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1.
J Transl Med ; 19(1): 383, 2021 09 08.
Artigo em Inglês | MEDLINE | ID: mdl-34496870

RESUMO

BACKGROUND: Antiangiogenic therapy has increasingly become an important strategy for the treatment of colorectal cancer. Recent studies have shown that the tumour microenvironment (TME) promotes tumour angiogenesis. Bufalin is an active antitumour compound whose efficacy has been indicated by previous studies. However, there are very few studies on the antiangiogenic effects of bufalin. METHODS: Herein, human umbilical vein endothelial cell (HUVEC) tube formation, migration and adhesion tests were used to assess angiogenesis in vitro. Western blotting and quantitative PCR were used to detect relevant protein levels and mRNA expression levels. A subcutaneous xenograft tumour model and a hepatic metastasis model were established in mice to investigate the influence of bufalin on angiogenesis mediated by the TME in vivo. RESULTS: We found that angiogenesis mediated by cells in the TME was significantly inhibited in the presence of bufalin. The results demonstrated that the proangiogenic genes in HUVECs, such as VEGF, PDGFA, E-selectin and P-selectin, were downregulated by bufalin and that this downregulation was mediated by inhibition of the STAT3 pathway. Overexpression of STAT3 reversed the inhibitory effects of bufalin on angiogenesis. Furthermore, there was little reduction in angiogenesis when bufalin directly acted on the cells in the tumour microenvironment. CONCLUSION: Our findings demonstrate that bufalin suppresses tumour microenvironment-mediated angiogenesis by inhibiting the STAT3 signalling pathway in vascular endothelial cells, revealing that bufalin may be used as a new antiangiogenic adjuvant therapy medicine to treat colorectal cancer.


Assuntos
Neoplasias Hepáticas , Microambiente Tumoral , Inibidores da Angiogênese/farmacologia , Animais , Bufanolídeos , Movimento Celular , Proliferação de Células , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Neoplasias Hepáticas/tratamento farmacológico , Camundongos , Camundongos Nus , Neovascularização Patológica/tratamento farmacológico , Fator de Transcrição STAT3/metabolismo
2.
Mater Sci Eng C Mater Biol Appl ; 128: 112274, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34474833

RESUMO

Corneal neovascularization (CNV) is one of the main factors that induce blindness worldwide. To effectively inhibit CNV, a novel nanohybrid has been developed by incorporating anti-VEGF bevacizumab (BEV)-loaded mesoporous silica nanoparticles (BEV@MSN) into the thermogel matrix with anti-inflammation cyclosporine A (CsA) (BEV@MSN-CsA@Thermogel). This nanohybrid regulates the in vitro release of both bevacizumab and cyclosporine A in a sustainable way for up to four weeks to enhance CNV inhibition through the synergistic anti-VEGF and anti-inflammation. The carrier materials (i.e. silica and thermogel) in this nanohybrid do not show any cytotoxicity to human Tenon's fibroblasts, corneal epithelial cells and corneal endothelial cells. BEV@MSN-CsA@Thermogel effectively prevents proliferation, migration, and tube-like structure formation of human umbilical vein endothelial cells. Moreover, subconjunctival injection of BEV@MSN-CsA@Thermogel significantly inhibits corneal neovascularization in terms of the CNV area, the new vessel length, the corneal opaque area, the corneal inflammation and abnormal fibrosis in a rabbit model. This nanohybrid is thus a promising alternative for effective CNV treatment.


Assuntos
Neovascularização da Córnea , Preparações Farmacêuticas , Animais , Bevacizumab/farmacologia , Neovascularização da Córnea/tratamento farmacológico , Modelos Animais de Doenças , Células Endoteliais da Veia Umbilical Humana , Humanos , Coelhos , Dióxido de Silício , Fator A de Crescimento do Endotélio Vascular
3.
Rev Assoc Med Bras (1992) ; 67(4): 555-560, 2021 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-34495060

RESUMO

OBJECTIVE: In this study, we aimed at investigating the role of isoleucyl-tRNA synthetase in the growth, migration, and angiogenesis of human umbilical vein endothelial cells and the underlying molecular mechanism. METHODS: To assess the role of isoleucyl-tRNA synthetase, we silenced isoleucyl-tRNA synthetase in human umbilical vein endothelial cells using lentiviral 2 specific short hairpin RNAs (short hairpin RNAs 1 and 2) and examined silencing efficiency using real time quantitative polymerase chain reaction and western blot analyses. Short hairpin RNAs 1-isoleucyl-tRNA synthetase had greater knockdown efficiency, it was used in the entire downstream analysis. Short hairpin RNAs 1- isoleucyl-tRNA synthetase silencing effects on cell proliferation, cell colony generation, cell migration, as well as angiogenesis were assessed using cell counting kit-8, colony development, cell migration, and angiogenesis tube formation assays, respectively. RESULTS: Compared to the control group, anti-isoleucyl-tRNA synthetase short hairpin RNAs significantly silenced isoleucyl-tRNA synthetase expression in human umbilical vein endothelial cells, and suppressed their proliferation, migration, and angiogenic capacity. To characterize the underlying mechanism, western blot analyses showed that isoleucyl-tRNA synthetase knockdown suppressed phosphorylation of extracellular-regulated kinase ½ and protein-serine- threonine kinase, as well as expression of vascular endothelial growth factor, GSK-3ß, and ß-catenin. CONCLUSIONS: We have shown, for the first time, the critical role of isoleucyl-tRNA synthetase in human umbilical vein endothelial cells. Our data show that isoleucyl-tRNA synthetase knockdown suppresses human umbilical vein endothelial cell proliferation, migration, and angiogenesis. We have also shown that isoleucyl-tRNA synthetase knockdown suppresses phosphorylation of extracellular-regulated kinase ½ and protein-serine- threonine kinase, as well as expression of vascular endothelial growth factor, GSK-3ß, and ß-catenin. Together, these data highlight isoleucyl-tRNA synthetase as a potential antitumor anti-angiogenic target.


Assuntos
Fator A de Crescimento do Endotélio Vascular , Proliferação de Células , Células Cultivadas , Glicogênio Sintase Quinase 3 beta , Células Endoteliais da Veia Umbilical Humana , Humanos
4.
Int J Mol Sci ; 22(16)2021 Aug 11.
Artigo em Inglês | MEDLINE | ID: mdl-34445337

RESUMO

In fibrotic diseases, myofibroblasts derive from a range of cell types including endothelial-to-mesenchymal transition (EndMT). Increasing evidence suggests that miRNAs are key regulators in biological processes but their profile is relatively understudied in EndMT. In human umbilical vein endothelial cells (HUVEC), EndMT was induced by treatment with TGFß2 and IL1ß. A significant decrease in endothelial markers such as VE-cadherin, CD31 and an increase in mesenchymal markers such as fibronectin were observed. In parallel, miRNA profiling showed that miR-126-3p was down-regulated in HUVECs undergoing EndMT and over-expression of miR-126-3p prevented EndMT, maintaining CD31 and repressing fibronectin expression. EndMT was investigated using lineage tracing with transgenic Cdh5-Cre-ERT2; Rosa26R-stop-YFP mice in two established models of fibrosis: cardiac ischaemic injury and kidney ureteric occlusion. In both cardiac and kidney fibrosis, lineage tracing showed a significant subpopulation of endothelial-derived cells expressed mesenchymal markers, indicating they had undergone EndMT. In addition, miR-126-3p was restricted to endothelial cells and down-regulated in murine fibrotic kidney and heart tissue. These findings were confirmed in patient kidney biopsies. MiR-126-3p expression is restricted to endothelial cells and is down-regulated during EndMT. Over-expression of miR-126-3p reduces EndMT, therefore, it could be considered for miRNA-based therapeutics in fibrotic organs.


Assuntos
Transdiferenciação Celular/genética , Rim/patologia , MicroRNAs/fisiologia , Miocárdio/patologia , Animais , Células Cultivadas , Células Endoteliais/patologia , Células Endoteliais/fisiologia , Fibrose/genética , Células Endoteliais da Veia Umbilical Humana/metabolismo , Células Endoteliais da Veia Umbilical Humana/patologia , Células Endoteliais da Veia Umbilical Humana/fisiologia , Humanos , Rim/metabolismo , Células-Tronco Mesenquimais/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Miocárdio/metabolismo , Miofibroblastos/metabolismo , Miofibroblastos/patologia
5.
Molecules ; 26(15)2021 Jul 25.
Artigo em Inglês | MEDLINE | ID: mdl-34361636

RESUMO

Naturally-occurring halloysite nanotubes (HNTs) have many advantages for constructing target-specific delivery of phototherapeutic agents. Here, HNTs were labeled with fluorescein isothiocyanate (FITC) and loaded with the type-II photosensitizer indocyanine green (ICG) for phototherapy. HNTs-FITC-ICG was structurally stable due to presence of HNTs as the nanocarrier and protective agent. The nanocarrier was further wrapped with red blood cell membrane (RBCM) to enhance the biocompatibility. The HNTs-FITC-ICG-RBCM nanocarrier show high cytocompatibility and hemocompatibility. Due to the photothermal effect of ICG, a significant temperature rising was achieved by irradiation of the nanocarrier using 808 nm laser. The photothermal temperature rising was used to kill the cancer cells effectively. The HNTs-FITC-ICG-RBCM nanocarrier was further linked with anti-EpCAM to endow it with targeting therapy performance against breast cancer, and the anti-EpCAM-conjugated nanocarrier exhibited significantly tumor-specific accumulation. The RBCM-coated and biocompatible HNTs nanocarrier is a promising candidate for target-specific therapy of cancer.


Assuntos
Membrana Celular/química , Argila/química , Materiais Revestidos Biocompatíveis , Portadores de Fármacos , Nanotubos/química , Neoplasias , Terapia Fototérmica , Animais , Materiais Revestidos Biocompatíveis/química , Materiais Revestidos Biocompatíveis/farmacologia , Portadores de Fármacos/química , Portadores de Fármacos/farmacologia , Células Endoteliais da Veia Umbilical Humana , Humanos , Células MCF-7 , Neoplasias/metabolismo , Neoplasias/patologia , Neoplasias/terapia , Coelhos
6.
Molecules ; 26(16)2021 Aug 16.
Artigo em Inglês | MEDLINE | ID: mdl-34443534

RESUMO

Thrombosis is a disease that seriously endangers human health, with a high rate of mortality and disability. However, current treatments with thrombolytic drugs (such as recombinant tissue-plasminogen activator) and the oral anticoagulants (such as dabigatran and rivaroxaban) are reported to have a tendency of major or life-threatening bleeding, such as intracranial hemorrhage or massive gastrointestinal bleed with non-specific antidotes. In contrast, lumbrokinase is very specific to fibrin as a substrate and does not cause excessive bleeding. It can dissolve the fibrin by itself or convert plasminogen to plasmin by inducing endogenous t-PA activity to dissolve fibrin clots. Therefore, searching for potentially new therapeutic molecules from earthworms is significant. In this study, we first collected a strong fibrinolytic extract (PvQ) from the total protein of the Pheretima vulgaris with AKTA pure protein purification systems; its fibrinolytic bioactivity was verified by the fibrin plate assay and zebrafish thrombotic model of vascular damage. Furthermore, according to the cell culture model of human umbilical vein endothelial cells (HUVECs), the PvQ was proven to exhibit the ability to promote the secretion of tissue-type plasminogen activator (t-PA), which further illustrated that it has an indirect thrombolytic effect. Subsequently, extensive chromatographic techniques were applied to reveal the material basis of the extract. Fortunately, six novel earthworm fibrinolytic enzymes were obtained from the PvQ, and the primary sequences of those functional proteins were determined by LC-MS/MStranscriptome cross-identification and the Edman degradation assay. The secondary structures of these six fibrinolytic enzymes were determined by circular dichroism spectroscopy and the three-dimensional structures of these proteases were predicted by MODELLER 9.23 based on multi-template modelling. In addition, those six genes encoding blood clot-dissolving proteins were cloned from P. vulgaris by RT-PCR amplification, which further determined the accuracy of proteins primary sequences identifications and laid the foundation for subsequent heterologous expression.


Assuntos
Fibrinolíticos/isolamento & purificação , Fibrinolíticos/farmacologia , Oligoquetos/química , Peptídeo Hidrolases/farmacologia , Trombose/patologia , Sequência de Aminoácidos , Animais , Sequência de Bases , Sobrevivência Celular/efeitos dos fármacos , Bases de Dados de Proteínas , Eritrócitos/efeitos dos fármacos , Fibrinólise/efeitos dos fármacos , Fibrinolíticos/química , Células Endoteliais da Veia Umbilical Humana/citologia , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Humanos , Modelos Moleculares , Peptídeo Hidrolases/química , Peptídeo Hidrolases/genética , Estrutura Secundária de Proteína , Estrutura Terciária de Proteína , Ativador de Plasminogênio Tecidual/metabolismo , Peixe-Zebra
7.
Int J Mol Sci ; 22(16)2021 Aug 08.
Artigo em Inglês | MEDLINE | ID: mdl-34445234

RESUMO

Abiotrophia defectiva is a nutritionally variant streptococci that is found in the oral cavity, and it is an etiologic agent of infective endocarditis. We have previously reported the binding activity of A. defectiva to fibronectin and to human umbilical vein endothelial cells (HUVECs). However, the contribution of some adhesion factors on the binding properties has not been well delineated. In this study, we identified DnaK, a chaperon protein, as being one of the binding molecules of A. defectiva to fibronectin. Recombinant DnaK (rDnaK) bound immobilized fibronectin in a concentration-dependent manner, and anti-DnaK antiserum reduced the binding activity of A. defectiva with both fibronectin and HUVECs. Furthermore, DnaK were observed on the cell surfaces via immune-electroscopic analysis with anti-DnaK antiserum. Expression of IL-8, CCL2, ICAM-1, and VCAM-1 was upregulated with the A. defectiva rDnaK treatment in HUVECs. Furthermore, TNF-α secretion of THP-1 macrophages was also upregulated with the rDnaK. We observed these upregulations in rDnaK treated with polymyxin B, but not in the heat-treated rDnaK. The findings show that A. defectiva DnaK functions not only as an adhesin to HUVECs via the binding to fibronectin but also as a proinflammatory agent in the pathogenicity to cause infective endocarditis.


Assuntos
Abiotrophia/metabolismo , Aderência Bacteriana , Proteínas de Bactérias/metabolismo , Fibronectinas/metabolismo , Proteínas de Choque Térmico HSP70/metabolismo , Células Endoteliais da Veia Umbilical Humana/metabolismo , Abiotrophia/genética , Proteínas de Bactérias/genética , Proteínas de Choque Térmico HSP70/genética , Células Endoteliais da Veia Umbilical Humana/microbiologia , Humanos , Inflamação/genética , Inflamação/metabolismo , Inflamação/microbiologia
8.
Int J Mol Sci ; 22(16)2021 Aug 23.
Artigo em Inglês | MEDLINE | ID: mdl-34445773

RESUMO

Inadequate vessel maintenance or growth causes ischemia in diseases such as myocardial infarction, stroke, and neurodegenerative disorders. Therefore, developing an effective strategy to salvage ischemic tissues using a novel compound is urgent. Drug repurposing has become a widely used method that can make drug discovery more efficient and less expensive. Additionally, computational virtual screening tools make drug discovery faster and more accurate. This study found a novel drug candidate for pro-angiogenesis by in silico virtual screening. Using Gene Expression Omnibus (GEO) microarray datasets related to angiogenesis studies, differentially expressed genes were identified and characteristic direction signatures extracted from GEO2EnrichR were used as input data on L1000CDS2 to screen pro-angiogenic molecules. After a thorough review of the candidates, a list of compounds structurally similar to TWS-119 was generated using ChemMine Tools and its clustering toolbox. ChemMine Tools and ChemminR structural similarity search tools for small-molecule analysis and clustering were used for second screening. A molecular docking simulation was conducted using AutoDock v.4 to evaluate the physicochemical effect of secondary-screened chemicals. A cell viability or toxicity test was performed to determine the proper dose of the final candidate, ellipticine. As a result, we found ellipticine, which has pro-angiogenic effects, using virtual computational methods. The noncytotoxic concentration of ellipticine was 156.25 nM. The phosphorylation of glycogen synthase kinase-3ß was decreased, whereas the ß-catenin expression was increased in human endothelial cells treated with ellipticine. We concluded that ellipticine at sublethal dosage could be successfully repositioned as a pro-angiogenic substance by in silico virtual screening.


Assuntos
Elipticinas/farmacologia , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Neovascularização Patológica/tratamento farmacológico , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Descoberta de Drogas/métodos , Reposicionamento de Medicamentos/métodos , Expressão Gênica/efeitos dos fármacos , Glicogênio Sintase Quinase 3 beta/metabolismo , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Simulação de Acoplamento Molecular/métodos , Simulação de Dinâmica Molecular , Neovascularização Patológica/metabolismo , Ligação Proteica/efeitos dos fármacos , beta Catenina/metabolismo
9.
J Agric Food Chem ; 69(36): 10536-10549, 2021 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-34460247

RESUMO

An emerging inference is that vascular cells transfer their biological cargo to recipient cells by secretion of extracellular vesicles (EVs). This study explored the effects of EVs produced from VSMCs with Ang II (EVs-A) or LSW + Ang II on HUVECs. The EVs-A increase ROS production, activate inflammation, and upregulate the expression of adhesion molecules. Among the EVs-A, miR-22, miR-143, miR-144, and miR-155 were significantly downregulated, while VSMCs pre-incubated with LSW could produce improved EVs. RNA sequencing revealed differential expression of genes associated with endothelial dysfunction, including the TNF signaling pathway, NOD-like receptor signaling pathway, NF-κB signaling pathway, and fluid shear stress and atherosclerosis pathway. Finally, we found that LSW could improve endothelial function by repairing the expression of miRNAs in VSMCs. It also suggests a potential mechanism for the injury action of endogenous peptide Ang II and protective effects of exogenous peptide LSW on vascular endothelial cells.


Assuntos
Angiotensina II , Vesículas Extracelulares , Angiotensina II/farmacologia , Anti-Hipertensivos , Células Endoteliais da Veia Umbilical Humana , Humanos , Soja
10.
Int J Mol Sci ; 22(15)2021 Jul 30.
Artigo em Inglês | MEDLINE | ID: mdl-34360944

RESUMO

Endothelial and epithelial barrier function is crucial for the maintenance of physiological processes. The barrier paracellular permeability depends on the composition and spatial distribution of the cell-to-cell tight junctions (TJ). Here, we provide an experimental workflow that yields several layers of physiological data in the setting of a single endothelial cell monolayer. Human umbilical vein endothelial cells were grown on Transwell filters. Transendothelial electrical resistance (TER) and 10 kDa FITC dextran flux were measured using Alanyl-Glutamine (AlaGln) as a paracellular barrier modulator. Single monolayers were immunolabelled for Zonula Occludens-1 (ZO-1) and Claudin-5 (CLDN5) and used for automated immunofluorescence imaging. Finally, the same monolayers were used for single molecule localization microscopy (SMLM) of ZO-1 and CLDN5 at the nanoscale for spatial clustering analysis. The TER increased and the paracellular dextran flux decreased after the application of AlaGln and these functional changes of the monolayer were mediated by an increase in the ZO-1 and CLDN5 abundance in the cell-cell interface. At the nanoscale level, the functional and protein abundance data were accompanied by non-random increased clustering of CLDN5. Our experimental workflow provides multiple data from a single monolayer and has wide applicability in the setting of paracellular studies in endothelia and epithelia.


Assuntos
Permeabilidade Capilar , Junções Íntimas/metabolismo , Claudina-5/metabolismo , Dextranos/metabolismo , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Proteína da Zônula de Oclusão-1/metabolismo
11.
Int J Mol Sci ; 22(15)2021 Jul 30.
Artigo em Inglês | MEDLINE | ID: mdl-34360960

RESUMO

BACKGROUND/AIMS: Estrogen has been reported to have beneficial effects on vascular biology through direct actions on endothelium. Together with transcription factors, miRNAs are the major drivers of gene expression and signaling networks. The objective of this study was to identify a comprehensive regulatory network (miRNA-transcription factor-downstream genes) that controls the transcriptomic changes observed in endothelial cells exposed to estradiol. METHODS: miRNA/mRNA interactions were assembled using our previous microarray data of human umbilical vein endothelial cells (HUVEC) treated with 17ß-estradiol (E2) (1 nmol/L, 24 h). miRNA-mRNA pairings and their associated canonical pathways were determined using Ingenuity Pathway Analysis software. Transcription factors were identified among the miRNA-regulated genes. Transcription factor downstream target genes were predicted by consensus transcription factor binding sites in the promoter region of E2-regulated genes by using JASPAR and TRANSFAC tools in Enrichr software. RESULTS: miRNA-target pairings were filtered by using differentially expressed miRNAs and mRNAs characterized by a regulatory relationship according to miRNA target prediction databases. The analysis identified 588 miRNA-target interactions between 102 miRNAs and 588 targets. Specifically, 63 upregulated miRNAs interacted with 295 downregulated targets, while 39 downregulated miRNAs were paired with 293 upregulated mRNA targets. Functional characterization of miRNA/mRNA association analysis highlighted hypoxia signaling, integrin, ephrin receptor signaling and regulation of actin-based motility by Rho among the canonical pathways regulated by E2 in HUVEC. Transcription factors and downstream genes analysis revealed eight networks, including those mediated by JUN and REPIN1, which are associated with cadherin binding and cell adhesion molecule binding pathways. CONCLUSION: This study identifies regulatory networks obtained by integrative microarray analysis and provides additional insights into the way estradiol could regulate endothelial function in human endothelial cells.


Assuntos
Estradiol/farmacologia , Estrogênios/farmacologia , Redes Reguladoras de Genes , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Transcriptoma , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , MicroRNAs/genética , MicroRNAs/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
12.
FASEB J ; 35(9): e21808, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34390515

RESUMO

The link between serum uric acid (SUA) and the risk of venous thromboembolism (VTE) is well established. Recent data suggested a causative role of UA in endothelial cells (ECs) dysfunction. However, the molecular mechanism of high UA on thrombogenesis is unknown. We investigate whether high UA induce phosphatidylserine (PS) externalization and microparticle (MP) shedding in cultured EC, and contribute to UA-induced hypercoagulable state. In the present study, we demonstrate that UA induces PS exposure and EMP release of EC in a concentration- and time-dependent manner, which enhances the procoagulant activity (PCA) of EC and inhibited over 90% by lactadherin in vitro. Furthermore, hyperuricemic rat model was used to evaluate the development of thrombi following by flow stasis in the inferior vena cava (IVC). Hyperuricemia group is more likely to form large and hard thrombi compared with control. Importantly, we found that TMEM16F expression is significantly upregulated in UA-treated EC, which is crucial for UA-induced PS exposure and MP formation. Additionally, UA increases the generation of reactive oxygen species (ROS), lipid peroxidation, and cytosolic Ca2+ concentration in EC, which might contribute to increased TMEM16F expression. Using confocal microscopy, we also observed disruption of the actin cytoskeleton, suggesting that depolymerization of actin filaments might be required for TMEM16F activation and followed by PS exposure and membrane blebbing in UA-treated EC. Our results demonstrate a thrombotic role of EC in hyperuricemia through TMEM16F-mediated PS exposure and MPs release.


Assuntos
Anoctaminas/metabolismo , Micropartículas Derivadas de Células/metabolismo , Células Endoteliais/metabolismo , Hiperuricemia/metabolismo , Fosfatidilserinas/metabolismo , Proteínas de Transferência de Fosfolipídeos/metabolismo , Citoesqueleto de Actina/metabolismo , Animais , Cálcio/metabolismo , Células Cultivadas , Células Endoteliais da Veia Umbilical Humana , Humanos , Hiperuricemia/sangue , Peroxidação de Lipídeos/fisiologia , Masculino , Ratos , Ratos Sprague-Dawley , Espécies Reativas de Oxigênio/metabolismo , Ácido Úrico/sangue
13.
FASEB J ; 35(9): e21824, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34370353

RESUMO

Crosstalk between multiple components underlies the formation of mature vessels. Although the players involved in angiogenesis have been identified, mechanisms underlying the crosstalk between them are still unclear. Using the ex vivo aortic ring assay, we set out to dissect the interactions between two key angiogenic signaling pathways, vascular endothelial growth factor (VEGF) and transforming growth factor ß (TGFß), with members of the lysyl oxidase (LOX) family of matrix modifying enzymes. We find an interplay between VEGF, TGFß, and the LOXs is essential for the formation of mature vascular smooth muscle cells (vSMC)-coated vessels. RNA sequencing analysis further identified an interaction with the endothelin-1 pathway. Our work implicates endothelin-1 downstream of TGFß in vascular maturation and demonstrate the complexity of processes involved in generating vSMC-coated vessels.


Assuntos
Endotelina-1/metabolismo , Neovascularização Patológica/metabolismo , Proteína-Lisina 6-Oxidase/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Animais , Diferenciação Celular/fisiologia , Células Cultivadas , Feminino , Células Endoteliais da Veia Umbilical Humana , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Morfogênese/fisiologia , Miócitos de Músculo Liso/metabolismo , Transdução de Sinais/fisiologia , Fator A de Crescimento do Endotélio Vascular/metabolismo
14.
Stem Cell Res Ther ; 12(1): 432, 2021 08 03.
Artigo em Inglês | MEDLINE | ID: mdl-34344474

RESUMO

BACKGROUND: Bone tissue engineering is a new concept bringing hope for the repair of large bone defects, which remains a major clinical challenge. The formation of vascularized bone is key for bone tissue engineering. Growth of specialized blood vessels termed type H is associated with bone formation. In vivo and in vitro studies have shown that low level laser therapy (LLLT) promotes angiogenesis, fracture healing, and osteogenic differentiation of stem cells by increasing reactive oxygen species (ROS). However, whether LLLT can couple angiogenesis and osteogenesis, and the underlying mechanisms during bone formation, remains largely unknown. METHODS: Mouse bone marrow mesenchymal stem cells (BMSCs) combined with biphasic calcium phosphate (BCP) grafts were implanted into C57BL/6 mice to evaluate the effects of LLLT on the specialized vessel subtypes and bone regeneration in vivo. Furthermore, human BMSCs and human umbilical vein endothelial cells (HUVECs) were co-cultured in vitro. The effects of LLLT on cell proliferation, angiogenesis, and osteogenesis were assessed. RESULTS: LLLT promoted the formation of blood vessels, collagen fibers, and bone tissue and also increased CD31hiEMCNhi-expressing type H vessels in mBMSC/BCP grafts implanted in mice. LLLT significantly increased both osteogenesis and angiogenesis, as well as related gene expression (HIF-1α, VEGF, TGF-ß) of grafts in vivo and of co-cultured BMSCs/HUVECs in vitro. An increase or decrease of ROS induced by H2O2 or Vitamin C, respectively, resulted in an increase or decrease of HIF-1α, and a subsequent increase and decrease of VEGF and TGF-ß in the co-culture system. The ROS accumulation induced by LLLT in the co-culture system was significantly decreased when HIF-1α was inhibited with DMBPA and was followed by decreased expression of VEGF and TGF-ß. CONCLUSIONS: LLLT enhanced vascularized bone regeneration by coupling angiogenesis and osteogenesis. ROS/HIF-1α was necessary for these effects of LLLT. LLLT triggered a ROS-dependent increase of HIF-1α, VEGF, and TGF-ß and resulted in subsequent formation of type H vessels and osteogenic differentiation of mesenchymal stem cells. As ROS also was a target of HIF-1α, there may be a positive feedback loop between ROS and HIF-1α, which further amplified HIF-1α induction via the LLLT-mediated ROS increase. This study provided new insight into the effects of LLLT on vascularization and bone regeneration in bone tissue engineering.


Assuntos
Terapia com Luz de Baixa Intensidade , Osteogênese , Animais , Regeneração Óssea , Células Endoteliais da Veia Umbilical Humana , Humanos , Peróxido de Hidrogênio , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Camundongos , Camundongos Endogâmicos C57BL , Neovascularização Fisiológica
15.
Int J Mol Sci ; 22(15)2021 Jul 28.
Artigo em Inglês | MEDLINE | ID: mdl-34360851

RESUMO

Endothelial cells can acquire a mesenchymal phenotype through a process called Endothelial-to-Mesenchymal transition (EndMT). This event is found in embryonic development, but also in pathological conditions. Blood vessels lose their ability to maintain vascular homeostasis and ultimately develop atherosclerosis, pulmonary hypertension, or fibrosis. An increase in inflammatory signals causes an upregulation of EndMT transcription factors, mesenchymal markers, and a decrease in endothelial markers. In our study, we show that the induction of EndMT results in an increase in long non-coding RNA AERRIE expression. JMJD2B, a known EndMT regulator, induces AERRIE and subsequently SULF1. Silencing of AERRIE shows a partial regulation of SULF1 but showed no effect on the endothelial and mesenchymal markers. Additionally, the overexpression of AERRIE results in no significant changes in EndMT markers, suggesting that AERRIE is marginally regulating mesenchymal markers and transcription factors. This study identifies AERRIE as a novel factor in EndMT, but its mechanism of action still needs to be elucidated.


Assuntos
Transição Epitelial-Mesenquimal , RNA Longo não Codificante/fisiologia , Sulfotransferases/metabolismo , Células Endoteliais da Veia Umbilical Humana , Humanos , Histona Desmetilases com o Domínio Jumonji/metabolismo
16.
Molecules ; 26(16)2021 Aug 10.
Artigo em Inglês | MEDLINE | ID: mdl-34443423

RESUMO

Chronic liver inflammation has become a major global health concern. In the absence of clinical surrogate markers to diagnose inflammatory liver disease, the intervention with effective drugs in modern medicine tends to be late. In Sri Lanka, traditional medical practitioners prescribe herbal preparations from Osbeckia octandra for the prevention and treatment of liver disorders. To test the efficacy of such treatments, we have administered thioacetamide (TAA) to male Wistar rats to induce chronic liver damage (disease control; DC) and examined how various leaf extracts: crude leaf suspension (CLS), boiled leaf extract (BLE), sonicated leaf extract (SLE), methanol leaf extract (MLE) and hexane leaf extract (HLE) of O. octandra ameliorate TAA-induced liver disease. The CLS, BLE and SLE treatments in cirrhotic rats significantly attenuated disease-related changes, such as liver weight and hepato-enzymes. The mRNA levels of Tnf-α were significantly decreased by 3.6, 10 and 3.9 times in CLS, BLE and SLE compared to DC. The same treatments resulted in significantly lower (19.5, 4.2 and 2.4 times) α-Sma levels compared to DC. In addition, Tgf-ß1 and Vegf-R2 mRNA expressions were significantly lower with the treatments. Moreover, BLE expressed a strong anti-angiogenic effect. We conclude that CLS, BLE and SLE from O. octandra have potent hepatic anti-fibrotic effects in TAA-induced liver cirrhosis.


Assuntos
Cirrose Hepática Experimental/tratamento farmacológico , Melastomataceae/química , Neovascularização Patológica/tratamento farmacológico , Extratos Vegetais/uso terapêutico , Folhas de Planta/química , Citocinas/genética , Citocinas/metabolismo , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Mediadores da Inflamação/metabolismo , Fígado/enzimologia , Fígado/patologia , Cirrose Hepática Experimental/sangue , Neovascularização Patológica/sangue , Tamanho do Órgão/efeitos dos fármacos , Extratos Vegetais/farmacologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Tioacetamida , Regulação para Cima/efeitos dos fármacos , Água , Perda de Peso/efeitos dos fármacos
17.
Int J Mol Sci ; 22(16)2021 Aug 18.
Artigo em Inglês | MEDLINE | ID: mdl-34445582

RESUMO

Exosomes secreted by adipose-derived stem cells (ADSCs) enhance angiogenesis and wound healing. However, in clinical settings, wounds may be infected by various bacteria or pathogens. We investigated whether human ADSCs stimulated with lipopolysaccharide (LPS) secrete exosomes (ADSC-LPS-exo) that augment the angiogenesis of human umbilical vein endothelial cells (HUVECs). ExoQuick-TC exosome precipitation solution was used to purify exosomes from human ADSC culture media in the presence or absence of 1 µg/mL LPS treatment for 24 h. The uptake of ADSC-LPS-exo significantly induced the activation of cAMP response element binding protein (CREB), activating protein 1 (AP-1), and nuclear factor-κB (NF-κB) signaling pathways and increased the migration of and tube formation in HUVECs. RNA interference with CREB, AP-1, or NF-κB1 significantly reduced the migration of and tube formation in HUVECs treated with ADSC-LPS-exo. An experiment with an antibody array for 25 angiogenesis-related proteins revealed that only interleukin-8 expression was significantly upregulated in HUVECs treated with ADSC-LPS-exo. In addition, proteomic analysis revealed that eukaryotic translation initiation factor 4E, amyloid beta A4 protein, integrin beta-1, and ras-related C3 botulinum toxin substrate 1 may be potential candidates involved in ADSC-LPS-exo-mediated enhanced angiogenesis.


Assuntos
Movimento Celular , Exossomos/fisiologia , Células Endoteliais da Veia Umbilical Humana/fisiologia , Lipopolissacarídeos/farmacologia , Células-Tronco Mesenquimais/fisiologia , Neovascularização Fisiológica , Proliferação de Células , Células Cultivadas , Células Endoteliais da Veia Umbilical Humana/citologia , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Humanos , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Transdução de Sinais
18.
ACS Appl Mater Interfaces ; 13(33): 39126-39134, 2021 Aug 25.
Artigo em Inglês | MEDLINE | ID: mdl-34383476

RESUMO

The usage of exogenous antioxidant materials to relieve oxidative stress offers an important strategy for the therapy of oxidative stress-induced injuries. However, the fabrication processes toward the antioxidant materials usually require the involvement of extra metal ions and organic agents, as well as sophisticated purification steps, which might cause tremendous environmental stress and induce unpredictable side effects in vivo. To address these issues, herein, we proposed a novel strategy to fabricate green nanoparticles for efficiently modulating oxidative stress, which was facilely prepared from tea polyphenol extracts (originated from green tea) via a green enzymatic polymerization-based chemistry method. The resulting nanoparticles possessed a uniform spherical morphology and good stability in water and biomedium and demonstrated excellent radical scavenging properties. These nanoparticle scavengers could effectively prevent intracellular oxidative damage, accelerate wound recovery, and protect the kidneys from reactive oxygen species damaging in the acute kidney injury model. We hope this work will inspire the further development of more types of green nanoparticles for antioxidant therapies via similar synthetic strategies using green biomass materials.


Assuntos
Injúria Renal Aguda/prevenção & controle , Antioxidantes/química , Nanopartículas/química , Estresse Oxidativo/efeitos dos fármacos , Polifenóis/química , Chá/química , Células 3T3 , Células A549 , Animais , Antioxidantes/farmacologia , Catecóis/química , Sobrevivência Celular/efeitos dos fármacos , Feminino , Sequestradores de Radicais Livres/metabolismo , Química Verde , Peroxidase do Rábano Silvestre/química , Células Endoteliais da Veia Umbilical Humana , Humanos , Peróxido de Hidrogênio/química , Camundongos , Espécies Reativas de Oxigênio/metabolismo , Nanomedicina Teranóstica , Cicatrização/efeitos dos fármacos
19.
J Agric Food Chem ; 69(35): 10350-10357, 2021 Sep 08.
Artigo em Inglês | MEDLINE | ID: mdl-34448567

RESUMO

As a membrane protein, the activity of angiotensin I-converting enzyme (ACE) can be modulated via regulation of its localization in the cell membrane with food-derived peptides. This study aimed to explore the effect of egg white peptides on the cell membrane localization and activity of ACE in human umbilical vein endothelial cells. ACE activity was found to be related to lipid rafts by using methyl-ß-cyclodextrin (MßCD). QVPLW and LCAY can inhibit ACE activity by preventing ACE recruitment into lipid rafts, with in situ IC50 values of 238.46 ± 11.35 µM and 31.55 ± 2.64 µM in the control groups, as well as 45.43 ± 6.15 µM and 34.63 ± 1.59 µM in the MßCD groups, respectively. QVPLW and LCAY may alter the cell membrane properties, including the fluidity, potential, and permeability, and eventually promote the transposition of ACE.


Assuntos
Clara de Ovo , Peptidil Dipeptidase A , Inibidores da Enzima Conversora de Angiotensina/farmacologia , Células Endoteliais da Veia Umbilical Humana , Humanos , Microdomínios da Membrana , Peptídeos/farmacologia
20.
Int J Mol Sci ; 22(15)2021 Jul 26.
Artigo em Inglês | MEDLINE | ID: mdl-34360750

RESUMO

Tissue regeneration depends on the complex processes of angiogenesis, inflammation and wound healing. Regarding muscle tissue, glucocorticoids (GCs) inhibit pro-inflammatory signalling and angiogenesis and lead to muscle atrophy. Our hypothesis is that the synthetic GC dexamethasone (dex) impairs angiogenesis leading to muscle atrophy or inhibited muscle regeneration. Therefore, this study aims to elucidate the effect of dexamethasone on HUVECs under different conditions in mono- and co-culture with myoblasts to evaluate growth behavior and dex impact with regard to muscle atrophy and muscle regeneration. Viability assays, qPCR, immunofluorescence as well as ELISAs were performed on HUVECs, and human primary myoblasts seeded under different culture conditions. Our results show that dex had a higher impact on the tube formation when HUVECs were maintained with VEGF. Gene expression was not influenced by dex and was independent of cells growing in a 2D or 3D matrix. In co-culture CD31 expression was suppressed after incubation with dex and gene expression analysis revealed that dex enhanced expression of myogenic transcription factors, but repressed angiogenic factors. Moreover, dex inhibited the VEGF mediated pro angiogenic effect of myoblasts and inhibited expression of angiogenic inducers in the co-culture model. This is the first study describing a co-culture of human primary myoblast and HUVECs maintained under different conditions. Our results indicate that dex affects angiogenesis via inhibition of VEGF release at least in myoblasts, which could be responsible not only for the development of muscle atrophy after dex administration, but also for inhibition of muscle regeneration after vascular damage.


Assuntos
Dexametasona/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Mioblastos Esqueléticos/metabolismo , Neovascularização Fisiológica/efeitos dos fármacos , Fator A de Crescimento do Endotélio Vascular/metabolismo , Técnicas de Cocultura , Células Endoteliais da Veia Umbilical Humana/citologia , Humanos , Mioblastos Esqueléticos/citologia
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