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Antidepressants, specifically Selective Serotonin Re-uptake Inhibitors (SSRIs), that alter serotonin metabolism are currently the most commonly prescribed drugs for the treatment of depression. There is some evidence to suggest these drugs contribute to birth defects. As jaw development is often altered in craniofacial birth defects, the purpose of this study was to interrogate the effects of in utero SSRI exposure in a preclinical model of mandible development.Wild type C57BL6 mice were used to produce litters that were exposed in utero to an SSRI, Citalopram (500 μg/day). Murine mandibles from P15 pups were analyzed for a change in shape and composition.Analysis indicated an overall shape change with total mandibular length and ramus height being shorter in exposed pups as compared to controls. Histomorphometric analysis revealed that first molar length was longer in exposed pups while third molar length was shorter in exposed as compared to control. Histological investigation of molars and surrounding periodontium revealed no change in collagen content of the molar in exposed pups, some alteration in collagen composition in the periodontium, increased alkaline phosphatase in molars and periodontium, and decreased mesenchymal cell marker presence in exposed mandibles.The results of this study reveal SSRI exposure may interrupt mandible growth as well as overall dental maturation in a model of development giving insight into the expectation that children exposed to SSRIs may require orthodontic intervention.

Several teaching resources are used to enhance the learning of anatomy. The purpose of this study was to examine the preference of medical students on the use of various resources to learn anatomy and their link to 12 learning outcomes. A selected response item questionnaire was administered that asked students to rank six laboratory teaching resources from most to least preferred, and rate how useful these six resources were towards achieving 12 learning outcomes. These learning outcomes covered many of the learning domains such as demonstrating an understanding of anatomy, visualizing structures, appreciating clinical correlations, and understanding anatomical variations. Medical students ranked cadaveric prosections paired with an active learning clinical tutorial as the highest rank and most useful resource for learning anatomy, followed by dissection videos, electronic resources, and printed material, followed by plastinated specimens and plastic models. Overall, cadaveric prosections were also rated as the most helpful teaching resource in achieving various learning outcomes. In conclusion, anatomy teachers should provide prosections coupled with clinical tutorials as well as electronic resources as students prefer these and think they help them learn anatomy. Future studies will investigate the impact of using these resources on students’ performance. Supplementary Information The online version contains supplementary material available at 10.1007/s40670-021-01436-2.

Introduction Alveolar bone grafting aims to restore bony continuity of the alveolus and provide optimal periodontal support for teeth adjacent to the cleft. We created a survey of cleft surgeons to assess the current standard of care regarding this procedure. Methods A multiple choice survey was implemented using Qualtrics software and emailed to a list of 708 surgeons from the ACPA membership directory. Correlation between various provider factors and treatment practices was assessed with Fisher's exact test and likelihood ratio tests. Results The response rate was 17.5%. Eighty-seven percent of providers preferred to perform grafts prior to secondary canine eruption while 10% favored before central incisor eruption. Eighty-one percent favored palatal expansion prior to bone grafting. Wide variability existed regarding the time to initiate postoperative orthodontics; 43% waited 4 to 6 months. Sixty-four percent of surgeons now utilize cone beam CT to assess graft take. The majority of respondents utilized cancellous bone autograft (92%) from the anterior iliac crest (97%) as graft material. Seventy percent used three or more modalities for post-operative pain control management. Early career surgeons (0–5 years) appeared more likely to use non-autologous materials ( p Conclusion Alveolar bone grafting prior to secondary canine eruption remains the most common strategy but other protocols are employed. Surgeons utilize multiple modalities for radiographic evaluation and most often use autologous cancellous bone as the primary grafting material. There is no true consensus on the perioperative timing and sequencing of orthodontic manipulation while principles of multimodal perioperative pain control appear widely accepted.

Evidence suggests alterations of thyroid hormone levels can disrupt normal bone development. Most data suggest the major targets of thyroid hormones to be the Htra1/Igf1 pathway. Recent discovery by our group suggests involvement of targets WNT pathway, specifically overexpression of antagonist Sfrp4 in the presence of exogenous thyroid hormone.Here we aimed to model these interactions in vitro using primary and isotype cell lines to determine if thyroid hormone drives increased Sfrp4 expression in cells relevant to craniofacial development. Transcriptional profiling, bioinformatics interrogation, protein and function analyses were used.Affymetrix transcriptional profiling found Sfrp4 overexpression in primary cranial suture derived cells stimulated with thyroxine in vitro. Interrogation of the SFRP4 promoter identified multiple putative binding sites for thyroid hormone receptors. Experimentation with several cell lines demonstrated that thyroxine treatment induced Sfrp4 expression, demonstrating that Sfrp4 mRNA and protein levels are not tightly coupled. Transcriptional and protein analyses demonstrate thyroid hormone receptor binding to the proximal promoter of the target gene Sfrp4 in murine calvarial pre-osteoblasts. Functional analysis after thyroxine hormone stimulation for alkaline phosphatase activity shows that pre-osteoblasts increase alkaline phosphatase activity compared to other cell types, suggesting cell type susceptibility. Finally, we added recombinant SFRP4 to pre-osteoblasts in combination with thyroxine treatment and observed a significant decrease in alkaline phosphatase positivity.Taken together, these results suggest SFRP4 may be a key regulatory molecule that prevents thyroxine driven osteogenesis. These data corroborate clinical findings indicating a potential for SFRP4 as a diagnostic or therapeutic target for hyperostotic craniofacial disorders.

BACKGROUND The submucous cleft palate can be overt or occult and may require surgical repair. The double-opposing Z-plasty (Furlow repair) is the authors' center's preferred approach. This study evaluated complication rates, differences in outcome between overt and occult types, and patient factors associated with surgical failure. METHODS This retrospective study reviewed documentation on all patients who underwent Furlow Z-plasty for submucous cleft palate at a single center between 2004 and 2018. Speech pathology was quantified using the Pittsburgh Weighted Speech Score. RESULTS A total of 351 patients were included (125 overt and 226 occult cases). Furlow Z-plasty was successful (postoperative Pittsburgh Weighted Speech Score

Timing of craniofacial suture fusion is important for the determination of demographics and primate ontogeny. There has been much work concerning the timing of fusion of calvarial sutures over the last century, but little comprehensive work focusing on facial sutures. Here we assess the relationships of facial suture fusion across ontogeny among select catarrhines. Fusion timing patterns for 5 facial sutures were examined in 1,599 crania of Homo, Pan, Gorilla, Pongo, Hylobatidae, Papio, and Macaca. Calvarial volume (early ontogeny) and dental eruption (late ontogeny) were used as indicators of stage of development. General linear models, test for homogeneity of slopes, and ANOVA were used to determine differences in timing of fusion by taxon. For calvarial volume, taxonomic groups segregated by regression slopes, with models for Homo indicating sutural fusion throughout ontogeny, Pongo, Macaca, and Papio representing earlier and more complete suture fusion, and Pan, Gorilla, and Hylobatidae indicating very early facial suture fusion. Similar patterns are observed when dental eruption is used for developmental staging. Only Gorilla and Hylobatidae are observed to, generally, fuse all facial suture sites in adulthood. Finally, Homo appears to be unique in its delay and patency of sutures into late ontogeny. The taxonomic patterns of facial suture closure identified in this study likely reflect important evolutionary shifts in facial growth and development in catarrhines.

Introduction: Oronasal fistulae following palatoplasty may affect patients’ quality of life by impacting their ability to eat, speak, and maintain oral hygiene. We aimed to quantify the impact of previous oronasal fistula repair on patients’ quality of life using patient-reported outcome psychometric tools. Methods: A cross-sectional study of 8- to 9-year-old patients with cleft palate and/or lip was completed. Patients who had a cleft team clinic between September 2018 and August 2019 were recruited. Participants were divided into 2 groups (no fistula, prior fistula repair). Differences in the individual CLEFT-Q and Child Oral Health Impact Profile-Short Form 19 (COHIP-SF 19) Oral Health scores between the 2 groups were evaluated using a multivariate analysis controlling for Veau classification and syndromic diagnosis. Results: Sixty patients with a history of cleft palate were included. Forty-two (70%) patients had an associated cleft lip. Thirty-two (53.3%) patients had no history of fistula and 28 (46.7%) patients had undergone a fistula repair. CLEFT-Q Dental, Jaw, and Speech Function were all higher in patients without a history of a fistula repair; however, none of these differences were statistically significant. The COHIP-SF 19 Oral Health score demonstrated a significantly lower score in the fistula group, indicating poorer oral health ( P = .05). Conclusions: One would expect that successful repair of a fistula would result in improved function and patient satisfaction, but the consistent trend toward lower CLEFT-Q scores and significantly increased COHIP-SF 19 Oral Health scores in our study group suggests that residual effects linger and that the morbidity of a fistula may not be completely treated with a secondary correction.

Living primates show a complex trend in reduction of nasal cavity spaces and structures due to moderate to severe constraint on interorbital breadth. Here we describe the ontogeny of the posterior end of the primate cartilaginous nasal capsule, the thimble shaped posterior nasal cupula (PNC), which surrounds the hind end of the olfactory region. We used a histologically sectioned sample of strepsirrhine primates and two non-primates (Tupaia belangeri, Rousettus leschenaulti), and histochemical and immunohistochemical methods to study the PNC in a perinatal sample. At birth, most strepsirrhines possess only fragments of PNC, and these lack a perichondrium. Fetal specimens of several species reveal a more complete PNC, but the cartilage exhibits uneven or weak reactivity to type II collagen antibodies. Moreover, there is relatively less matrix than in the septal cartilage, resulting in clustering of chondrocytes, some of which are in direct contact with adjacent connective tissues. In one primate (Varecia spp.) and both non-primates, the PNC has a perichondrium at birth. In older, infant Varecia and Rousettus, the perichondrium of the PNC is absent, and PNC fragmentation at its posterior pole has occurred in the former. Loss of the perichondrium for the PNC appears to precede resorption of the posterior end of the nasal capsule. These results suggest that the consolidation of the basicranial and facial skeletons happens ontogenetically earlier in primates than other mammals. We hypothesize that early loss of cartilage at the sphenoethmoidal articulation limits chondral mechanisms for nasal complexity, such as interstitial expansion or endochondral ossification.

Cranial synchondroses are cartilaginous joints between basicranial bones or between basicranial bones and septal cartilage, and have been implicated as having a potential active role in determining craniofacial form. However, few studies have examined them histologically. Using histological and immunohistochemical methods, we examined all basicranial joints in serial sagittal sections of newborn heads from nine genera of primates (five anthropoids, four strepsirrhines). Each synchondrosis was examined for characteristics of active growth centers, including a zonal distribution of proliferating and hypertrophic chondrocytes, as well as corresponding changes in matrix characteristics (i.e., density and organization of Type II collagen). Results reveal three midline and three bilateral synchondroses possess attributes of active growth centers in all species (sphenooccipital, intrasphenoidal, presphenoseptal). One midline synchondrosis (ethmoseptal) and one bilateral synchondrosis (alibasisphenoidal synchondrosis [ABS]) are active growth centers in some but not all newborn primates. ABS is oriented more anteriorly in monkeys compared to lemurs and bushbabies. The sphenoethmoidal synchondrosis (SES) varies at birth: in monkeys, it is a suture-like joint (i.e., fibrous tissue between the two bones); however, in strepsirrhines, the jugum sphenoidale is ossified while the mesethmoid remains cartilaginous. No species possesses an SES that has the organization of a growth plate. Overall, our findings demonstrate that only four midline synchondroses have the potential to actively affect basicranial angularity and facial orientation during the perinatal timeframe, while the SES of anthropoids essentially transitions toward a "suture-like" function, permitting passive growth postnatally. Loss of cartilaginous continuity at SES and reorientation of ABS distinguish monkeys from strepsirrhines.

Introduction:Surgical outcomes for patients with complete cleft lips are not as ideal as those achieved for milder phenotypes. We hypothesized that in addition to the greater width of the cleft, patients with complete cleft lip and palate exhibit a greater degree of hypoplasia and asymmetry.Methods:Stereophotographs of 14 infants with unrepaired unilateral complete and 14 with incomplete cleft lips were measured using Vectra imaging software (Canfield Imaging). Unpaired t tests were used to compare measured asymmetry between groups. Measurements included nasion to endocathion, sn-sbal, subnasale to alare (sn-al), subnasale to crista philtra, subalare to crista philtra (sbal-cphi), chelion to crista philtra, lateral lip element fullness, medial lip element fullness (mef), and non-cleft lip fullness. Duplicate measurements allowed Pearson correlation to be used to determine intra-rater reliability. Statistical significance was set at P < .05.Results:Degree of asymmetry of the nasal base, sn-al, and sn-sbal was significantly greater for patients with complete clefts ( P = .0001, P = .0001). Hypoplasia of the lateral lip element was seen when comparing lateral and mef ( P = .04, P = .004) and lateral lip height (sbal-cphi’’; P = .002). The degree of cupid’s bow asymmetry did not differ between groups ( P = .23). Intrarater reliability was high for all facial measures, ranging from 0.70 to 0.99.Conclusions:More severe, complete cleft lips demonstrate statistically significant greater asymmetry in surgically relevant dimensions. There was greater width of the nasal base. Vertical asymmetry of cupid’s bow was unaffected by cleft severity, but that asymmetry was greater in patients with complete clefts due to hypoplasia of the lateral lip element. This may contribute to the less favorable results in these patients.

Once thought to have revolutionized therapeutic intervention in surgery, Recombinant Human Bone Morphogenic Protein 2 (rhBMP2) is now in its second decade of sustained controversy over the side effects associated with its use. Side effects associated with clinical use of rhBMP2 (Infuse, Medtronic Inc) include a marked inflammatory response, pain, therapeutic failures, ectopic bone, tissue degradation, and death. What is missing, despite the depth of literature on the subject, is a direct interrogation of rhBMP2, specifically for inflammation. Here we set out to determine if rhBMP2 alters traditional macrophage markers associated with pro-inflammatory responses, and pro-reparative responses to injury. Based on our previous work, we hypothesized there would be no direct effect of the peptide on macrophage polarization. Here we utilized commercially available murine macrophages, RAW 264.7, and treated these cells with rhBMP2 in standard growth media or macrophage polarizing media (M1 and M2) at several doses of the peptide. Our readouts were cell viability, apoptosis, gene expression of M1 and M2 markers, and ELISA for M1 marker iNOS, and M2 marker Arg1. Our data give very little evidence to support an alteration in macrophage phenotype by rhBMP2 alone, or alteration of the phenotype when cultured in enriched M1 or M2 media. These results further suggest that other factors associated with the clinical use of Infuse, likely supraphysiological rhBMP2 doses and off label usage, are more likely the culprit for poor outcomes. This further reinforces the utility of rhBMP2 and other peptides in tissue engineering therapies when conditions are tightly controlled.

The pressing need for SARS-CoV-2 controls has led to a reassessment of strategies to identify and develop natural product inhibitors of zoonotic, highly virulent, and rapidly emerging viruses. This review article addresses how contemporary approaches involving computational chemistry, natural product (NP) and protein databases, and mass spectrometry (MS) derived target-ligand interaction analysis can be utilized to expedite the interrogation of NP structures while minimizing the time and expense of extraction, purification, and screening in BioSafety Laboratories (BSL)3 laboratories. The unparalleled structural diversity and complexity of NPs is an extraordinary resource for the discovery and development of broad-spectrum inhibitors of viral genera, including Betacoronavirus, which contains MERS, SARS, SARS-CoV-2, and the common cold. There are two key technological advances that have created unique opportunities for the identification of NP prototypes with greater efficiency: (1) the application of structural databases for NPs and target proteins and (2) the application of modern MS techniques to assess protein-ligand interactions directly from NP extracts. These approaches, developed over years, now allow for the identification and isolation of unique antiviral ligands without the immediate need for BSL3 facilities. Overall, the goal is to improve the success rate of NP-based screening by focusing resources on source materials with a higher likelihood of success, while simultaneously providing opportunities for the discovery of novel ligands to selectively target proteins involved in viral infection.

The human temporomandibular joint (TMJ) lateral capsule ligament (LCL) complex is debated as a fibrous capsule with distinct ligaments or ligamentous thickening, necessitating further evaluation of the complex and its role in TMJ anatomy and mechanics. This study explores the ultrastructural arrangement, biomechanical tensile properties, and biochemical composition of the human LCL complex including region-specific differences to explore the presence of a distinct temporomandibular ligament and sex-specific differences to inform evaluations of potential etiological mechanisms. LCL complex ultrastructural arrangement, biomechanical properties, and biochemical composition were determined using cadaveric samples. Statistical modeling assessed sex- and region-specific effects on LCL complex tissue properties. Collagen fiber coherency, collagen fiber bundle size, and elastin fiber count did not differ between sexes, but females trended higher in elastin fiber count. LCL complex water and sGAG content did not differ between sexes or regions, but collagen content was higher in the anterior region (311.0 ± 185.6 μg/mg) compared to the posterior region (221.0 ± 124.9 μg/mg) (p = 0.045) across sexes and in males (339.6 ± 170.6 μg/mg) compared to females (204.5 ± 130.7 μg/mg) (p = 0.006) across regions. Anterior failure stress (1.1 ± 0.7 MPa) was larger than posterior failure stress (0.6 ± 0.4 MPa) (p = 0.024). Regional differences confirm the presence of a mechanically and compositionally distinct temporomandibular ligament. Baseline sex-specific differences are critical for etiological investigations of sex disparities in TMJ disorders. These results have important biomechanical and clinical ramifications, providing critical baseline tissue material properties, informing the development of TMJ musculoskeletal models, and identifying new areas for etiologic investigations for temporomandibular disorders.

BACKGROUND Large calvarial defects represent a major reconstructive challenge, as they do not heal spontaneously. Infection causes inflammation and scarring, further reducing the healing capacity of the calvaria. Bone morphogenetic protein-2 (BMP2) has been shown to stimulate osteogenesis but has significant side effects in high doses. BMP2 has not been tested in combination with antiinflammatory cytokines such as interleukin-10. METHODS Sixteen New Zealand White rabbits underwent 15 × 15-mm flap calvarectomies. The flap was incubated in Staphylococcus aureus and replaced, and infection and scarring were allowed to develop. The flap was subsequently removed and the wound debrided. A 15 × 15-mm square of acellular dermal matrix biopatterned with low-dose BMP2, interleukin-10, or a combination was implanted. Computed tomographic scans were taken over 42 days. Rabbits were then killed and histology was performed. RESULTS Defects treated with BMP2 showed significantly (p < 0.05) greater osseous regeneration than untreated controls. Interleukin-10 did not significantly augment the healing achieved with BMP2, and interleukin-10 alone did not significantly increase healing compared with controls. Histology showed evidence of bone formation in defects treated with BMP2. Untreated controls and defects treated with interleukin-10 alone showed only fibrous tissue in the defect site. CONCLUSIONS Low-dose BMP2 delivered directly to the scarred calvarial defect augments bony healing. Interleukin-10 at the dose applied did not significantly augment healing alone or in combination with BMP2. Healing had not finished at 42 days and analysis at later time points or the use of higher doses of BMP2 may yield greater healing.

Resources detailing the scope, details, and duration for teaching and learning surgical model development in research are poorly described. Situated learning and instructional scaffolding are useful skill-building tools. Herein, we discuss educational theory in the context of a training paradigm for surgical researchers, using our experience with a nonunion femoral fracture model as an example. Stages of learning include cognitive, associative, and autonomous stages. In surgical training, the cognitive stage involves the acquisition of basic knowledge, including anatomy, surgical approach, instrumentation, and suturing, which can be taught by using books, videos, skeletons, and cadavers. To these basic skills, the associative stage adds advanced techniques—including anesthesia, asepsis, hemostasis, and the full surgical procedure—through mentored nonsurvival surgical experiences. After a mentor has assured competence, trainees perform supervised and then independent survival surgeries to complete the autonomous stage. Through these stages, instructional scaffolding is applied in the context of a situated learning environment in which trainees learn in a layered approach through their own experiences. Thus, the proposed training paradigm is structured to teach trainees how to think and act as surgeons so they can adapt and grow, rather than only to ensure technical competency in a specific model. Development and mastery of complex surgical models may require as long as 6 mo to achieve optimal outcomes, depending on the preexisting skill of the research surgeons, technical difficulty, and the stage of model evolution.

Large bone injuries, defects, and chronic wounds present a major problem for medicine. Several therapeutic strategies are used clinically to precipitate bone including a combination therapy delivering osteoinductive bone morphogenetic protein 2 (rhBMP-2) via an osteoconductive scaffold (absorbable collagen sponge, ACS) (i.e. INFUSE). Adverse side effects reportedly associated with rhBMP2 administration include rampant inflammation and clinical failures. Although acute inflammation is necessary for proper healing in bone, inflammatory cascade dysregulation can result in sustained tissue damage and poor healing. We hypothesized that a subclinical dose of rhBMP2 modeled in the murine calvarial defect would not precipitate alterations to inflammatory markers during acute phases of bone wound healing. We utilized the 5mm critical size calvarial defect in C57BL6 wild-type mice which were subsequently treated with ACS and a subclinical dose of rhBMP2 shown to be optimal for healing. Three and seven-day post-operative time points were used to assess the role that rhBMP-2 plays in modulating inflammation versus ACS alone by cytokine array and histological interrogation. Data revealed that rhBMP-2 delivery resulted in substantial modulation of several markers associated with inflammation, most of which decreased to levels similar to control by the seven day time point. Additionally, while rhBMP-2 administration increased macrophage response, this peptide had little noticeable effect on traditional markers of macrophage polarization (M1-iNOS, M2-Arg1). These results suggest that rhBMP-2 delivered at a lower dose does not precipitate rampant inflammation. Thus, an assessment of dosing for rhBMP-2 therapies may lead to better healing outcomes and less surgical failure.

Background Bone is a highly vascularized and resilient organ with innate healing abilities, however some bone injuries overwhelm these attributes and require intervention, such as bone tissue engineering strategies. Combining biomaterials and growth factors, such as bone morphogenetic protein 2 (BMP2), is one of the most commonly used tissue engineering strategies. However, use of BMP2 has been correlated with negative clinical outcomes including aberrant inflammatory response, poor quality bone, and ectopic bone. Methods In the present study, a novel poly-n-acetyl glucosamine (pGlcNAc, trade name Talymed) scaffold was utilized in addition to the commonly used acellular collagen sponge (ACS) BMP2 delivery system in a murine calvarial defect model to investigate whether the innate properties of Talymed can reduce the noted negative bone phenotypes associated with BMP2 treatment. Results Comparison of murine calvarial defect healing between ACS with and without Talymed revealed that there was no measurable healing benefit for the combined treatment. Healing was most effective utilizing the traditional acellular collagen sponge with a reduced dose of BMP2. Conclusions The results of this investigation lead to the conclusion that excessive dosing of BMP2 may be responsible for the negative clinical side effects observed with this bone tissue engineering strategy. Rather than augmenting the currently used ACS BMP2 bone wound healing strategy with an additional anti-inflammatory scaffold, reducing the dose of BMP2 used in the traditional delivery system results in optimal healing without the published negative side effects of BMP2 treatment.

Background aims Previous studies identified a circulating human osteoblastic population that expressed osteocalcin (OCN), increased following fracture and pubertal growth, and formed mineralized colonies in vitro and bone in vivo. A subpopulation expressed CD34, a hematopoietic/endothelial marker. These findings led to our hypothesis that hematopoietic-derived CD34+OCN+ cells exist in the circulation of mice and are modulated after fracture. Methods Flow cytometry was used to identify CD34+OCN+ cells in male B6.SJL-PtprcaPepcb/BoyJ and Vav-Cre/mTmG (VavR) mice. Non-stabilized tibial fractures were created by three-point bend. Fractures were longitudinally imaged by micro-computed tomography, and immunofluorescent staining was used to evaluate CD34+OCN+ cells within fracture callus. AMD3100 (10 mg/kg) was injected subcutaneously for 3 days and the CD34+OCN+ population was evaluated by flow cytometry. Results Circulating CD34+OCN+ cells were identified in mice and confirmed to be of hematopoietic origin (CD45+; Vav1+) using two mouse models. Both circulating and bone marrow-derived CD34+OCN+ cells peaked three weeks post-non-stabilized tibial fracture, suggesting association with cartilage callus transition to bone and early mineralization. Co-expression of CD34 and OCN in the fracture callus at two weeks post-fracture was observed. By three weeks, there was 2.1-fold increase in number of CD34+OCN+ cells, and these were observed throughout the fracture callus. AMD3100 altered CD34+OCN+ cell levels in peripheral blood and bone marrow. Discussion Together, these data demonstrate a murine CD34+OCN+ circulating population that may be directly involved in fracture repair. Future studies will molecularly characterize CD34+OCN+ cells, determine mechanisms regulating their contribution, and examine if their number correlates with improved fracture healing outcomes.

INTRODUCTION Direct anthropometric and three-dimensional (3D) photogrammetry measurements have been used extensively in cleft/craniofacial surgery to assess morphological changes and surgical outcomes. Craniofacial procedures alter the sagittal projection of periorbital bony prominences. Mulliken described a method of measuring their projection relative to the corneal plane but is impractical in clinical practice. Three-dimensional photogrammetry may offer a solution; however, the cornea is not visualized on this. The authors propose to develop new normative measurements of facial projection relative to the pupil. METHODS Five 3D photographs were taken of 5 individuals using Vectra M5 camera. Facial projection measurements were taken of the sagittal projection of the bilateral periorbital landmarks and nasal radix relative to the pupil using Mirror 3D analysis. Standard deviations (SD) were determined for each subject and laterality. Chi-square tests confirmed all SD

The sphenoid bone articulates with multiple basicranial, facial, and calvarial bones, and in humans its synchondroses are known to contribute to elongation of the skull base and possibly to cranial base angulation. Its early development (embryological, early fetal) has frequently been studied in a comparative context. However, the perinatal events in morphogenesis of the sphenoid have been explored in very few primates. Using a cross-sectional age sample of non-human primates (n=39; 22 platyrrhines; 17 strepsirrhines), we used microcomputed tomographic (µCT) and histological methods to track age changes in the sphenoid bone. In the midline, the sphenoid expands its dimensions at three growth centers, including the sphenooccipital, intrasphenoidal (ISS) and presphenoseptal (PSept) synchondroses. Bilaterally, the alisphenoid is enlarged via appositional bone growth that radiates outward from cartilaginous parts of the alisphenoid during midfetal stages. The alisphenoid remains connected to the basitrabecular process of the basisphenoid via the alibasisphenoidal synchondrosis (ABS). Reactivity to proliferating cell-nuclear antigen is observed in all synchondroses, indicating active growth perinatally. Between mid-fetal and birth ages in Saguinus geoffroyi, all synchondroses decrease in the breadth of proliferating columns of chondrocytes. In most primates, the ABS is greatly diminished by birth, and is likely the earliest to fuse, although at least some cartilage may remain by at least one-month of age. Unlike humans, no non-human primate in our sample exhibits perinatal fusion of ISS. A dichotomy among primates is the orientation of the ABS, which is more rostrally directed in platyrrhines. Based on fetal Saguinus geoffroyi specimens, the ABS was initially oriented within a horizontal plane, and redirects inferiorly during late fetal and perinatal stages. These changes occur in tandem with forward orientation of the orbits in platyrrhines, combined with downward growth of the midface. Thus, we postulate that active growth centers direct the orientation of the midface and orbit before birth.

Breast cancer is a polygenic disorder and is the leading cause of cancer related mortality among women. Early-onset breast cancer (EOBC) is diagnosed in women prior to 45 years-of-age and is associated with worse clinical outcomes, a more aggressive disease phenotype, and poor prognosis for disease-free survival. While substantial progress has been made in defining the genetics of breast cancer, EOBC remains less well understood. In the current study we perform a retrospective analysis of data derived from The Two Sister Study. The use of alternate strategies for handling age-at-diagnosis in conjunction with haplotype-based methods yielded novel findings that help to explain the heritability of EOBC. These findings are validated through comparison against discordant sibs from The Two Sister Study as well as using data derived The Cancer Genome Atlas (TCGA).

Aims Data suggest pharmacological treatment of depression with selective serotonin reuptake inhibitors (SSRI) may impair bone health. Our group has previously modeled compromised craniofacial healing after treatment with sertraline, a commonly prescribed SSRI, and hypothesized potential culprits: alterations in bone cells, collagen, and/or inflammation. Here we interrogate bone lineage cell alterations due to sertraline treatment as a potential cause of the noted compromised bone healing. Main methods Murine pre-osteoblast, pre-osteoclast, osteoblast, and osteoclast cells were treated with clinically relevant concentrations of the SSRI. Studies focused on serotonin pathway targets, cell viability, apoptosis, differentiation, and the osteoblast/osteoclast feedback loop. Key findings All cells studied express neurotransmitters (e.g. serotonin transporter, SLC6A4, SSRI target) and G-protein-coupled receptors associated with the serotonin pathway. Osteoclasts presented the greatest native expression of Slc6a4 with all cell types exhibiting decreases in Slc6a4 expression after SSRI treatment. Pre-osteoclasts exhibited alteration to their differentiation pathway after treatment. Pre-osteoblasts and osteoclasts showed reduced apoptosis after treatment but showed no significant differences in functional assays. RANKL:OPG mRNA and protein ratios were decreased in the osteoblast lineage. Osteoclast lineage cells treated with sertraline demonstrated diminished TRAP positive cells when pre-exposed to sertraline prior to RANKL-induced differentiation. Significance These data suggest osteoclasts are a likely target of bone homeostasis disruption due to sertraline treatment, most potently through the osteoblast/clast feedback loop.

Although bone repair is typically an efficient process, an inadequate healing response can occur, with approximately 5-20% of fractures developing nonunion. Even with improved healing strategies and external fixation devices, overall rate of nonunion has not been significantly reduced, particularly for atrophic nonunion. Atrophic nonunion is characterized by sparse or no callus formation and is difficult to treat clinically, resulting in long-term pain and functional limitation. Reliable preclinical models are needed to study the pathophysiology of atrophic nonunion to create better treatment options. The MouseNail kit (RISystem, Landquart, Switzerland) provides a highly standardized approach in which stabilized segmental bone defects are achieved through interlocked intramedullary nailing. However, reliably performing this surgery is technically challenging, particularly while maintaining strict asepsis. Skilled and aseptic surgical execution is important and necessary because it ensures optimal animal welfare and reproducibility. Therefore, the aim of this paper is to describe:•Novel modifications to the MouseNail kit that allow for: 1) a completely aseptic surgical environment, including description of a hanging limb orthopedic aseptic preparation and 2) a reduction in fracture gap size necessary for induction of atrophic nonunion.•Pre- to post-operative recommendations to facilitate successful performance of murine orthopedic survival surgery., Graphical Abstract Image, graphical abstract

Large craniofacial defects present a substantial clinical challenge that often requires the use of osteoconductive matrices and osteoinductive cues (i.e., bone morphogenetic proteins [BMP2]) to augment healing. While these methods have improved clinical outcomes, a better understanding of how the osteogenic fronts surrounding the defect, the underlying dura mater, and the cranial suture area contribute to healing may lead to more targeted therapies to enhance bone regeneration. We hypothesized that healing within a large bone defect will be precipitated from cells within the remaining or available suture mesenchyme abutting the edges of a murine critical sized defect. To investigate this hypothesis, 39 adult, wild-type mice were randomly arranged into groups (9 or 10 per group) by time (4 and 8 weeks) and treatment (control, acellular collagen sponge alone, or acellular collagen sponge loaded with a clinically relevant scaled dosage of BMP2). The skulls were then subjected to microcomputed tomography and histological analysis to assess bone regeneration in regions of interest within the defect area. A regional assessment of healing indicated that BMP2 drives greater healing than control and that healing emanates from the surgical margin, particularly from the margin associated with undisrupted suture mesenchyme. Though BMP2 treatment drove an increase in cell presence within the healing defect, there was no regional orientation of craniofacial stem cells or vascularity. Overall, these data reinforce that osteoconductive matrices in conjunction with osteoinductive peptides result in better healing of large calvarial defects. This healing is characterized as emanating from the surgical margin where there is an abundant supply of vasculature and progenitor cells.

It is suggested that craniosynostosis is caused by a heterogeneous set of effects including gene mutations, teratogenic exposure during critical periods of development and gene/environment interactions. Distinguishing between sufficient, additive and interactive effects is important to the study of gene/environment interactions and allows for segregation of environmental exposures effecting susceptible populations. Through the identification of sufficient and interactive effects, efforts in prevention of craniosynostosis may be successful. Here, we provide a brief review focusing on defining these categorized exposures and relevant literature that has interrogated gene/environment interactions for craniosynostosis.

Objective Fusion of the cranial sutures is thought to depend on signaling among perisutural tissues. Mapping regional variations in gene expression would improve current models of craniosynostosis. Laser capture microdissection (LCM) isolates discrete cell populations for gene expression analysis. LCM has rarely been used in the study of mineralized tissue. This study sought to evaluate the potential use of LCM for mapping of regional gene expression within the cranial suture. Design Coronal sutures were isolated from 10-day-old wild-type and craniosynostotic (CS) New Zealand White rabbits, and LCM was used to isolate RNA from the sutural ligament (SL), osteogenic fronts (OF), dura mater, and periosteum. Relative expression levels for Fibroblast Growth Factor 2 (FGF2), Fibroblast Growth Factor Receptor 2 (FGFR2), Transforming Growth Factor Beta 2 (TGFβ-2), Transforming Growth Factor Beta 3 (TGFβ-3), Bone Morphogenetic Protein 2 (BMP-2), Bone Morphogenetic Protein 4 (BMP-4), and Noggin were determined using quantitative real-time PCR. Results A fivefold increase in TGFβ2 expression was detected in the CS SL relative to wild type, whereas 152-fold less TGFβ-3 was detected within the OF of CS animals. Noggin expression was increased by 10-fold within the CS SL, but reduced by 13-fold within the CS dura. Reduced expression of FGF2 was observed within the CS SL and dura, whereas increased expression of FGFR2 was observed within the CS SL. Reduced expression of BMP-2 was observed in the CS periosteum, and elevated expression of BMP-4 was observed in the CS SL and dura. Conclusions LCM provides an effective tool for measuring regional variations in cranial suture gene expression. More precise measurements of regional gene expression with LCM may facilitate efforts to correlate gene expression with suture morphogenesis and pathophysiology.

Background Craniosynostosis ranges in severity from single suture involvement with prenatal onset to multiple suture involvement with postnatal onset. The present study was designed to test the hypothesis that increasing homozygosity may be responsible for more severe phenotypic expression by examining the relationship between inbreeding and phenotypic expression in synostotic rabbits. Methods Data were obtained from 173 litters and 209 rabbits with familial craniosynostosis. Five distinct phenotypes were identified (normal n = 62; unicoronal delayed onset synostosis (DOS) n = 47; bicoronal DOS n - 21; unicoronal early onset synostosis (EOS) n - 26, and bicoronal EOS n = 53). Wright's coefficients of inbreeding (CI) were calculated using CompuPed software. Radiographs were taken at 10, 25, 42, 84, and 126 days of age to assess coronal suture, craniofacial, and skeletal growth. The relationship between CI and growth data was assessed using correlation coefficients. Results Mean CIs ranged from 15.68 (±2.22) in normal rabbits to 25.89 (±5.03) in bicoronal DOS, to 36.29 (±2.10) in unicoronal EOS to 42.85 (±2.10) in bicoronal EOS rabbits. Significant differences were noted among groups (F - 11.48; P < .001). Significant negative correlations were noted between CI and sutural and craniofacial growth at 25 (r = -.45, P < .001; and r = -.66, P < .001) through 126 (r = -.40, P < .001 and r = -.46, P < .001) days of age. Conclusions While the synostotic phenotype is inherited in an autosomal dominant fashion in these rabbits, increasing homozygosity is associated with more severely affected phenotypes. These findings suggest that an accumulation of additional, modifier genes may determine the severity of the synostotic phenotype in rabbits.

The Centers for Disease Control and Prevention, National Birth Defects Study suggests that environmental exposures including maternal thyroid diseases, maternal nicotine use, and use of selective serotonin reuptake inhibitors (SSRIs) may exacerbate incidence and or severity of craniofacial abnormalities including craniosynostosis. Premature fusion of a suture(s) of the skull defines the birth defect craniosynostosis which occurs in 1:1800-2500 births. A proposed mechanism of craniosynostosis is the disruption of proliferation and differentiation of cells in the perisutural area. Here, we hypothesize that pharmacological exposures including excess thyroid hormone, nicotine, and SSRIs lead to an alteration of stem cells within the sutures resulting in premature fusion. In utero exposure to nicotine and citalopram (SSRI) increased the risk of premature suture fusion in a wild-type murine model. Gli1+ stem cells were reduced, stem cell populations were depleted, and homeostasis of the suture mesenchyme was altered with exposure. Thus, although these pharmacological exposures can deplete calvarial stem cell populations leading to craniosynostosis, depletion of stem cells is not a unifying mechanism for pharmacological exposure associated craniosynostosis. Keywords: Craniosynostosis, Stem cells, Gli1, Teratogens

Previous attempts to study the effect of prophylactic antibiotics on the outcomes of cleft palate surgery have been hampered by the need for a very large sample size to provide adequate power to discern a potentially small therapeutic effect. This limitation can be overcome by querying large databases created by health care governing bodies.Data from the Pediatric Health Information System database were used for this analysis. Patients, aged 6 to 18 months, who had undergone primary palatoplasty (International Classification of Diseases, Ninth Revision, code 27.62) between 2004 and 2009 were included. Subsequent repair of an oronasal fistula between 2004 and 2015 was identified by International Classification of Diseases, Ninth Revision, procedure code 21.82. Pharmacy billing records were used to determine antibiotic administration. Associations between antibiotic administration and fistula repair were assessed using random-intercept logistic regression adjusting for age, sex, race, and cleft type.Seven thousand one hundred sixty patients were available for analysis; of these, 460 (6.4 percent) had a subsequent repair of an oronasal fistula. Fistula rates were 5.9, 11.4, and 5.2 percent among patients given preoperative antibiotics, only postoperative antibiotics, and no antibiotics, respectively (p0.001). Multivariable analysis results showed that the odds of having an oronasal fistula among patients who were administered preoperative antibiotics did not differ significantly (statistically) from patients who did not receive antibiotics (OR, 0.88; 95 percent CI, 0.59 to 1.31).The treatment goal of primary palatoplasty is the successful repair of the cleft without an oronasal fistula. Administration of preoperative antibiotics did not significantly reduce the odds of subsequent fistula repair within the same Pediatric Health Information System institution following primary palatoplasty.Therapeutic, III.

Background:The use of subgaleal drains following primary cranioplasty for craniosynostosis has undergone limited investigation. Proposed benefits include prevention of seroma, detection of postoperative bleeding, and cerebrospinal fluid leak. We conducted a systematic review of the literature and surveyed craniofacial surgeons to ascertain the current evidence pertaining to drain use following primary cranioplasty for craniosynostosis and to determine surgical practice patterns.Methods:PubMed and Embase databases were searched to identify relevant articles. Abstracts were reviewed by 2 investigators, and a Cohen κ statistic was calculated. Patient demographic and outcome data were extracted and compared. A 9-question survey was e-mailed to active and associate members of the American Society of Craniofacial Surgeons.Results:A total of 7395 unique citations were identified. Only 2 retrospective chart reviews met inclusion criteria. All objective parameters demonstrated no difference between patients with and without drains. A subjective benefit of limiting facial swelling was proposed without objective analysis. Fifty (32.5%) of the 154 craniofacial surgeons responded to the survey. Forty-two percent used postoperative drains. A significant association ( P = .01) was found between the belief that drains limited facial swelling and their use.Conclusions:The literature examining postoperative drain use in primary cranioplasty for craniosynostosis is restricted. The current studies show no definite benefit to drain use but are limited in their assessment of key outcomes. There is wide variability among surgeons regarding drain use, and this seems to be motivated by belief and tradition.

Despite the link between adverse birth outcomes due to pre- and peri-natal nicotine exposure, research suggests 11% of US women continue to smoke or use alternative nicotine products throughout pregnancy. Maternal smoking has been linked to incidence of craniofacial anomalies. We hypothesized that pre-natal nicotine exposure may directly alter craniofacial development independent of the other effects of cigarette smoking. To test this hypothesis, we administered pregnant C57BL6 mice drinking water supplemented with 0, 50, 100 or 200 μg/ml nicotine throughout pregnancy. On postnatal day 15 pups were sacrificed and skulls underwent micro-computed tomography (µCT) and histological analyses. Specific nicotinic acetylcholine receptors, α3, α7, β2, β4 were identified within the calvarial growth sites (sutures) and centers (synchondroses). Exposing murine calvarial suture derived cells and isotype cells to relevant circulating nicotine levels alone and in combination with nicotinic receptor agonist and antagonists resulted in cell specific effects. Most notably, nicotine exposure increased proliferation in calvarial cells, an effect that was modified by receptor agonist and antagonist treatment. Currently it is unclear what component(s) of cigarette smoke is causative in birth defects, however these data indicate that nicotine alone is capable of disrupting growth and development of murine calvaria.

Background Craniosynostosis, the premature fusion of one or more of the cranial sutures, is estimated to occur in 1:1800 to 2500 births. Genetic murine models of craniosynostosis exist, but often imperfectly model human patients. Case, cohort, and surveillance studies have identified excess thyroid hormone as an agent that can either cause or exacerbate human cases of craniosynostosis. Methods Here we investigate the influence of in utero and in vitro exogenous thyroid hormone exposure on a murine model of craniosynostosis, Twist 1 +/−. Results By 15 days post-natal, there was evidence of coronal suture fusion in the Twist 1 +/− model, regardless of exposure. With the exception of craniofacial width, there were no significant effects of exposure; however, the Twist 1 +/− phenotype was significantly different from the wild-type control. Twist 1 +/− cranial suture cells did not respond to thyroxine treatment as measured by proliferation, osteogenic differentiation, and gene expression of osteogenic markers. However, treatment of these cells did result in modulation of thyroid associated gene expression. Conclusion Our findings suggest the phenotypic effects of the genetic mutation largely outweighed the effects of thyroxine exposure in the Twist 1 +/− model. These results highlight difficultly in experimentally modeling gene–environment interactions for craniosynostotic phenotypes. Birth Defects Research (Part A) 106:803–813, 2016. © 2016 Wiley Periodicals, Inc.

Postoperative reossification is a common clinical correlate following surgery. It has been suggested that an underexpression of transforming growth factor-β3 (TGF-β3) may be related to craniosynostosis and postoperative reossification. Adding TGF-β3 may delay reossification and improve postoperative growth. The present study was designed to test this hypothesis. Thirty 10-day-old New Zealand white rabbits with hereditary coronal suture synostosis were divided into three groups: (1) suturectomy controls (n = 14), (2) suturectomy treated with bovine serum albumin (n = 8), and (3) suturectomy treated with TGF-β3 protein (n = 8). At 10 days of age, a 3-mm × 15-mm coronal suturectomy was performed, and serial three-dimensional (3D) computed tomography (CT) scans and cephalographs were taken at 10, 25, 42, and 84 days of age. Calvaria were harvested at 84 days of age for histomorphometric analysis. Mean differences were analyzed using a group by age analysis of variance. Analysis of the 3D CT scan data revealed that sites treated with TGF-β3 had significantly (P < .05) greater defect areas and significantly (P < .05) greater intracranial volumes through 84 days of age compared with controls. Histomorphometry showed that sites treated with TGF-β3 had patent suturectomy sites and significantly (P < .001) less new bone in the suturectomy site compared with controls. Serial radiograph data revealed significant (P < .05) differences in craniofacial growth from 25 to 84 days in TGF-β3-treated rabbits compared with controls. Data show that TGF-β3 administration delayed reossification and improved craniofacial growth in this rabbit model. These findings also suggest that this molecular-based therapy may have potential clinical use.

BACKGROUND Management of the previously infected craniofacial defect remains a significant clinical challenge, posing obstacles such as wound healing complications, lack of donor site availability, and predisposition to failure of the repair. Optimal therapy would reconstruct like with like, without donor site morbidity. The purpose of this study was to compare the efficacy of recombinant human bone morphogenetic protein-2 (rhBMP-2)-mediated bone regeneration with the current standard of autologous bone graft for repair of previously infected calvarial defects. METHODS Nineteen adult New Zealand white rabbits underwent subtotal calvariectomy. Bone flaps were inoculated with Staphylococcus aureus and replanted. After 1 week of infection, bone flaps were removed, and wounds were debrided, followed by 10 days of antibiotic treatment. After 6 weeks, animals underwent scar debridement followed by definitive reconstruction in 1 of 4 groups: empty control (n = 3), vehicle control (buffer solution on absorbable collagen sponge [ACS], n = 3), autologous bone graft (n = 3), or rhBMP-2 repair (rhBMP-2/ACS, n = 10). Animals underwent computed tomography imaging at 0, 2, 4, and 6 weeks postoperatively, followed by euthanization and histological analysis. Percent healing was determined by 3-dimensional analysis. A (time × group) 2-way analysis of variance was performed on healing versus treatment group and postoperative time. RESULTS At 6 weeks postoperatively, rhBMP-2/ACS and autologous bone graft resulted in 93% and 68% healing, respectively, whereas the empty and vehicle control treatment resulted in 27% and 26% healing (P < 0.001). Histologically, compared to autologous bone graft, bone in the rhBMP-2/ACS group was more cellular and more consistently continuous with wound margins. CONCLUSIONS The rhBMP-2 therapy is effective in achieving radiographic coverage of previously infected calvarial defects.

Progressive curricular changes in medical education over the past two decades have resulted in the diaspora of gross anatomy content into integrated curricula while significantly reducing total contact hours. Despite the development of a wide range of alternative teaching modalities, gross dissection remains a critical component of medical education. The challenge posed to modern anatomists is how to maximize and integrate the time spent dissecting under the current curricular changes. In this study, an alternative approach to the dissection of the pelvis and perineum is presented in an effort to improve content delivery and student satisfaction. The approach involves removal of the perineum en bloc from the cadaver followed by excision of the pubic symphysis, removal and examination of the bladder and associated structures, examination and bisection of the midline pelvic organs in situ, and midsagittal hemisection of the pelvis for identification of the neurovasculature. Results indicate that this novel dissecting approach increases the number of structures identified by 46% ± 14% over current dissecting methods. Survey results indicate that students were better able to integrate lecture and laboratory concepts, understand the concepts, and successfully identify more structures using the new approach (P < 0.05). The concept of anatomic efficiency is introduced and proposed as a standard quantitative measure of gross dissection proficiency across programs and institutions. These findings provide evidence that innovative solutions to anatomy education can be found that help to maintain critical content and student satisfaction in a modern medical curriculum.

Bone wound healing is a highly dynamic and precisely controlled process through which damaged bone undergoes repair and complete regeneration. External factors can alter this process, leading to delayed or failed bone wound healing. The findings of recent studies suggest that the use of selective serotonin reuptake inhibitors (SSRIs) can reduce bone mass, precipitate osteoporotic fractures and increase the rate of dental implant failure. With 10% of Americans prescribed antidepressants, the potential of SSRIs to impair bone healing may adversely affect millions of patients’ ability to heal after sustaining trauma. Here, we investigate the effect of the SSRI sertraline on bone healing through pre-treatment with (10 mg·kg-1 sertraline in drinking water, n = 26) or without (control, n = 30) SSRI followed by the creation of a 5-mm calvarial defect. Animals were randomized into three surgical groups: (a) empty/sham, (b) implanted with a DermaMatrix scaffold soak-loaded with sterile PBS or (c) DermaMatrix soak-loaded with 542.5 ng BMP2. SSRI exposure continued until sacrifice in the exposed groups at 4 weeks after surgery. Sertraline exposure resulted in decreased bone healing with significant decreases in trabecular thickness, trabecular number and osteoclast dysfunction while significantly increasing mature collagen fiber formation. These findings indicate that sertraline exposure can impair bone wound healing through disruption of bone repair and regeneration while promoting or defaulting to scar formation within the defect site.

Many variables serve to alter the process of bone remodelling and diminish regeneration including the size and nature of the wound bed and health status of the individual. To overcome these inhibitory factors, tissue-engineered osteoconductive scaffolds paired with various growth factors have been utilized clinically. However, many limitations still remain, for example, bone morphogenetic protein 2 (BMP2) can lead to rampant inflammation, ectopic bone formation, and graft failure. Here, we studied the ability for a nanofiber scaffold (Talymed) to accelerate BMP2 growth factor-induced bone healing compared with the traditional absorbable collagen sponge (ACS) delivery system. One hundred fifty-five adult wild type mice were arranged in 16 groups by time, 4 and 8 weeks, and treatment, ACS or Talymed, loaded with control, low, medium, or high dosages of BMP2. Skulls were subjected to microCT, biomechanical, and histological analysis to assess bone regeneration. The use of Talymed within the defect site was found to decrease the bone volume, bone formation rate, and alkaline phosphatase activity compared with ACS/BMP2 combinations. Interestingly, though Talymed regenerated less bone, the regenerate was found to have a greater hardness value than that of bone within the ACS groups. However, the difference in bone hardness between scaffolds was not detectable by 8 weeks. Based on these results, we found that the nanofiber scaffold generated a better quality of bone regenerate at 4 weeks but, due to the lack of overall bone formation and the inhibition of normal remodelling processes, was not as efficacious as the current clinical standard ACS/BMP2 therapy.

An estimated 3% of US pregnancies are affected by maternal thyroid dysfunction, with between one and three of every 1000 pregnancies being complicated by overactive maternal thyroid levels. Excess thyroid hormones are linked to neurological impairment and excessive craniofacial variation, affecting both endochondral and intramembranous bone. Using a geometric morphometric approach, this study evaluates the role of in utero thyroxine overexposure on the growth of offspring mandibles in a sample of 241 mice. Canonical variate analysis utilized 16 unilateral mandibular landmarks obtained from 3D micro-computed tomography to assess shape changes between unexposed controls (n = 63) and exposed mice (n = 178). By evaluating shape changes in the mandible among three age groups (15, 20 and 25 days postnatal) and different dosage levels (low, medium and high), this study found that excess maternal thyroxine alters offspring mandibular shape in both age- and dosage-dependent manners. Group differences in overall shape were significant (P < 0.001), and showed major changes in regions of the mandible associated with muscle attachment (coronoid process, gonial angle) and regions of growth largely governed by articulation with the cranial base (condyle) and occlusion (alveolus). These results compliment recent studies demonstrating that maternal thyroxine levels can alter the cranial base and cranial vault of offspring, contributing to a better understanding of both normal and abnormal mandibular development, as well as the medical implications of craniofacial growth and development.

Previous studies identified a circulating human osteoblastic population that expressed osteocalcin (OCN), increased following fracture and pubertal growth, and formed mineralized colonies in vitro and bone in vivo. A subpopulation expressed CD34, a hematopoietic/endothelial marker. These findings led to our hypothesis that hematopoietic-derived CD34Flow cytometry was used to identify CD34Circulating CD34Together, these data demonstrate a murine CD34

Bone has the potential for spontaneous healing. This process, however, often fails in patients with comorbidities. Tissue engineering combining functional cells, biomaterials and osteoinductive cues may provide alternative treatment strategies. We have recently demonstrated that stromal cell-derived factor-1β (SDF-1β) works in concert with bone morphogenetic protein-2 (BMP-2) to potentiate osteogenic differentiation of bone marrow-derived mesenchymal stem/stromal cells (BMSCs). Here, we test the hypothesis that SDF-1β overexpressed in Tet-Off-SDF-1β BMSCs, delivered on acellular dermal matrix (ADM), synergistically augments BMP-2-induced healing of critical-sized mouse calvarial defects. BMSC therapies alone showed limited bone healing, which was increased with co-delivery of BMP-2. This was further enhanced in Tet-Off-SDF-1β BMSCs + BMP-2. Only limited BMSC retention on ADM constructs was observed after 4 weeks in vivo, which was increased with BMP-2 co-delivery. In vitro cell proliferation studies showed that supplementing BMP-2 to Tet-Off BMSCs significantly increased the cell number during the first 24 h. Consequently, the increased cell numbers decreased the detectable BMP-2 levels in the medium, but increased cell-associated BMP-2. The data suggest that SDF-1β provides synergistic effects supporting BMP-2-induced, BMSC-mediated bone formation and appears suitable for optimization of bone augmentation in combination therapy protocols. Copyright © 2015 John Wiley & Sons, Ltd.

Thyroid hormone is important for skull bone growth, which primarily occurs at the cranial sutures and synchondroses. Thyroid hormones regulate metabolism and act in all stages of cartilage and bone development and maintenance by interacting with growth hormone and regulating insulin-like growth factor. Aberrant thyroid hormone levels and exposure during development are exogenous factors that may exacerbate susceptibility to craniofacial abnormalities potentially through changes in growth at the synchondroses of the cranial base. To elucidate the direct effect of in utero therapeutic thyroxine exposure on the synchondroses in developing mice, we provided scaled doses of the thyroid replacement drug, levothyroxine, in drinking water to pregnant C57BL6 wild-type dams. The skulls of resulting pups were subjected to micro-computed tomography analysis revealing less bone volume relative to tissue volume in the synchondroses of mouse pups exposed in utero to levothyroxine. Histological assessment of the cranial base area indicated more active synchondroses as measured by metabolic factors including Igf1. The cranial base of the pups exposed to high levels of levothyroxine also contained more collagen fiber matrix and an increase in markers of bone formation. Such changes due to exposure to exogenous thyroid hormone may drive overall morphological changes. Thus, excess thyroid hormone exposure to the fetus during pregnancy may lead to altered craniofacial growth and increased risk of anomalies in offspring.

Background Selective serotonin reuptake inhibitor (SSRI) use in pregnancy has been linked to craniofacial birth defects. Little is known about the effects of serotonin or SSRIs on craniofacial development. Here, we provide evidence that citalopram (SSRI) alters the osteogenic profile of murine calvarial cells and leads to craniofacial dysmorphology. Methods We used mouse calvarial pre-osteoblast cells (MC3T3-E1) to study the biochemical profile (microarray and quantitative reverse transcription polymerase chain reactions) after treatment with a titrated dose of citalopram. We used C57BL-6 wild-type breeders to produce litters treated with a clinical dose of citalopram during the third trimester of pregnancy. We used micro-computed tomography and morphometric measures to determine effects on craniofacial development. Results Controls included untreated cells and age matched untreated litters. We observed decreases in proliferation and increases in alkaline phosphatase activity after citalopram exposure. We confirmed altered expression of genes linked to osteogenesis including Ocn and significant increase in expression of Alp after 7 days of treatment. Our data suggest altered expression of several genes related to craniofacial development (Fgf2, Fgfr2, Tgfβr2 Irs1, Igf1) and statistically significant changes in expression for (Col2a1, Gdf6, Hmox1, and Notch1). We also observed changes in regulation of the serotonin pathway (Sert, Tph1, Tph2, Htr2a, Lrp5) after treatment with citalopram. After in utero exposure to citalopram, mice displayed shorter narrow snouts, more globular skulls and several craniofacial anomalies. Conclusion Our results provide confirmatory evidence that citalopram exposure is associated with cellular and morphological alterations of the craniofacial complex, which may have important implications for use during pregnancy. Birth Defects Research (Part A), 2014. © 2014 Wiley Periodicals, Inc. Birth Defects Research (Part A) 100:912–923, 2014. © 2014 Wiley Periodicals, Inc.