RESUMO
Glycolytic metabolic reprogramming in cancer is regulated by both cancer intrinsic variations like isocitrate dehydrogenase 1 (IDH1) status and non-cancerous microenvironment components like tumor associated macrophages (TAMs). However, the detailed mechanism remains elusive. Here, we identify hexosaminidase B (HEXB) as a key regulator for glycolysis in glioblastoma (GBM). HEXB intercellularly manipulates TAMs to promote glycolysis in GBM cells, while intrinsically enhancing cancer cell glycolysis. Mechanistically, HEXB elevation augments tumor HIF1α protein stability through activating ITGB1/ILK/YAP1; Subsequently, HIF1α promotes HEXB and multiple glycolytic gene transcription in GBM cells. Genetic ablation and pharmacological inhibition of HEXB elicits substantial therapeutic effects in preclinical GBM models, while targeting HEXB doesn't induce significant reduction in IDH1 mutant glioma and inhibiting IDH1 mutation-derived 2-hydroxyglutaric acid (2-HG) significantly restores HEXB expression in glioma cells. Our work highlights a HEXB driven TAMs-associated glycolysis-promoting network in GBM and provides clues for developing more effective therapies against it.
Assuntos
Neoplasias Encefálicas , Carcinogênese , Glioblastoma , Glicólise , Subunidade alfa do Fator 1 Induzível por Hipóxia , Isocitrato Desidrogenase , beta-N-Acetil-Hexosaminidases , Humanos , Glioblastoma/genética , Glioblastoma/patologia , Glioblastoma/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Isocitrato Desidrogenase/genética , Isocitrato Desidrogenase/metabolismo , Linhagem Celular Tumoral , Animais , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patologia , Neoplasias Encefálicas/genética , Carcinogênese/genética , Camundongos , beta-N-Acetil-Hexosaminidases/metabolismo , beta-N-Acetil-Hexosaminidases/genética , Microambiente Tumoral/imunologia , Regulação Neoplásica da Expressão Gênica , Macrófagos Associados a Tumor/metabolismo , Macrófagos Associados a Tumor/imunologia , Integrina beta1/metabolismo , Integrina beta1/genética , Glutaratos/metabolismo , Mutação , Proteínas de Sinalização YAP/metabolismoRESUMO
Silicosis is a systemic disease caused by long-term exposure to high concentrations of free silica dust particles in the workplace. It is characterized by a persistent inflammatory response, fibroblast proliferation, and excessive collagen deposition, leading to pulmonary interstitial fibrosis. Epithelial interstitial transformation (EMT) can cause epithelial cells to lose their tight junctions, cell polarity, and epithelial properties, thereby enhancing the properties of interstitial cells, which can lead to the progression of fibrosis and the formation of scar tissue. Integrin 1 (ITGB1) is considered an important factor for promoting EMT and tumor invasion in a variety of tumors and also plays an important role in the progression of fibrotic diseases. Therefore, ITGB1 can be used as a potential target for the treatment of silicosis. In this study, we found that silica exposure induced epithelial-mesenchymal transformation in rats and that the expression of integrin ITGB1 was elevated along with the EMT. We used CRISPR/Cas9 technology to construct integrin ITGB1 knockdown cell lines for in vitro experiments. We compared the expression of the EMT key proteins E-cadherin and vimentin in the ITGB1 knockdown cells and wild-type cells simultaneously stimulated by silica and detected the aggregation point distribution of E-cadherin and vimentin in the cells using laser confocal microscopy. Our results showed that ITGB1 knockout inhibited the ITGB1/ILK/Snail signaling pathway and attenuated the EMT occurrence compared to control cells. These results suggested that ITGB1 is associated with silica-induced EMT and may be a potential target for the treatment of silicosis.
Assuntos
Transição Epitelial-Mesenquimal , Integrina beta1 , Fibrose Pulmonar , Dióxido de Silício , Animais , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Transição Epitelial-Mesenquimal/genética , Dióxido de Silício/toxicidade , Dióxido de Silício/efeitos adversos , Integrina beta1/genética , Integrina beta1/metabolismo , Fibrose Pulmonar/induzido quimicamente , Fibrose Pulmonar/genética , Fibrose Pulmonar/patologia , Ratos , Silicose/patologia , Silicose/genética , Masculino , Caderinas/metabolismo , Caderinas/genéticaRESUMO
Coagulation factor XII (FXII) conveys various functions as an active protease that promotes thrombosis and inflammation, and as a zymogen via surface receptors like urokinase-type plasminogen activator receptor (uPAR). While plasma levels of FXII are increased in diabetes mellitus and diabetic kidney disease (DKD), a pathogenic role of FXII in DKD remains unknown. Here we show that FXII is locally expressed in kidney tubular cells and that urinary FXII correlates with kidney dysfunction in DKD patients. F12-deficient mice (F12-/-) are protected from hyperglycemia-induced kidney injury. Mechanistically, FXII interacts with uPAR on tubular cells promoting integrin ß1-dependent signaling. This signaling axis induces oxidative stress, persistent DNA damage and senescence. Blocking uPAR or integrin ß1 ameliorates FXII-induced tubular cell injury. Our findings demonstrate that FXII-uPAR-integrin ß1 signaling on tubular cells drives senescence. These findings imply previously undescribed diagnostic and therapeutic approaches to detect or treat DKD and possibly other senescence-associated diseases.
Assuntos
Senescência Celular , Nefropatias Diabéticas , Fator XII , Integrina beta1 , Receptores de Ativador de Plasminogênio Tipo Uroquinase , Animais , Feminino , Humanos , Masculino , Camundongos , Nefropatias Diabéticas/metabolismo , Nefropatias Diabéticas/patologia , Nefropatias Diabéticas/genética , Fator XII/metabolismo , Fator XII/genética , Integrina beta1/metabolismo , Integrina beta1/genética , Túbulos Renais/metabolismo , Túbulos Renais/patologia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Estresse Oxidativo , Receptores de Ativador de Plasminogênio Tipo Uroquinase/metabolismo , Receptores de Ativador de Plasminogênio Tipo Uroquinase/genética , Transdução de SinaisRESUMO
BACKGROUND: The involvement of tetraspanins in cancer development has been widely implicated. In this study, the function and molecular mechanisms of tetraspanin 3 (TSPAN3) in non-small cell lung cancer (NSCLC) cells were explored. METHODS: Tissue samples from patients diagnosed with NSCLC were analyzed by immunohistochemistry, western blotting, and real-time polymerase chain reaction (PCR) to indicate the involvement of TSPAN3 in cancer progression. In the meantime, we also performed exhaustive mechanistic studies using A549 and H460 cells in vitro through a variety of methods including western blotting, real-time PCR, immunofluorescent staining, coimmunoprecipitation, cell proliferation assay, and nocodazole (NZ) washout assay. Proper statistical analysis was implemented wherever necessary in this study. RESULTS: TSPAN3 was found to be highly expressed in lung cancer cells and tissues. Moreover, high levels of TSPAN3 positively correlated with poor differentiation, lymph node involvement, advanced pathological tumor-node-metastasis stage, and poor prognosis in patients with NSCLC. TSPAN3 showed potential to promote the proliferation of NSCLC cells in vitro and in vivo. Specifically, TSPAN3 was found to interact with ß1 integrin via the LEL domain, thereby facilitating the sorting of ß1 integrin into Rab11a endosomes and promoting ß1 integrin recycling and upregulation. CONCLUSIONS: Our findings reveal TSPAN3 may represent a potentially valuable therapeutic target for NSCLC.
Assuntos
Carcinoma Pulmonar de Células não Pequenas , Proliferação de Células , Integrina beta1 , Neoplasias Pulmonares , Tetraspaninas , Humanos , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Carcinoma Pulmonar de Células não Pequenas/patologia , Carcinoma Pulmonar de Células não Pequenas/genética , Proliferação de Células/genética , Tetraspaninas/metabolismo , Tetraspaninas/genética , Integrina beta1/metabolismo , Integrina beta1/genética , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/genética , Feminino , Masculino , Linhagem Celular Tumoral , Pessoa de Meia-Idade , Animais , Células A549 , Camundongos Nus , Endossomos/metabolismo , Regulação Neoplásica da Expressão Gênica , Proteínas rab de Ligação ao GTP/metabolismo , Proteínas rab de Ligação ao GTP/genética , Camundongos , Camundongos Endogâmicos BALB CRESUMO
BACKGROUND: Excess fibrotic remodeling causes cardiac dysfunction in ischemic heart disease, driven by MAP (mitogen-activated protein) kinase-dependent TGF-ß1 (transforming growth factor-ß1) activation by coagulation signaling of myeloid cells. How coagulation-inflammatory circuits can be specifically targeted to achieve beneficial macrophage reprogramming after myocardial infarction (MI) is not completely understood. METHODS: Mice with permanent ligation of the left anterior descending artery were used to model nonreperfused MI and analyzed by single-cell RNA sequencing, protein expression changes, confocal microscopy, and longitudinal monitoring of recovery. We probed the role of the tissue factor (TF)-FVIIa (activated factor VII)-integrin ß1-PAR2 (protease-activated receptor 2) signaling complex by utilizing genetic mouse models and pharmacological intervention. RESULTS: Cleavage-insensitive PAR2R38E and myeloid cell integrin ß1-deficient mice had improved cardiac function after MI compared with controls. Proximity ligation assays of monocytic cells demonstrated that colocalization of FVIIa with integrin ß1 was diminished in monocyte/macrophage FVII-deficient mice after MI. Compared with controls, F7fl/fl CX3CR1 (CX3C motif chemokine receptor 1)Cre mice showed reduced TGF-ß1 and MAP kinase activation, as well as cardiac dysfunction after MI, despite unaltered overall recruitment of myeloid cells. Single-cell mRNA sequencing of CD45 (cluster of differentiation 45)+ cells 3 and 7 days after MI uncovered a trajectory from recruited monocytes to inflammatory TF+/TREM (triggered receptor expressed on myeloid cells) 1+ macrophages requiring F7. As early as 7 days after MI, macrophage F7 deletion led to an expansion of reparative Olfml 3 (olfactomedin-like protein 3)+ macrophages and, conversely, to a reduction of TF+/TREM1+ macrophages, which were also reduced in PAR2R38E mice. Short-term treatment from days 1 to 5 after nonreperfused MI with a monoclonal antibody inhibiting the macrophage TF-FVIIa-PAR2 signaling complex without anticoagulant activity improved cardiac dysfunction, decreased excess fibrosis, attenuated vascular endothelial dysfunction, and increased survival 28 days after MI. CONCLUSIONS: Extravascular TF-FVIIa-PAR2 complex signaling drives inflammatory macrophage polarization in ischemic heart disease. Targeting this signaling complex for specific therapeutic macrophage reprogramming following MI attenuates cardiac fibrosis and improves cardiovascular function.
Assuntos
Macrófagos , Camundongos Endogâmicos C57BL , Infarto do Miocárdio , Receptor PAR-2 , Remodelação Ventricular , Animais , Macrófagos/metabolismo , Infarto do Miocárdio/metabolismo , Infarto do Miocárdio/patologia , Infarto do Miocárdio/genética , Receptor PAR-2/metabolismo , Receptor PAR-2/genética , Receptor PAR-2/deficiência , Camundongos , Fator VIIa/metabolismo , Masculino , Transdução de Sinais , Camundongos Knockout , Fator de Crescimento Transformador beta1/metabolismo , Integrina beta1/metabolismo , Integrina beta1/genética , Tromboplastina/metabolismo , Tromboplastina/genética , FibroseRESUMO
Primordial germ cells (PGCs) are the precursors of gametes and the sole mechanism by which animals transmit genetic information across generations. In the mouse embryo, the transcriptional and epigenetic regulation of PGC specification has been extensively characterized. However, the initial event that triggers the soma-germline segregation remains poorly understood. Here, we uncover a critical role for the basement membrane in regulating germline entry. We show that PGCs arise in a region of the mouse embryo that lacks contact with the basement membrane, and the addition of exogenous extracellular matrix (ECM) inhibits both PGC and PGC-like cell (PGCLC) specification in mouse embryos and stem cell models, respectively. Mechanistically, we demonstrate that the engagement of ß1 integrin with laminin blocks PGCLC specification by preventing the Wnt signaling-dependent down-regulation of the PGC transcriptional repressor, Otx2. In this way, the physical segregation of cells away from the basement membrane acts as a morphogenetic fate switch that controls the soma-germline bifurcation.
Assuntos
Células Germinativas , Células-Tronco Pluripotentes , Animais , Camundongos , Células Germinativas/metabolismo , Células Germinativas/citologia , Células-Tronco Pluripotentes/metabolismo , Células-Tronco Pluripotentes/citologia , Transdução de Sinais , Integrinas/metabolismo , Integrinas/genética , Membrana Basal/metabolismo , Via de Sinalização Wnt , Diferenciação Celular , Matriz Extracelular/metabolismo , Laminina/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Integrina beta1/metabolismo , Integrina beta1/genética , Fatores de Transcrição Otx/metabolismo , Fatores de Transcrição Otx/genética , Embrião de Mamíferos/metabolismo , Embrião de Mamíferos/citologiaRESUMO
Integrins are fundamental for cell adhesion and the formation of focal adhesions (FA). Accordingly, these receptors guide embryonic development, tissue maintenance, and haemostasis but are also involved in cancer invasion and metastasis. A detailed understanding of the molecular interactions that drive integrin activation, FA assembly, and downstream signalling cascades is critical. Here, we reveal a direct association of paxillin, a marker protein of FA sites, with the cytoplasmic tails of the integrin ß1 and ß3 subunits. The binding interface resides in paxillin's LIM3 domain, where based on the NMR structure and functional analyses, a flexible, 7-amino acid loop engages the unstructured part of the integrin cytoplasmic tail. Genetic manipulation of the involved residues in either paxillin or integrin ß3 compromises cell adhesion and motility of murine fibroblasts. This direct interaction between paxillin and the integrin cytoplasmic domain identifies an alternative, kindlin-independent mode of integrin outside-in signalling particularly important for integrin ß3 function.
Assuntos
Paxilina , Ligação Proteica , Paxilina/metabolismo , Animais , Camundongos , Domínios Proteicos , Adesão Celular/fisiologia , Adesões Focais/metabolismo , Humanos , Movimento Celular , Integrina beta3/metabolismo , Integrina beta3/genética , Integrina beta3/química , Fibroblastos/metabolismo , Cadeias beta de Integrinas/metabolismo , Cadeias beta de Integrinas/química , Cadeias beta de Integrinas/genética , Integrina beta1/metabolismo , Transdução de SinaisRESUMO
Integrin activation resulting in enhanced adhesion to the extracellular matrix plays a key role in fundamental cellular processes. Although integrin activation has been extensively studied in circulating cells such as leukocytes and platelets, much less is known about the regulation and functional impact of integrin activation in adherent cells such as smooth muscle. Here, we show that two different asthmagenic cytokines, IL-13 and IL-17A, activate type I and IL-17 cytokine receptor families, respectively, to enhance adhesion of airway smooth muscle. These cytokines also induce activation of ß1 integrins detected by the conformation-specific antibody HUTS-4. Moreover, HUTS-4 binding is increased in the smooth muscle of patients with asthma compared to nonsmokers without lung disease, suggesting a disease-relevant role for integrin activation in smooth muscle. Indeed, integrin activation induced by the ß1-activating antibody TS2/16, the divalent cation manganese, or the synthetic peptide ß1-CHAMP that forces an extended-open integrin conformation dramatically enhances force transmission in smooth muscle cells and airway rings even in the absence of cytokines. We demonstrate that cytokine-induced activation of ß1 integrins is regulated by a common pathway of NF-κB-mediated induction of RhoA and its effector Rho kinase, which in turn stimulates PIP5K1γ-mediated synthesis of PIP2 at focal adhesions, resulting in ß1 integrin activation. Taken together, these data identify a pathway by which type I and IL-17 cytokine receptor family stimulation induces functionally relevant ß1 integrin activation in adherent smooth muscle and help to explain the exaggerated force transmission that characterizes chronic airway diseases such as asthma.
Assuntos
Asma , Integrina beta1 , Interleucina-13 , Interleucina-17 , Músculo Liso , NF-kappa B , Quinases Associadas a rho , Humanos , Integrina beta1/metabolismo , Interleucina-17/metabolismo , Músculo Liso/metabolismo , NF-kappa B/metabolismo , Quinases Associadas a rho/metabolismo , Interleucina-13/metabolismo , Asma/metabolismo , Transdução de Sinais , Adesão Celular , Miócitos de Músculo Liso/metabolismo , AnimaisRESUMO
High invasive capacity and acquired tyrosine kinase inhibitors (TKI) resistance of kidney renal clear cell carcinoma (KIRC) cells remain obstacles to prolonging the survival time of patients with advanced KIRC. In the present study, we reported that sine oculis homeobox 1 (SIX1) was upregulated in sunitinib-resistant KIRC cells and metastatic KIRC tissues. Subsequently, we found that SIX1 mediated metastasis and sunitinib resistance via Focal adhesion (FA) signaling, and knockdown of SIX1 enhanced the antitumor efficiency of sunitinib in KIRC. Mechanistically, Integrin subunit beta 1 (ITGB1), an upstream gene of FA signaling, was a direct transcriptional target of SIX1. In addition, we showed that DExH-box helicase 9 (DHX9) was an important mediator for SIX1-induced ITGB1 transcription, and silencing the subunits of SIX1/DHX9 complex significantly reduced transcription of ITGB1. Downregulation of SIX1 attenuated nuclear translocation of DHX9 and abrogated the binding of DHX9 to ITGB1 promoter. Collectively, our results unveiled a new signal axis SIX1/ITGB1/FAK in KIRC and identified a novel therapeutic strategy for metastatic KIRC patients.
Assuntos
Carcinoma de Células Renais , RNA Helicases DEAD-box , Resistencia a Medicamentos Antineoplásicos , Adesões Focais , Regulação Neoplásica da Expressão Gênica , Proteínas de Homeodomínio , Integrina beta1 , Neoplasias Renais , Metástase Neoplásica , Transdução de Sinais , Sunitinibe , Humanos , Resistencia a Medicamentos Antineoplásicos/genética , Carcinoma de Células Renais/genética , Carcinoma de Células Renais/patologia , Carcinoma de Células Renais/tratamento farmacológico , Carcinoma de Células Renais/metabolismo , Sunitinibe/farmacologia , Sunitinibe/uso terapêutico , Neoplasias Renais/patologia , Neoplasias Renais/genética , Neoplasias Renais/tratamento farmacológico , Neoplasias Renais/metabolismo , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Linhagem Celular Tumoral , Integrina beta1/genética , Integrina beta1/metabolismo , Animais , Adesões Focais/genética , Adesões Focais/metabolismo , RNA Helicases DEAD-box/genética , RNA Helicases DEAD-box/metabolismo , Camundongos , Transcrição Gênica , Integrinas/metabolismo , Integrinas/genética , Quinase 1 de Adesão FocalRESUMO
BACKGROUND: Platelets prevent bleeding in a variety of inflammatory settings, the adhesion receptors and activation pathways involved being highly context-dependent and functionally redundant. In some situations, platelets recruited to inflammatory sites act independently of aggregation. The mechanisms underlying stable platelet adhesion in inflamed microvessels remain incompletely understood, in particular, whether and if so, how ß1 and ß3 integrins are involved. METHODS: The impact of isolated or combined platelet deficiency in ß1 and ß3 integrins on inflammation-associated hemostasis was investigated in 3 models of acute inflammation: immune complex-based cutaneous reverse passive Arthus reaction, intranasal lipopolysaccharide-induced lung inflammation, and cerebral ischemia-reperfusion following transient (2-hour) occlusion of the middle cerebral artery. RESULTS: Mice with platelet-directed inactivation of Itgb1 (PF4Cre-ß1-/-) displayed no bleeding in any of the inflammation models, while mice defective in platelet Itgb3 (PF4Cre-ß3-/-) exhibited bleeding in all 3 models. Remarkably, the bleeding phenotype of PF4Cre-ß3-/- mice was exacerbated in the reverse passive Arthus model by the concomitant deletion of ß1 integrins, PF4Cre-ß1-/-/ß3-/- animals presenting increased bleeding. Intravital microscopy in reverse passive Arthus experiments highlighted a major defect in the adhesion of PF4Cre-ß1-/-/ß3-/- platelets to inflamed microvessels. Unlike PF4Cre-ß1-/- and PF4Cre-ß3-/- mice, PF4Cre-ß1-/-/ß3-/- animals developed early hemorrhagic transformation 6 hours after transient middle cerebral artery occlusion. PF4Cre-ß1-/-/ß3-/- mice displayed no more bleeding in lipopolysaccharide-induced lung inflammation than PF4Cre-ß3-/- animals. CONCLUSIONS: Altogether, these results show that the requirement for and degree of functional redundancy between platelet ß1 and ß3 integrins in inflammation-associated hemostasis vary with the inflammatory situation.
Assuntos
Plaquetas , Modelos Animais de Doenças , Hemorragia , Integrina beta1 , Integrina beta3 , Camundongos Endogâmicos C57BL , Camundongos Knockout , Animais , Masculino , Camundongos , Plaquetas/metabolismo , Hemorragia/genética , Hemorragia/sangue , Hemostasia , Infarto da Artéria Cerebral Média/genética , Infarto da Artéria Cerebral Média/patologia , Infarto da Artéria Cerebral Média/sangue , Infarto da Artéria Cerebral Média/metabolismo , Inflamação/genética , Inflamação/metabolismo , Inflamação/sangue , Integrina beta1/metabolismo , Integrina beta1/genética , Integrina beta3/genética , Integrina beta3/metabolismo , Lipopolissacarídeos , Adesividade Plaquetária , Pneumonia/genética , Pneumonia/sangue , Pneumonia/metabolismo , Pneumonia/patologia , Traumatismo por Reperfusão/genética , Traumatismo por Reperfusão/metabolismo , Traumatismo por Reperfusão/sangueRESUMO
BACKGROUND: Irreversible pulmonary fibrosis induced by paraquat is the most prevalent cause of death in patients with paraquat poisoning. Pulmonary fibrosis is characterized by abnormal deposition of extracellular matrix (ECM). Currently, the role of fibrotic ECM microenvironment in paraquat-induced pulmonary fibrosis has not been established. METHODS: Rat pulmonary fibrosis model was induced by paraquat, ATN-161 (an integrin-ß1 antagonist) was given to investigate their effect on Rat survival and pulmonary fibrosis. Lungs were decellularized to generate normal and fibrotic acellular ECM scaffolds using Triton and SDS. Fibroblasts were cocultured with ECM scaffolds to established 3D culture systems to investigate the relationship between fibrotic ECM and the differentiation of fibroblasts. Then we explored the effect of fibrotic ECM microenvironment systematically promoting on integrin-ß1/FAK/ERK1/2 pathway and established 3D culture systems to investigate the relationship between fibrotic ECM and the differentiation of fibroblasts. RESULTS: Antagonism of integrin-ß1 could alleviate paraquat-induced pulmonary fibrosis and ameliorate survival status of rats. Compared to normal ECM, fibrotic extracellular microenvironment promoted the differentiation of fibroblasts to myofibroblasts. Antagonism of integrin-ß1 could also ameliorate the promotion of fibrotic extracellular microenvironment on differentiation of fibroblasts to myofibroblasts. Fibrotic ECM microenvironment promotes fibroblasts transforming into myofibroblasts through integrin-ß1/FAK/ERK1/2 signaling pathway. Moreover, this phenomenon holds independent on exogenous integrin-ß1. CONCLUSIONS: Activation of integrin-ß1/FAK/ERK1/2 pathway aggravates paraquat-induced pulmonary fibrosis depend on fibrotic ECM and integrin-ß1 may be a prospective therapeutic target for paraquat-induced pulmonary fibrosis in the future.
Assuntos
Matriz Extracelular , Fibroblastos , Integrina beta1 , Sistema de Sinalização das MAP Quinases , Paraquat , Fibrose Pulmonar , Ratos Sprague-Dawley , Animais , Paraquat/toxicidade , Fibrose Pulmonar/induzido quimicamente , Fibrose Pulmonar/patologia , Fibrose Pulmonar/metabolismo , Matriz Extracelular/metabolismo , Masculino , Fibroblastos/efeitos dos fármacos , Fibroblastos/patologia , Fibroblastos/metabolismo , Ratos , Integrina beta1/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Quinase 1 de Adesão Focal/metabolismo , Pulmão/patologia , Pulmão/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Miofibroblastos/metabolismo , Miofibroblastos/patologia , Miofibroblastos/efeitos dos fármacos , Células Cultivadas , Modelos Animais de DoençasRESUMO
Talin2 is localized to large focal adhesions and is indispensable for traction force generation, invadopodium formation, cell invasion as well as metastasis. Talin2 has a higher affinity toward ß-integrin tails than talin1. Moreover, disruption of the talin2-ß-integrin interaction inhibits traction force generation, invadopodium formation and cell invasion, indicating that a strong talin2-ß-integrin interaction is required for talin2 to fulfill these functions. Nevertheless, the role of talin2 in mediation of these processes remains unknown. Here we show that talin2 binds to the N-terminus of non-muscle myosin IIA (NMIIA) through its F3 subdomain. Moreover, talin2 co-localizes with NMIIA at cell edges as well as at some cytoplasmic spots. Talin2 also co-localizes with cortactin, an invadopodium marker. Furthermore, overexpression of NMIIA promoted the talin2 head binding to the ß1-integrin tail, whereas knockdown of NMIIA reduced fibronectin and matrix metalloproteinase secretion as well as inhibited cell attachment on fibronectin-coated substrates. These results suggest that talin2 binds to NMIIA to control the secretion of extracellular matrix proteins and this interaction modulates cell adhesion.
Assuntos
Adesão Celular , Fibronectinas , Miosina não Muscular Tipo IIA , Ligação Proteica , Talina , Animais , Humanos , Cortactina/metabolismo , Fibronectinas/metabolismo , Adesões Focais/metabolismo , Integrina beta1/metabolismo , Miosina não Muscular Tipo IIA/metabolismo , Podossomos/metabolismo , Talina/metabolismo , CamundongosRESUMO
OBJECTIVE: The purpose of this study was to investigate the role and possible mechanism of lncRNA XIST in renal fibrosis and to provide potential endogenous targets for renal fibrosis in obstructive nephropathy (ON). METHODS: The study included 50 cases of ON with renal fibrosis (samples taken from patients undergoing nephrectomy due to ON) and 50 cases of normal renal tissue (samples taken from patients undergoing total or partial nephrectomy due to accidental injury, congenital malformations, and benign tumors). Treatment of human proximal renal tubular epithelium (HK-2) cells with TGF-ß1 simulated renal fibrosis in vitro. Cell viability and proliferation were measured by CCK-8 and EdU, and cell migration was measured by transwell. XIST, miR-124-3p, ITGB1, and epithelial-mesenchymal transition (EMT)-related proteins (E-cadherin, α-SMA, and fibronectin) were detected by PCR and immunoblot. The targeting relationship between miR-124-3p and XIST or ITGB1 was verified by starBase and dual luciferase reporter gene experiments. In addition, The left ureter was ligated in mice as a model of unilateral ureteral obstruction (UUO), and the renal histopathology was observed by HE staining and Masson staining. RESULTS: ON patients with renal fibrosis had elevated XIST and ITGB1 levels and reduced miR-124-3p levels. The administration of TGF-ß1 exhibited a dose-dependent promotion of HK-2 cell viability, proliferation, migration, and EMT. Conversely, depleting XIST or enhancing miR-124-3p hindered HK-2 cell viability, proliferation, migration, and EMT in TGF-ß1-damaged HK-2 cells HK-2 cells. XIST functioned as a miR-124-3p sponge. Additionally, miR-124-3p negatively regulated ITGB1 expression. Elevating ITGB1 weakened the impact of XIST depletion on TGF-ß1-damaged HK-2 cells. Down-regulating XIST improved renal fibrosis in UUO mice. CONCLUSION: XIST promotes renal fibrosis in ON by elevating miR-124-3p and reducing ITGB1 expressions.
Assuntos
Transição Epitelial-Mesenquimal , Fibrose , Nefropatias , MicroRNAs , RNA Longo não Codificante , RNA Longo não Codificante/genética , MicroRNAs/genética , MicroRNAs/metabolismo , Humanos , Fibrose/genética , Fibrose/patologia , Fibrose/metabolismo , Animais , Camundongos , Nefropatias/genética , Nefropatias/patologia , Nefropatias/metabolismo , Transição Epitelial-Mesenquimal/genética , Integrina beta1/metabolismo , Integrina beta1/genética , Proliferação de Células , Masculino , Movimento Celular/genética , Rim/patologia , Rim/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , Fator de Crescimento Transformador beta1/genética , Linhagem Celular , Feminino , Obstrução Ureteral/patologia , Obstrução Ureteral/metabolismo , Obstrução Ureteral/genéticaRESUMO
The blood-brain barrier (BBB) acquires unique properties to regulate neuronal function during development. The formation of the BBB, which occurs in tandem with angiogenesis, is directed by the Wnt/ß-catenin signaling pathway. Yet the exact molecular interplay remains elusive. Our study reveals the G protein-coupled receptor GPR126 as a critical target of canonical Wnt signaling, essential for the development of the BBB's distinctive vascular characteristics and its functional integrity. Endothelial cell-specific deletion of the Gpr126 gene in mice induced aberrant vascular morphogenesis, resulting in disrupted BBB organization. Simultaneously, heightened transcytosis in vitro compromised barrier integrity, resulting in enhanced vascular permeability. Mechanistically, GPR126 enhanced endothelial cell migration, pivotal for angiogenesis, acting through an interaction between LRP1 and ß1 integrin, thereby balancing the levels of ß1 integrin activation and recycling. Overall, we identified GPR126 as a specifier of an organotypic vascular structure, which sustained angiogenesis and guaranteed the acquisition of the BBB properties during development.
Assuntos
Barreira Hematoencefálica , Integrina beta1 , Receptores Acoplados a Proteínas G , Animais , Camundongos , Barreira Hematoencefálica/metabolismo , Permeabilidade Capilar , Movimento Celular , Células Endoteliais/metabolismo , Integrina beta1/metabolismo , Integrina beta1/genética , Proteína-1 Relacionada a Receptor de Lipoproteína de Baixa Densidade/metabolismo , Proteína-1 Relacionada a Receptor de Lipoproteína de Baixa Densidade/genética , Camundongos Knockout , Neovascularização Fisiológica , Receptores Acoplados a Proteínas G/metabolismo , Receptores Acoplados a Proteínas G/genética , Via de Sinalização Wnt , Masculino , FemininoRESUMO
The paracrine actions of adipokine plasminogen activator inhibitor-1 (PAI-1) are implicated in obesity-associated tumorigenesis. Here, we show that PAI-1 mediates extracellular matrix (ECM) signaling via epigenetic repression of DKK1 in endometrial epithelial cells (EECs). While the loss of DKK1 is known to increase ß-catenin accumulation for WNT signaling activation, this epigenetic repression causes ß-catenin release from transmembrane integrins. Furthermore, PAI-1 elicits the disengagement of TIMP2 and SPARC from integrin-ß1 on the cell surface, lifting an integrin-ß1-ECM signaling constraint. The heightened interaction of integrin-ß1 with type 1 collagen (COL1) remodels extracellular fibrillar structures in the ECM. Consequently, the enhanced nanomechanical stiffness of this microenvironment is conducive to EEC motility and neoplastic transformation. The formation of extensively branched COL1 fibrils is also observed in endometrial tumors of patients with obesity. The findings highlight PAI-1 as a contributor to enhanced integrin-COL1 engagement and extensive ECM remodeling during obesity-associated neoplastic development.
Assuntos
Matriz Extracelular , Integrina beta1 , Obesidade , Inibidor 1 de Ativador de Plasminogênio , beta Catenina , Humanos , Obesidade/metabolismo , Obesidade/patologia , Feminino , Inibidor 1 de Ativador de Plasminogênio/metabolismo , beta Catenina/metabolismo , Integrina beta1/metabolismo , Matriz Extracelular/metabolismo , Neoplasias do Endométrio/metabolismo , Neoplasias do Endométrio/patologia , Inibidor Tecidual de Metaloproteinase-2/metabolismo , Animais , Osteonectina/metabolismo , Osteonectina/genética , Colágeno/metabolismo , Endométrio/metabolismo , Endométrio/patologia , Colágeno Tipo I/metabolismo , Membrana Celular/metabolismo , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Peptídeos e Proteínas de Sinalização IntercelularRESUMO
Cementum is a vital component of periodontium, yet its regeneration remains a challenge. Pentraxin 3 (PTX3) is a multifunctional glycoprotein involved in extracellular matrix remodeling and bone metabolism regulation. However, the role of PTX3 in cementum formation and cementoblast differentiation has not been elucidated. In this study, we initially observed an increase in PTX3 expression during cementum formation and cementoblast differentiation. Then, overexpression of PTX3 significantly enhanced the differentiation ability of cementoblasts. While conversely, PTX3 knockdown exerted an inhibitory effect. Moreover, in Ptx3-deficient mice, we found that cementum formation was hampered. Furthermore, we confirmed the presence of PTX3 within the hyaluronan (HA) matrix, thereby activating the ITGB1/FAK/YAP1 signaling pathway. Notably, inhibiting any component of this signaling pathway partially reduced the ability of PTX3 to promote cementoblast differentiation. In conclusion, our study indicated that PTX3 promotes cementum formation and cementoblast differentiation, which is partially dependent on the HA/ITGB1/FAK/YAP1 signaling pathway. This research will contribute to our understanding of cementum regeneration after destruction.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal , Diferenciação Celular , Cemento Dentário , Transdução de Sinais , Proteínas de Sinalização YAP , Animais , Cemento Dentário/metabolismo , Proteínas de Sinalização YAP/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Camundongos , Proteína C-Reativa/metabolismo , Integrina beta1/metabolismo , Componente Amiloide P Sérico/metabolismo , Componente Amiloide P Sérico/genética , Camundongos Endogâmicos C57BL , Quinase 1 de Adesão Focal/metabolismo , Quinase 1 de Adesão Focal/genética , CementogêneseRESUMO
Dermal fibroblasts deposit type I collagen, the dominant extracellular matrix molecule found in skin, during early postnatal development. Coincident with this biosynthetic program, fibroblasts proteolytically remodel pericellular collagen fibrils by mobilizing the membrane-anchored matrix metalloproteinase, Mmp14. Unexpectedly, dermal fibroblasts in Mmp14-/- mice commit to a large-scale apoptotic program that leaves skin tissues replete with dying cells. A requirement for Mmp14 in dermal fibroblast survival is recapitulated in vitro when cells are embedded within, but not cultured atop, three-dimensional hydrogels of crosslinked type I collagen. In the absence of Mmp14-dependent pericellular proteolysis, dermal fibroblasts fail to trigger ß1 integrin activation and instead actuate a TGF-ß1/phospho-JNK stress response that leads to apoptotic cell death in vitro as well as in vivo. Taken together, these studies identify Mmp14 as a requisite cell survival factor that maintains dermal fibroblast viability in postnatal dermal tissues.
Assuntos
Apoptose , Sobrevivência Celular , Fibroblastos , Metaloproteinase 14 da Matriz , Animais , Metaloproteinase 14 da Matriz/metabolismo , Metaloproteinase 14 da Matriz/genética , Fibroblastos/metabolismo , Camundongos , Camundongos Knockout , Colágeno Tipo I/metabolismo , Colágeno Tipo I/genética , Integrina beta1/metabolismo , Integrina beta1/genética , Fator de Crescimento Transformador beta1/metabolismo , Derme/metabolismo , Derme/citologia , Células Cultivadas , Matriz Extracelular/metabolismo , Camundongos Endogâmicos C57BL , Pele/metabolismoRESUMO
Cancer cells can aberrantly express various markers, including transferrin receptor 1 (CD71) and ß1-integrin molecules. Their role in invasion, migration and metastasis has been demonstrated. Determination of their expression in breast cancer (BC) may be an important point to characterize the clinical course of the tumor and prognosis of the disease. OBJECTIVE: To study of transferrin receptor 1 (CD71) expression by primary breast cancer cells in correlation with tumor cell phenotype. MATERIAL AND METHODS: Determination of BC phenotype: immunohistochemical staining method (immunofluorescence). Antibodies to ER (estrogen receptors), KL-1 (pancytokeratin), CD71 (transferrin receptor), CD29 (ß1-integrins). CD45, CD3, CD4, CD8, CD20 infiltration was also evaluated. ZEISS microscope (AXIOSKOP; Germany), method of G.J. Hammerling et al. Statistical processing: IBM-SPSS Statistics v.21. RESULTS: 63% of BC cases had CD71+ phenotype. CD71-mosaic tumors were observed in 14.4%. ß1-integrin expression was monomorphic in 51.6% of cases and mosaic in 38.7%. 85% of ER-positive tumors were CD71-positive with a monomorphic type of reaction; p=0.014. Among ER-negative tumors, CD71-negative reactions were 2-fold more frequent and the monomorphic type was less frequent. ER-positive tumors were CD29-positive in 73%; p=0.031. 45.5% of ER+ tumors were CD29-monomorphic. Among ER-negative tumors, the frequency of CD29-monomorphic tumors was 55%. Significant infiltration by CD3+ cells was predominant in CD71-positive tumors; p=0.016. In the CD29-monomorphic phenotype, CD45+ infiltration was 31.3%, and in the mosaic phenotype, 67.1%. CONCLUSION: BC aberrantly expresses transferrin receptors, ß1-integrins. CD71 expression is associated with ER expression. ER-positive tumors are often monomorphic for CD71. Prominent CD3+ infiltration was present in CD71+ tumors. Expression of ß1-integrins correlated with ER+ status and weak immune infiltration.
Assuntos
Antígenos CD , Neoplasias da Mama , Integrina beta1 , Receptores de Estrogênio , Receptores da Transferrina , Humanos , Receptores da Transferrina/metabolismo , Neoplasias da Mama/patologia , Neoplasias da Mama/imunologia , Neoplasias da Mama/metabolismo , Neoplasias da Mama/genética , Feminino , Integrina beta1/metabolismo , Receptores de Estrogênio/metabolismo , Pessoa de Meia-Idade , Regulação Neoplásica da Expressão Gênica , Adulto , Idoso , Biomarcadores Tumorais/metabolismo , Proteínas de Neoplasias/biossíntese , Proteínas de Neoplasias/imunologiaRESUMO
More than 90% of hepatocellular carcinoma (HCC) cases develop in the presence of fibrosis or cirrhosis, making the tumor microenvironment (TME) of HCC distinctive due to the intricate interplay between cancer-associated fibroblasts (CAFs) and cancer stem cells (CSCs), which collectively regulate HCC progression. However, the mechanisms through which CSCs orchestrate the dynamics of the tumor stroma during HCC development remain elusive. Our study unveils a significant upregulation of Sema3C in fibrotic liver, HCC tissues, peripheral blood of HCC patients, as well as sorafenib-resistant tissues and cells, with its overexpression correlating with the acquisition of stemness properties in HCC. We further identify NRP1 and ITGB1 as pivotal functional receptors of Sema3C, activating downstream AKT/Gli1/c-Myc signaling pathways to bolster HCC self-renewal and tumor initiation. Additionally, HCC cells-derived Sema3C facilitated extracellular matrix (ECM) contraction and collagen deposition in vivo, while also promoting the proliferation and activation of hepatic stellate cells (HSCs). Mechanistically, Sema3C interacted with NRP1 and ITGB1 in HSCs, activating downstream NF-kB signaling, thereby stimulating the release of IL-6 and upregulating HMGCR expression, consequently enhancing cholesterol synthesis in HSCs. Furthermore, CAF-secreted TGF-ß1 activates AP1 signaling to augment Sema3C expression in HCC cells, establishing a positive feedback loop that accelerates HCC progression. Notably, blockade of Sema3C effectively inhibits tumor growth and sensitizes HCC cells to sorafenib in vivo. In sum, our findings spotlight Sema3C as a novel biomarker facilitating the crosstalk between CSCs and stroma during hepatocarcinogenesis, thereby offering a promising avenue for enhancing treatment efficacy and overcoming drug resistance in HCC.