rodent-model-masseter-volumetric-muscle-loss-for-studying

Rodent Model of Masseter Volumetric Muscle Loss for Studying Bioengineering Materials

Craniofacial volumetric muscle loss (VML) injuries can occur as a result of severe trauma, surgical excision, inflammation, and congenital or other ...

University of California San Francisco (UCSF)

Identification of human satellite cells that fulfill muscle stem cell criteria is an unmet need in regenerative medicine. This hurdle limits understanding how closely muscle stem cell properties are conserved among mice and humans and hampers translational efforts in muscle regeneration. Here, we report that PAX7 satellite cells exist at a consistent frequency of 2-4 cells/mm of fiber in muscles of the human trunk, limbs, and head. Xenotransplantation into mice of 50-70 fiber-associated, or 1,000-5,000 FACS-enriched CD56(+)/CD29(+) human satellite cells led to stable engraftment and formation of human-derived myofibers. Human cells with characteristic PAX7, CD56, and CD29 expression patterns populated the satellite cell niche beneath the basal lamina on the periphery of regenerated fibers. After additional injury, transplanted satellite cells robustly regenerated to form hundreds of human-derived fibers. Together, these findings conclusively delineate a source of bona-fide endogenous human muscle stem cells that will aid development of clinical applications.

Human Satellite Cell Transplantation and Regeneration from Diverse Skeletal Muscles.

Although human induced pluripotent stem cells (hiPSCs) hold great potential for the study of human diseases affecting disparate cell types, they have been underutilized in seeking mechanistic insights into the pathogenesis of congenital craniofacial disorders. Craniofrontonasal syndrome (CFNS) is a rare X-linked disorder caused by mutations in EFNB1 and characterized by craniofacial, skeletal, and neurological anomalies. Heterozygous females are more severely affected than hemizygous males, a phenomenon termed cellular interference that involves mosaicism for EPHRIN-B1 function. Although the mechanistic basis for cellular interference in CFNS has been hypothesized to involve Eph/ephrin-mediated cell segregation, no direct evidence for this has been demonstrated. Here, by generating hiPSCs from CFNS patients, we demonstrate that mosaicism for EPHRIN-B1 expression induced by random X inactivation in heterozygous females results in robust cell segregation in human neuroepithelial cells, thus supplying experimental evidence that Eph/ephrin-mediated cell segregation is relevant to pathogenesis in human CFNS patients.

EPHRIN-B1 Mosaicism Drives Cell Segregation in Craniofrontonasal Syndrome hiPSC-Derived Neuroepithelial Cells.

Satellite cells are mononucleated cells of the skeletal muscle lineage that exist beneath the basal lamina juxtaposed to the sarcolemma of skeletal muscle fibers. It is widely accepted that satellite cells mediate skeletal muscle regeneration. Within the satellite cell pool of adult muscle are skeletal muscle stem cells (MuSCs), also called satellite stem cells, which fulfill criteria of tissue stem cells: They proliferate and their progeny either occupies the adult MuSC niche during self-renewal or differentiates to regenerate mature muscle fibers. Here, we describe robust methods for the isolation of enriched populations of human satellite cells containing MuSCs from fresh human muscle, utilizing mechanical and enzymatic dissociation and purification by fluorescence-activated cell sorting. We also describe a process for xenotransplantation of human satellite cells into mouse muscle by injection into irradiated, immunodeficient, mouse leg muscle with concurrent notexin or bupivacaine muscle injury to increase engraftment efficiency. The engraftment of human MuSCs and the formation of human muscle can then be analyzed by histological and immunofluorescence staining, or subjected to in vivo experimentation.

Human Satellite Cell Isolation and Xenotransplantation.

Investigation of human muscle regeneration requires robust methods to purify and transplant muscle stem and progenitor cells that collectively constitute the human satellite cell (HuSC) pool. Existing approaches have yet to make HuSCs widely accessible for researchers, and as a result human muscle stem cell research has advanced slowly. Here, we describe a robust and predictable HuSC purification process that is effective for each human skeletal muscle tested and the development of storage protocols and transplantation models in dystrophin-deficient and wild-type recipients. Enzymatic digestion, magnetic column depletion, and 6-marker flow-cytometric purification enable separation of 10 highly enriched HuSCs per gram of muscle. Cryostorage of HuSCs preserves viability, phenotype, and transplantation potential. Development of enhanced and species-specific transplantation protocols enabled serial HuSC xenotransplantation and recovery. These protocols and models provide an accessible system for basic and translational investigation and clinical development of HuSCs.

High-Yield Purification, Preservation, and Serial Transplantation of Human Satellite Cells.

Little is known about the risk factors for facial fractures among homeless patients. We investigated the association between homelessness, mechanism of injury, and type of facial fracture in patients treated at an urban trauma center.

A Cross-sectional Study of the Association between Homelessness and Facial Fractures.

Stem cell behavior is tightly regulated by spatiotemporal signaling from the niche, which is a four-dimensional microenvironment that can instruct stem cells to remain quiescent, self-renew, proliferate, or differentiate. In this review, we discuss recent advances in understanding the signaling cues provided by the stem cell niche in two contrasting adult tissues, the rapidly cycling intestinal epithelium and the slowly renewing skeletal muscle. Drawing comparisons between these two systems, we discuss the effects of niche-derived growth factors and signaling molecules, metabolic cues, the extracellular matrix and biomechanical cues, and immune signals on stem cells. We also discuss the influence of the niche in defining stem cell identity and function in both normal and pathophysiologic states.

From gut to glutes: The critical role of niche signals in the maintenance and renewal of adult stem cells.

Craniofacial clinics are composed of multidisciplinary teams of providers to deliver coordinated and comprehensive patient care. The coronavirus disease of 2019 (COVID-19) pandemic has disrupted this model, as social distancing guidelines have precluded in-person patient appointments and forced clinics to reconsider their method of care delivery. The University of California, San Francisco, Craniofacial Center has continued to serve patients during this acute period, adopting a hybrid model in which the vast majority of patients are seen through telehealth and a limited number of patients are evaluated in-person. Surveyed patients and families reported high rates of satisfaction, with time savings cited as a particular benefit. Furthermore, most felt comfortable using the video technology required for their appointment. This experience has demonstrated to us that multidisciplinary craniofacial evaluations can be effectively delivered in a telehealth format and has informed our conception of idealized clinic structure. Moving forward, we intend to utilize telehealth visits for selected components of craniofacial evaluations in an effort to maximize efficiency and minimize burden, including addressing barriers to accessing care. Benefits of a hybrid model will include decongestion of clinics and waiting areas, allowing social distancing, addressing clinic space limits, and increased efficiency by eliminating the need for patient and family movement. Demonstration of the safety and efficacy of telehealth visits, combined with regulatory reform that improves reimbursement and allows for appointments across state lines, will be critical for this model to persist beyond the pandemic.

Transitioning Multidisciplinary Craniofacial Care to Telehealth during the COVID-19 Pandemic: A Single Center Experience.

This study sought to identify disparities in the timing of alveolar bone grafting (ABG) surgery and the replacement strategy for missing maxillary lateral incisors for patients with clefts. A retrospective record review identified patients who underwent ABG. Multivariable regression analyzed the independent contribution of each variable. This institutional study was performed at the University of California, San Francisco. Patients who presented under age 12 and underwent secondary ABG between 2012 and 2020 (n = 160). The age at secondary ABG and the recommended dental replacement treatment for each patient, either dental implantation or canine substitution. The average age at ABG was 10.8 ± 2.1 years, 106 (66.3%) patients were not White, and 80 (50.0%) had private insurance. Independent predictors of older age at ABG included an income below $ 50 000 as estimated from ZIP code (β = 15.0 months, 95% CI, 5.7-24.3,  = .002) and identifying as a race other than White (β = 10.1 months, 95% CI, 2.1-18.0,  = .01). After ABG, patients were more likely to undergo dental implantation over canine substitution if they were female (odds ratio [OR] = 4.3, 95% CI, 1.3-17.1,  = .02) or had private insurance (OR = 12.5, 95% CI, 2.2-143.2,  = .01). Patients who were low-income or not White experienced delays in ABG, whereas dental implantation was more likely to be recommended for patients with private insurance. Understanding the sources of disparities in dental reconstruction of cleft deformities may reveal opportunities to improve equity.

Disparities in Timing of Alveolar Bone Grafting and Dental Reconstruction in Patients With Clefts.

Strip craniectomy with orthotic helmet therapy (SCOT) is an increasingly supported treatment for metopic craniosynostosis, although the long-term efficacy of deformity correction remains poorly defined. We compared the longterm outcomes of SCOT versus open cranial vault reconstruction (OCVR).

Superior Long-term Appearance of Strip Craniectomy Compared with Cranial Vault Reconstruction in Metopic Craniosynostosis.

The purpose of this study was to identify racial and socioeconomic disparities in craniosynostosis evaluation and treatment, from referral to surgery. Patients diagnosed with craniosynostosis between 2012 and 2020 at a single center were identified. Chart review was used to collect demographic variables, age at referral to craniofacial care, age at diagnosis, age at surgery, and surgical technique (open versus limited incision). Multivariable linear and logistic regression models with lasso regularization assessed the independent effect of each variable. A total of 298 patients were included. Medicaid insurance was independently associated with a delay in referral of 83 days [95% confidence interval (CI) 4-161, P=0.04]. After referral, patients were diagnosed a median of 21 days later (interquartile range 7-40), though this was significantly prolonged in patients who were not White (β 23 d, 95% CI 9-38, P=0.002), had coronal synostosis (β 24 d, 95% CI 2-46, P=0.03), and had multiple suture synostosis (β 47 d, 95% CI 27-67, P<0.001). Medicaid insurance was also independently associated with diagnosis over 3 months of age (risk ratio 1.3, 95% CI 1.1-1.4, P=0.002) and undergoing surgery over 1 year of age (risk ratio 3.9, 95% CI 1.1-9.4, P=0.04). In conclusion, Medicaid insurance was associated with a 3-month delay in referral to craniofacial specialists and increased risk of diagnosis over 3 months of age, limiting surgical treatment options in this group. Patients with Medicaid also faced a 4-fold greater risk of delayed surgery, which could result in neurodevelopmental sequelae.

Racial and Socioeconomic Disparities in Prompt Craniosynostosis Workup and Treatment.

To examine linguistic disparities between English- and Spanish-speaking patients in access to care, satisfaction, and telehealth appointment attendance.

Article	Year
Human Satellite Cell Transplantation and Regeneration from Diverse Skeletal Muscles. Stem cell reports\| PubMed ID: 26352798	2015
EPHRIN-B1 Mosaicism Drives Cell Segregation in Craniofrontonasal Syndrome hiPSC-Derived Neuroepithelial Cells. Stem cell reports\| PubMed ID: 28238796	2017
Human Satellite Cell Isolation and Xenotransplantation. Methods in molecular biology (Clifton, N.J.)\| PubMed ID: 28842905	2017
High-Yield Purification, Preservation, and Serial Transplantation of Human Satellite Cells. Stem cell reports\| PubMed ID: 29478895	2018
A Cross-sectional Study of the Association between Homelessness and Facial Fractures. Plastic and reconstructive surgery. Global open\| PubMed ID: 31624672	2019
From gut to glutes: The critical role of niche signals in the maintenance and renewal of adult stem cells. Current opinion in cell biology\| PubMed ID: 32036295	2020
Transitioning Multidisciplinary Craniofacial Care to Telehealth during the COVID-19 Pandemic: A Single Center Experience. Plastic and reconstructive surgery. Global open\| PubMed ID: 33133981	2020
Disparities in Timing of Alveolar Bone Grafting and Dental Reconstruction in Patients With Clefts. The Cleft palate-craniofacial journal : official publication of the American Cleft Palate-Craniofacial Association\| PubMed ID: 35044260	2023
Superior Long-term Appearance of Strip Craniectomy Compared with Cranial Vault Reconstruction in Metopic Craniosynostosis. Plastic and reconstructive surgery. Global open\| PubMed ID: 35169528	2022
Racial and Socioeconomic Disparities in Prompt Craniosynostosis Workup and Treatment. The Journal of craniofacial surgery\| PubMed ID: 36409867	2022
Analyzing Linguistic Disparities in Telehealth Care Outcomes at a Multidisciplinary Craniofacial Center. The Cleft palate-craniofacial journal : official publication of the American Cleft Palate-Craniofacial Association\| PubMed ID: 37605606	2023

Jason Pomerantz

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