RESUMO
14-3-3 proteins are highly conserved regulatory proteins that interact with hundreds of structurally diverse clients and act as central hubs of signaling networks. However, how 14-3-3 paralogs differ in specificity and how they regulate client protein function are not known for most clients. Here, we map the interactomes of all human 14-3-3 paralogs and systematically characterize the effect of disrupting these interactions on client localization. The loss of 14-3-3 binding leads to the coalescence of a large fraction of clients into discrete foci in a client-specific manner, suggesting a central chaperone-like function for 14-3-3 proteins. Congruently, the engraftment of 14-3-3 binding motifs to nonclients can suppress their aggregation or phase separation. Finally, we show that 14-3-3s negatively regulate the localization of the RNA-binding protein SAMD4A to cytoplasmic granules and inhibit its activity as a translational repressor. Our work suggests that 14-3-3s have a more prominent role as chaperone-like molecules than previously thought.
Assuntos
Proteínas 14-3-3 , Proteínas de Choque Térmico HSP90 , Humanos , Proteínas 14-3-3/genética , Proteínas 14-3-3/metabolismo , Proteínas de Choque Térmico HSP90/metabolismo , Chaperonas Moleculares/metabolismo , Ligação ProteicaRESUMO
The primary cilium decorates most eukaryotic cells and regulates tissue morphogenesis and maintenance. Structural or functional defects of primary cilium result in ciliopathies, congenital human disorders affecting multiple organs. Pathogenic variants in the ciliogenesis and planar cell polarity effectors (CPLANE) genes FUZZY, INTU and WDPCP disturb ciliogenesis, causing severe ciliopathies in humans and mice. Here, we show that the loss of Fuzzy in mice results in defects of primary cilia, accompanied by increased RhoA activity and excessive actin polymerization at the basal body. We discovered that, mechanistically, Fuzzy interacts with and recruits the negative actin regulator ARHGAP35 (also known as p190A RhoGAP) to the basal body. We identified genetic interactions between the two genes and found that a mutant ArhGAP35 allele increases the severity of phenotypic defects observed in Fuzzy-/- mice. Based on our findings, we propose that Fuzzy regulates ciliogenesis by recruiting ARHGAP35 to the basal body, where the latter likely restricts actin polymerization and modifies the actin network. Our study identifies a mechanism whereby CPLANE proteins control both actin polymerization and primary cilium formation.
Assuntos
Actinas , Ciliopatias , Proteínas Ativadoras de GTPase , Camundongos , Humanos , Animais , Actinas/metabolismo , Cílios/metabolismo , PolimerizaçãoRESUMO
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.
Assuntos
Suturas Cranianas , Crânio , Camundongos , Animais , Células-Tronco , Crista Neural , MesodermaRESUMO
Down syndrome (DS), trisomy of human chromosome 21 (Hsa21), occurs in 1 in 800 live births and is the most common human aneuploidy. DS results in multiple phenotypes, including craniofacial dysmorphology, which is characterised by midfacial hypoplasia, brachycephaly and micrognathia. The genetic and developmental causes of this are poorly understood. Using morphometric analysis of the Dp1Tyb mouse model of DS and an associated mouse genetic mapping panel, we demonstrate that four Hsa21-orthologous regions of mouse chromosome 16 contain dosage-sensitive genes that cause the DS craniofacial phenotype, and identify one of these causative genes as Dyrk1a. We show that the earliest and most severe defects in Dp1Tyb skulls are in bones of neural crest (NC) origin, and that mineralisation of the Dp1Tyb skull base synchondroses is aberrant. Furthermore, we show that increased dosage of Dyrk1a results in decreased NC cell proliferation and a decrease in size and cellularity of the NC-derived frontal bone primordia. Thus, DS craniofacial dysmorphology is caused by an increased dosage of Dyrk1a and at least three other genes.
Assuntos
Síndrome de Down , Camundongos , Humanos , Animais , Síndrome de Down/genética , Crânio , Mapeamento Cromossômico , Fenótipo , Modelos Animais de DoençasRESUMO
Although rare, childhood (paediatric) cancers are a major cause of death in young children. Unlike many adult cancers, paediatric cancers, such as neuroblastoma (NB), are developmental diseases that rarely show genetic predispositions. NB is the most common extracranial solid tumour in children, accounting for â¼15% of paediatric cancer deaths. This heterogeneous cancer arises from undifferentiated neural crest-derived progenitor cells. As neural crest cells are multipotent and migratory, they are often considered the embryonic paradigm of cancer stem cells. However, very little is known about the events that trigger tumour initiation and progression. Here, we discuss recent insights into sympathoadrenal lineage specification, as well as genetic factors associated with NB. With this in mind, we consider the molecular underpinnings of NB in the context of developmental trajectories of the neural crest lineage. This allows us to compare distinct subtypes of the disease and gene-function interactions during sensitive phases of neural crest development.
Assuntos
Crista Neural , Neuroblastoma , Biomarcadores , Diferenciação Celular , Criança , Pré-Escolar , Humanos , Células-Tronco Neoplásicas/patologia , Neuroblastoma/genética , Neuroblastoma/patologia , NeurogêneseRESUMO
Musculoskeletal geometry and muscle volumes vary widely in the population and are intricately linked to the performance of tasks ranging from walking and running to jumping and sprinting. As an alternative to experimental approaches, where it is difficult to isolate factors and establish causal relationships, simulations can be used to independently vary musculoskeletal geometry and muscle volumes, and develop a fundamental understanding. However, our ability to understand how these parameters affect task performance has been limited due to the high computational cost of modelling the necessary complexity of the musculoskeletal system and solving the requisite multi-dimensional optimization problem. For example, sprinting and running are fundamental to many forms of sport, but past research on the relationships between musculoskeletal geometry, muscle volumes, and running performance has been limited to observational studies, which have not established cause-effect relationships, and simulation studies with simplified representations of musculoskeletal geometry. In this study, we developed a novel musculoskeletal simulator that is differentiable with respect to musculoskeletal geometry and muscle volumes. This simulator enabled us to find the optimal body segment dimensions and optimal distribution of added muscle volume for sprinting and marathon running. Our simulation results replicate experimental observations, such as increased muscle mass in sprinters, as well as a mass in the lower end of the healthy BMI range and a higher leg-length-to-height ratio in marathon runners. The simulations also reveal new relationships, for example showing that hip musculature is vital to both sprinting and marathon running. We found hip flexor and extensor moment arms were maximized to optimize sprint and marathon running performance, and hip muscles the main target when we simulated strength training for sprinters. Our simulation results provide insight to inspire future studies to examine optimal strength training. Our simulator can be extended to other athletic tasks, such as jumping, or to non-athletic applications, such as designing interventions to improve mobility in older adults or individuals with movement disorders.
Assuntos
Desempenho Atlético , Treinamento Resistido , Corrida , Humanos , Idoso , Corrida/fisiologia , Músculo Esquelético/fisiologia , Caminhada/fisiologia , Desempenho Atlético/fisiologiaRESUMO
The primary cilium is a hair-like projection that controls cell development and tissue homeostasis. Although accumulated studies identify the molecular link between cilia and cilia-related diseases, the underlying etiology of ciliopathies has not been fully understood. In this paper, we determine the function of Rab34, a small GTPase, as a key regulator for controlling ciliogenesis and type I collagen trafficking in craniofacial development. Mechanistically, Rab34 is required to form cilia that control osteogenic proliferation, survival, and differentiation via cilia-mediated Hedgehog signaling. In addition, Rab34 is indispensable for regulating type I collagen trafficking from the ER to the Golgi. These results demonstrate that Rab34 has both ciliary and non-ciliary functions to regulate osteogenesis. Our study highlights the critical function of Rab34, which may contribute to understanding the novel etiology of ciliopathies that are associated with the dysfunction of RAB34 in humans.
Assuntos
Cílios , Osteogênese , Proteínas rab de Ligação ao GTP , Cílios/metabolismo , Proteínas rab de Ligação ao GTP/metabolismo , Proteínas rab de Ligação ao GTP/genética , Animais , Camundongos , Humanos , Crânio/metabolismo , Proteínas Hedgehog/metabolismo , Diferenciação Celular , Colágeno Tipo I/metabolismo , Colágeno Tipo I/genética , Transdução de Sinais , Desenvolvimento Ósseo , Ossos Faciais/metabolismo , Ossos Faciais/crescimento & desenvolvimento , Ossos Faciais/embriologia , Proliferação de Células , Transporte Proteico , Complexo de Golgi/metabolismoRESUMO
The primary cilium is an essential organelle that is important for normal cell signalling during development and homeostasis but its role in pituitary development has not been reported. The primary cilium facilitates signal transduction for multiple pathways, the best-characterised being the SHH pathway, which is known to be necessary for correct pituitary gland development. FUZ is a planar cell polarity (PCP) effector that is essential for normal ciliogenesis, where the primary cilia of Fuz-/- mutants are shorter or non-functional. FUZ is part of a group of proteins required for recruiting retrograde intraflagellar transport proteins to the base of the organelle. Previous work has reported ciliopathy phenotypes in Fuz-/- homozygous null mouse mutants, including neural tube defects, craniofacial abnormalities, and polydactyly, alongside PCP defects including kinked/curly tails and heart defects. Interestingly, the pituitary gland was reported to be missing in Fuz-/- mutants at 14.5 dpc but the mechanisms underlying this phenotype were not investigated. Here, we have analysed the pituitary development of Fuz-/- mutants. Histological analyses reveal that Rathke's pouch (RP) is initially induced normally but is not specified and fails to express LHX3, resulting in hypoplasia and apoptosis. Characterisation of SHH signalling reveals reduced pathway activation in Fuz-/- mutant relative to control embryos, leading to deficient specification of anterior pituitary fate. Analyses of the key developmental signals FGF8 and BMP4, which are influenced by SHH, reveal abnormal patterning in the ventral diencephalon, contributing further to abnormal RP development. Taken together, our analyses suggest that primary cilia are required for normal pituitary specification through SHH signalling.
Assuntos
Polaridade Celular , Cílios , Animais , Camundongos , Cílios/fisiologia , Proteínas Hedgehog/metabolismo , Camundongos Knockout , Hipófise/metabolismo , Proteínas/metabolismoRESUMO
Walking balance is central to independent mobility, and falls due to loss of balance are a leading cause of death for people 65 years of age and older. Bipedal gait is typically unstable, but healthy humans use corrective torques to counteract perturbations and stabilize gait. Exoskeleton assistance could benefit people with neuromuscular deficits by providing stabilizing torques at lower-limb joints to replace lost muscle strength and sensorimotor control. However, it is unclear how applied exoskeleton torques translate to changes in walking kinematics. This study used musculoskeletal simulation to investigate how exoskeleton torques applied to the ankle and subtalar joints alter center of mass kinematics during walking. We first created muscle-driven walking simulations using OpenSim Moco by tracking experimental kinematics and ground reaction forces recorded from five healthy adults. We then used forward integration to simulate the effect of exoskeleton torques applied to the ankle and subtalar joints while keeping muscle excitations fixed based on our previous tracking simulation results. Exoskeleton torque lasted for 15% of the gait cycle and was applied between foot-flat and toe-off during the stance phase, and changes in center of mass kinematics were recorded when the torque application ended. We found that changes in center of mass kinematics were dependent on both the type and timing of exoskeleton torques. Plantarflexion torques produced upward and backward changes in velocity of the center of mass in mid-stance and upward and smaller forward velocity changes near toe-off. Eversion and inversion torques primarily produced lateral and medial changes in velocity in mid-stance, respectively. Intrinsic muscle properties reduced kinematic changes from exoskeleton torques. Our results provide mappings between ankle plantarflexion and inversion-eversion torques and changes in center of mass kinematics which can inform designers building exoskeletons aimed at stabilizing balance during walking. Our simulations and software are freely available and allow researchers to explore the effects of applied torques on balance and gait.
Assuntos
Tornozelo , Exoesqueleto Energizado , Adulto , Humanos , Torque , Fenômenos Biomecânicos/fisiologia , Caminhada/fisiologia , Marcha/fisiologiaRESUMO
BACKGROUND: The transition into residential aged care is frequently associated with a reduction in physical activity, social engagement, and emotional wellbeing. Our aim was to evaluate the impact of a 26-day international cycling competition (Road Worlds Competition for Seniors), incorporating elements of exercise, audiovisual cycling footage, social engagement, and gamification, on the physical, psychological, and social well-being of aged care residents. We aimed to use findings to inform the development of a multi-modal intervention model to maximise wellbeing for older adults. METHODS: Residents (N = 32) participated in a mixed-methods single-group intervention pilot study that compared pre-and post-competition measures for the following wellbeing domains; physical, psychological, and social. In addition, interviews were conducted with residents (n = 27) and staff (n = 6) to explore their experiences. RESULTS: Measures identified significant improvements across multiple wellbeing domains, including functional fitness, depression, self-efficacy, and social network sizes. Findings from the interview data indicated that the multimodal components involved in the program delivery were valued by staff and residents who enjoyed the gamification, audiovisual cycling footage, social engagement, opportunities for reminiscence, and camaraderie between peers, staff, and volunteers. CONCLUSIONS: Findings highlight a constellation of benefits across physical, psychological, and social domains of wellbeing and inform a model for innovative multidimensional programs in residential aged care. The benefits for residents with varying physical and cognitive abilities support the use of creative strategies that maximise inclusion and engagement for residents.
Assuntos
Instituição de Longa Permanência para Idosos , Humanos , Masculino , Feminino , Idoso , Projetos Piloto , Idoso de 80 Anos ou mais , Ciclismo/psicologia , Ciclismo/fisiologia , Exercício Físico/psicologia , Exercício Físico/fisiologia , Avaliação de Programas e Projetos de Saúde , Terapia por Exercício/métodos , Terapia por Exercício/psicologiaRESUMO
INTRODUCTION: Occupational therapists have vital roles in inpatient rehabilitation to focus on independence in activities of daily living. Occupational therapy interventions are uniquely designed to address goals of service users and can be delivered individually or in group formats. Group interventions promote service users' mutual support and enable therapists to increase frequency and intensity of service provision. Student-led programs have become an attractive model, benefiting students while providing positive treatment outcomes for service users. There is an emerging body of literature that explores service users' and students' satisfaction with student-led group models of practice within inpatient rehabilitation and occupational performance outcomes of service users participating in student-led programs. This study aimed to explore the satisfaction of service users and students in addition to the self-reported occupational performance outcomes of a student-led activities of daily living group program in inpatient rehabilitation. METHODS: Data from 33 service users and seven students were collected retrospectively. The intervention involved a student-led activities of daily living group program, consisting of three groups: breakfast, morning tea, and home readiness group. All service users and students completed unique satisfaction surveys at the conclusion of their participation in the student-led program. Service users completed a self-reported activities of daily living performance measure pre- and post-program. Findings were reported in descriptive statistics, and pre- and post-program data were compared with the Wilcoxon signed-rank test. RESULTS: All students were satisfied with the student-led program. Majority of service users were satisfied with all components of the student-led program. Median scores for self-reported performance increased significantly following the student-led program (P < 0.001). CONCLUSION: This study highlighted that service users and students were satisfied with the service delivery of a student-led activities of daily living group program. The program was effective in addressing self-reported performance for service users in inpatient rehabilitation. The findings from this study have potential to inform clinical practice on the implementation of student-led programs in occupational therapy settings.
Assuntos
Atividades Cotidianas , Pacientes Internados , Terapia Ocupacional , Satisfação do Paciente , Humanos , Terapia Ocupacional/organização & administração , Terapia Ocupacional/métodos , Feminino , Masculino , Pessoa de Meia-Idade , Adulto , Estudos Retrospectivos , Idoso , Adulto Jovem , Estudantes de Ciências da Saúde/psicologiaRESUMO
Proper temporal and spatial activation of stem cells relies on highly coordinated cell signaling. The primary cilium is the sensory organelle that is responsible for transmitting extracellular signals into a cell. Primary cilium size, architecture, and assembly-disassembly dynamics are under rigid cell cycle-dependent control. Using mouse incisor tooth epithelia as a model, we show that ciliary dynamics in stem cells require the proper functions of a cholesterol-binding membrane glycoprotein, Prominin-1 (Prom1/CD133), which controls sequential recruitment of ciliary membrane components, histone deacetylase, and transcription factors. Nuclear translocation of Prom1 and these molecules is particularly evident in transit amplifying cells, the immediate derivatives of stem cells. The absence of Prom1 impairs ciliary dynamics and abolishes the growth stimulation effects of sonic hedgehog (SHH) treatment, resulting in the disruption of stem cell quiescence maintenance and activation. We propose that Prom1 is a key regulator ensuring appropriate response of stem cells to extracellular signals, with important implications for development, regeneration, and diseases.
Assuntos
Antígeno AC133/metabolismo , Cílios/metabolismo , Incisivo/citologia , Antígeno AC133/genética , Animais , Núcleo Celular/metabolismo , Células Cultivadas , Humanos , Incisivo/metabolismo , Camundongos , Modelos Biológicos , Mutagênese Sítio-Dirigida , Transporte Proteico , Transdução de Sinais , Células-Tronco/citologia , Células-Tronco/metabolismoRESUMO
The Hedgehog pathway gene Gli1 has been proposed to mark a subpopulation of skeletal stem cells (SSCs) in craniofacial bone. Skeletal stem cells (SSCs) are multi-potent cells crucial for the development and homeostasis of bone. Recent studies on long bones have suggested that skeletal stem cells in endochondral or intramembranous ossification sites have different differentiation capacities. However, this has not been well-defined in neural crest derived bones. Generally, the long bones are derived from mesoderm and follow an endochondral ossification model, while most of the cranial bones are neural crest (NC) in origin and follow an intramembranous ossification model. The mandible is unique: It is derived from the neural crest lineage but makes use of both modes of ossification. Early in fetal development, the mandibular body is generated by intramembranous ossification with subsequent endochondral ossification forming the condyle. The identities and properties for SSCs in these two sites remain unknown. Here, we use genetic lineage tracing in mouse to identify cells expressing the Hedgehog responsive gene Gli1, which is thought to mark the tissue resident SSCs. We track the Gli1+ cells, comparing cells within the perichondrium to those in the periosteum covering the mandibular body. In juvenile mice, these have distinct differentiation and proliferative potential. We also assess the presence of Sox10+ cells, thought to mark neural crest stem cells, but find no substantial population associated with the mandibular skeleton, suggesting that Sox10+ cells have limited contribution to maintaining postnatal mandibular bone. All together, our study indicates that the Gli1+ cells display distinct and limited differentiation capacity dependent on their regional associations.
Assuntos
Proteínas Hedgehog , Osteogênese , Camundongos , Animais , Proteína GLI1 em Dedos de Zinco/metabolismo , Proteínas Hedgehog/metabolismo , Mandíbula/metabolismo , Crânio , Crista NeuralRESUMO
BACKGROUND: Volar plate injuries are a common hand injury and complications associated with this injury such as a fixed flexion deformity, persistent pain and oedema can have a significant impact on a person's function. The literature reports these injuries are treated using various splinting materials such as thermoplastic, in varying degrees of proximal interphalangeal joint flexion or buddy loops. Despite volar plate injuries being reported as common, optimal non-surgical treatment of these injuries remains unclear. This study aims to investigate whether a dorsal blocking orthosis in a neutral position (00) is more effective than buddy loops for a volar plate injury to the proximal interphalangeal joint in preventing a fixed flexion deformity, reducing pain, managing oedema, and promoting function. METHODS: This study is a single-centre, prospective parallel-group, single blinded (assessor), randomised clinical trial. Patients between 18-65 years, who have sustained a volar plate injury to a single digit, have adequate cognitive functioning and give written informed consent will be invited to participate in this study. Patients will be randomised to either the control group where they will be fitted with buddy loops and commence early active motion exercises or the experimental group where they will receive a dorsal thermoplastic orthosis in a neutral position and commence early active motion exercises. The primary outcome measure is passive proximal interphalangeal joint extension and secondary outcome measures include passive range of motion, total passive motion, active range of motion, total active motion, grip strength, oedema, pain, function and adherence to treatment. Assessments will be completed until 8 weeks following commencement of treatment. The sample size calculation indicates that 23 patients is required in each group. With an expected dropout rate of 25% a total of 32 patients will be enrolled in each group. DISCUSSION: This study will assist in trying to improve treatment of volar plate injuries and assist in reducing complications associated with volar plate injuries, potentially reducing the need for prolonged hand therapy. TRIAL REGISTRATION: This trial has been registered with the Australian New Zealand Clinical Trials Registry (ACTRN12622001425785p). Ethical approval has been granted by the South Eastern Sydney Local Health District ethical committee (2022/ETH01697).
Assuntos
Braquetes , Contratura , Humanos , Estudos Prospectivos , Austrália , Aparelhos Ortopédicos , Extremidades , Ensaios Clínicos Controlados Aleatórios como AssuntoRESUMO
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.
Assuntos
Cateninas/genética , Fenda Labial/genética , Fissura Palatina/genética , Anormalidades Craniofaciais/genética , Ectrópio/genética , Cardiopatias Congênitas/genética , Anormalidades Dentárias/genética , Adolescente , Adulto , Animais , Anodontia/diagnóstico por imagem , Anodontia/genética , Anodontia/fisiopatologia , Criança , Pré-Escolar , Fenda Labial/diagnóstico por imagem , Fenda Labial/fisiopatologia , Fissura Palatina/diagnóstico por imagem , Fissura Palatina/fisiopatologia , Anormalidades Craniofaciais/diagnóstico por imagem , Anormalidades Craniofaciais/fisiopatologia , Modelos Animais de Doenças , Ectrópio/diagnóstico por imagem , Ectrópio/fisiopatologia , Feminino , Predisposição Genética para Doença , Cardiopatias Congênitas/diagnóstico por imagem , Cardiopatias Congênitas/fisiopatologia , Humanos , Masculino , Camundongos , Anormalidades Dentárias/diagnóstico por imagem , Anormalidades Dentárias/fisiopatologia , Xenopus , Adulto Jovem , delta CateninaRESUMO
The tongue is a highly specialised muscular organ with a complex anatomy required for normal function. We have utilised multiple genetic approaches to investigate local temporospatial requirements for sonic hedgehog (SHH) signalling during tongue development. Mice lacking a Shh cis-enhancer, MFCS4 (ShhMFCS4/-), with reduced SHH in dorsal tongue epithelium have perturbed lingual septum tendon formation and disrupted intrinsic muscle patterning, with these defects reproduced following global Shh deletion from E10.5 in pCag-CreERTM; Shhflox/flox embryos. SHH responsiveness was diminished in local cranial neural crest cell (CNCC) populations in both mutants, with SHH targeting these cells through the primary cilium. CNCC-specific deletion of orofaciodigital syndrome 1 (Ofd1), which encodes a ciliary protein, in Wnt1-Cre; Ofdfl/Y mice led to a complete loss of normal myotube arrangement and hypoglossia. In contrast, mesoderm-specific deletion of Ofd1 in Mesp1-Cre; Ofdfl/Y embryos resulted in normal intrinsic muscle arrangement. Collectively, these findings suggest key temporospatial requirements for local SHH signalling in tongue development (specifically, lingual tendon differentiation and intrinsic muscle patterning through signalling to CNCCs) and provide further mechanistic insight into the tongue anomalies seen in patients with disrupted hedgehog signalling.
Assuntos
Padronização Corporal , Proteínas Hedgehog/metabolismo , Crista Neural/citologia , Transdução de Sinais , Língua/embriologia , Alelos , Animais , Proliferação de Células , Elementos Facilitadores Genéticos , Feminino , Deleção de Genes , Regulação da Expressão Gênica no Desenvolvimento , Proteínas Hedgehog/genética , Heterozigoto , Ligantes , Mesoderma/metabolismo , Camundongos , Morfogênese/genética , Fenótipo , Proteínas/metabolismo , Tendões/metabolismo , Fatores de Tempo , Fator de Crescimento Transformador beta/metabolismo , Proteína Wnt1/metabolismoRESUMO
The maintenance of human skeletal stem cells (hSSCs) and their progeny in bone defects is a major challenge. Here, we report on a transplantable bandage containing a three-dimensional Wnt-induced osteogenic tissue model (WIOTM). This bandage facilitates the long-term viability of hSSCs (8 weeks) and their progeny, and enables bone repair in an in vivo mouse model of critical-sized calvarial defects. The newly forming bone is structurally comparable to mature cortical bone and consists of human and murine cells. Furthermore, we show that the mechanism of WIOTM formation is governed by Wnt-mediated asymmetric cell division of hSSCs. Covalently immobilizing Wnts onto synthetic materials can polarize single dividing hSSCs, orient the spindle and simultaneously generate a Wnt-proximal hSSC and a differentiation-prone Wnt-distal cell. Our results provide insight into the regulation of human osteogenesis and represent a promising approach to deliver human osteogenic constructs that can survive in vivo and contribute to bone repair.
Assuntos
Osso e Ossos/citologia , Divisão Celular , Osteogênese , Crânio/citologia , Células-Tronco/citologia , Engenharia Tecidual/métodos , Proteínas Wnt/metabolismo , Animais , Humanos , Camundongos , Crânio/fisiologiaRESUMO
Muscle stem cells (MuSCs) are requisite for skeletal muscle regeneration and homeostasis. Proper functioning of MuSCs, including activation, proliferation, and fate decision, is determined by an orchestrated series of events and communication between MuSCs and their niche. A multitude of biochemical stimuli are known to regulate MuSC fate and function. However, in addition to biochemical factors, it is conceivable that MuSCs are subjected to mechanical forces during muscle stretch-shortening cycles because of myofascial connections between MuSCs and myofibers. MuSCs respond to mechanical forces in vitro, but it remains to be proven whether physical forces are also exerted on MuSCs in their native niche and whether they contribute to the functioning and fate of MuSCs. MuSC deformation in their native niche resulting from mechanical loading of ex vivo myofiber bundles was visualized utilizing mT/mG double-fluorescent Cre-reporter mouse and multiphoton microscopy. MuSCs were subjected to 1 h pulsating fluid shear stress (PFSS) with a peak shear stress rate of 6.5 Pa/s. After PFSS treatment, nitric oxide, messenger RNA (mRNA) expression levels of genes involved in regulation of MuSC proliferation and differentiation, ERK 1/2, p38, and AKT activation were determined. Ex vivo stretching of extensor digitorum longus and soleus myofiber bundles caused compression as well as tensile and shear deformation of MuSCs in their niche. MuSCs responded to PFSS in vitro with increased nitric oxide production and an upward trend in iNOS mRNA levels. PFSS enhanced gene expression of c-Fos, Cdk4, and IL-6, whereas expression of Wnt1, MyoD, Myog, Wnt5a, COX2, Rspo1, Vangl2, Wnt10b, and MGF remained unchanged. ERK 1/2 and p38 MAPK signaling were also upregulated after PFSS treatment. We conclude that MuSCs in their native niche are subjected to force-induced deformations due to myofiber stretch-shortening. Moreover, MuSCs are mechanoresponsive, as evidenced by PFSS-mediated expression of factors by MuSCs known to promote proliferation.
Assuntos
Músculo Esquelético , Mioblastos , Animais , Diferenciação Celular , Expressão Gênica , Camundongos , Estresse MecânicoRESUMO
The vertebrate tongue is a complex muscular organ situated in the oral cavity and involved in multiple functions including mastication, taste sensation, articulation and the maintenance of oral health. Although the gross embryological contributions to tongue formation have been known for many years, it is only relatively recently that the molecular pathways regulating these processes have begun to be discovered. In particular, there is now evidence that the Hedgehog, TGF-Beta, Wnt and Notch signaling pathways all play an important role in mediating appropriate signaling interactions between the epithelial, cranial neural crest and mesodermal cell populations that are required to form the tongue. In humans, a number of congenital abnormalities that affect gross morphology of the tongue have also been described, occurring in isolation or as part of a developmental syndrome, which can greatly impact on the health and well-being of affected individuals. These anomalies can range from an absence of tongue formation (aglossia) through to diminutive (microglossia), enlarged (macroglossia) or bifid tongue. Here, we present an overview of the gross anatomy and embryology of mammalian tongue development, focusing on the molecular processes underlying formation of the musculature and connective tissues within this organ. We also survey the clinical presentation of tongue anomalies seen in human populations, whilst considering their developmental and genetic etiology.
Assuntos
Tecido Conjuntivo/embriologia , Músculos/embriologia , Crista Neural/embriologia , Língua/embriologia , Animais , Tecido Conjuntivo/anatomia & histologia , Tecido Conjuntivo/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Mamíferos/anatomia & histologia , Mamíferos/embriologia , Mamíferos/genética , Músculos/citologia , Músculos/metabolismo , Crista Neural/citologia , Crista Neural/metabolismo , Organogênese/genética , Transdução de Sinais/genética , Língua/citologia , Língua/metabolismoRESUMO
Neuroblastoma is a common extracranial solid tumour of childhood, responsible for 15% of cancer-related deaths in children. Prognoses vary from spontaneous remission to aggressive disease with extensive metastases, where treatment is challenging. Tumours are thought to arise from sympathoadrenal progenitor cells, which derive from an embryonic cell population called neural crest cells that give rise to diverse cell types, such as facial bone and cartilage, pigmented cells, and neurons. Tumours are found associated with mature derivatives of neural crest, such as the adrenal medulla or paraspinal ganglia. Sympathoadrenal progenitor cells express anaplastic lymphoma kinase (ALK), which encodes a tyrosine kinase receptor that is the most frequently mutated gene in neuroblastoma. Activating mutations in the kinase domain are common in both sporadic and familial cases. The oncogenic role of ALK has been extensively studied, but little is known about its physiological role. Recent studies have implicated ALK in neural crest migration and sympathetic neurogenesis. However, very few downstream targets of ALK have been identified. Here, we describe pathological activation of ALK in the neural crest, which promotes proliferation and migration, while preventing differentiation, thus inducing the onset of neuroblastoma. Understanding the effects of ALK activity on neural crest cells will help find new targets for neuroblastoma treatment.