Craniofacial Bone Research Articles

Optimization and Implication of Adipose-Derived Stem Cells in Craniofacial Bone Regeneration and Repair

Adipose-derived stem cells (ADSCs) have emerged as a promising resource for craniofacial bone regeneration due to their high abundance and easy accessibility, significant osteogenic potential, versatile applications, and potential for personalized medicine, which underscore their importance in this field. This article reviews the current progress of preclinical studies that describe the careful selection of specific ADSC subpopulations, key signaling pathways involved, and usage of various strategies to enhance the osteogenic potential of ADSCs. Additionally, clinical case reports regarding the application of ADSCs in the repair of calvarial defects, cranio-maxillofacial defects, and alveolar bone defects are also discussed.

019. Craniofacial Fracture and Secretome: A Systematic Review

Background: The challange in craniofacial fracture management is restore the morphology and physiology of the carniofacial features to pre-injury remains a challenge. Tissue engginering such as secretome become a great hope as a modern and comprehensive therapy for handling craniofacial fractures. To evaluate the effectiveness of application of secretome in craniofacial bone regeneration in animal and human models. Methods: We conducted in August 2024 through several online databases, including PubMed (MEDLINE), The Cochrane Library, and Google Scholar. The inclusion criteria include methodological research (in vivo, in vitro, or clinical studies) which provided a complete set of data and full-text articles. We omitted studies of non-craniofacial bone fracture repair. COCHRANE RoB 2.0 checklist to analyze the risk of bias. Results: Twenty articles selected, 16 investigated animal models and four conducted clinical studies. Two animal studies researched on rabbits (n=34), one on dogs (n=4) and 13 on rats (n=281). There are 48 participants on the clinical studies. Secretome in animal and humans model increase titer which had effect to bone regeneration such as IGF, VEGF, TGF-B-1, osteocalcin, Runx2, PCNA, ANG1, ANG2, MMP8, BMP2, IBSP, MMP13 and ICAM-1. In radiology and histology evaluation, secretome reported increase rate of bone regeneration. Conclusion: Secretome promotes the expression of cytokines required for bone healing. secretome can enhace craniofacial bone regeneration by speeding up healing and increasing the areas of repair. Analysis is done through morphological studies, CT scan results, and histological findings.

A Novel’s Evidence of MSG-Induced Craniofacial Defects in Chick Embryo Models

Monosodium glutamate (MSG), commonly used as an additive food enhancer, has been reported to have teratogenic effects on the embryo during development. The present study aimed to investigate the effect of MSG-induced teratogenicity, focusing on craniofacial formation in the chick embryo model. One hundred eighty fertilized eggs were divided into control and MSG groups. The chemicals were administered, and the results were investigated in 3-day-old embryos (ED-3), ED-6, and ED-10. The morphology and histology of craniofacial structures were studied using a stereomicroscope and light microscopy. The neural crest cells (NCCs) were investigated by the immunostaining technique. The results showed that a dose of 3 mg of MSG induced anterior neuropore opening and eye malformation in ED-3, and this was clearly demonstrated in ED-6. MSG also caused craniofacial bone malformation and delayed calcification in ED-10. Moreover, MSG induced apoptosis of NCCs and reduced the proliferation of NCCs in the craniofacial structures. These findings are the 1st report to demonstrate the teratogenicity of high doses of MSG-induced craniofacial defects in chick embryos during organogenesis. HIGHLIGHTS Monosodium glutamate exposure causes craniofacial defects in chick embryos. The functions of the neural crest cells were abrogated due to monosodium glutamate-induced teratogenesis, which led to birth defects. The experimental protocols benefit teratogenicity studies, and the results can be applied to future studies to elucidate the causes of birth defects during organogenesis. GRAPHICAL ABSTRACT

Distinct role of Klotho in long bone and craniofacial bone: skeletal development, repair and regeneration.

Bone defects are highly prevalent diseases caused by trauma, tumors, inflammation, congenital malformations and endocrine abnormalities. Ideally effective and side effect free approach to dealing with bone defects remains a clinical conundrum. Klotho is an important protein, which plays an essential role in regulating aging and mineral ion homeostasis. More recently, research revealed the function of Klotho in regulating skeleton development and regeneration. Klotho has been identified in mesenchymal stem cells, osteoblasts, osteocytes and osteoclasts in different skeleton regions. The specific function and regulatory mechanisms of Klotho in long bone and craniofacial bone vary due to their different embryonic development, ossification and cell types, which remain unclear and without conclusion. Moreover, studies have confirmed that Klotho is a multifunctional protein that can inhibit inflammation, resist cancer and regulate the endocrine system, which may further accentuate the potential of Klotho to be the ideal molecule in inducing bone restoration clinically. Besides, as an endogenous protein, Klotho has a promising potential for clinical therapy without side effects. In the current review, we summarized the specific function of Klotho in long bone and craniofacial skeleton from phenotype to cellular alternation and signaling pathway. Moreover, we illustrated the possible future clinical application for Klotho. Further research on Klotho might help to solve the existing clinical difficulties in bone healing and increase the life quality of patients with bone injury and the elderly.

Delivery of a Jagged1-PEG-MAL hydrogel with pediatric human bone cells regenerates critically sized craniofacial bone defects.

Current treatments for congenital and acquired craniofacial (CF) bone abnormalities are limited and costly. Conventional methods involve surgical correction, short-term stabilization, and long-term bone grafting, which may include problematic allografts and limited autografts. While bone morphogenetic protein 2 (BMP2) has been used for bone regeneration, it can cause bone overgrowth and life-threatening inflammation. Bone marrow-derived mesenchymal stem cell therapies, though promising, are not Food and Drug Administration approved and are resource intensive. Thus, there is a need for effective, affordable, and less side-effect-prone bone regenerative therapies. Previous research demonstrated that JAGGED1 induces osteoblast commitment in murine cranial neural crest cells through a NOTCH-dependent non-canonical pathway involving JAK2-STAT5. We hypothesize that delivery of JAGGED1 and induction of its downstream NOTCH non-canonical signaling in pediatric human osteoblasts constitutes an effective bone regenerative treatment. Delivering pediatric human bone-derived osteoblast-like cells to an in vivo murine bone loss model of a critically sized cranial defect, we identified that JAGGED1 promotes human pediatric osteoblast commitment and bone formation through p70 S6K phosphorylation. This approach highlights the potential of JAGGED1 and its downstream activators as innovative treatments for pediatric CF bone loss.

Delivery of a Jagged1-PEG-MAL hydrogel with pediatric human bone cells regenerates critically sized craniofacial bone defects

Current treatments for congenital and acquired craniofacial (CF) bone abnormalities are limited and costly. Conventional methods involve surgical correction, short-term stabilization, and long-term bone grafting, which may include problematic allografts and limited autografts. While bone morphogenetic protein 2 (BMP2) has been used for bone regeneration, it can cause bone overgrowth and life-threatening inflammation. Bone marrow-derived mesenchymal stem cell therapies, though promising, are not Food and Drug Administration approved and are resource intensive. Thus, there is a need for effective, affordable, and less side-effect-prone bone regenerative therapies. Previous research demonstrated that JAGGED1 induces osteoblast commitment in murine cranial neural crest cells through a NOTCH-dependent non-canonical pathway involving JAK2–STAT5. We hypothesize that delivery of JAGGED1 and induction of its downstream NOTCH non-canonical signaling in pediatric human osteoblasts constitutes an effective bone regenerative treatment. Delivering pediatric human bone-derived osteoblast-like cells to an in vivo murine bone loss model of a critically sized cranial defect, we identified that JAGGED1 promotes human pediatric osteoblast commitment and bone formation through p70 S6K phosphorylation. This approach highlights the potential of JAGGED1 and its downstream activators as innovative treatments for pediatric CF bone loss.

Primary jugular foramen fibrous dysplasia: surgically nuanced video of extradural infratemporal transjugular approach. Illustrative case.

Fibrous dysplasia of the bone is a disease caused by a somatic GNAS mutation that affects craniofacial bones and can have a mass effect on different neurovascular structures. The authors present the first case of primary jugular foramen fibrous dysplasia with occlusion of the transverse and sigmoid sinuses. A 33-year-old man presented with a history of dizziness and occasional dysphagia over the past year. Magnetic resonance imaging showed a uniform enhanced mass in the left jugular foramen, with complete blockage of the adjacent transverse and sigmoid sinuses. The computed tomography scan revealed a cystic bone lesion of the jugular foramen. The patient underwent a gross-total removal of the tumor through an infratemporal transjugular approach with complete preservation of the lower cranial nerves. This first reported case of primary jugular foramen fibrous dysplasia highlights the importance of considering this diagnosis when evaluating jugular fossa lesions. Understanding the anatomy of the infratemporal and jugular fossae, along with proficiency in microsurgical techniques, is essential for removing such tumors while preserving cranial nerve functions and the patient's quality of life. https://thejns.org/doi/10.3171/CASE24396.

Enhancing craniofacial bone tissue engineering strategy: integrating rapid wet chemically synthesised bioactive glass with photopolymerized resins

BackgroundCraniofacial bone regeneration represents a dynamic area within tissue engineering and regenerative medicine. Central to this field, is the continual exploration of new methodologies for template fabrication, leveraging established bio ceramic materials, with the objective of restoring bone integrity and facilitating successful implant placements.MethodsPhotopolymerized templates were prepared using three distinct bio ceramic materials, specifically a wet chemically synthesized bioactive glass and two commercially sourced hydroxyapatite variants. These templates underwent comprehensive characterization to assess their physicochemical and mechanical attributes, employing techniques including Fourier transform infrared spectroscopy, scanning electron microscopy, and nano-computed tomography. Evaluation of their biocompatibility was conducted through interaction with primary human osteoblasts (hOB) and subsequent examination using scanning electron microscopy.ResultsThe results demonstrated that composite showed intramolecular hydrogen bonding interactions with the photopolymer, while computerized tomography unveiled the porous morphology and distribution within the templates. A relatively higher porosity percentage (31.55 ± 8.70%) and compressive strength (1.53 ± 0.11 MPa) was noted for bioactive glass templates. Human osteoblast cultured on bioactive glass showed higher viability compared to other specimens. Scanning micrographs of human osteoblast on templated showed cellular adhesion and the presence of filopodia and lamellipodia.ConclusionIn summary these templates have the potential to be used for alveolar bone regeneration in critical size defect. Photopolymerization of bioceramics may be an interesting technique for scaffolds fabrication for bone tissue engineering application but needs more optimization to overcome existing issues like the ideal ratio of the photopolymer to bioceramics.

Hierarchical Integration of Curcumin-Loaded CaCO3 Nanoparticles and Black Phosphorus Nanosheets in Core/Shell Nanofiber for Cranial Defect Repair.

Reconstruction and healing of large craniofacial bone defects are major clinical challenges due to high risk of chronic inflammation and reduced cell mineralization levels. Herein, a core-shell nanofiber-based implant with significant pro-osteogenesis capability for treating skull defects is reported, which is hierarchically integrated with curcumin-loaded calcium carbonate nanoparticles (CaCO3@Cur NPs) in the outer layers and black phosphorus nanosheets (BPNSs) in the core compartments. The radical alignment of the integrated nanocomponents allows the sequential in situ release of the therapeutic agents in a controlled manner after implantation. Curcumin can repolarize M1 macrophages into M2 phenotypes for anti-inflammation purposes. Meanwhile, the released calcium and phosphate ions can promote the biomineralization of hydroxyapatite at the defect site and facilitate bone regeneration. Evaluations on cranial defect-bearing rat models demonstrated that the electrospun fibers in the present study substantially promoted restoration of the damaged skulls and inhibited inflammation in the wound bed. This strategy provides a new idea for the treatment of skull defects in the clinic.

Challenges in managing ossifying fibroma of the maxilla: A case report

Ossifying fibroma (OF) is a rare benign bone tumor that tends to grow aggressively, particularly in craniofacial bones such as the upper and lower jaw. The diagnosis of OF is often challenging, due to its rare nature and ability to invade adjacent anatomical structures. We present a case of OF in a 35-year-old woman with recurrent complaints of a lump on the right side of the face. Imaging evaluation showed widespread lesions in the right maxilla. Multidisciplinary discussion was required to determine the best approach in the management of this case, focusing on complete excision of the tumor while preserving function and aesthetics. In the context of recurrent or extensive disease, reconstructive techniques using local or distant flaps can be important to restore postoperative structure and function. Close postoperative surveillance is required to monitor for recurrence and facilitate timely intervention. With a multidisciplinary approach and appropriate management strategies, challenging OF cases can be effectively managed, although more research is needed to improve understanding and outcomes in this condition.

Deciphering the spatial distribution of Gli1-lineage cells in dental, oral, and craniofacial regions

Abstract The craniofacial bone, crucial for protecting brain tissue and supporting facial structure, undergoes continuous remodeling through mesenchymal (MSCs) or skeletal stem cells (SSCs) in their niches. Gli1 is an ideal marker for labeling MSCs and osteoprogenitors in this region, and Gli1-lineage cells are identified as pivotal for bone growth, development, repair, and regeneration. Despite its significance, the distribution of Gli1-lineage cells across the dental, oral, and craniofacial (DOC) regions remains to be systematically explored. Utilizing tissue-clearing and light sheet fluorescence microscopy (LSFM) with a Gli1CreER; tdTomatoAi14 mouse model, we mapped the spatial distribution of Gli1-lineage cells throughout the skull, focusing on calvarial bones, sutures, bone marrow, teeth, periodontium, jaw bones, and the temporomandibular joint (TMJ). We found Gli1-lineage cells widespread in these areas, underscoring their significance in DOC regions. Additionally, we observed their role in repairing calvarial bone defects, providing novel insights into craniofacial biology and stem cell niches and enhancing our understanding of stem cells and their progeny’s behavior in vivo.

Cyclophilin D, regulator of the mitochondrial permeability transition, impacts bone development and fracture repair

Mitochondrial Permeability Transition Pore (MPTP) and its key positive regulator, Cyclophilin D (CypD), control activity of cell oxidative metabolism important for differentiation of stem cells of various lineages including osteogenic lineage. Our previous work (Sautchuk et al., 2022) showed that CypD gene, Ppif, is transcriptionally repressed during osteogenic differentiation by regulatory Smad transcription factors in BMP canonical pathway, a major driver of osteoblast (OB) differentiation. Such a repression favors closure of the MPTP, priming OBs to higher usage of mitochondrial oxidative metabolism. The physiological role of CypD/MPTP regulation was demonstrated by its inverse correlation with BMP signaling in aging and bone fracture healing in addition to the negative effect of CypD gain-of-function (GOF) on bone maintenance. Here we show evidence that CypD GOF also negatively affects bone development and growth as well as fracture healing in adult mice. Developing craniofacial and long bones presented with delayed ossification and decreased growth rate, respectively, whereas in fracture, bony callus volume was diminished. Given that Genome Wide Association Studies showed that PPIF locus is associated with both body height and bone mineral density, our new data provide functional evidence for the role of PPIF gene product, CypD, and thus MPTP in bone growth and repair.

Sclerostin Antibody-Loaded Dense Collagen Hydrogels Promote Critical-Size Bone Defect Repair.

The management of extensive bone loss remains a clinical challenge. Numerous studies are underway to develop a combination of biomaterials, biomolecules, and stem cells to address this challenge. In particular, the systemic administration of antibodies against sclerostin, a regulator of bone formation, was recently shown to enhance the bone repair efficiency of dense collagen hydrogels (DCHs) hosting murine dental pulp stem cells (mDPSCs). The aim of the present study was to assess whether these antibodies, encapsulated and released from DCHs, could promote craniofacial bone repair by the local inhibition of sclerostin. In vitro studies showed that antibody loading modified neither the hydrogel structure nor the viability of seeded mDPSCs. When implanted in a mouse calvaria critical-size bone defect, antibody-loaded DCHs showed repair capabilities similar to those of acellular unloaded DCHs combined with antibody injections. Importantly, the addition of mDPSCs provided no further benefit. Altogether, the local delivery of antisclerostin antibodies from acellular dense collagen scaffolds is highly effective for bone repair. The drastic reduction in the required amount of antibody compared to systemic injection should reduce the cost of the procedure, making the strategy proposed here a promising therapeutic approach for large bone defect repair.

Pediatric Scapular Cavernous Hemangioma Presenting with Kasabach–Merritt Syndrome and Controlled by Preoperative Angioembolization: A Case Report

AbstractIntraosseous hemangioma is a common and benign vascular tumor with a propensity to occur in any bone of the body. Although the most common skeletal sites are the craniofacial bones and the spine, hemangioma of the scapula is a rare occurrence, which is rarer in the pediatric population. Kasabach–Merritt syndrome (KMS) is characterized by the combination of a rapidly growing vascular tumor, thrombocytopenia, microangiopathic hemolytic anemia, and consumptive coagulopathy. This condition can cause massive bleeding even after minor trauma and immediate operative management is usually contraindicated.We report a rare case of KMS in an 8-year-old girl with cavernous hemangioma of the scapula presenting with progressively increasing shoulder mass, thrombocytopenia, and serially falling hemoglobin levels. She was treated with preoperative angioembolization to manage the consumptive coagulopathy followed by surgery poststabilization. Intraosseous hemangiomas, while benign, can show aggressive features on imaging and angioembolization can be a life-saving tool in the management of vascular tumors presenting with consumptive coagulopathy. Intraosseous hemangiomas, while benign, can show aggressive features on imaging and angioembolization can be a life-saving tool in the management of vascular tumors presenting with consumptive coagulopathy.

MR Imaging in Covid-19-Associated Invasive Fungal Sinusitis.

We witnessed a sharp peak in the incidence of invasive fungal sinusitis, particularly mucormycosis, in patients with history of coronavirus disease 2019 (COVID-19) infection in India. Rhino-orbito-cerebral mucormycosis (ROCM) is a fulminant rapidly progressive disease. Early diagnosis significantly improves patient survival and outcomes. Hence, neuroimaging plays a very important role. We studied the magnetic resonance (MR) imaging manifestations of invasive fungal sinusitis and established an imaging protocol, which helps in early diagnosis of the disease per se as well as its complications. We evaluated the differences between COVID-19-associated and non-COVID-19-associated ROCM. We retrospectively analyzed the MR imaging manifestations of 91 histopathologically proven cases of post-COVID-19-invasive fungal sinusitis. We observed stage I disease limited to sinuses in 25.2%, stage II disease with intraorbital spread in 23%, and stage III disease with intracranial spread in 51.6% of our patients. Dural involvement was the commonest and earliest sign of stage III disease. Direct parenchymal invasion from the adjacent paranasal sinuses was the commonest pattern of cerebral involvement, involving basifrontal lobe (14.2%) followed by anteromedial temporal lobe (5.4%). We observed orbital and intracranial complications including subperiosteal orbital abscess (1%), cavernous sinus involvement (29.6%), angioinvasion (15.3%), perineural spread (9.8%), and osteomyelitis of skull base and craniofacial bones (45%). Contrary to non-COVID-19-associated ROCM, we did not observe any case with superior ophthalmic vein/dural venous sinus thrombosis or basilar artery angioinvasion in our study. In our study, stage III disease was most commonly due to direct parenchymal invasion into frontal and temporal lobes from the adjacent frontal and sphenoid sinuses, respectively. The commonest vascular complications in our study were cavernous sinus involvement followed by angioinvasion into the cavernous ICA leading to watershed infarcts.

Malignant transformation of craniofacial fibrous dysplasia: A clinicopathological, immunohistochemical and molecular analysis of 15 cases in one single institution

ObjectiveMalignant transformation of craniofacial fibrous dysplasia (FD) is not common and its clinicopathological as well as molecular characteristics remain largely unknown with limited literature reports. Study designPatients diagnosed with FD including McCune-Albright syndrome (MAS), polyostotic fibrous dysplasia (PFD), and monostotic fibrous dysplasia (MFD), accompanied by malignant transformation at our institution over the past 18 years (2005–2023) were retrospectively screened and analyzed to investigate the epidemiology and clinicopathological features of these tumors. ResultsThree hundred and five patients were diagnosed as FD in our hospital from 2005 to 2023, with 176 females (57.7 %) and 129 males (42.3 %). The average age at diagnosis was 28.35 years, ranging from 7 to 70 years. A total number of 15 (4. 9 %) cases of FD with malignant transformation were selected. Among these 15 patients, the age of the initial diagnosis of FD ranged from 6 to 54 years (mean age 28.87 ± 16.77), and the ages when malignant transformation occurred ranged from 18 to 57 years (mean age 38.53 ± 13.05). Among 15 patients, 12 patients were female (80 %) and 3 were male (20 %). Fifteen cases included MSA in 2 patients, PFD in 4 patients, and MFD in 9 patients. Of the anatomical sites in craniofacial bones, the most common site of the lesion was the maxilla, followed by the mandible. Malignant neoplasm arising in FD were osteosarcoma (12/15), chondrosarcoma (1/15) and high-grade sarcoma of uncertain differentiation (2/15). The 3- and 5-year overall survival rate was 33.3 % (5/15) and 20 % (3/15) respectively. In secondary osteosarcoma from FD, MDM2 and CDK4 positivity were 33.3 % and 41.7 % respectively, and only one case was MDM2-amplified and CDK4-amplified. ConclusionMalignant transformation in fibrous dysplasia was an exceedingly rare event and with a female predominance. The overall survival rate was poor. Osteosarcoma was the most common malignant neoplasm arising in FD. MDM2 and CDK4 expression may aid in the diagnosis of secondary osteosarcoma in FD.

Fibrous Dysplasia: Clinicopathological Behavior and Its Management

Fibrous dysplasia (FD) is a rare disorder of bone in which normal bone is replaced by abnormal fibro-osseous tissue. It more commonly involves the long bones, craniofacial bones, ribs, and pelvis. The involvement of single bone is called as monostotic type of FD and if multiple bones are involved it is called polyostotic type of FD. Its occurrence is predominantly observed in teenagers, and it usually becomes static after adulthood. The incidence of monostotic FD (MFD) is four times more than that of polyostotic FD (PFD). The posterior region of the jaw bone is more prone and rarely crosses the midline. The malignant potential is 0.5% for cases that remain untreated. Here, we present a case of MFD involving the posterior region of the right side of the mandible in a 37-year-old female patient. The clinical diagnostic approach, different imaging modalities, and histological examination methods for definitive diagnosis have been elaborated. During the regular follow-up, the MFD lesion showed no obvious signs of progression or malignancy features.

Mandible development under gestational protein restriction: cellular and molecular mechanisms.

Insufficient evidence regarding how maternal undernutrition affects craniofacial bone development persists. With its unique focus on the impact of gestational protein restriction on calvaria and mandible osteogenesis, this study aims to fill, at least in part, this gap. Female mice were mated and randomized into NP (normal protein) or LP (low protein) groups. On the 18th gestational day (GD), male embryos were collected and submitted to microtomography (µCT), scanning electron microscopy (SEM), energy-dispersive X-ray spectroscopy (EDS), PCR, and autophagy dynamic analyses. The study shows that the LP offspring exhibited lower body mass than the NP group, with µCT analysis revealing no volumetric differences in fetus's head. EDS analysis showed lower calcium and higher phosphorus percentages in mandibles and calvaria. SEM assessment evidenced higher hydroxyapatite crystal-like (HC) deposition on the calvaria surface in LP fetus. Conversely, lower HC deposition was observed on the mandible surface, suggesting delayed matrix mineralization in LP fetuses with a higher percentage of collagen fibers in the mandible bone. The autophagy process was reduced in the mesenchyme of LP fetuses. PCR array analysis of 84 genes revealed 27 genes with differential expression in the LP progeny-moreover, increased mRNA levels of Akt1, Mtor, Nfkb, and Smad1 in the LP offspring. In conclusion, the results suggest that gestational protein restriction anticipated bone differentiation in utero, before 18GD, where this process is reduced compared to the control, leading to the reduction in bone area at 15 postnatal day previously observed. These findings provide insights into the molecular and cellular mechanisms of mandible development and suggest potential implications for the Developmental Origins of Health and Disease (DOHaD).

ENPP1 enzyme replacement therapy improves ectopic calcification but does not rescue skeletal phenotype in a mouse model for craniometaphyseal dysplasia.

Craniometaphyseal dysplasia (CMD) is a rare genetic bone disorder, characterized by progressive thickening of craniofacial bones and flared metaphyses of long bones. Craniofacial hyperostosis leads to the obstruction of neural foramina and neurological symptoms such as facial palsy, blindness, deafness, or severe headache. Mutations in ANKH (mouse ortholog ANK), a transporter of small molecules such as citrate and ATP, are responsible for autosomal dominant CMD. Knock-in (KI) mice carrying an ANKF377del mutation (AnkKI/KI ) replicate many features of human CMD. Pyrophosphate (PPi) levels in plasma are significantly reduced in AnkKI/KI mice. PPi is a potent inhibitor of mineralization. To examine the extent to which restoration of circulating PPi levels may prevent the development of a CMD-like phenotype, we treated AnkKI/KI mice with the recombinant human ENPP1-Fc protein IMA2a. ENPP1 hydrolyzes ATP into AMP and PPi. Male and female Ank+/+ and AnkKI/KI mice (n ≥ 6/group) were subcutaneously injected with IMA2a or vehicle weekly for 12wk, starting at the age of 1wk. Plasma ENPP1 activity significantly increased in AnkKI/KI mice injected with IMA2a (Vehicle/IMA2a: 28.15 ± 1.65/482.7 ± 331.2 mOD/min; p <.01), which resulted in the successful restoration of plasma PPi levels (Ank+/+ /AnkKI/KI vehicle treatment/AnkKI/KI IMA2a: 0.94 ± 0.5/0.43 ± 0.2/1.29 ± 0.8μM; p <.01). We examined the skeletal phenotype by X-Ray imaging and μCT. IMA2a treatment of AnkKI/KI mice did not significantly correct CMD features such as the abnormal shape of femurs, increased bone mass of mandibles, hyperostotic craniofacial bones, or the narrowed foramen magnum. However, μCT imaging showed ectopic calcification near basioccipital bones at the level of the foramen magnum and on joints of AnkKI/KI mice. Interestingly, IMA2a treatment significantly reduced the volume of calcified nodules at both sites. Our data demonstrate that IMA2a is sufficient to restore plasma PPi levels and reduce ectopic calcification but fails to rescue skeletal abnormalities in AnkKI/KI mice under our treatment conditions.

Beyond bone volume: Understanding tissue-level quality in healing of maxillary vs. femoral defects

Currently, principles of tissue engineering and implantology are uniformly applied to all bone sites, disregarding inherent differences in collagen, mineral composition, and healing rates between craniofacial and long bones. These differences could potentially influence bone quality during the healing process. Evaluating bone quality during healing is crucial for understanding local mechanical properties in regeneration and implant osseointegration. However, site-specific changes in bone quality during healing remain poorly understood. In this study, we assessed newly formed bone quality in sub-critical defects in the maxilla and femur, while impairing collagen cross-linking using β-aminopropionitrile (BAPN). Our findings revealed that femoral healing bone exhibited a 73 % increase in bone volume but showed significantly greater viscoelastic and collagen changes compared to surrounding bone, leading to increased deformation during long-term loading and poorer bone quality in early healing. In contrast, the healing maxilla maintained equivalent hardness and viscoelastic constants compared to surrounding bone, with minimal new bone formation and consistent bone quality. However, BAPN-impaired collagen cross-linking induced viscoelastic changes in the healing maxilla, with no further changes observed in the femur. These results challenge the conventional belief that increased bone volume correlates with enhanced tissue-level bone quality, providing crucial insights for tissue engineering and site-specific implant strategies. The observed differences in bone quality between sites underscore the need for a nuanced approach in assessing the success of regeneration and implant designs and emphasize the importance of exploring site-specific tissue engineering interventions. Statement of significanceAccurate measurement of bone quality is crucial for tissue engineering and implant therapies. Bone quality varies between craniofacial and long bones, yet it's often overlooked in the healing process. Our study is the first to comprehensively analyze bone quality during healing in both the maxilla and femur. Surprisingly, despite significant volume increase, femur healing bone had poorer quality compared to the surrounding bone. Conversely, maxilla healing bone maintained consistent quality despite minimal bone formation. Impaired collagen diminished maxillary healing bone quality, but had no further effect on femur bone quality. These findings challenge the notion that more bone volume equals better quality, offering insights for improving tissue engineering and implant strategies for different bone sites.