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Mesenchymal stem cells (MSCs) reside in niches to maintain tissue homeostasis and contribute to repair and regeneration. Although the physiological functions of blood and lymphatic vasculature are well studied, their regulation of MSCs as niche components remains largely unknown. Using adult mouse incisors as a model, we uncover the role of Trp53 in regulating vascular composition through THBS2 to maintain mesenchymal tissue homeostasis. Loss of Trp53 in GLI1+ progeny increases arteries and decreases other vessel types. Platelet-derived growth factors from arteries deposit in the MSC region and interact with PDGFRA and PDGFRB. Significantly, PDGFRA+ and PDGFRB+ cells differentially contribute to defined cell lineages in the adult mouse incisor. Collectively, our results highlight Trp53's importance in regulating the vascular niche for MSCs. They also shed light on how different arterial cells provide unique cues to regulate MSC subpopulations and maintain their heterogeneity. Furthermore, they provide mechanistic insight into MSC-vasculature crosstalk.
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Incisivo , Células Madre Mesenquimatosas , Transducción de Señal , Proteína p53 Supresora de Tumor , Animales , Células Madre Mesenquimatosas/metabolismo , Células Madre Mesenquimatosas/citología , Ratones , Proteína p53 Supresora de Tumor/metabolismo , Incisivo/citología , Incisivo/metabolismo , Factor de Crecimiento Derivado de Plaquetas/metabolismo , Receptor beta de Factor de Crecimiento Derivado de Plaquetas/metabolismo , Receptor alfa de Factor de Crecimiento Derivado de Plaquetas/metabolismoRESUMEN
This study aims to investigate the effect of Buyang Huanwu Decoction on blood flow recovery and arteriogenesis after hindlimb ischemia in mice via the platelet-derived growth factor(PDGF) signaling pathway. Forty C57BL/6 mice were randomized into model(clean water, 10 mL·kg~(-1)·d~(-1)), beraprost sodium(positive control, 18 µg·kg~(-1)·d~(-1)), and low-, medium-, and high-dose(10, 20, and 40 g·kg~(-1)·d~(-1), respectively) Buyang Huanwu Decoction groups(n=8). The hindlimb ischemia model was established by femoral artery ligation. The mice were administrated with corresponding agents by gavage daily for 14 days after ligation. For laser Doppler perfusion imaging, the mice were anesthetized and measured under a Periscan PSI imager. The density of capillary and arterio-le in the ischemic gastrocnemius was measured using immunofluorescence staining of the frozen tissue sections. Western blot was employed to determine the expression of PDGF subunit B(PDGFB), phosphorylated mitogen extracellular kinase(p-MEK), MEK, phosphorylated extracellular signal-regulated kinase(p-ERK), and ERK. Real-time PCR was employed to determine the mRNA level of PDGFB. The Buyang Huanwu Decoction-containing serum was used to treat the vascular smooth muscle cells(VSMCs) in hypoxia at doses of 10% and 20%. The proliferation and migration of VSMCs was assessed in vitro. The results showed that compared with the model group, beraprost sodium and Buyang Huanwu Decoction enhanced the blood flow recovery, increased the capillary and arteriole density, and up-regulated the protein levels of PDGFB, p-MEK, p-ERK, and mRNA levels of PDGFB, with the medium-dose Buyang Huanwu Decoction demonstrating the most significant effect. The 10% Buyang Huanwu Decoction-containing serum enhanced the proliferation and migration of VSMCs. Our findings demonstrate that Buyang Huanwu Decoction up-regulates PDGFB transcription and activates PDGF signaling pathway to promote arteriogenesis and blood flow recovery in ischemic gastrocnemius.
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Medicamentos Herbarios Chinos , Ratas , Ratones , Animales , Ratas Sprague-Dawley , Proteínas Proto-Oncogénicas c-sis , Ratones Endogámicos C57BL , Medicamentos Herbarios Chinos/uso terapéutico , Transducción de Señal , Isquemia/tratamiento farmacológico , Miembro Posterior/metabolismo , ARN Mensajero/metabolismo , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismoRESUMEN
Platelet-derived growth factor (PDGF)-induced changes in vascular smooth muscle cells (VSMCs) stimulate vascular remodeling, resulting in vascular diseases such as pulmonary arterial hypertension. VSMCs communicate with endothelial cells through extracellular vesicles (EVs) carrying cargos, including microRNAs. To understand the molecular mechanisms through which PDGF-stimulated pulmonary artery smooth muscle cells (PASMCs) interact with pulmonary artery endothelial cells (PAECs) under pathological conditions, we investigated the crosstalk between PASMCs and PAECs via extracellular vesicle miR-409-5p under PDGF stimulation. miR-409-5p expression was upregulated in PASMCs upon PDGF signaling, and it was released into EVs. The elevated expression of miR-409-5p was transported to PAECs and led to their impaired function, including reduced NO release, which consequentially resulted in enhanced PASMC proliferation. We propose that the positive regulatory loop of PASMC-extracellular vesicle miR-409-5p-PAEC is a potential mechanism underlying the proliferation of PASMCs under PDGF stimulation. Therefore, miR-409-5p may be a novel therapeutic target for the treatment of vascular diseases, including pulmonary arterial hypertension.
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Vesículas Extracelulares , MicroARNs , Hipertensión Arterial Pulmonar , Enfermedades Vasculares , Humanos , Arteria Pulmonar , Células Endoteliales , MicroARNs/genética , Miocitos del Músculo Liso , Factor de Crecimiento Derivado de PlaquetasRESUMEN
Pancreatic ductal adenocarcinoma (PDAC) is associated with a vast stromal reaction that arises mainly from cancer-associated fibroblasts (CAFs) and promotes both immune escape and tumor growth. Here, we used a mouse model with deletion of the activin A receptor ALK4 in the context of the KrasG12D mutation, which strongly drives collagen deposition that leads to tissue stiffness. By ligand-receptor analysis of single-cell RNA-sequencing data, we identified that, in stiff conditions, neoplastic ductal cells instructed CAFs through sustained platelet-derived growth factor (PDGF) signaling. Tumor-associated tissue rigidity resulted in the emergence of stiffness-induced CAFs (siCAFs) in vitro and in vivo. Similar results were confirmed in human data. siCAFs were able to strongly inhibit CD8+ T-cell responses in vitro and in vivo, promoting local immunosuppression. More importantly, targeting PDGF signaling led to diminished siCAF and reduced tumor growth. Our data show for the first time that early paracrine signaling leads to profound changes in tissue mechanics, impacting immune responses and tumor progression. Our study highlights that PDGF ligand neutralization can normalize the tissue architecture independent of the genetic background, indicating that finely tuned stromal therapy may open new therapeutic avenues in pancreatic cancer.
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[This corrects the article DOI: 10.3389/fmolb.2023.1149828.].
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Introduction: Atopic dermatitis (AD) is a common allergic eczema that affects up to 10% of adults in developed countries. Immune cells in the epidermis, namely, Langerhans cells (LCs), contribute to the pathogenesis of AD, although their exact role(s) in disease remain unclear. Methods: We performed immunostaining on human skin and peripheral blood mononuclear cells (PBMCs) and visualized primary cilium. Result and discussion: We show that human dendritic cells (DCs) and LCs have a previously unknown primary cilium-like structure. The primary cilium was assembled during DC proliferation in response to the Th2 cytokine GM-CSF, and its formation was halted by DC maturation agents. This suggests that the role of primary cilium is to transduce proliferation signaling. The platelet-derived growth factor receptor alpha (PDGFRα) pathway, which is known for transducing proliferation signals in the primary cilium, promoted DC proliferation in a manner dependent on the intraflagellar transport (IFT) system. We also examined the epidermal samples from AD patients, and observed aberrantly ciliated LCs and keratinocytes in immature and proliferating states. Our results identify a potential relationship between the primary cilium and allergic skin barrier disorders, and suggest that targeting the primary cilium may contribute to treating AD.
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Platelet-derived growth factors (PDGFs) regulate embryonic development, tissue regeneration, and wound healing through their binding to PDGF receptors, PDGFRα and PDGFRß. However, the role of PDGF signaling in regulating muscle development and regeneration remains elusive, and the cellular and molecular responses of myogenic cells are understudied. Here, we explore the PDGF-PDGFR gene expression changes and their involvement in skeletal muscle myogenesis and myogenic fate. By surveying bulk RNA sequencing and single-cell profiling data of skeletal muscle stem cells, we show that myogenic progenitors and muscle stem cells differentially express PDGF ligands and PDGF receptors during myogenesis. Quiescent adult muscle stem cells and myoblasts preferentially express PDGFRß over PDGFRα. Remarkably, cell culture- and injury-induced muscle stem cell activation altered PDGF family gene expression. In myoblasts, PDGF-AB and PDGF-BB treatments activate two pro-chemotactic and pro-mitogenic downstream transducers, RAS-ERK1/2 and PI3K-AKT. PDGFRs inhibitor AG1296 inhibited ERK1/2 and AKT activation, myoblast migration, proliferation, and cell cycle progression induced by PDGF-AB and PDGF-BB. We also found that AG1296 causes myoblast G0/G1 cell cycle arrest. Remarkably, PDGF-AA did not promote a noticeable ERK1/2 or AKT activation, myoblast migration, or expansion. Also, myogenic differentiation reduced the expression of both PDGFRα and PDGFRß, whereas forced PDGFRα expression impaired myogenesis. Thus, our data highlight PDGF signaling pathway to stimulate satellite cell proliferation aiming to enhance skeletal muscle regeneration and provide a deeper understanding of the role of PDGF signaling in non-fibroblastic cells.
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Desarrollo de Músculos , Fosfatidilinositol 3-Quinasas , División Celular , Proliferación Celular/fisiología , Fosfatidilinositol 3-Quinasas/metabolismo , Receptor beta de Factor de Crecimiento Derivado de Plaquetas/genética , Receptor beta de Factor de Crecimiento Derivado de Plaquetas/metabolismoRESUMEN
BACKGROUND: Medulloblastoma (MB) is the most common malignant pediatric brain tumor that originates in the cerebellum and brainstem. Frequent somatic mutations and deregulated expression of epigenetic regulators in MB highlight the substantial role of epigenetic alterations. 5-hydroxymethylcytosine (5hmC) is a highly abundant cytosine modification in the developing cerebellum and is regulated by ten-eleven translocation (TET) enzymes. RESULTS: We investigate the alterations of 5hmC and TET enzymes in MB and their significance to cerebellar cancer formation. We show total abundance of 5hmC is reduced in MB, but identify significant enrichment of MB-specific 5hmC marks at regulatory regions of genes implicated in stem-like properties and Nanog-binding motifs. While TET1 and TET2 levels are high in MBs, only knockout of Tet1 in the smoothened (SmoA1) mouse model attenuates uncontrolled proliferation, leading to a favorable prognosis. The pharmacological Tet1 inhibition reduces cell viability and platelet-derived growth factor signaling pathway-associated genes. CONCLUSIONS: These results together suggest a potential key role of 5hmC and indicate an oncogenic nature for TET1 in MB tumorigenesis, suggesting it as a potential therapeutic target for MBs.
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Susceptibilidad a Enfermedades , Meduloblastoma/etiología , Meduloblastoma/metabolismo , Oxigenasas de Función Mixta/genética , Oxigenasas de Función Mixta/metabolismo , Proteínas Proto-Oncogénicas/genética , Proteínas Proto-Oncogénicas/metabolismo , 5-Metilcitosina/análogos & derivados , Animales , Biomarcadores de Tumor , Biología Computacional/métodos , Islas de CpG , Metilación de ADN , Bases de Datos de Ácidos Nucleicos , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Epigénesis Genética , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Humanos , Meduloblastoma/mortalidad , Meduloblastoma/patología , Ratones , Ratones Transgénicos , Motivos de Nucleótidos , PronósticoRESUMEN
In pulmonary arterial hypertension (PAH), excessive proliferation of pulmonary artery smooth muscle cells (PASMCs) causes vascular medial thickening. Medial thickening is a histopathological hallmark of pulmonary vascular remodeling, the central disease process driving PAH progression. Pulmonary vascular remodeling causes stenosis and/or obstruction of small pulmonary arteries. This leads to increased pulmonary vascular resistance, elevated pulmonary arterial pressure, and ultimately right heart failure. To improve the survival of PAH patients, which remains at approximately 60% at 3 years after diagnosis, the development of novel PAH-targeted drugs is desired. To this end, a detailed understanding of the mechanisms underlying excessive PASMC proliferation and the medial thickening that ensues is necessary. However, a lack of in vitro models that recapitulate medial thickening impedes our deeper understanding of the pathogenetic mechanisms involved. In the present study, we applied 3-dimensional (3D) cell culture technology to develop a novel in vitro model of the pulmonary artery medial layer using human PAH patient-derived PASMCs. The addition of platelet-derived growth factor (PDGF)-BB, a mitogen known to promote excessive PASMC proliferation in PAH, resulted in increased thickness of the 3D-PAH media tissues. Conversely, administration of the PDGF receptor inhibitor imatinib or other clinical PAH drugs inhibited this medial thickening-inducing effect of PDGF-BB. Altogether, by using 3D cell culture technology, we report the generation of an in vitro model of medial thickening in PAH, which had hitherto not been successfully modeled in vitro. This model is potentially useful for assessing the ability of candidate PAH drugs to suppress medial thickening.
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Melatonin has been reported to participate in bone metabolism in recent studies. However, the underlying mechanism in melatonin-mediated osteoblastic differentiation remains largely unknown. The aim of this study is to investigate the role of melatonin in osteoblastic differentiation. In the present study, additional melatonin significantly promoted osteoblastic differentiation of MC3T3-E1 cells as evidenced by increased messenger RNA (mRNA) levels of osteogenic markers, alkaline phosphatase (ALP), collagen type I α1 chain, osteocalcin, and runt-related transcription factor 2 (Runx2). It was noteworthy that the expression level of platelet-derived growth factor subunit B (PDGFB) and content of its homodimer PDGF-BB were remarkably increased after melatonin administration. Moreover, the mRNA levels of phosphorylated PDGFRß (PDGF receptor ß) and Akt, a serine/threonine-specific protein kinase, were significantly upregulated in melatonin-treated MC3T3-E1 cells determined by a real-time polymerase chain reaction. Besides, by performing alizarin red staining, osteoblastic differentiation of MC3T3-E1 cells was conspicuously promoted by melatonin, which could be partially attenuated by crenolanib, a PDGFR inhibitor. Similarly, results from immunofluorescence and western blot assay showed that melatonin-induced upregulation of Runx2 and phosphorylated Akt was suppressed by crenolanib. Akt inhibition by MK-2206 also suppressed osteoblastic differentiation. Furthermore, by in vivo assay, additional melatonin promoted osteoblastic differentiation in mice with femoral fracture, and obvious callus formation was observed in melatonin-treated mice 5 weeks after fracture. Melatonin supplement also inhibited osteoclastic differentiation in mice. All statistical analysis was performed using GraphPad Prism and a P < 0.05 was deemed to be significant. To summarize, we demonstrate that melatonin promotes osteoblastic differentiation in MC3T3-E1 cells and enhances fracture healing in mouse femoral fracture model and regulates PDGF/AKT signaling pathway.
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Antioxidantes/farmacología , Diferenciación Celular , Melatonina/farmacología , Osteoblastos/citología , Osteogénesis , Factor de Crecimiento Derivado de Plaquetas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Fosfatasa Alcalina/metabolismo , Animales , Proliferación Celular , Células Cultivadas , Masculino , Ratones , Ratones Endogámicos C57BL , Osteoblastos/efectos de los fármacos , Osteoblastos/metabolismo , Fosforilación , Factor de Crecimiento Derivado de Plaquetas/genética , Proteínas Proto-Oncogénicas c-akt/genética , Transducción de SeñalRESUMEN
Receptor tyrosine kinase signaling pathways are key regulators for the formation of the primitive endoderm (PrE) and the epiblast (Epi) from the inner cell mass (ICM) of the mouse preimplantation embryo. Among them, FGF signaling is critical for PrE cell specification, whereas PDGF signaling is critical for the survival of committed PrE cells. Here, we investigated possible functional redundancies among FGF, PDGF, and KIT signaling and showed that only PDGF signaling is involved in PrE cell survival. In addition, we analyzed the effectors downstream of PDGFRα. Our results suggest that the role of PDGF signaling in PrE cell survival is mediated through PI3K-mTOR and independently from p53. Lastly, we uncovered a role for PI3K-mTOR signaling in the survival of Epi cells. Taken together, we propose that survival of ICM cell lineages relies on the regulation of PI3K-mTOR signaling through the regulation of multiple signaling pathways. Stem Cells 2019;37:888-898.
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Masa Celular Interna del Blastocisto/metabolismo , Linaje de la Célula/genética , Endodermo/metabolismo , Regulación del Desarrollo de la Expresión Génica , Fosfatidilinositol 3-Quinasas/genética , Factor de Crecimiento Derivado de Plaquetas/genética , Serina-Treonina Quinasas TOR/genética , Animales , Blastocisto , Masa Celular Interna del Blastocisto/citología , Supervivencia Celular , Endodermo/citología , Endodermo/crecimiento & desarrollo , Femenino , Factores de Crecimiento de Fibroblastos/genética , Factores de Crecimiento de Fibroblastos/metabolismo , Masculino , Ratones , Fosfatidilinositol 3-Quinasas/metabolismo , Factor de Crecimiento Derivado de Plaquetas/metabolismo , Proteínas Proto-Oncogénicas c-kit/genética , Proteínas Proto-Oncogénicas c-kit/metabolismo , Receptor alfa de Factor de Crecimiento Derivado de Plaquetas/genética , Receptor alfa de Factor de Crecimiento Derivado de Plaquetas/metabolismo , Transducción de Señal , Serina-Treonina Quinasas TOR/metabolismo , Proteína p53 Supresora de Tumor/genética , Proteína p53 Supresora de Tumor/metabolismoRESUMEN
The blood-brain barrier (BBB) constitutes a neurovascular unit formed by microvascular endothelial cells, pericytes, and astrocytes. Brain pericytes are important regulators of BBB integrity, permeability, and blood flow. Pericyte loss has been implicated in injury; however, how the crosstalk among pericytes, endothelial cells, and astrocytes ultimately leads to BBB dysfunction in traumatic brain injury (TBI) remains elusive. In this study, we demonstrate the importance of pericyte-endothelium interaction in maintaining the BBB function. TBI causes the platelet-derived growth factor-B (PDGF-B)/PDGF receptor-ß signaling impairment that results in loss of interaction with endothelium and leads to neurovascular dysfunction. Using in vivo mild (7â¯psi) and moderate (15â¯psi) fluid percussion injury (FPI) in mice, we demonstrate the expression of various pericyte markers including PDGFR-ß, NG2 and CD13 that were significantly reduced with a subsequent reduction in the expression of various integrins; adherent junction protein, N-cadherin; gap junction protein, connexin-43; and tight junction proteins such as occludin, claudin-5, ZO-1, and JAM-a. Impairment of pericyte-endothelium interaction increases the BBB permeability to water that is marked by a significant increase in aquaporin4 expression in injured animals. Similarly, pericyte-endothelium integrity impairment in FPI animals greatly increases the permeability of small-molecular-weight sodium fluorescein and high-molecular-weight-tracer Evans blue across the BBB. In addition, the injury-inflicted animals show significantly higher levels of S100ß and NSE in the blood samples compared with controls. In conclusion, our data provide an insight that brain trauma causes an early impairment of pericyte-endothelium integrity and results in BBB dysregulation that initiates pathological consequences associated with TBI.
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Barrera Hematoencefálica/patología , Lesiones Traumáticas del Encéfalo/patología , Endotelio Vascular/patología , Pericitos/patología , Animales , Barrera Hematoencefálica/metabolismo , Lesiones Traumáticas del Encéfalo/metabolismo , Permeabilidad Capilar/fisiología , Endotelio Vascular/metabolismo , Ratones , Pericitos/metabolismo , Receptor Cross-Talk/fisiologíaRESUMEN
Malignant peripheral nerve sheath tumors (MPNSTs) are highly aggressive Schwann cell (SC)-lineage-derived sarcomas. Molecular events driving SC-to-MPNST transformation are incompletely understood. Here, we show that human MPNSTs exhibit elevated HIPPO-TAZ/YAP expression, and that TAZ/YAP hyperactivity in SCs caused by Lats1/2 loss potently induces high-grade nerve-associated tumors with full penetrance. Lats1/2 deficiency reprograms SCs to a cancerous, progenitor-like phenotype and promotes hyperproliferation. Conversely, disruption of TAZ/YAP activity alleviates tumor burden in Lats1/2-deficient mice and inhibits human MPNST cell proliferation. Moreover, genome-wide profiling reveals that TAZ/YAP-TEAD1 directly activates oncogenic programs, including platelet-derived growth factor receptor (PDGFR) signaling. Co-targeting TAZ/YAP and PDGFR pathways inhibits tumor growth. Thus, our findings establish a previously unrecognized convergence between Lats1/2-TAZ/YAP signaling and MPNST pathogenesis, revealing potential therapeutic targets in these untreatable tumors.
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Proteínas Adaptadoras Transductoras de Señales/genética , Fosfoproteínas/genética , Proteínas Serina-Treonina Quinasas/genética , Células de Schwann/citología , Animales , Proteínas de Ciclo Celular , Diferenciación Celular/genética , Proliferación Celular/genética , Transformación Celular Neoplásica , Humanos , Ratones , Transducción de Señal/genética , Factores de Transcripción , Proteínas Señalizadoras YAPRESUMEN
To investigate whether genetic variants of platelet-derived growth factor (PDGF) signaling pathway genes are associated with survival of cutaneous melanoma (CM) patients, we assessed associations of single-nucleotide polymorphisms in PDGF pathway with melanoma-specific survival in 858 CM patients of M.D. Anderson Cancer Center (MDACC). Additional data of 409 cases from Harvard University were also included for further analysis. We identified 13 SNPs in four genes (COL6A3, NCK2, COL5A1 and PRKCD) with a nominal P < 0.05 and false discovery rate (FDR) < 0.2 in MDACC dataset. Based on linkage disequilibrium, functional prediction and minor allele frequency, a representative SNP in each gene was selected. In the meta-analysis using MDACC and Harvard datasets, there were two SNPs associated with poor survival of CM patients: rs6707820 C>T in NCK2 (HR = 1.87, 95% CI = 1.35-2.59, Pmeta= 1.53E-5); and rs2306574 T>C in PRKCD (HR = 1.73, 95% CI = 1.33-2.24, Pmeta= 4.56E-6). Moreover, CM patients in MDACC with combined risk genotypes of these two loci had markedly poorer survival (HR = 2.47, 95% CI = 1.58-3.84, P < 0.001). Genetic variants of rs6707820 C>T in NCK2 and rs2306574 T>C in PRKCD of the PDGF signaling pathway may be biomarkers for melanoma survival.
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Connective tissues-skeleton, dermis, pericytes, fascia-are a key cell source for regenerating the patterned skeleton during axolotl appendage regeneration. This complexity has made it difficult to identify the cells that regenerate skeletal tissue. Inability to identify these cells has impeded a mechanistic understanding of blastema formation. By tracing cells during digit tip regeneration using brainbow transgenic axolotls, we show that cells from each connective tissue compartment have distinct spatial and temporal profiles of proliferation, migration, and differentiation. Chondrocytes proliferate but do not migrate into the regenerate. In contrast, pericytes proliferate, then migrate into the blastema and give rise solely to pericytes. Periskeletal cells and fibroblasts contribute the bulk of digit blastema cells and acquire diverse fates according to successive waves of migration that choreograph their proximal-distal and tissue contributions. We further show that platelet-derived growth factor signaling is a potent inducer of fibroblast migration, which is required to form the blastema.
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Ambystoma mexicanum/fisiología , Tejido Conectivo/fisiología , Extremidades/fisiología , Imagenología Tridimensional , Regeneración/fisiología , Células Madre/citología , Animales , Animales Modificados Genéticamente , Huesos/fisiología , Movimiento Celular , Proliferación Celular , Condrocitos/citología , Células Clonales , Dermis/citología , Fibroblastos/citología , Modelos Biológicos , Pericitos/citología , Factor de Crecimiento Derivado de Plaquetas/metabolismo , Transducción de Señal , Factores de TiempoRESUMEN
The primary cilium is a cellular organelle that coordinates signaling pathways critical for cell proliferation, differentiation, survival, and homeostasis. Intraflagellar transport (IFT) plays a pivotal role in assembling primary cilia. Disruption and/or dysfunction of IFT components can cause multiple diseases, including skeletal dysplasia. However, the mechanism by which IFT regulates skeletogenesis remains elusive. Here, we show that a neural crest-specific deletion of intraflagellar transport 20 (Ift20) in mice compromises ciliogenesis and intracellular transport of collagen, which leads to osteopenia in the facial region. Whereas platelet-derived growth factor receptor alpha (PDGFRα) was present on the surface of primary cilia in wild-type osteoblasts, disruption of Ift20 down-regulated PDGFRα production, which caused suppression of PDGF-Akt signaling, resulting in decreased osteogenic proliferation and increased cell death. Although osteogenic differentiation in cranial neural crest (CNC)-derived cells occurred normally in Ift20-mutant cells, the process of mineralization was severely attenuated due to delayed secretion of type I collagen. In control osteoblasts, procollagen was easily transported from the endoplasmic reticulum (ER) to the Golgi apparatus. By contrast, despite having similar levels of collagen type 1 alpha 1 (Col1a1) expression, Ift20 mutants did not secrete procollagen because of dysfunctional ER-to-Golgi trafficking. These data suggest that in the multipotent stem cells of CNCs, IFT20 is indispensable for regulating not only ciliogenesis but also collagen intracellular trafficking. Our study introduces a unique perspective on the canonical and noncanonical functions of IFT20 in craniofacial skeletal development.
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Desarrollo Óseo/fisiología , Anomalías Craneofaciales/fisiopatología , Huesos Faciales/fisiología , Flagelos/fisiología , Cresta Neural/fisiología , Cráneo/fisiología , Animales , Transporte Biológico Activo/fisiología , Proteínas Portadoras , Células Cultivadas , Anomalías Craneofaciales/patología , Huesos Faciales/citología , Flagelos/patología , Regulación del Desarrollo de la Expresión Génica/fisiología , Ratones , Modelos Biológicos , Morfogénesis/fisiología , Osteoblastos/fisiología , Osteoblastos/ultraestructura , Cráneo/citologíaRESUMEN
Osteosarcoma (OS) is the most frequent primary solid malignant tumor of bone. Its prognosis remains poor in the substantial proportion of patients who do not respond to chemotherapy and novel therapeutic options are therefore needed. We previously established a mouse model that mimics the aggressive behavior of human OS. Enzyme-linked immunosorbent assay-based screening of such mouse tumor lysates identified platelet-derived growth factor-BB (PDGF-BB) as an abundant soluble factor, the gene for which was expressed dominantly in surrounding non-malignant cells of the tumor, whereas that for the cognate receptor (PDGF receptor ß) was highly expressed in OS cells. Platelet-derived growth factor-BB induced activation of both MEK-ERK and phosphatidylinositol 3-kinase-protein kinase B signaling pathways and promoted survival in OS cells deprived of serum, and these effects were blocked by the PDGF receptor inhibitor imatinib. However, these actions of PDGF-BB and imatinib were mostly masked in the presence of serum. Whereas imatinib alone did not manifest an antitumor effect in mice harboring OS tumors, combined treatment with imatinib and adriamycin exerted a synergistic antiproliferative effect on OS cells in vivo. These results suggest that treatment of OS with imatinib is effective only when cell survival is dependent on PDGF signaling or when imatinib is combined with another therapeutic intervention that renders the tumor cells susceptible to imatinib action, such as by inducing cellular stress.
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Antineoplásicos/farmacología , Benzamidas/farmacología , Doxorrubicina/farmacología , Piperazinas/farmacología , Pirimidinas/farmacología , Receptor beta de Factor de Crecimiento Derivado de Plaquetas/metabolismo , Animales , Becaplermina , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Sinergismo Farmacológico , Femenino , Humanos , Mesilato de Imatinib , Ratones Endogámicos C57BL , Osteosarcoma , Proteínas Proto-Oncogénicas c-sis/metabolismo , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
CITED2 is a transcriptional co-factor with important roles in many organs of the developing mammalian embryo. Complete deletion of this gene causes severe malformation of the placenta, and results in significantly reduced embryonic growth and death from E14.5. The placenta is a complex organ originating from cells derived from three lineages: the maternal decidua, the trophectoderm, and the extra-embryonic mesoderm. Cited2 is expressed in many of these cell types, but its exact role in the formation of the placenta is unknown. Here we use a conditional deletion approach to remove Cited2 from overlapping subsets of trophectoderm and extra-embryonic mesoderm. We find that Cited2 in sinusoidal trophoblast giant cells and syncytiotrophoblasts is likely to have a non-cell autonomous role in patterning of the pericytes associated with the embryonic capillaries. This function is likely to be mediated by PDGF signaling. Furthermore, we also identify that loss of Cited2 in syncytiotrophoblasts results in the subcellular mislocalization of one of the major lactate transporters in the placenta, SLC16A3 (MCT4). We hypothesize that the embryonic growth retardation observed in Cited2 null embryos is due in part to a disorganized embryonic capillary network, and in part due to abnormalities of the nutrient transport functions of the feto-maternal interface.
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Tipificación del Cuerpo , Capilares/embriología , Placenta/irrigación sanguínea , Placenta/embriología , Circulación Placentaria/genética , Proteínas Represoras/genética , Transactivadores/genética , Trofoblastos/enzimología , Actinas/biosíntesis , Animales , Proteínas Portadoras/metabolismo , Desarrollo Embrionario , Femenino , Eliminación de Gen , Ratones , Ratones Endogámicos C57BL , Transportadores de Ácidos Monocarboxílicos/biosíntesis , Proteínas Musculares/biosíntesis , Neovascularización Fisiológica/genética , Neovascularización Fisiológica/fisiología , Pericitos/citología , Pericitos/fisiología , Molécula-1 de Adhesión Celular Endotelial de Plaqueta/biosíntesis , Factor de Crecimiento Derivado de Plaquetas/genética , Factor de Crecimiento Derivado de Plaquetas/metabolismo , Embarazo , Proteínas Proto-Oncogénicas c-sis/biosíntesis , Receptor beta de Factor de Crecimiento Derivado de Plaquetas/biosíntesis , Simportadores/biosíntesisRESUMEN
At the end of the preimplantation period, the inner cell mass (ICM) of the mouse blastocyst is composed of two distinct cell lineages, the pluripotent epiblast (EPI) and the primitive endoderm (PrE). The current model for their formation involves initial co-expression of lineage-specific markers followed by mutual-exclusive expression resulting in a salt-and-pepper distribution of lineage precursors within the ICM. Subsequent to lineage commitment, cell rearrangements and selective apoptosis are thought to be key processes driving and refining the emergence of two spatially distinct compartments. Here, we have addressed a role for Platelet Derived Growth Factor (PDGF) signaling in the regulation of programmed cell death during early mouse embryonic development. By combining genetic and pharmacological approaches, we demonstrate that embryos lacking PDGF activity exhibited caspase-dependent selective apoptosis of PrE cells. Modulating PDGF activity did not affect lineage commitment or cell sorting, suggesting that PDGF is involved in the fine-tuning of patterning information. Our results also indicate that PDGF and fibroblast growth factor (FGF) tyrosine kinase receptors exert distinct and non-overlapping functions in PrE formation. Taken together, these data uncover an early role of PDGF signaling in PrE cell survival at the time when PrE and EPI cells are segregated.