes seen in juvenile OIM mice replicate the midfacial hypoplasia seen in human children with OI. This mouse model can be used to investigate the structural changes underlying the human OI phenotype and potential therapeutic interventions. These results can be used to inform future investigations of Col1 in craniofacial development.Item Effects of Osteogenesis Imperfecta on Dental Tissue Volumes in Mice(2022) Moore, Jacob; Handler, Emma; Menegaz, Rachel A.; Gonzales, Lauren; Organ, JasonOsteogenesis imperfecta (OI), commonly known as brittle bone disease, is associated with lifelong dental problems, including increased dental fractures, discolored teeth, and malocclusion. OI is a disorder of the type I collagen protein. Insufficient amounts or misshapen forms of this protein lead to disruptions in the microstructure of bone and teeth tissues. Dentin, the hard tissue which comprises the bulk of the tooth and absorbs shock forces during chewing, develops on a type I collagen matrix. Thus, collagen abnormalities in OI lead to disorganized and less stable dentin. Further, teeth in people with OI frequently exhibit dentin hypertrophy, where increased amounts of dentin are deposited at the interior of the tooth, shortly after dental eruption. Enamel, unlike dentin, develops on a matrix of non-collagenous proteins, and is thought to develop normally in OI. However, abnormalities in the underlying dentin in OI can lead to enamel fractures. Finally, the teeth in people with OI have often been noted to be smaller than those of people without OI. Because dental development occurs early in life, there is a lack of data surrounding the developmental processes and associated issues in dental development of children with OI. Mice are commonly used models for dental development, yet, this process has not yet been studied in mice. The objective of this study is to compare dental tissue volumes of teeth from mice with OI (oim) and wild type mice (wt) at different developmental stages to determine the degree of volume and gross dental size differences during late growth stages. Three-dimensional models of upper and lower first molars and incisors were created from microCT scans from oim and wt mice. Scans were taken at weaning age (four weeks after birth; "W4") and young adulthood (sixteen weeks after birth; "W16"). Dental tissue volumes were measured using 3D Slicer and normalized to mandibular centroid size. Mann-Whitney U tests were used to compare tissue volumes between genotypes and age groups. At W4 and W16, oim mice had significantly lower dentin volumes and total tooth volumes for upper incisors than wt mice (p < 0.05), with no significant difference between groups for other tooth types at either timepoint. At W16, total tooth volume was significantly lower in oim mice for molars before adjusting for mandible size (p < 0.05). For both oim and wt groups, W16 mice had significantly greater dentin, enamel, and total tissue volumes for lower and upper incisors compared to W4 mice (p < 0.05), as well as greater dentin volumes for lower molars (p < 0.05). These results demonstrate that the trend of smaller teeth in humans with OI also holds for the oim mouse. These differences are present at both the juvenile and young adult life stages. This affirms the oim mouse as a possible model for dental development in humans with OI. Further studies are needed to determine the developmental program of these volume differences at earlier growth stages.Item Effects of Osteogenesis Imperfecta on the Cochlea and Sensorineural Hearing(2022) Huston, Lila Athena; Menegaz, Rachel A.; Handler, Emma; Organ, Jason; Gonzales, LaurenBackground: Osteogenesis imperfecta (OI), a developmental disorder of type I collagen, is known to cause hearing loss in ~ 60% of the diseased population. Identified forms include conductive hearing loss (17.4% of OI patients), involving loss of function within the ossicular chain, and sensorineural hearing loss (25.8%), resulting from damage to the cochlea, with the most predominant form being mixed hearing loss (56.8%), involving damage to both the cochlea and ossicles. While OI-related pathologies have largely focused on the middle ear, the pathological appearance of the cochlea (the organ most often compromised in OI-related hearing loss) has gained little focus. In this study, we examine OI-related pathologies on the cochlea in a mouse model for the severe type III OI, to document 1) any visible variation between WT and OI variants, and 2) assess the encroachment of the otic capsule onto the cochlea by analyzing differences in duct volumes. We hypothesize that cochlea in mice with OI will have less consistent morphology overall than their WT counterparts due to abnormal growth of the bony capsule. Methods: 16 week old OIM mice (B6C3Fe a/a-Col1a2oim/J) (n=6) were compared to unaffected wildtype (WT) littermates (n=6) with no known hearing defects. High-resolution micro-CT scans were created for all specimens and 3D models and volumes of the cochlea were generated using 3D Slicer software. Two-tailed Mann-Whitney U-tests were used to investigate differences between 1) right and left ears of the same mouse to examine intraindividual symmetry and 2) differences in volumes between WT and OI cochlea. Results: No major morphologic differences between OI and WT were observed, except for minor areas of higher ossification at the base of the cochlea, mostly within the OI sample. Within WT specimens, we observed little intraindividual difference in the cochlear volume (0-3%). Within OI specimens, significant differences were observed in cochlear volume between right and left ears in the same animal (4-15%; p< 0.05), indicating potential unilateral effects. When average WT and OI volumes were compared, there was much overlap between the two samples although the OI volumes had a significantly larger range than the WT range (Mann-Whitney U, p< 0.05). Discussion: Overall, our results indicate that mice with OI are much more likely to have evidence of unilateral cochlear volume losses, despite very little difference in overall shape appearance, possibly due to bony capsule encroachment. This find indicates an extremely high potential for sensorineural and mixed hearing loss in OI-bred mice and elucidates at least one mechanism behind how this type of hearing loss might be occurring. Little is known about the pathological appearance of the cochlea in OI, leading to difficulty in managing hearing loss. Further investigation of the etiology and progression of cochlear pathologies will allow for better outcomes in hearing for those patients afflicted with OI-related hearing loss.Item Masticatory muscle morphology in early postnatal mice with osteogenesis imperfecta(2024-03-21) Ansari, Zahra; Miller, Courtney; Emmanuel, Tanusha; Handler, Emma; Gonzales, Lauren; Organ, Jason; Menegaz, Rachel A.Purpose: Osteogenesis imperfecta (OI) is a connective tissue disorder resulting from mutations in COL1A1 or COL1A2, responsible for encoding type I collagen alpha chains. While OI is primarily distinguished by manifestations of bone fragility, including recurrent fractures and bone deformities, muscle abnormalities have also been documented in those affected by OI. While prior research has shown postcranial muscle weakness in mouse models of OI, it remains unclear whether this also applies to feeding musculature and if these differences are present at birth or develop postnatally. This study investigates the development of the masticatory muscles during the early postnatal period in a mouse model. Our hypothesis posits that mice affected by OI will exhibit decreased muscle mass, and therefore potentially weaker muscles, in comparison to unaffected mice. Methods: Cranial tissues from OIM mice (B6C3Fe a/a-Col1a2oim/J) and unaffected wild type (WT) littermates were collected at day of birth (P0) and postnatal day 14 (P14). Tissues were fixed and stained with 1.25% buffered Lugol’s solution, then micro-CT scanned with a reconstruction of 0.02 mm3 voxels. 3D Slicer software was used to isolate and measure the volumes of the superficial masseter, deep masseter, and temporalis muscles. Muscle volumes were compared between genotypes using a Mann-Whitney U test. Results: At birth, no significant differences were observed in body mass or muscle volumes between OIM and WT mice. A trend was observed for OIM mice to have lower superficial masseter volumes compared to WT mice at P0, but this difference was not significant. At P14, OIM mice have significantly lower body weights (p=0.002). Data collection is ongoing for volumetric muscle data from the P14 stage. After birth, body masses diverge rapidly between OIM and WT mice. These growth curves suggest poor feeding performance during the suckling stage in OIM mice. Although masticatory muscle volumes (similar to body mass) start out similar between genotypes at birth, a trend for decreased superficial masseter volume in OIM mice suggests feeding musculature will also lag behind unaffected mice during early postnatal growth. Conclusion: The production of strain above an osteogenic threshold by feeding musculature is critical to typical craniofacial growth during early life. Weaker masticatory muscles may produce lower (yet still osteogenic) levels of strain, contributing to the midfacial hypoplasia seen in OIM mice. A better understanding of muscle development during this critical growth period will provide insight on feeding disorders seen in OI, and the development of the craniofacial phenotype in pediatric patients with OI.Item Microstructure & Macrostructure Interrelationship in the Growing Hard Palate(2022) Miller, Courtney; Steele, Ashley T.; Organ, Jason; Menegaz, Rachel A.Bone strength is the result of microstructure (bone material properties) and macrostructure (bone size and shape), and deficiencies in either can produce skeletal fragilities with an increased likelihood of injury. The micro- and macro-architecture work together during re/modeling, ensuring that the skeleton is resistant to repetitive loading and preventing fracture. Bone is particularly responsive to loading during the rapid growth occurring in early life. Within the craniofacial complex, the hard palate is a unique structure that undergoes continuous loading due to continuous resting/active tongue pressure and bite forces. Because of this, palatal growth is theorized to drive midfacial growth. The aim of this study is to investigate longitudinal changes in the palate when the relationship between microstructure and macrostructure is perturbed. Here we use the OIM mouse (B6C3FE a/a-Col1a2OIM/J), a strain with a mutation to the structural protein type I collagen, resulting in increased bone fragility and improper biomineralization. We hypothesize that OIM and unaffected wild-type (WT) mice will be most similar in bone micro- and macro-structure in regions of the palate that experience high loading and therefore high rates of bone modeling/remodeling. Mice were micro-CT scanned at two timepoints: week 4 (juvenile) and week 16 (adult). BMD was collected at 3 regions (anterior, mid, and posterior palate). Mann-Whitney U tests were used to compare BMD between genotypes. 28 fixed and 10 sliding landmarks were placed across the palate. Geometric morphologic analyses were conducted to determine variation between genotypes in overall shape and curvature throughout growth. Average BMD decreased along an anteroposterior gradient for all mice. While juvenile OIM mice had lower palate BMD than WT mice at all regions, adult OIM mice had significantly lower BMD at only the anterior and posterior regions but not the midpalate. Procrustes ANOVA revealed significant differences in palatal shape between the genotypes at both the juvenile (p=0.001) and adult (p=0.0015) stages. Principal component analyses revealed that juvenile OIM mice had a shorter anterior palate and broader palate compared to WT mice, while adult OIM mice had a shorter posterior palate than WT mice. OI mice had flatter palatal arches in the coronal plane as juveniles, and in the midsagittal plane as juveniles and adults. Results show significant differences in palatal BMD and morphology between genotypes throughout growth, with fewer significant differences at the adult stage. Increased loading of the anterior palate during incisive gnawing may result in the convergence of bone macrostructure among adult mice, while increased strain at the midpalatal suture may necessitate increased BMD even in the presence of a biomineralization defect. Further research is needed to understand the functional significance of the anteroposterior palatal gradient and its relation to soft tissue attachments, particularly given the difference in oral behaviors between juveniles and adults.Item Neurocranial Growth in the OIM Mouse Model of Osteogenesis Imperfecta(2022) Husain, Tooba S.; Miller, Courtney; Steele, Ashley T.; Gonzales, Lauren; Handler, Emma; Organ, Jason; Menegaz, Rachel A.Osteogenesis imperfecta (OI) is a disorder of type I collagen characterized by abnormal bone formation and weakened bone architecture. Human patients with OI have larger cranial vaults (macrocephaly), altered cranial base morphology including basilar invagination and platybasia (skull base flattening), and midfacial underdevelopment. The neurocranial changes may affect both underlying nervous tissue and growth patterns of the facial skeleton. However, we still do not fully understand how and when these divergent morphologies occur. The aims of this study are: (1) to investigate the integrated development of the skull and the brain in amouse model of OI; and (2) to identify the developmental trajectories of these structures to facilitate future therapeutic interventions. We hypothesize that compared to unaffected mice, mice with OI will have decreased brain volumes due to an overall reduction in cranial size and decreased cranial base angles (CBA) due to platybasia. To test these hypotheses, we used the osteogenesis imperfecta murine (OIM or B6C3FE a/a-Col1a2/J), a model for the severe type III OI in humans, and unaffected wild-type (WT) littermates. Mice were imaged using in vivo micro-computed tomography (micro-CT) at the juvenile (week 4; 10 OIM/14 WT) and adult (week 16; 9OIM/11 WT) stages. All measurements were taken in 3D Slicer software. 82 cranial landmarks were used to calculate centroid size, an estimate of overall head size. The segmentations tool was used to create virtual endocasts as a proxy for brain volume. The angle tool was used to measure CBA in the midsagittal plane using threelandmarks: foramen cecum, midsphenoidal synchondrosis, and basion. Mann-Whitney U tests were used to compare centroid sizes, brain volumes, and CBA between the genotypes. Both juvenile (p=0.008) and adult (p=0.003) OIM mice were found to have absolutely smaller brains than WT mice. However, OIM mice also have significantly smaller cranial centroid sizes compared to WT mice (p=0.003, p< 0.001). When scaled to cranial size, juvenile mice had relatively larger brain volumes (p=0.016) butadult OIM relative brain volumes were not significantly different from WT. No significant difference was seen in CBA at the juvenile (p=0.065) or adult (p=0.171) stages, however a trend was observed for decreased CBA at the adult stage. These results suggest that neurocranial dysmorphologies in OI may be more severe at earlier stages of postnatal development. Previous analyses of these mice have documented relative skeletal macrocephaly in both juvenile and adults, however here we document an increase in relative endocranial volume only at the juvenile stage. A reduction in CBA during growth, possibly due to platybasia, may underlie this decoupling between external and internal cranial morphology. Future work will investigate the effect of CBA on facial growth and midfacial underdevelopment in these mice. A better understanding of the integration and growth trajectory of the neurocranium is foundational for formulating treatments to manage basicranial instabilities in patients with OI. Support or Funding Information Funding was provided by an Indiana University Collaborative Research Grant, Ralph W. and Grace Showalter Trust, and a UNTHSC Physiology & Anatomy SEED Grant.Item Postweaning Craniofacial Growth in the OIM Mouse Model of Osteogenesis Imperfecta(2022) Steele, Ashley T.; Mitchell, D. Rex; Organ, Jason; Menegaz, Rachel A.Osteogenesis imperfecta (OI) type III is a severe genetic disorder of type I collagen (Col1) resulting in bone fragility, reduced stature, and impaired craniofacial growth resulting in midface hypoplasia, dental malocclusions, and macrocephaly. While the adult OI murine (OIM) mouse model exhibits craniofacial phenotypes similar to patients with OI, little is known about the developmental trajectories of these phenotypes. To investigate the mechanisms by which Col1 mutations alter postnatal craniofacial growth, we analyzed the phenotype of the OIM mouse from the age of weaning until adulthood (skeletal maturity). OIM and wild-type (WT) littermates were scanned in-vivo with a Skyscan 1176 micro-CT system at 4 weeks (weaning) and 16 weeks (adulthood). 3D landmarks were collected using 3D Slicer software. Centroid size (a proxy for craniomandibular size) was compared using Mann-Whitney U tests. Morphological analysis for shape variation, including Generalized Procrustes analysis (GPA) and principal component analyses (PCA), were performed using the "geomorph" package in R. Procrustes ANOVAs were used to test for significant differences in craniomandibular shape between the genotypes. Morphological disparity was estimated as the Procrustes variance and statistically compared using the morphol.disparity function in R. Craniomandibular centroid sizes were significantly smaller in the OIM mice than the WT mice at both weeks 4 and 16 (p< 0.010). When the effects of size were accounted for by the GPA, significant shape differences were present (p< 0.002) throughout growth. For their size, both juvenile and adult OIM mice had shortened midfaces and increased cranial vault dimensions (relative macrocephaly) compared to WT littermates. Morphological differences were seen around the rostrum, temporal crests, and zygomatic arches. Marked vertical expansion of the neurocranium at the sagittal and coronal sutures presented with a concomitant basicranial shortening in the juvenile OIM mice only. Compared to WT, both juvenile and adult OIM mice had mandibles that were longitudinally shortened and mediolaterally wide. Morphological differences were seen around the incisal ramus and angular process at both ages, and in the coronoid process of adult OIM mice. Significantly different (p=0.001) morphological disparity between the genotypes existed only at the juvenile stage, demonstrating an overall decrease in variance throughout postnatal development period. This suggests that mice with greater variances in cranial shape experienced higher rates of attrition. This is potentially related to increased craniofacial fracture rates seen in the OIM mice. These results suggest that while the OIM craniofacial phenotype differs significantly from the WT throughout postweaning growth, functional constraints (such as feeding performance) may limit the degree of potential phenotypic divergence. Furthermore, while previous work in human patients with OI has linked the development of macrocephaly to basicranial shortening during early skull development, our results provide limited support for this hypothesis due to the absence of basicranial changes in adult OIM mice. Future investigations will examine perinatal and pre-weaning growth in the OIM mouse model to better understand the development of these craniofacial dysmorphologies, and to identify optimal growth windows during which interventions might recover bone quality and growth trajectories in patients with OI.Item Pre-weaning craniofacial development in mice with Osteogenesis Imperfecta(2024-03-21) Miller, Courtney; Emmanuel, Tanusha; Gonzales, Lauren; Handler, Emma; Organ, Jason; Menegaz, Rachel A.The craniofacial region plays a pivotal role in various physiological functions, including mastication, speech, and respiration. Early life behaviors have a profound role in shaping adult structure and function. In the early stages of life, all mammals undergo the transition from suckling to mastication, a period coinciding with rapid cranial biomineralization. Osteogenesis imperfecta (OI), a genetic disorder that impacts the production of type I collagen, disrupts biomineralization, leading to craniofacial growth differences affecting overall quality of life. This study investigates the preweaning craniofacial growth trajectory in mice OI (the OIM mouse) compared to unaffected wild type (WT mice). We hypothesize that mice with OI will exhibit smaller overall size and greater craniofacial variation than WT mice due to the abnormal collagen synthesis during skull development. Micro-CT based geometric morphometric analyses of the OIM mouse model (B6C3Fe a/a-Col1a2oim/J) were used to compare craniofacial size and shape differences at birth (P0; n=27 OIM / 20 WT) and postnatal days 7 (P7; n=21/21) and 14 (P14; n=16/20). The SlicerMorph package for 3D Slicer software was used to generate landmark point clouds for the cranium and mandible. Dimension ratios were calculated as width/length for the crania. Principal component analysis with Procrustes ANOVA were used to examine differences between genotypes at each time point, and a canonical variate analysis (CVA) used to identify shape features that maximize the distinction between genotypes across all time points. Results reveal the development of significant differences in both shape and size between the genotypes following birth. At birth, size and shape are similar between genotypes. However, by P7 and P14, OIM mice are significantly (p<0.05) smaller and display pronounced shape changes (p<0.001) characterized by larger neurocranium and shorter viscerocranium. Additionally, OIM mice have significant mandibular alterations by P7 (p<0.001) - shorter ramus, more posterior position of the coronoid, and shorter and wider dental arcade. All of these changes align with the suckling developmental stage, suggesting changes in the ratio of growth between the neurocranium and the viscerocranium during early life. Widening the neurocranium while shortening the viscerocranium during this critical developmental stage alters the masticatory muscle line of action, consequently, influences the health of individuals with OI. These findings underscore the suckling stage’s significance in shaping the foundational structures for later life, providing insights into OI craniofacial development, and suggest potential benefits to directing interventions toward an earlier time point for more effective treatment of OI.