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1.
Clinics (Sao Paulo) ; 75: e1865, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32935825

RESUMO

OBJECTIVES: Hypoxia leads to endothelial cell inflammation, apoptosis, and damage, which plays an important role in the complications associated with ischemic cardiovascular disease. As an oxidoreductase, p66Shc plays an important role in the regulation of reactive oxygen species (ROS) production and apoptosis. Ketamine is widely used in clinics. This study was designed to assess the potential protective effect of ketamine against hypoxia-induced injury in human umbilical vein endothelial cells (HUVECs). Moreover, we explored the potential mechanism by which ketamine protected against hypoxia-induced endothelial injury. METHODS: The protective effects of ketamine against hypoxia-induced injury was assessed using cell viability and adhesion assays, quantitative polymerase chain reaction, and western blotting. RESULTS: Our data showed that hypoxia reduced HUVEC viability, increased the adhesion between HUVECs and monocytes, and upregulated the expression of endothelial adhesion molecules at the protein and mRNA levels. Moreover, hypoxia increased ROS accumulation and upregulated p66Shc expression. Furthermore, hypoxia downregulated sirt1 expression in HUVECs. Alternatively, ketamine was shown to reverse the hypoxia-mediated reduction of cell viability and increase in the adhesion between HUVECs and monocytes, ameliorate hypoxia-induced ROS accumulation, and suppress p66Shc expression. Moreover, EX527, a sirt1 inhibitor, reversed the protective effects of ketamine against the hypoxia-mediated reduction of cell viability and increase in adhesion between HUVECs and monocytes. CONCLUSION: Ketamine reduces hypoxia-induced p66Shc expression and attenuates ROS accumulation via upregulating sirt1 in HUVECs, thus attenuating hypoxia-induced endothelial cell inflammation and apoptosis.


Assuntos
Apoptose/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Hipóxia , Ketamina/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Sobrevivência Celular , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Estresse Oxidativo , Proteína 1 de Transformação que Contém Domínio 2 de Homologia de Src , Veias Umbilicais
2.
Ecotoxicol Environ Saf ; 202: 110932, 2020 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-32800216

RESUMO

Adverse health effects arising from exposure to fine particulates have become a major concern. Angiogenesis is a vital physiological process for the growth and development of cells and structures in the human body, whereby excessive or insufficient vessel growth could contribute to pathogenesis of diseases. We therefore evaluated indirect effects of carbon black (CB) and inhalable airborne particles on the angiogenic ability of unexposed Human Umbilical Vein Endothelial Cells (HUVECs) by co-culturing HUVECs with pre-exposed Small Airway Epithelial Cells (SAECs). As endothelial cells are major components of blood vessels and potential targets of fine particles, we investigated if lung epithelial cells exposed to ambient PM2.5 surrogates could induce bystander effects on neighboring unexposed endothelial cells in an alveolar-capillary co-culture lung model. Epithelial exposure to CB at a non-toxic dose of 25 µg/mL reduced endothelial tube formation and cell adhesion in co-cultured HUVECs, and decreased expression of angiogenic genes in SAECs. Similarly, exposure of differentiated SAECs to PM2.5 surrogates reduced cell reproductive ability, adhesion and tube formation of neighboring HUVECs. This indicates epithelial exposure to CB and urban PM2.5 surrogates both compromised the angiogenic ability of endothelial cells through bystander effects, thereby potentially perturbing the ventilation-perfusion ratio and affecting lung function.


Assuntos
Poluentes Atmosféricos/toxicidade , Material Particulado/toxicidade , Testes de Toxicidade , Técnicas de Cocultura , Células Epiteliais , Células Endoteliais da Veia Umbilical Humana/citologia , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Pulmão/metabolismo , Neovascularização Patológica , Fuligem
3.
Nat Commun ; 11(1): 3571, 2020 07 16.
Artigo em Inglês | MEDLINE | ID: mdl-32678094

RESUMO

Pathogenic bacteria of the genus Bartonella can induce vasoproliferative lesions during infection. The underlying mechanisms are unclear, but involve secretion of an unidentified mitogenic factor. Here, we use functional transposon-mutant screening in Bartonella henselae to identify such factor as a pro-angiogenic autotransporter, called BafA. The passenger domain of BafA induces cell proliferation, tube formation and sprouting of microvessels, and drives angiogenesis in mice. BafA interacts with vascular endothelial growth factor (VEGF) receptor-2 and activates the downstream signaling pathway, suggesting that BafA functions as a VEGF analog. A BafA homolog from a related pathogen, Bartonella quintana, is also functional. Our work unveils the mechanistic basis of vasoproliferative lesions observed in bartonellosis, and we propose BafA as a key pathogenic factor contributing to bacterial spread and host adaptation.


Assuntos
Bartonella/patogenicidade , Neovascularização Patológica/metabolismo , Transdução de Sinais , Sistemas de Secreção Tipo V/metabolismo , Fatores de Crescimento do Endotélio Vascular/metabolismo , Fatores de Virulência/metabolismo , Animais , Bartonella/classificação , Bartonella/genética , Proliferação de Células , Perfilação da Expressão Gênica , Células Endoteliais da Veia Umbilical Humana/citologia , Células Endoteliais da Veia Umbilical Humana/metabolismo , Células Endoteliais da Veia Umbilical Humana/microbiologia , Humanos , Camundongos , Neovascularização Patológica/genética , Neovascularização Patológica/microbiologia , Domínios Proteicos , Sistemas de Secreção Tipo V/química , Sistemas de Secreção Tipo V/genética , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Fatores de Virulência/química , Fatores de Virulência/genética
4.
Nat Commun ; 11(1): 3697, 2020 07 29.
Artigo em Inglês | MEDLINE | ID: mdl-32728101

RESUMO

As the number of genomics datasets grows rapidly, sample mislabeling has become a high stakes issue. We present CrosscheckFingerprints (Crosscheck), a tool for quantifying sample-relatedness and detecting incorrectly paired sequencing datasets from different donors. Crosscheck outperforms similar methods and is effective even when data are sparse or from different assays. Application of Crosscheck to 8851 ENCODE ChIP-, RNA-, and DNase-seq datasets enabled us to identify and correct dozens of mislabeled samples and ambiguous metadata annotations, representing ~1% of ENCODE datasets.


Assuntos
Sequenciamento de Nucleotídeos em Larga Escala , Desequilíbrio de Ligação/genética , Bases de Dados de Ácidos Nucleicos , Genótipo , Células HEK293 , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Células K562 , Escore Lod , Anotação de Sequência Molecular
5.
Int J Nanomedicine ; 15: 4151-4169, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32606670

RESUMO

Purpose: Focused ultrasound (FUS) is a noninvasive method to produce thermal and mechanical destruction along with an immune-stimulatory effect against cancer. However, FUS ablation alone appears insufficient to generate consistent antitumor immunity. In this study, a multifunctional nanoparticle was designed to boost FUS-induced immune effects and achieve systemic, long-lasting antitumor immunity, along with imaging and thermal enhancement. Materials and Methods: PEGylated PLGA nanoparticles encapsulating astragalus polysaccharides (APS) and gold nanorods (AuNRs) were constructed by a simple double emulsion method, characterized, and tested for cytotoxicity. The abilities of PA imaging and thermal-synergetic ablation efficiency were analyzed in vitro and in vivo. The immune-synergistic effect on dendritic cell (DC) differentiation in vitro and the immune response in vivo were also evaluated. Results: The obtained APS/AuNR/PLGA-PEG nanoparticles have an average diameter of 255.00±0.1717 nm and an APS-loading efficiency of 54.89±2.07%, demonstrating their PA imaging capability and high biocompatibility both in vitro and in vivo. In addition, the as-prepared nanoparticles achieved a higher necrosis cell rate and induced apoptosis rate in an in vitro cell suspension assay, greater necrosis area and decreased energy efficiency factor (EEF) in an in vivo rabbit liver assay, and remarkable thermal-synergic performance. In particular, the nanoparticles upregulated the expression of MHC-II, CD80 and CD86 on cocultured DCs in vitro, followed by declining phagocytic function and enhanced interleukin (IL)-12 and interferon (INF)-γ production. Furthermore, they boosted the production of tumor necrosis factor (TNF)-α, IFN-γ, IL-4, IL-10, and IgG1 (P< 0.001) but not IgG2a. Immune promotion peaked on day 3 after FUS in vivo. Conclusion: The multifunctional APS/AuNR/PLGA-PEG nanoparticles can serve as an excellent synergistic agent for FUS therapy, facilitating real-time imaging, promoting thermal ablation effects, and boosting FUS-induced immune effects, which have the potential to be used for further clinical FUS treatment.


Assuntos
Astrágalo (Planta)/química , Neoplasias da Mama/terapia , Ouro/química , Nanopartículas Multifuncionais/química , Nanotubos/química , Polissacarídeos/química , Terapia por Ultrassom , Animais , Antígenos CD/metabolismo , Apoptose , Morte Celular , Diferenciação Celular , Linhagem Celular Tumoral , Proliferação de Células , Citocinas/metabolismo , Células Dendríticas/citologia , Feminino , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Imunoglobulina G/metabolismo , Linfócitos do Interstício Tumoral/imunologia , Camundongos Endogâmicos BALB C , Fagocitose , Técnicas Fotoacústicas , Poliésteres/síntese química , Poliésteres/química , Polietilenoglicóis/síntese química , Polietilenoglicóis/química , Coelhos , Nanomedicina Teranóstica , Fator A de Crescimento do Endotélio Vascular/metabolismo
6.
Int J Nanomedicine ; 15: 4311-4324, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32606679

RESUMO

Purpose: By providing a stem cell microenvironment with particular bioactive constituents in vivo, synthetic biomaterials have been progressively successful in stem cell-based tissue regeneration by enhancing the engraftment and survival of transplanted cells. Designs with bioactive motifs to influence cell behavior and with D-form amino acids to modulate scaffold stability may be critical for the development and optimization of self-assembling biomimetic hydrogel scaffolds for stem cell therapy. Materials and Methods: In this study, we linked naphthalene (Nap) covalently to a short D-form peptide (Nap-DFDFG) and the C domain of insulin-like growth factor-1 (IGF-1C) as a functional hydrogel-based scaffolds, and we hypothesized that this hydrogel could enhance the therapeutic efficiency of human placenta-derived mesenchymal stem cells (hP-MSCs) in a murine acute kidney injury (AKI) model. Results: The self-assembling peptide was constrained into a classical ß-sheet structure and showed hydrogel properties. Our results revealed that this hydrogel exhibited increased affinity for IGF-1 receptor. Furthermore, cotransplantation of the ß-IGF-1C hydrogel and hP-MSCs contributed to endogenous regeneration post-injury and boosted angiogenesis in a murine AKI model, leading to recovery of renal function. Conclusion: This hydrogel could provide a favorable niche for hP-MSCs and thereby rescue renal function in an AKI model by promoting cell survival and angiogenesis. In conclusion, by covalently linking the desired functional groups to D-form peptides to create functional hydrogels, self-assembling ß-sheet peptide hydrogels may serve as a promising platform for tissue-engineering and stem cell therapy.


Assuntos
Lesão Renal Aguda/tratamento farmacológico , Hidrogéis/química , Fator de Crescimento Insulin-Like I/química , Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais/citologia , Peptídeos/química , Lesão Renal Aguda/fisiopatologia , Animais , Materiais Biocompatíveis/química , Sobrevivência Celular , Feminino , Fibrose , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Hidrogéis/síntese química , Rim/patologia , Rim/fisiopatologia , Camundongos Transgênicos , Neovascularização Fisiológica , Placenta/citologia , Gravidez , Conformação Proteica em Folha beta , Domínios Proteicos
7.
Am J Physiol Heart Circ Physiol ; 319(2): H443-H455, 2020 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-32618511

RESUMO

Neuregulin-1 (NRG1) is a paracrine growth factor, secreted by cardiac endothelial cells (ECs) in conditions of cardiac overload/injury. The current concept is that the cardiac effects of NRG1 are mediated by activation of erythroblastic leukemia viral oncogene homolog (ERBB)4/ERBB2 receptors on cardiomyocytes. However, recent studies have shown that paracrine effects of NRG1 on fibroblasts and macrophages are equally important. Here, we hypothesize that NRG1 autocrine signaling plays a role in cardiac remodeling. We generated EC-specific Erbb4 knockout mice to eliminate endothelial autocrine ERBB4 signaling without affecting paracrine NRG1/ERBB4 signaling in the heart. We first observed no basal cardiac phenotype in these mice up to 32 wk. We next studied these mice following transverse aortic constriction (TAC), exposure to angiotensin II (ANG II), or myocardial infarction in terms of cardiac performance, myocardial hypertrophy, myocardial fibrosis, and capillary density. In general, no major differences between EC-specific Erbb4 knockout mice and control littermates were observed. However, 8 wk following TAC both myocardial hypertrophy and fibrosis were attenuated by EC-specific Erbb4 deletion, albeit these responses were normalized after 20 wk. Similarly, 4 wk after ANG II treatment, myocardial fibrosis was less pronounced compared with control littermates. These observations were supported by RNA-sequencing experiments on cultured endothelial cells showing that NRG1 controls the expression of various hypertrophic and fibrotic pathways. Overall, this study shows a role of endothelial autocrine NRG1/ERBB4 signaling in the modulation of hypertrophic and fibrotic responses during early cardiac remodeling. This study contributes to understanding the spatiotemporal heterogeneity of myocardial autocrine and paracrine responses following cardiac injury.NEW & NOTEWORTHY The role of NRG1/ERBB signaling in endothelial cells is not completely understood. Our study contributes to the understanding of spatiotemporal heterogeneity of myocardial autocrine and paracrine responses following cardiac injury and shows a role of endothelial autocrine NRG1/ERBB4 signaling in the modulation of hypertrophic and fibrotic responses during early cardiac remodeling.


Assuntos
Comunicação Autócrina , Cardiomiopatias/metabolismo , Células Endoteliais/metabolismo , Hipertrofia Ventricular Esquerda/metabolismo , Infarto do Miocárdio/metabolismo , Miócitos Cardíacos/metabolismo , Neuregulina-1/metabolismo , Receptor ErbB-4/metabolismo , Função Ventricular Esquerda , Remodelação Ventricular , Animais , Cardiomiopatias/genética , Cardiomiopatias/patologia , Cardiomiopatias/fisiopatologia , Células Cultivadas , Modelos Animais de Doenças , Fibrose , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Hipertrofia Ventricular Esquerda/genética , Hipertrofia Ventricular Esquerda/patologia , Hipertrofia Ventricular Esquerda/fisiopatologia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Infarto do Miocárdio/genética , Infarto do Miocárdio/patologia , Infarto do Miocárdio/fisiopatologia , Miócitos Cardíacos/patologia , Neovascularização Fisiológica , Comunicação Parácrina , Receptor ErbB-4/deficiência , Receptor ErbB-4/genética , Transdução de Sinais
8.
Arterioscler Thromb Vasc Biol ; 40(9): 2244-2264, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32640900

RESUMO

OBJECTIVE: Nanog is expressed in adult endothelial cells (ECs) at a low-level, however, its functional significance is not known. The goal of our study was to elucidate the role of Nanog in adult ECs using a genetically engineered mouse model system. Approach and Results: Biochemical analyses showed that Nanog is expressed in both adult human and mouse tissues. Primary ECs isolated from adult mice showed detectable levels of Nanog, Tert (telomerase reverse transcriptase), and eNos (endothelial nitric oxide synthase). Wnt3a (Wnt family member 3A) increased the expression of Nanog and hTERT (human telomerase reverse transcriptase) in ECs and increased telomerase activity in these cells. In a chromatin immunoprecipitation experiment, Nanog directly bound to the hTERT and eNOS promoter/enhancer DNA elements, thereby regulating their transcription. Administration of low-dose tamoxifen to ROSAmT/mG::Nanogfl/+::Cdh5CreERT2 mice induced deletion of a single Nanog allele, simultaneously labeling ECs with green fluorescent protein and resulting in decreased Tert and eNos levels. Histological and morphometric analyses of heart tissue sections prepared from these mice revealed cell death, microvascular rarefaction, and increased fibrosis in cardiac vessels. Accordingly, EC-specific Nanog-haploinsufficiency resulted in impaired EC homeostasis and angiogenesis. Conversely, re-expression of cDNA encoding the hTERT in Nanog-depleted ECs, in part, restored the effect of loss of Nanog. CONCLUSIONS: We showed that low-level Nanog expression is required for normal EC homeostasis and angiogenesis in adulthood.


Assuntos
Proliferação de Células , Senescência Celular , Vasos Coronários/metabolismo , Células Endoteliais/metabolismo , Endotélio Vascular/metabolismo , Proteína Homeobox Nanog/metabolismo , Animais , Apoptose , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Senescência Celular/efeitos dos fármacos , Vasos Coronários/efeitos dos fármacos , Vasos Coronários/patologia , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/patologia , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/patologia , Feminino , Fibrose , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteína Homeobox Nanog/deficiência , Proteína Homeobox Nanog/genética , Neovascularização Fisiológica , Óxido Nítrico Sintase Tipo III/genética , Óxido Nítrico Sintase Tipo III/metabolismo , Telomerase/genética , Telomerase/metabolismo , Ativação Transcricional , Via de Sinalização Wnt , Proteína Wnt3A/farmacologia
9.
Life Sci ; 258: 118106, 2020 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-32682916

RESUMO

AIMS: Endothelial dysfunction is a hallmark of hypertension. Herein, we assessed the effect of quercetin, a common dietary antioxidant, on endothelial function of spontaneously hypertensive rats (SHRs), and investigated the underlying molecular mechanisms. MAIN METHODS: The Wistar-Kyoto (WKY) and SHR rats were administered vehicle (1% w/v methyl cellulose) or quercetin (10 mg/kg body weight) by oral gavage once a day for 6 weeks. Systolic blood pressure (SBP) and diastolic blood pressure (DBP) were measured with a tail-cuff system. Functional of rat mesenteric arterioles was assessed by the temperature-controlled myograph. A dose-response curve was generated by the cumulative addition of acetylcholine (ACh) or sodium nitroprusside (SNP). NO production in the culture medium was assessed by measuring the concentration of nitrite, a stable metabolite of NO, using a modified Griess reagent. KEY FINDINGS: Quercetin improved endothelial function and decreased blood pressure in SHRs. Endothelial autophagy, an important cellular homeostatic process, was increased in the early phase of treatment, and decreased in the late phase of treatment. Quercetin promoted autophagy in cultured endothelial cells under both normal and oxidative stress conditions. Pharmacological inhibition of autophagy aggravated endothelial dysfunction in quercetin-treated endothelial cells under oxidative stress, and attenuated the antihypertensive and endothelial protective effects of quercetin in SHRs. SIGNIFICANCE: Quercetin protects endothelial function in hypertensive rats through promotion of autophagy. Thus, autophagy could serve as a potential therapeutic target for hypertension.


Assuntos
Autofagia/efeitos dos fármacos , Endotélio Vascular/patologia , Endotélio Vascular/fisiopatologia , Hipertensão/fisiopatologia , Quercetina/farmacologia , Animais , Pressão Sanguínea/efeitos dos fármacos , Endotélio Vascular/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Masculino , Artérias Mesentéricas/efeitos dos fármacos , Artérias Mesentéricas/fisiopatologia , Estresse Oxidativo/efeitos dos fármacos , Ratos Endogâmicos SHR , Ratos Endogâmicos WKY
10.
Life Sci ; 258: 118124, 2020 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-32702443

RESUMO

AIMS: Ketogenic diet (KD) has been proposed to be an effective lifestyle intervention for metabolic syndrome. However, the effects of KD on hypertension have not been well investigated. The present study aimed to investigate the effects and underling mechanisms of KD on hypertension in spontaneously hypertensive rats (SHRs). MATERIALS AND METHODS: SHRs were subjected to normal diet or KD for 4 weeks, starting at the age of 10 weeks. Then, the blood pressure and vascular function were assessed. Next, the eNOS expression, inflammatory factors and relative signaling pathway were examined. Human umbilical vein endothelial cells were used to investigate the underlying mechanism account for the effect of ketone on inflammation and eNOS expression. KEY FINDINGS: Compared with the normal diet, KD was indicated to aggravate hypertension and impaire endothelium-dependent relaxation in mesenteric arteries of SHRs. eNOS and CD31 expression in mesenteric arteries were also significantly suppressed by KD. In addition, KD markedly increased the activation of NF-κB pathway and the expression of IL1-ß and TNF-α. In vitro, results showed that inhibition of NF-κB could rescue the adverse effects of ketone body and TGF-ß on eNOS expression and inflammation response. SIGNIFICANCE: Our study indicated that KD impaired endothelium-dependent relaxation in mesenteric arteries and aggravated the development of hypertension in SHRs, suggesting that it should be more cautious to apply KD into clinical application in hypertensive individuals.


Assuntos
Dieta Cetogênica , Endotélio Vascular/patologia , Endotélio Vascular/fisiopatologia , Hipertensão/metabolismo , Hipertensão/patologia , NF-kappa B/metabolismo , Animais , Biomarcadores/metabolismo , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Corpos Cetônicos/metabolismo , Masculino , Artérias Mesentéricas/metabolismo , Artérias Mesentéricas/patologia , Óxido Nítrico Sintase Tipo III/metabolismo , Molécula-1 de Adesão Celular Endotelial a Plaquetas/metabolismo , Ratos Endogâmicos SHR , Vasodilatação , Perda de Peso
11.
Life Sci ; 257: 118013, 2020 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-32603818

RESUMO

AIMS: Emerging literature illustrates critical roles of long noncoding RNAs (lncRNAs) in the progression of atherosclerosis. However, the biological functions and mechanism by which lncRNAs regulate the atherosclerosis remain unclear. MATERIALS AND METHODS: Human umbilical vein endothelial cells (HUVECs) were treated with oxidative low-density lipoprotein (ox-LDL). RNA and protein levels were respectively measured using RT-qPCR and western blot. Molecular interaction was detected using luciferase reporter assay and chromatin immunoprecipitation (ChIP). Proliferation and migration were measured using CCK-8 and wound healing assay. KEY FINDINGS: Here, results unveiled that lncRNA SNHG7 was remarkedly up-regulated in ox-LDL exposed HUVECs. Gain and loss of function experiments showed that the SNHG7 repressed the proliferation and migration of HUVECs. Mechanistically, transcription factor E2F1 was found to target the promoter region of lncRNA SNHG7 and accelerated its expression. Moreover, miR-186-5p was found to bind with the 3'-UTR of SNHG7, meanwhile miR-186-5p also bound with the MMP2 mRNA 3'-UTR. SIGNIFICANCE: In conclusion, these results show the essential roles of E2F1/SNHG7/miR-186-5p/MMP2 axis on the proliferation and migration of endothelial cells, providing a potential therapeutic target for atherosclerosis.


Assuntos
Aterosclerose/metabolismo , Células Endoteliais/metabolismo , RNA Longo não Codificante/metabolismo , Regiões 3' não Traduzidas , Apoptose/genética , Aterosclerose/fisiopatologia , Movimento Celular/genética , Proliferação de Células/genética , Progressão da Doença , Fator de Transcrição E2F1/metabolismo , Células Endoteliais/fisiologia , Técnicas de Silenciamento de Genes , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Lipoproteínas LDL , Metaloproteinase 2 da Matriz/genética , Metaloproteinase 2 da Matriz/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , RNA Longo não Codificante/genética , RNA Longo não Codificante/fisiologia
12.
Anticancer Res ; 40(6): 3191-3201, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32487613

RESUMO

BACKGROUND/AIM: Although it has been accepted that the tandem repeat galectin-8 (Gal-8) is linked to angiogenesis, the underlying mechanisms in endothelial cells has remained poorly understood. In this study we aimed to investigate the effect of Gal-8 on selected biological processes linked to angiogenesis in in vitro and in vivo models. MATERIALS AND METHODS: In detail, we assessed how exogenously added human recombinant Gal-8 (with or without vascular endothelial growth factor - VEGF) affects selected steps involved in vessel formation in human umbilical vein endothelial cells (HUVECs) as well as using the chick chorioallantoic membrane (CAM) assay. Gene expression profiling of HUVECs was performed to extend the scope of our investigation. RESULTS: Our findings demonstrate that Gal-8 in combination with VEGF enhanced cell proliferation and migration, two cellular events linked to angiogenesis. However, Gal-8 alone did not exhibit any significant effects on cell proliferation or on cell migration. The molecular analysis revealed that Gal-8 in the presence of VEGF influenced cytokine-cytokine receptor interactions, HIF-1 and PI3K/AKT signaling pathways. Gal-8 alone also targeted cytokine-cytokine receptor interactions, but with a different expression profile as well as a modulated focal adhesion and TNF signaling. CONCLUSION: Gal-8 promotes a pro-angiogenic phenotype possibly in a synergistic manner with VEGF.


Assuntos
Membrana Corioalantoide/irrigação sanguínea , Membrana Corioalantoide/efeitos dos fármacos , Galectinas/farmacologia , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Fator A de Crescimento do Endotélio Vascular/metabolismo , Animais , Movimento Celular/efeitos dos fármacos , Embrião de Galinha , Membrana Corioalantoide/metabolismo , Galectinas/metabolismo , Perfilação da Expressão Gênica , Células Endoteliais da Veia Umbilical Humana/citologia , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Técnicas In Vitro , Neovascularização Fisiológica/efeitos dos fármacos
13.
Science ; 368(6496): 1205-1210, 2020 06 12.
Artigo em Inglês | MEDLINE | ID: mdl-32527825

RESUMO

Cell migration is driven by local membrane protrusion through directed polymerization of F-actin at the front. However, F-actin next to the plasma membrane also tethers the membrane and thus resists outgoing protrusions. Here, we developed a fluorescent reporter to monitor changes in the density of membrane-proximal F-actin (MPA) during membrane protrusion and cell migration. Unlike the total F-actin concentration, which was high in the front of migrating cells, MPA density was low in the front and high in the back. Back-to-front MPA density gradients were controlled by higher cofilin-mediated turnover of F-actin in the front. Furthermore, nascent membrane protrusions selectively extended outward from areas where MPA density was reduced. Thus, locally low MPA density directs local membrane protrusions and stabilizes cell polarization during cell migration.


Assuntos
Actinas/metabolismo , Movimento Celular , Extensões da Superfície Celular , Actinas/química , Actinas/genética , Membrana Celular , Polaridade Celular , Proteínas de Fluorescência Verde/química , Proteínas de Fluorescência Verde/genética , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos
14.
Gen Physiol Biophys ; 39(3): 249-258, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32525818

RESUMO

Nitric oxide is known as relaxing factor because it acts as a vasodilator, increases blood flow, and inhibits platelet aggregation and adhesion, on the other hand nitric oxide can modulate cellular and physiological processes to limit oxidative injury, limiting processes such as leukocyte adhesion. As the complete mechanism of myricetin and its cardiovascular benefits is not completely understood, the aim of this study was to investigate the antihypertensive activity of myricetin in human umbilical vein endothelial cell (HUVEC). Angiotensin converting enzyme (ACE) activity, nitric oxide production, reactive oxygen species (ROS) scavenger activity, cellular calcium concentration, and endothelial nitric oxide synthase (eNOS) activity and protein expression was investigated in HUVEC treated with different concentration of myricetin (1-60 µM). Myricetin increased nitric oxide production in HUVEC through decreased ROS levels and increased nitric oxide production and eNOS activation. Activation of eNOS enzyme was achieved by an increase of cellular calcium concentration. At the same examined concentration of myricetin, the activity of ACE was significantly inhibited. These findings indicate that myricetin may be helpful for lowering blood pressure; this could be achieved through dietary intervention or by the production of new antihypertensive treatments from a natural product.


Assuntos
Inibidores da Enzima Conversora de Angiotensina/farmacologia , Flavonoides/farmacologia , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Óxido Nítrico/biossíntese , Células Cultivadas , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Óxido Nítrico Sintase Tipo II/metabolismo , Óxido Nítrico Sintase Tipo III , Peptidil Dipeptidase A/metabolismo
15.
DNA Cell Biol ; 39(7): 1264-1273, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32584608

RESUMO

Transforming growth factor-beta 1 (TGF-ß1) plays important roles in the endothelial-to-mesenchymal transition (EndMT). Recently, long noncoding RNAs (lncRNAs) have been identified to be involved in the physiological and pathological processes of human diseases. However, the role of endothelial lncRNAs in the TGF-ß1-mediated control of angiogenesis and its underlying mechanism remains largely unclear. In this study, we first demonstrated that TGF-ß1 induced EndMT; promoted cell viability, proliferation, migration, and tube formation of human umbilical vein endothelial cells (HUVECs). Second, our study displayed that TGF-ß1 upregulated the lncRNA UCA1 expression in HUVECs, knocked down UCA1 with small interfering RNAs, and inhibited the function of TGF-ß1 in HUVECs. Third, our study showed that UCA1 was located in the cytoplasm and absorbed miR-455 in TGF-ß1-treated HUVECs. Further, the miR-455 inhibitor restored the role of the inhibited UCA1 in HUVECs treated with TGF-ß1. Fourth, our study revealed that miR-455 inhibited ZEB1 expression, and overexpression of ZEB1 restored the role of miR-455 in HUVECs treated with TGF-ß1. Finally, our study revealed that UCA1 exerted its role via regulating the UCA1/miR-455/ZEB1 regulatory axis in HUVECs treated with TGF-ß1. Collectively, our study identified the role of the UCA1/miR-455/ZEB1 pathway in HUVECs treated with TGF-ß1 and indicated the potential therapeutic role of this regulatory axis in angiogenesis.


Assuntos
Células Endoteliais/citologia , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Mesoderma/citologia , MicroRNAs/genética , RNA Longo não Codificante/genética , Fator de Crescimento Transformador beta1/farmacologia , Homeobox 1 de Ligação a E-box em Dedo de Zinco/genética , Sequência de Bases , Regulação da Expressão Gênica/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/citologia , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Regulação para Cima/efeitos dos fármacos
16.
Int J Nanomedicine ; 15: 3291-3302, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32494130

RESUMO

Background: Inhaled nanoparticles can cross pulmonary air-blood barrier into circulation and cause vascular endothelial injury and progression of cardiovascular disease. However, the molecular mechanism underlying the vascular toxicity of copper oxide nanoparticles (CuONPs) remains unclear. We have recently demonstrated that the release of copper ions and the accumulation of superoxide anions contributed to CuONPs-induced cell death in human umbilical vein endothelial cells (HUVECs). Herein, we further demonstrate the mechanism underlying copper ions-induced cell death in HUVECs. Methods and Results: CuONPs were suspended in culture medium and vigorously vortexed for several seconds before exposure. After treatment with CuONPs, HUVECs were collected, and cell function assays were conducted to elucidate cellular processes including cell viability, oxidative stress, DNA damage and cell signaling pathways. We demonstrated that CuONPs uptake induced DNA damage in HUVECs as evidenced by γH2AX foci formation and increased phosphorylation levels of ATR, ATM, p53 and H2AX. Meanwhile, we showed that CuONPs exposure induced oxidative stress, indicated by the increase of cellular levels of superoxide anions, the upregulation of protein levels of heme oxygenase-1 (HO-1) and glutamate-cysteine ligase modifier subunit (GCLM), the elevation of the levels of malondialdehyde (MDA), but the reduction of glutathione to glutathione disulfide ratio. We also found that antioxidant N-acetyl-L-cysteine (NAC) could ameliorate CuONPs-induced oxidative stress and cell death. Interestingly, we demonstrated that p38 mitogen-activated protein kinase (MAPK) signaling pathway was activated in CuONPs-treated HUVECs, while p38α MAPK knockdown by siRNA significantly rescued HUVECs from CuONPs-induced DNA damage and cell death. Importantly, we showed that copper ions chelator tetrathiomolybdate (TTM) could alleviate CuONPs-induced oxidative stress, DNA damage, p38 MAPK pathway activation and cell death in HUVECs. Conclusion: We demonstrated that CuONPs induced oxidative DNA damage and cell death via copper ions-mediated p38 MAPK activation in HUVECs, suggesting that the release of copper ions was the upstream activator for CuONPs-induced vascular endothelial toxicity, and the copper ions chelator TTM can alleviate CuONPs-associated cardiovascular disease.


Assuntos
Cobre/toxicidade , Dano ao DNA , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/patologia , Nanopartículas/toxicidade , Estresse Oxidativo/efeitos dos fármacos , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Morte Celular/efeitos dos fármacos , Ativação Enzimática/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Modelos Biológicos , Molibdênio/química , Nanopartículas/ultraestrutura
17.
PLoS Biol ; 18(6): e3000734, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32502201

RESUMO

Cerebral cavernous malformations (CCMs) are vascular lesions predominantly developing in the central nervous system (CNS), with no effective treatments other than surgery. Loss-of-function mutation in CCM1/krev interaction trapped 1 (KRIT1), CCM2, or CCM3/programmed cell death 10 (PDCD10) causes lesions that are characterized by abnormal vascular integrity. Vascular endothelial cadherin (VE-cadherin), a major regulator of endothelial cell (EC) junctional integrity is strongly disorganized in ECs lining the CCM lesions. We report here that microRNA-27a (miR-27a), a negative regulator of VE-cadherin, is elevated in ECs isolated from mouse brains developing early CCM lesions and in cultured ECs with CCM1 or CCM2 depletion. Furthermore, we show miR-27a acts downstream of kruppel-like factor (KLF)2 and KLF4, two known key transcription factors involved in CCM lesion development. Using CD5-2 (a target site blocker [TSB]) to prevent the miR-27a/VE-cadherin mRNA interaction, we present a potential therapy to increase VE-cadherin expression and thus rescue the abnormal vascular integrity. In CCM1- or CCM2-depleted ECs, CD5-2 reduces monolayer permeability, and in Ccm1 heterozygous mice, it restores dermal vessel barrier function. In a neonatal mouse model of CCM disease, CD5-2 normalizes vasculature and reduces vascular leakage in the lesions, inhibits the development of large lesions, and significantly reduces the size of established lesions in the hindbrain. Furthermore, CD5-2 limits the accumulation of inflammatory cells in the lesion area. Our work has established that VE-cadherin is a potential therapeutic target for normalization of the vasculature and highlights that targeting miR-27a/VE-cadherin interaction by CD5-2 is a potential novel therapy for the devastating disease, CCM.


Assuntos
Antígenos CD/metabolismo , Caderinas/metabolismo , Hemangioma Cavernoso do Sistema Nervoso Central/genética , MicroRNAs/metabolismo , Animais , Regulação para Baixo/genética , Hemangioma Cavernoso do Sistema Nervoso Central/patologia , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Fatores de Transcrição Kruppel-Like/metabolismo , Masculino , Camundongos Endogâmicos C57BL , MicroRNAs/genética , Rombencéfalo/irrigação sanguínea , Rombencéfalo/patologia , Regulação para Cima/genética , Proteína rhoA de Ligação ao GTP/metabolismo
18.
Nat Commun ; 11(1): 3188, 2020 06 24.
Artigo em Inglês | MEDLINE | ID: mdl-32581225

RESUMO

Nitric oxide (NO) has been implicated in a variety of physiological and pathological processes. Monitoring cellular levels of NO requires a sensor to feature adequate sensitivity, transient recording ability and biocompatibility. Herein we report a single-atom catalysts (SACs)-based electrochemical sensor for the detection of NO in live cellular environment. The system employs nickel single atoms anchored on N-doped hollow carbon spheres (Ni SACs/N-C) that act as an excellent catalyst for electrochemical oxidation of NO. Notably, Ni SACs/N-C shows superior electrocatalytic performance to the commonly used Ni based nanomaterials, attributing from the greatly reduced Gibbs free energy that are required for Ni SACs/N-C in activating NO oxidation. Moreover, Ni SACs-based flexible and stretchable sensor shows high biocompatibility and low nanomolar sensitivity, enabling the real-time monitoring of NO release from cells upon drug and stretch stimulation. Our results demonstrate a promising means of using SACs for electrochemical sensing applications.


Assuntos
Técnicas Biossensoriais/métodos , Níquel/química , Óxido Nítrico/análise , Nitrogênio/química , Técnicas Biossensoriais/instrumentação , Carbono/química , Catálise , Eletroquímica , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Nanoestruturas/química , Nanoestruturas/ultraestrutura , Óxido Nítrico/metabolismo , Oxirredução
19.
PLoS One ; 15(6): e0235116, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32569321

RESUMO

Here, we examine known GTPase regulators of vesicle trafficking events to assess whether they affect endothelial cell (EC) lumen and tube formation. We identify novel roles for the small GTPases Rab3A, Rab3B, Rab8A, Rab11A, Rab27A, RalA, RalB and caveolin-1 in co-regulating membrane trafficking events that control EC lumen and tube formation. siRNA suppression of individual GTPases such as Rab3A, Rab8A, and RalB markedly inhibit tubulogenesis, while greater blockade is observed with combinations of siRNAs such as Rab3A and Rab3B, Rab8A and Rab11A, and RalA and RalB. These combinations of siRNAs also disrupt very early events in lumen formation including the formation of intracellular vacuoles. In contrast, knockdown of the endocytosis regulator, Rab5A, fails to inhibit EC tube formation. Confocal microscopy and real-time videos reveal that caveolin-1 strongly labels intracellular vacuoles and localizes to the EC apical surface as they fuse to form the luminal membrane. In contrast, Cdc42 and Rab11A localize to a perinuclear, subapical region where intracellular vacuoles accumulate and fuse during lumen formation. Our new data demonstrates that EC tubulogenesis is coordinated by a series of small GTPases to control polarized membrane trafficking events to generate, deliver, and fuse caveolin-1-labeled vacuoles to create the apical membrane surface.


Assuntos
Caveolina 1/metabolismo , Células Endoteliais da Veia Umbilical Humana/metabolismo , Vacúolos/metabolismo , Proteínas rab de Ligação ao GTP/metabolismo , Membrana Celular/metabolismo , Colágeno/metabolismo , Exocitose , Proteínas de Fluorescência Verde/metabolismo , Humanos , Modelos Biológicos , Transporte Proteico , RNA Interferente Pequeno/metabolismo , Proteínas rac de Ligação ao GTP/metabolismo , Proteínas ral de Ligação ao GTP/metabolismo , Quinases da Família src/metabolismo
20.
Life Sci ; 256: 117976, 2020 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-32561397

RESUMO

AIMS: We have previously reported that Centchroman (CC), an oral contraceptive drug, inhibits breast cancer progression and metastasis. In this study, we investigated whether CC inhibits local invasion of tumor cells and/or their metastatic colonization with detailed underlying mechanisms. MAIN METHODS: The effect of CC on the experimental metastasis and spontaneous metastasis was demonstrated by using tail-vein and orthotopic 4T1-syngeneic mouse tumor models, respectively. The anti-angiogenic potential of CC was evaluated using well established in vitro and in vivo models. The role of RAC1/PAK1/ß-catenin signaling axis in the metastasis was investigated and validated using siRNA-mediated knockdown of PAK1 as well as by pharmacological PAK1-inhibitor. KEY FINDINGS: The oral administration of CC significantly suppressed the formation of metastatic lung nodules in the 4T1-syngeneic orthotopic as well as experimental metastatic models. More importantly, CC treatment suppressed the tube formation and migration capacities of human umbilical vein endothelial cells (HUVEC) and inhibited pre-existing vasculature as well as the formation of neovasculature. The suppression of migration and invasion capacities of metastatic breast cancer cells upon CC treatment was associated with the inhibition of small GTPases (Rac1 and Cdc42) concomitant with the downregulation of PAK1 and downstream ß-catenin signaling. In addition, CC upregulated the expression of miR-145, which is known to target PAK1. SIGNIFICANCE: This study warrants the repurposing of CC as a potential therapeutic agent against metastatic breast cancer.


Assuntos
Neoplasias da Mama/metabolismo , Centocromano/farmacologia , Antagonistas de Estrogênios/farmacologia , Neuropeptídeos/antagonistas & inibidores , beta Catenina/antagonistas & inibidores , Quinases Ativadas por p21/antagonistas & inibidores , Proteínas rac1 de Ligação ao GTP/antagonistas & inibidores , Animais , Neoplasias da Mama/tratamento farmacológico , Centocromano/uso terapêutico , Antagonistas de Estrogênios/uso terapêutico , Feminino , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Células MCF-7 , Camundongos , Camundongos Endogâmicos BALB C , Neovascularização Patológica/tratamento farmacológico , Neovascularização Patológica/metabolismo , Neuropeptídeos/metabolismo , Distribuição Aleatória , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , beta Catenina/metabolismo , Quinases Ativadas por p21/metabolismo , Proteínas rac1 de Ligação ao GTP/metabolismo
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