Cranial suture lineage and contributions to repair of the mouse skull.

The cranial sutures are proposed to be a stem cell niche, harbouring skeletal stem cells that are directly involved in development, homeostasis and healing. Like the craniofacial bones, the sutures are formed from both mesoderm and neural crest. During cranial bone repair, neural crest cells have been proposed to be key players; however, neural crest contributions to adult sutures are not well defined, and the relative importance of suture proximity is unclear. Here, we use genetic approaches to re-examine the neural crest-mesoderm boundaries in the adult mouse skull. These are combined with calvarial wounding experiments suggesting that suture proximity improves the efficiency of cranial repair. Furthermore, we demonstrate that Gli1+ and Axin2+ skeletal stem cells are present in all calvarial sutures examined. We propose that the position of the defect determines the availability of neural crest-derived progenitors, which appear to be a key element in the repair of calvarial defects.

Novel truncating mutations in CTNND1 cause a dominant craniofacial and cardiac syndrome.

CTNND1 encodes the p120-catenin (p120) protein, which has a wide range of functions, including the maintenance of cell-cell junctions, regulation of the epithelial-mesenchymal transition and transcriptional signalling. Due to advances in next-generation sequencing, CTNND1 has been implicated in human diseases including cleft palate and blepharocheilodontic (BCD) syndrome albeit only recently. In this study, we identify eight novel protein-truncating variants, six de novo, in 13 participants from nine families presenting with craniofacial dysmorphisms including cleft palate and hypodontia, as well as congenital cardiac anomalies, limb dysmorphologies and neurodevelopmental disorders. Using conditional deletions in mice as well as CRISPR/Cas9 approaches to target CTNND1 in Xenopus, we identified a subset of phenotypes that can be linked to p120-catenin in epithelial integrity and turnover, and additional phenotypes that suggest mesenchymal roles of CTNND1. We propose that CTNND1 variants have a wider developmental role than previously described and that variations in this gene underlie not only cleft palate and BCD but may be expanded to a broader velocardiofacial-like syndrome.

Inactivation of Osteoblast PKC Signaling Reduces Cortical Bone Mass and Density and Aggravates Renal Osteodystrophy in Mice with Chronic Kidney Disease on High Phosphate Diet.

Chronic kidney disease (CKD) frequently leads to hyperphosphatemia and hyperparathyroidism, mineral bone disorder (CKD-MBD), ectopic calcifications and cardiovascular mortality. PTH activates the osteoanabolic Gαs/PKA and the Gαq/11/PKC pathways in osteoblasts, the specific impact of the latter in CKD-MBD is unknown. We generated osteoblast specific Gαq/11 knockout (KO) mice and established CKD-MBD by subtotal nephrectomy and dietary phosphate load. Bone morphology was assessed by micro-CT, osteoblast function by bone planar scintigraphy at week 10 and 22 and by histomorphometry. Osteoblasts isolated from Gαq/11 KO mice increased cAMP but not IP3 in response to PTH 1-34, demonstrating the specific KO of the PKC signaling pathway. Osteoblast specific Gαq/11 KO mice exhibited increased serum calcium and reduced bone cortical thickness and mineral density at 24 weeks. CKD Gαq/11 KO mice had similar bone morphology compared to WT, while CKD Gαq/11-KO on high phosphate diet developed decreased metaphyseal and diaphyseal cortical thickness and area, as well as a reduction in trabecular number. Gαq/11-KO increased bone scintigraphic tracer uptake at week 10 and mitigated tracer uptake in CKD mice at week 22. Histological bone parameters indicated similar trends. Gαq/11-KO in osteoblast modulates calcium homeostasis, bone formation rate, bone morphometry, and bone mineral density. In CKD and high dietary phosphate intake, osteoblast Gαq/11/PKC KO further aggravates mineral bone disease.

Disrupted mitochondrial function in the Opa3L122P mouse model for Costeff Syndrome impairs skeletal integrity.

Mitochondrial dysfunction connects metabolic disturbance with numerous pathologies, but the significance of mitochondrial activity in bone remains unclear. We have, therefore, characterized the skeletal phenotype in the Opa3L122P mouse model for Costeff syndrome, in which a missense mutation of the mitochondrial membrane protein, Opa3, impairs mitochondrial activity resulting in visual and metabolic dysfunction. Although widely expressed in the developing normal mouse head, Opa3 expression was restricted after E14.5 to the retina, brain, teeth and mandibular bone. Opa3 was also expressed in adult tibiae, including at the trabecular surfaces and in cortical osteocytes, epiphyseal chondrocytes, marrow adipocytes and mesenchymal stem cell rosettes. Opa3L122P mice displayed craniofacial abnormalities, including undergrowth of the lower mandible, accompanied in some individuals by cranial asymmetry and incisor malocclusion. Opa3L122P mice showed an 8-fold elevation in tibial marrow adiposity, due largely to increased adipogenesis. In addition, femoral length and cortical diameter and wall thickness were reduced, the weakening of the calcified tissue and the geometric component of strength reducing overall cortical strength in Opa3L122P mice by 65%. In lumbar vertebrae reduced vertebral body area and wall thickness were accompanied by a proportionate reduction in marrow adiposity. Although the total biomechanical strength of lumbar vertebrae was reduced by 35%, the strength of the calcified tissue (σmax) was proportionate to a 38% increase in trabecular number. Thus, mitochondrial function is important for the development and maintenance of skeletal integrity, impaired bone growth and strength, particularly in limb bones, representing a significant new feature of the Costeff syndrome phenotype.

Subcutaneous extraskeletal osteosarcoma of the forearm: a case report and review of the literature.

Extraskeletal osteosarcoma (ESOS) originating in the subcutaneous tissue is a rare occurrence, accounting for less than 10 % of ESOS cases. Osteosarcoma of extraskeletal origin accounts for approximately 2-4 % of all osteosarcomas, and 1 % of soft tissue sarcomas. We report a case of an 80-year-old female with an isolated primary subcutaneous tumor of the forearm. After imaging, surgical excision, and pathological analysis, the diagnosis of a subcutaneous osteosarcoma was made. This report documents the clinical and pathological findings of subcutaneous ESOS in this case, along with a review of previous cases of subcutaneous ESOS.

Management of Grade 1 Open Distal Radius Fractures: A Survey of Practicing Surgeons.

Introduction Standard of care management for open fractures historically mandates emergent systemic antibiotic administration, followed by urgent irrigation and debridement in the operating room, regardless of injury severity. However, significant controversy exists regarding the specific implementation and importance of these commonly accepted guidelines. We aimed to define differences in the management of grade 1 open distal radius fractures. Methods An anonymous online survey was distributed to attending surgeon members of either the Orthopaedic Trauma Association (OTA) between January 2019 and April 2019 or the New York Society for Surgery of the Hand (NYSSH) in January 2019. Results A total of 68 attending surgeons responded to the survey. A total of 24 OTA members and 40 NYSSH members replied and were included in the study. Several factors influenced management in addition to the level of contamination. Of the surgeons, 68% stated that litigation was not a major factor of concern. When compared to surgeons who trained in trauma fellowships, more surgeons who trained in hand/upper extremity fellowships considered closed reduction alone as reasonable definitive treatment (when excluding antibiotic administration and debridement considerations, p = 0.024) and oral antibiotics as a supplement to IV antibiotics (p < 0.001). Of the surgeons, 90% would nonoperatively treat a patient who presented with a grade 1 open distal radius fracture greater than 72 hours after injury with stable and acceptable alignment on X-rays. Conclusion Some surgeons are willing to deviate from standard-of-care management protocols.

Caspase-7 participates in differentiation of cells forming dental hard tissues.

Apoptosis during tooth development appears dependent on the apoptotic executioner caspase-3, but not caspase-7. Instead, activated caspase-7 has been found in differentiated odontoblasts and ameloblasts, where it does not correlate with apoptosis. To further investigate these findings, the mouse incisor was used as a model. Analysis of caspase-7-deficient mice revealed a significant thinner layer of hard tissue in the adult incisor. Micro computed tomography scan confirmed this decrease in mineralized tissues. These data strongly suggest that caspase-7 might be directly involved in functional cell differentiation and regulation of the mineralization of dental matrices.

PTEN Loss Mediates Clinical Cross-Resistance to CDK4/6 and PI3Kα Inhibitors in Breast Cancer.

The combination of CDK4/6 inhibitors with antiestrogen therapies significantly improves clinical outcomes in ER-positive advanced breast cancer. To identify mechanisms of acquired resistance, we analyzed serial biopsies and rapid autopsies from patients treated with the combination of the CDK4/6 inhibitor ribociclib with letrozole. This study revealed that some resistant tumors acquired RB loss, whereas other tumors lost PTEN expression at the time of progression. In breast cancer cells, ablation of PTEN, through increased AKT activation, was sufficient to promote resistance to CDK4/6 inhibition in vitro and in vivo. Mechanistically, PTEN loss resulted in exclusion of p27 from the nucleus, leading to increased activation of both CDK4 and CDK2. Because PTEN loss also causes resistance to PI3Kα inhibitors, currently approved in the post-CDK4/6 setting, these findings provide critical insight into how this single genetic event may cause clinical cross-resistance to multiple targeted therapies in the same patient, with implications for optimal treatment-sequencing strategies. SIGNIFICANCE: Our analysis of serial biopsies uncovered RB and PTEN loss as mechanisms of acquired resistance to CDK4/6 inhibitors, utilized as first-line treatment for ER-positive advanced breast cancer. Importantly, these findings have near-term clinical relevance because PTEN loss also limits the efficacy of PI3Kα inhibitors currently approved in the post-CDK4/6 setting.This article is highlighted in the In This Issue feature, p. 1.

Growth and mineralogy in dental plates of the holocephalan Harriotta raleighana (Chondrichthyes): novel dentine and conserved patterning combine to create a unique chondrichthyan dentition.

ABSTRACT: The dentition in extant holocephalans (Chondrichthyes) comprises three pairs of continuously growing dental plates, rather than the separate teeth characterizing elasmobranchs. We investigated how different types of dentine in these plates, including hypermineralized dentine, are arranged, and how this is renewed aborally, in adult and juvenile dentitions of Harriotta raleighana (Rhinochimeridae). Individual plates were analysed using x-ray computed tomography (µCT), scanning electron microscopy (SEM) in back scattered mode with energy dispersive X-ray (EDX) analysis, and optical microscopy on hard tissue sections. RESULTS: Harriotta dental plates are made entirely of dentine tissue, mostly as trabecular dentine, bone itself being absent. Hypermineralized dentine forms in restricted ovoid and tritor spaces within trabecular dentine, inside a shell of outer and inner dentine layers. Trabecular dentine is ubiquitous but changes to sclerotic osteodentine near the oral surface by increasing density, remaining less mineralized than the hypermineralized dentine. All structures are renewed aborally, within a vascular dental pulp, a tissue suggested to be a source of stem cells for tissue renewal. Ca density profiles and concentrations of Mg, P, and Ca ions reveal extreme differences in the level and type of mineralization. Early mineralization in ovoids and tritors has very high levels of Mg, then a sudden increase in mineralization to a high total mineral content, whereas there is gradual change in trabecular dentine, remaining at a low level.Hypermineralized dentine fills the prepatterned ovoid, rod and tritor spaces, early at the aboral surface within the trabecular dentine. Deposition of the hypermineralized dentine (HD, proposed as new specific name, whitlockin replacing pleromin) is from surfaces that are lined with large specialized odontoblasts, (whitloblasts, instead of pleromoblasts) within cell body spaces connecting with extensive, ramifying tubules. Early mineralization occurs amongst this maze of tubules that penetrate far into the dentine, expanding into a mass of saccules and membranous bodies, dominating in the absence of other organic matrix. This early stage has hydroxyapatite, also significantly rich in Mg, initiated as a poorly crystalline phase. In the hypermineralized dentine, formation occurs as clusters of variably shaped crystals, arising from a sudden phase transition. CONCLUSIONS: In the hypermineralized dentine, high MgO + CaO + P2O5 suggests that almost pure Mg containing tricalciumphosphate (MgTCP: (ß-Ca3(PO4)2) (whitlockite) is present, with little or no hydroxyapatite. Serial replacement of tritors and ovoids is suggested to occur within the dental plate, probably representing a relic of patterning, as classically found in elasmobranch dentitions.

Impact of a Resident-Guided Rounding Initiative on the Hospital Consumer Assessment of Healthcare Providers and Systems Survey Scores in Orthopaedic Surgery Inpatients.

Patient-centered medicine is becoming the main focus of many healthcare systems, and the Hospital Consumer Assessment of Healthcare Providers and Systems (HCAHPS) survey is a tool used to track patient satisfaction. In this study, we evaluate the HCAHPS scores in orthopaedic surgery inpatients before and after implementation of a resident-guided rounding protocol. Analyses of the HCAHPS surveys for 154 orthopaedic surgical inpatients at one community hospital were compared 6 months before and after implementation of a resident-guided rounding initiative. Specific questions of the HCAHPS survey were analyzed using the top box, mean, and positive scores. Implementation of the rounding initiative resulted in an increase in the top box, mean, and positive scores for all questions evaluated; however, no significance was noted in the results, with the exception of the positive score for a staff cohesiveness question (P = 0.046). Physician and hospital recommendation questions showed a 5-point increase (91st to 96th percentile) compared with 42-point increase (21st to 63rd percentile) by publicly reported national data. Implementation of the rounding initiative resulted in increases in HCAHPS scores across multiple questions and domains; however, these were not significant. These results suggest that simple interventions can help increase the overall patient satisfaction and promote future investigations.

Characterisation of the genomic canine Fgf8 locus and screen for genetic variants in 4 dogs with different face types.

We are investigating the genetic basis of morphological differences in skull shape between domestic dogs of different breeds using a candidate gene approach to identify genes involved in the genetic regulation. One such candidate is Fgf8. Fgf8 is a signalling molecule important in the embryonic development and patterning of the craniofacial region. Mice conditional null for the expression of Fgf8 after E9.5 have a short foreface and a wide skull (Trumpp et al. 1999). Using a combination of bioinformatics and PCR cloning, we have characterised the genomic loci of the canine Fgf8 gene. Like the mouse homologue, it is composed of six exons and we also predict that like the mouse, there are eight alternative isoforms that are generated by alternative splicing events. We have identified a short 200 bp sequence upstream of the Fgf8 gene that is highly conserved between species and have predicted putative transcription factor binding sites using the Transfac database. Genetic analysis of 4 dogs with different skull types identified genetic variation. None of the variants however, were predicted to have any functional significance.

Effects of Photo-Depicted Pupil Diameter on Judgments of Others' Attentiveness and on Facial Recognition Memory.

Occasionally, individuals perceive that someone is no longer paying attention to the discussion at hand even when there are no overt cues of inattentiveness. As a preliminary study of this phenomenon, we examined whether pupil diameter might be implicitly used to infer others' attentiveness. Forty participants (27 women, 13 men, M age = 19.7 year, SD = 2.8) were presented with images of male faces with either large or small pupils, and, in the context of a personnel selection scenario, participants then judged the attentiveness of the person in the image. Images of faces with large pupils were judged as more attentive, compared with images of faces with small pupils. Face recognition memory performance was not affected by depicted pupil size. Our results are consistent with the proposal that pupillary fluctuations can be an index of perceived attention, and they provide preliminary evidence that pupil dilation may be implicitly relied upon to infer attentional states.

Expression of Claudin-3 during chick development.

Claudins are membrane proteins located within tight junctions. Using degenerate and gene specific primers the chick homologue of Claudin-3 was isolated. Here we show the expression of Claudin-3 transcripts in the developing chick embryo from Hamburger and Hamilton Stages (HH) 6-22. The early expression domains of Claudin 3 in the developing chick embryo include the mesoderm surrounding Hensen's node and the head fold. Between HH 9 and HH 11 expression domains include the anterior intestinal portal and otic vesicle. By HH 14, gene expression is observed in the pharyngeal endoderm and pouches, in addition to the continued expression in the otic vesicle. Expression in the more posterior pouches was also observed as development proceeded. At HH 20 expression is present in the mesonephric system and also the developing liver, lung bud and intestine.

Characterisation of the genomic structure of chick Fgf8.

The Fgf8 gene encodes a series of secreted signalling molecules important in the normal development of the face, brain and limbs. The genomic structure of the chick Fgf8 gene has been analysed and compared to the human and mouse sequences. Divergence between the chick, human and mouse genomic structure was observed. Data indicates that the long alternatively spliced form of exon 1b observed in mouse and exon 1c observed in human and mouse do not exist in the chick Fgf8 gene. RT-PCR analysis indicates that chick Fgf8, like its mouse and human counterpart is alternatively spliced. This data along with the genomic structure data indicates that in the chick there are only two isoforms of Fgf8. This is in contrast to the human and mouse, where evidence suggests that there are 4 and 8 isoforms, respectively. Approximately 400 bp of intron 1d is highly conserved between chick, human and mouse genomic sequences. Using TRANSFAC possible conserved regulatory element binding sites within this domain were identified.

Wearable cardioverter-defibrillator for prevention of sudden cardiac death after infected implantable cardioverter-defibrillator removal: A cost-effectiveness evaluation.

BACKGROUND: Prevention of sudden cardiac arrest (SCA) after removal of an infected implantable cardioverter-defibrillator (ICD) is a challenging clinical dilemma. The cost-effectiveness of the wearable cardioverter-defibrillator (WCD) in this setting remains uncertain. OBJECTIVE: The purpose of this study was to compare the cost-effectiveness of the WCD with discharge home, discharge to a skilled nursing facility, or inpatient monitoring for the prevention of SCA after infected ICD removal. METHODS: A decision model was developed to compare the cost-effectiveness of use of the WCD to several different strategies for patients who undergo ICD removal. One-way and 2-way sensitivity analyses were performed to account for uncertainties. RESULTS: In the base-case analysis, the incremental cost-effectiveness of the WCD strategy was $20,300 per life-year (LY) or $26,436 per quality-adjusted life-year (QALY) compared to discharge home without a WCD. Discharge to a skilled nursing facility and in-hospital monitoring resulted in higher costs and worse clinical outcomes. The incremental cost-effectiveness ratio was as low as $15,392/QALY if the WCD successfully terminated 95% of SCA events and exceeded the $50,000/QALY willingness-to-pay threshold if the efficacy was <69%.The WCD strategy remained cost-effective, assuming 5.6% 2-month SCA risk, as long as the time to reimplantation was at least 2 weeks. CONCLUSION: The WCD likely is cost-effective in protecting patients against SCA after infected ICD removal while waiting for ICD reimplantation compared to keeping patients in the hospital or discharging them home or to a skilled nursing facility.

Fuz mutant mice reveal shared mechanisms between ciliopathies and FGF-related syndromes.

Ciliopathies are a broad class of human disorders with craniofacial dysmorphology as a common feature. Among these is high arched palate, a condition that affects speech and quality of life. Using the ciliopathic Fuz mutant mouse, we find that high arched palate does not, as commonly suggested, arise from midface hypoplasia. Rather, increased neural crest expands the maxillary primordia. In Fuz mutants, this phenotype stems from dysregulated Gli processing, which in turn results in excessive craniofacial Fgf8 gene expression. Accordingly, genetic reduction of Fgf8 ameliorates the maxillary phenotypes. Similar phenotypes result from mutation of oral-facial-digital syndrome 1 (Ofd1), suggesting that aberrant transcription of Fgf8 is a common feature of ciliopathies. High arched palate is also a prevalent feature of fibroblast growth factor (FGF) hyperactivation syndromes. Thus, our findings elucidate the etiology for a common craniofacial anomaly and identify links between two classes of human disease: FGF-hyperactivation syndromes and ciliopathies.

Regionalisation of early head ectoderm is regulated by endoderm and prepatterns the orofacial epithelium.

The oral epithelium becomes regionalised proximodistally early in development, and this is reflected by the spatial expression of signalling molecules such as Fgf8 and Bmp4. This regionalisation is responsible for regulating the spatial expression of genes in the underlying mesenchyme. These genes are required for the spatial patterning of bone, cartilage orofacial development and, in mammals, teeth. The mechanism and timing of this important regionalisation during head epithelium development are not known. Using lipophilic dyes to fate map the oral epithelium in chick embryos, we show that the cells that will occupy the epithelium of the distal and the proximal mandible primordium already occupy different spatial locations in the developing head ectoderm prior to the formation of the first pharyngeal arch and neural crest migration. Moreover, the ectoderm cells fated to become proximal oral epithelium express Fgf8 and this expression requires the presence of endoderm. Thus, the first fundamental patterning process in jaw morphogenesis is controlled by the early separation of specific areas of ectoderm that are regulated by ectoderm-endoderm interactions, and does not involve neural crest cells.