RESUMO
Blood vessel growth and osteogenesis in the skeletal system are coupled; however, fundamental aspects of vascular function in osteoblast-to-osteocyte transition remain unclear. Our study demonstrates that vascular smooth muscle cells (VSMCs), but not endothelial cells, are sufficient to drive bone marrow mesenchymal stromal cell-derived osteoblast-to-osteocyte transition via ß-catenin signaling and exosome-mediated communication. We found that VSMC-derived exosomes are loaded with transcripts encoding proteins associated with the osteocyte phenotype and members of the WNT/ß-catenin signaling pathway. In contrast, endothelial cell-derived exosomes facilitated mature osteoblast differentiation by reprogramming the TGFB1 gene family and osteogenic transcription factors osterix (SP7) and RUNX2. Notably, VSMCs express significant levels of tetraspanins (CD9, CD63, and CD81) and drive the intracellular trafficking of exosomes with a lower membrane zeta potential than those from other cells. Additionally, the high ATP content within these exosomes supports mineralization mechanisms, as ATP is a substrate for alkaline phosphatase. Osteocyte function was further validated by RNA sequencing, revealing activity in genes related to intermittent mineralization and sonic hedgehog signaling, alongside a significant increase in TNFSF11 levels. Our findings unveil a novel role of VSMCs in promoting osteoblast-to-osteocyte transition, thus offering new insights into bone biology and homeostasis, as well as in bone-related diseases. Clinically, these insights could pave the way for innovative therapeutic strategies targeting VSMC-derived exosome pathways to treat bone-related disorders such as osteoporosis. By manipulating these signaling pathways, it may be possible to enhance bone regeneration and improve skeletal health in patients with compromised bone structure and function.
Assuntos
Exossomos , Músculo Liso Vascular , Osteoblastos , Osteócitos , Osteogênese , beta Catenina , Osteoblastos/metabolismo , Osteoblastos/citologia , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/citologia , Exossomos/metabolismo , Animais , beta Catenina/metabolismo , beta Catenina/genética , Osteócitos/metabolismo , Osteócitos/citologia , Camundongos , Osteogênese/genética , Osteogênese/fisiologia , Miócitos de Músculo Liso/metabolismo , Diferenciação Celular , Humanos , Via de Sinalização Wnt , Células-Tronco Mesenquimais/metabolismo , Células-Tronco Mesenquimais/citologia , Células Cultivadas , Transdução de Sinais , Camundongos Endogâmicos C57BLRESUMO
Force transmission through adherens junctions (AJs) is crucial for multicellular organization, wound healing and tissue regeneration. Recent studies shed light on the molecular mechanisms of mechanotransduction at the AJs. However, the canonical model fails to explain force transmission when essential proteins of the mechanotransduction module are mutated or missing. Here, we demonstrate that, in absence of α-catenin, ß-catenin can directly and functionally interact with vinculin in its open conformation, bearing physiological forces. Furthermore, we found that ß-catenin can prevent vinculin autoinhibition in the presence of α-catenin by occupying vinculin´s head-tail interaction site, thus preserving force transmission capability. Taken together, our findings suggest a multi-step force transmission process at AJs, where α-catenin and ß-catenin can alternatively and cooperatively interact with vinculin. This can explain the graded responses needed to maintain tissue mechanical homeostasis and, importantly, unveils a force-bearing mechanism involving ß-catenin and extended vinculin that can potentially explain the underlying process enabling collective invasion of metastatic cells lacking α-catenin.
Assuntos
Junções Aderentes , Mecanotransdução Celular , Vinculina , alfa Catenina , beta Catenina , Vinculina/metabolismo , Junções Aderentes/metabolismo , beta Catenina/metabolismo , alfa Catenina/metabolismo , alfa Catenina/genética , Animais , Humanos , Camundongos , Ligação ProteicaRESUMO
Tumor hypoxia has been associated with cancer progression, angiogenesis, and metastasis via modifications in the release and cargo composition of extracellular vesicles secreted by tumor cells. Indeed, hypoxic extracellular vesicles are known to trigger a variety of angiogenic responses via different mechanisms. We recently showed that hypoxia promotes endosomal signaling in tumor cells via HIF-1α-dependent induction of the guanine exchange factor ALS2, which activates Rab5, leading to downstream events involved in cell migration and invasion. Since Rab5-dependent signaling is required for endothelial cell migration and angiogenesis, we explored the possibility that hypoxia promotes the release of small extracellular vesicles containing ALS2, which in turn activate Rab5 in recipient endothelial cells leading to pro-angiogenic properties. In doing so, we found that hypoxia promoted ALS2 expression and incorporation as cargo within small extracellular vesicles, leading to subsequent transfer to recipient endothelial cells and promoting cell migration, tube formation, and downstream Rab5 activation. Consequently, ALS2-containing small extracellular vesicles increased early endosome size and number in recipient endothelial cells, which was followed by subsequent sequestration of components of the ß-catenin destruction complex within endosomal compartments, leading to stabilization and nuclear localization of ß-catenin. These events converged in the expression of ß-catenin target genes involved in angiogenesis. Knockdown of ALS2 in donor tumor cells precluded its incorporation into small extracellular vesicles, preventing Rab5-downstream events and endothelial cell responses, which depended on Rab5 activity and guanine exchange factor activity of ALS2. These findings indicate that vesicular ALS2, secreted in hypoxia, promotes endothelial cell events leading to angiogenesis. Finally, these events might explain how tumor angiogenesis proceeds in hypoxic conditions.
Assuntos
Movimento Celular , Vesículas Extracelulares , Fatores de Troca do Nucleotídeo Guanina , Transdução de Sinais , beta Catenina , Proteínas rab5 de Ligação ao GTP , Humanos , Proteínas rab5 de Ligação ao GTP/metabolismo , Proteínas rab5 de Ligação ao GTP/genética , beta Catenina/metabolismo , Vesículas Extracelulares/metabolismo , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Fatores de Troca do Nucleotídeo Guanina/genética , Neovascularização Patológica/metabolismo , Neovascularização Patológica/patologia , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Células Endoteliais da Veia Umbilical Humana/metabolismo , Linhagem Celular TumoralRESUMO
PURPOSE: Pre-eclampsia (PE) is a pregnancy-related complication. Eucommia is effective in the treatment of hypertensive disorders in pregnancy, but the specific effects and possible mechanisms of Eucommia granules (EG) in PE remain unknown. The aim of this study was to investigate the effects and possible mechanisms of EG in PE rats. METHODS: Pregnant Sprague Dawley rats were divided into five groups (n = 6): the control group, the model group, the low-dose group, the medium-dose group, and the high-dose group of EG. The PE model was established by subcutaneous injection of levonitroarginine methyl ester. Saline was given to the blank and model groups, and the Eucommia granules were given by gavage to the remaining groups. Blood pressure and urinary protein were detected. The body length and weight of the pups and the weight of the placenta were recorded. Superoxide dismutase (SOD) activity and levels of malondialdehyde (MDA), placental growth factor (PIGF), and soluble vascular endothelial growth factor receptor-1 (sFIt-1) were measured in the placenta. Pathological changes were observed by hematoxylin-eosin staining. Wnt/ß-catenin pathway-related protein expression was detected using Western blot. RESULTS: Compared with the model group, the PE rats treated with EG had lower blood pressure and urinary protein. The length and weight of the pups and placental weight were increased. Inflammation and necrosis in the placental tissue was improved. SOD level increased, MDA content and sFIt-1/PIGF ratio decreased, and Wnt/ß-catenin pathway-related protein expression level increased. Moreover, the results of EG on PE rats increased with higher doses of EG. CONCLUSIONS: EG may activate the Wnt/ß-catenin pathway and inhibit oxidative stress, inflammation, and vascular endothelial injury in PE rats, thereby improving the perinatal prognosis of preeclamptic rats. EG may inhibit oxidative stress, inflammation, and vascular endothelial injury through activation of the Wnt/ß-catenin pathway in preeclampsia rats, thereby improving perinatal outcomes in PE rats.
Assuntos
Pré-Eclâmpsia , Complicações na Gravidez , Humanos , Ratos , Feminino , Gravidez , Animais , Pré-Eclâmpsia/tratamento farmacológico , Pré-Eclâmpsia/metabolismo , Pré-Eclâmpsia/patologia , Placenta , Ratos Sprague-Dawley , Fator A de Crescimento do Endotélio Vascular/metabolismo , beta Catenina/metabolismo , Fator de Crescimento Placentário/metabolismo , Fator de Crescimento Placentário/farmacologia , Fator de Crescimento Placentário/uso terapêutico , Estresse Oxidativo , Complicações na Gravidez/metabolismo , Inflamação/patologia , Superóxido Dismutase/metabolismoRESUMO
Changes in the expression of nuclear ß-catenin are responsible for tumorigenesis. Beta-catenin acts synergistically with the TGF-ß/BMPs pathway. This interaction leads to greater dentin deposition and may explain the differences between distinct tooth morphologies and hamartomas. The aim of this study was to investigate the role of ß-catenin, BMP4 and TGF-ß in the development of odontomas. This cross-sectional, retrospective, immunohistochemical study evaluated 30 compound odontomas, 30 complex odontomas and 17 tooth germs. The results showed that BMP4 and TGF-ß were more immunoexpressed in the ectomesenchyme of complex odontomas (median = 33.7, p < 0.001; median = 76.4, p = 0.002, respectively). Higher immunoexpression of BMP4 and TGF-ß was also observed in the epithelium of tooth germs (median = 2.0, p < 0.001; median = 120.3, p < 0.001, respectively). TGF-ß and BMP4 showed a positive and significant correlation (p < 0.001). Both TGF-ß and BMP4 were positively correlated with nuclear ß-catenin in ectomesenchyme (p = 0.047 and p = 0.023, respectively). Developing teeth exhibited higher concentrations of the proteins studied in odontogenic epithelium, especially during the bud and cap stages. Higher immunoexpression in odontomas occurred mainly in the ectomesenchyme. We therefore suggest that changes in the ectomesenchyme can lead to the development of odontomas.
Assuntos
Odontoma , Animais , Odontoma/veterinária , beta Catenina/metabolismo , Fator de Crescimento Transformador beta , Estudos Retrospectivos , Estudos TransversaisRESUMO
INTRODUCTION: Studies addressing the methylation pattern in adamantinomatous craniopharyngioma (ACP) are lacking. OBJECTIVE: To identify methylation signatures in ACPs regarding clinical presentation and outcome. METHODS: Clinical and pathology data were collected from 35 patients with ACP (54% male; 18.1 years [2-68]). CTNNB1 mutations and methylation profile (MethylationEPIC/Array-Illumina) were analyzed in tumoral DNA. Unsupervised machine learning analysis of this comprehensive methylome sample was achieved using hierarchical clustering and multidimensional scaling. Statistical associations between clusters and clinical features were achieved using the Fisher test and global biological process interpretations were aided by Gene Ontology enrichment analyses. RESULTS: Two clusters were revealed consistently by all unsupervised methods (ACP-1: n = 18; ACP-2: n = 17) with strong bootstrap statistical support. ACP-2 was enriched by CTNNB1 mutations (100% vs 56%, P = .0006), hypomethylated in CpG island, non-CpG Island sites, and globally (P < .001), and associated with greater tumor size (24.1 vs 9.5 cm3, P = .04). Enrichment analysis highlighted pathways on signaling transduction, transmembrane receptor, development of anatomical structures, cell adhesion, cytoskeleton organization, and cytokine binding, and cell type-specific biological processes as regulation of oligodendrocytes, keratinocyte, and epithelial cells differentiation. CONCLUSION: Two clusters of patients with ACP were consistently revealed by unsupervised machine learning methods, with one of them significantly hypomethylated, enriched by CTNNB1 mutated ACPs, and associated with increased tumor size. Enrichment analysis reinforced pathways involved in tumor proliferation and in cell-specific tumoral microenvironment.
Assuntos
Craniofaringioma , Metilação de DNA , Epigênese Genética , Neoplasias Hipofisárias , beta Catenina , Humanos , Craniofaringioma/genética , Craniofaringioma/patologia , Masculino , Feminino , Neoplasias Hipofisárias/genética , Neoplasias Hipofisárias/patologia , Adolescente , Adulto , Criança , Pessoa de Meia-Idade , Adulto Jovem , beta Catenina/genética , beta Catenina/metabolismo , Pré-Escolar , Idoso , Mutação , Ilhas de CpG/genética , Regulação Neoplásica da Expressão GênicaRESUMO
INTRODUCTION AND OBJECTIVES: Cavin1 is a cell membrane caveolin, with controversial function in different tumors. Meanwhile, the role of Cavin1 in hepatocellular carcinoma (HCC) progression remains unclear. In this study, we attempted to elucidate the significance of Cavin1 in HCC occurrence and progression. MATERIALS AND METHODS: Cavin1 content was examined in HCC tissues and paired adjacent normal liver tissues by qRT-PCR and IHC among 81 HCC patients. The Cavin1-mediated regulation of HCC proliferation and metastasis was assessed through in vitro and in vivo experiments. Finally, using GSEA, we found out Cavin1 could be a potential regulator of the Wnt pathway. The alterations of the Wnt pathway-related proteins were identified by Western Blot analysis. RESULTS: Cavin1 was lower expressed in HCC, which implied poor survival outcomes in HCC patients. Phenotypic experiments revealed that Cavin1 strongly suppressed HCC proliferation and migration in vitro and in vivo. Besides, altered epithelial-mesenchymal transition (EMT)-related protein expressions were detected. Based on our GSEA analysis, Cavin1 activated the Wnt pathway, and Western Blot analysis revealed diminished ß-catenin, c-Myc, and MMP9 contents upon Cavin1 overexpression. CONCLUSIONS: Cavin1 suppresses HCC progression by modulating HCC proliferation and migration via inhibiting the Wnt/ß-catenin axis activation.
Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , Humanos , beta Catenina/genética , beta Catenina/metabolismo , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Regulação Neoplásica da Expressão Gênica , Neoplasias Hepáticas/patologia , Via de Sinalização WntRESUMO
PURPOSE: Prostate cancer (PCa) is the leading cause of death among men in 48 countries. Genetic alterations play a significant role in PCa carcinogenesis. For the hypothesis of this research, five unique polymorphisms (SNP) were investigated in different genes that showed to be associated in different ways with PCa: rs4430796, rs2735839, rs4792311, rs12329760, and rs28931588, respectively for the genes HNF1B, KLK3, ELAC2, TMPRSS2-ERG, and CTNNB1. PATIENTS AND METHODS: Blood samples from 426 subjects were evaluated: 290 controls (161 females and 129 males) and 136 PCa patients. SNP were determined by real-time polymerase chain reaction. TaqMan SNP genotyping assay. In the control samples, the SNPs were defined in association with the self-reported ethnicity, and in 218 control samples with markers with ancestry indicators. The genes were in Hardy-Weinberg equilibrium. One hundred and seventy control samples were matched by ethnicity for comparison with the PCa samples. RESULTS: The G allele at rs28931588 was monomorphic in both patients and controls studied. Significant differences were observed in allelic and genotypic frequencies between the control and Pca samples in rs2735839 (KLK3; p = 0.002 and χ2 = 8.73 and p = 0.01, respectively), by the global frequency and in the dominant model rs2735839_GG (odds ratio [OR] = 0.51, p = 0.02). AA and GA genotypes at rs4792311 (ELAC2) were more frequent in patients with Gleason 7(4 + 3), 8, and 9 (n = 37%-59.7%) compared to patients with Gleason 6 and 7(3 + 4) (n = 26%-40.0%) conferring a protective effect on the GG genotype (OR = 0.45, p = 0.02). The same genotype showed an OR = 2.71 (p = 0.01) for patients with low severity. The HNF1B-KLK3-ELAC2-TMPRSS2-ERG haplotypes: GAAT, AAAT, GAGT, and AAGT were more frequent in patients with Pca with OR ranging from 4.65 to 2.48. CONCLUSIONS: Higher frequencies of risk alleles were confirmed in the SNPs, KLK3 rs2735839_A, ELAC2 rs4792311_A, and TMPRSS2 rs12329760_T in patients with Pca. Rs2735839_A was associated with risk of Pca and rs4792311_A with severity and Gleason score of 7(4 + 3) or greater. There is a need for careful observation of rs2735839 and rs4792311 in association with the prostatic biopsy due to the increased risk of Pca.
Assuntos
Antígeno Prostático Específico , Neoplasias da Próstata , Masculino , Humanos , Calicreínas/genética , Predisposição Genética para Doença , Neoplasias da Próstata/patologia , Genótipo , Polimorfismo de Nucleotídeo Único , Regulador Transcricional ERG/genética , Fator 1-beta Nuclear de Hepatócito/genética , Proteínas de Neoplasias , beta Catenina/genéticaRESUMO
PURPOSE: Oral squamous cell carcinoma (OSCC) is commonly preceded by potentially malignant lesions, referred to as oral dysplasia. We recently reported that oral dysplasia is associated with aberrant activation of the Wnt/ß-catenin pathway, due to overexpression of Wnt ligands in a Porcupine (PORCN)-dependent manner. Pharmacologic inhibition of PORCN precludes Wnt secretion and has been proposed as a potential therapeutic approach to treat established cancers. Nevertheless, there are no studies that explore the effects of PORCN inhibition at the different stages of oral carcinogenesis. EXPERIMENTAL DESIGN: We performed a model of tobacco-induced oral cancer in vitro, where dysplastic oral keratinocytes (DOK) were transformed into oral carcinoma cells (DOK-TC), and assessed the effects of inhibiting PORCN with the C59 inhibitor. Similarly, an in vivo model of oral carcinogenesis and ex vivo samples derived from patients diagnosed with oral dysplasia and OSCC were treated with C59. RESULTS: Both in vitro and ex vivo oral carcinogenesis approaches revealed decreased levels of nuclear ß-catenin and Wnt3a, as observed by immunofluorescence and IHC analyses. Consistently, reduced protein and mRNA levels of survivin were observed after treatment with C59. Functionally, treatment with C59 in vitro resulted in diminished cell migration, viability, and invasion. Finally, by using an in vivo model of oral carcinogenesis, we found that treatment with C59 prevented the development of OSCC by reducing the size and number of oral tumor lesions. CONCLUSIONS: The inhibition of Wnt ligand secretion with C59 represents a feasible treatment to prevent the progression of early oral lesions toward OSCC.
Assuntos
Carcinoma de Células Escamosas , Neoplasias de Cabeça e Pescoço , Neoplasias Bucais , Humanos , Via de Sinalização Wnt , Carcinoma de Células Escamosas/tratamento farmacológico , Carcinoma de Células Escamosas/genética , beta Catenina/genética , beta Catenina/metabolismo , Neoplasias Bucais/tratamento farmacológico , Carcinoma de Células Escamosas de Cabeça e Pescoço , Carcinogênese/genética , Aciltransferases/metabolismo , Aciltransferases/farmacologia , Proteínas de Membrana/metabolismoRESUMO
Tissue homeostasis is crucial for multicellular organisms, wherein the loss of cells is compensated by generating new cells with the capacity for proliferation and differentiation. At the origin of these populations are the stem cells, which have the potential to give rise to cells with both capabilities, and persevere for a long time through the self-renewal and quiescence. Since the discovery of stem cells, an enormous effort has been focused on learning about their functions and the molecular regulation behind them. Wnt signaling is widely recognized as essential for normal and cancer stem cell. Moreover, ß-catenin-dependent Wnt pathway, referred to as canonical, has gained attention, while ß-catenin-independent Wnt pathways, known as non-canonical, have remained conspicuously less explored. However, recent evidence about non-canonical Wnt pathways in stem cells begins to lay the foundations of a conceivably vast field, and on which we aim to explain this in the present review. In this regard, we addressed the different aspects in which non-canonical Wnt pathways impact the properties of stem cells, both under normal conditions and also under disease, specifically in cancer.