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1.
J Colloid Interface Sci ; 559: 313-323, 2020 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-31675662

RESUMO

Antibiotic resistance is a common phenomenon observed during treatment with antibacterials. Use of nanozymes, especially those with synergistic enzyme-like activities, as antibacterials could overcome this problem, but their synthesis is limited by their high cost and/or complex production process. Herein, vanadium oxide nanodots (VOxNDs) were prepared via a one-step bottom-up ethanol-thermal method using vanadium trichloride as the precursor. VOxNDs alone possess bienzyme mimics of peroxidase and oxidase. Accordingly, highly efficient antibacterials against drug-resistant bacteria can be obtained through synergistic catalysis; the oxidase-like activity decomposes O2 to generate superoxide anion radical (O2-) and hydroxyl radicals (OH), and the intrinsic peroxidase-like activity can further induce the production of OH from external H2O2. Consequently, H2O2 concentration could decrease up to four magnitude orders with VOxNDs to achieve an antibacterial efficacy similar to that of H2O2 alone. Wound healing in vivo further confirms the high antibacterial efficiency, good biocompatibility, and application potential of the synergistic antibacterial system due to the "nano" structure of VOxNDs. The method of synthesis of nanodot antibacterials described in this paper is inexpensive, and the results of this study reveal the multi-enzymatic synergism of nanozymes.


Assuntos
Antibacterianos/química , Nanopartículas Metálicas/química , Óxidos/química , Compostos de Vanádio/química , Cicatrização/efeitos dos fármacos , Animais , Materiais Biocompatíveis/química , Materiais Biomiméticos/química , Catálise , Sobrevivência Celular/efeitos dos fármacos , Farmacorresistência Bacteriana/efeitos dos fármacos , Escherichia coli/efeitos dos fármacos , Células HeLa , Células Endoteliais da Veia Umbilical Humana , Humanos , Peróxido de Hidrogênio/química , Radical Hidroxila/química , Peroxidases/metabolismo , Ratos Sprague-Dawley , Staphylococcus aureus/efeitos dos fármacos
2.
APMIS ; 128(1): 10-19, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31642122

RESUMO

Atherogenesis is associated with chronic gut infections; however, the mechanisms are not clear. The aim of the study was to determine whether lipopolysaccharide of E. coli (E. coli LPS) may affect endothelial barrier and modify IL-10 expression in dendritic cells (DCs). Human umbilical vein endothelial cells (HUVECs) and monocyte-derived DCs were treated with E. coli LPS, apolipoprotein B100 (ApoB100) and 7-ketocholesterol (7-kCH) - harmful oxidized form of cholesterol. The effect of E. coli LPS, 7-kCH and ApoB100 on the barrier functions of HUVECs in real-time cell electric impedance sensing system (RTCA-DP) was assessed. Furthermore, the effect of 7-kCH and ApoB100 on barrier functions of HUVECs co-cultured with DCs previously treated with LPS was analyzed. Both E. coli LPS and 7-kCH decreased barrier functions of HUVECs and reduced tight junction protein mRNA expression, whereas ApoB100 increased endothelial barrier. In DCs, ApoB100 and E. coli LPS decreased IL-10 mRNA expression. In HUVECs co-cultured with DCs treated with LPS and subsequently pulsed with ApoB100 or 7-kCH, IL-10 mRNA expression was lower. E. coli LPS-exposed DCs diminished the protective effect of ApoB100 on endothelial integrity and led to the decrease in occludin mRNA expression. LPS potentially derived from gut microflora may destabilize endothelial barrier together with oxidized cholesterol and intensify the immunogenicity of ApoB100.


Assuntos
Células Dendríticas/efeitos dos fármacos , Escherichia coli/imunologia , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Interleucina-10/genética , Lipopolissacarídeos/imunologia , Junções Íntimas/efeitos dos fármacos , Apolipoproteína B-100/farmacologia , Células Cultivadas , Técnicas de Cocultura , Células Dendríticas/imunologia , Escherichia coli/química , Humanos , Cetocolesteróis/farmacologia , Lipopolissacarídeos/farmacologia , Ocludina/genética , Transdução de Sinais/efeitos dos fármacos , Proteínas de Junções Íntimas/genética
3.
Nan Fang Yi Ke Da Xue Xue Bao ; 39(11): 1265-1272, 2019 Nov 30.
Artigo em Chinês | MEDLINE | ID: mdl-31852645

RESUMO

OBJECTIVE: To investigate the effect of atorvastatin on the expression of lectin- like oxLDL receptor 1 (LOX-1) and endothelial nitric oxide synthase (eNOS) in collateral vessels of hypercholesterolemic rats. METHODS: Forty male SD rats were randomized equally into 4 groups: femoral ligation group (L), hypercholesterolemia + femoral ligation group (HL), hypercholesterolemia+atorvastatin+femoral ligation group (AL), and hypercholesterolemia+normal saline+femoral ligation group (NL). The rats in the latter 3 groups were fed atherogenic diet for 8 weeks. At the end of the 8 weeks, the rats were subjected to femoral artery ligation with or without intraperitoneal injection of atorvastatin (AL group) or saline (NL group). Two weeks later, all the rats were euthanized and the expressions of LOX-1 and eNOS in the collateral vessels were detected with immunofluorescence assay. In the in vitro experiment, cultured human umbilical vein endothelial cells (HUVECs) were transfected with LOX-1 siRNA followed by treatment with oxLDL and/or atorvastatin. The expressions of LOX-1 and eNOS in the cells were detected with realtime PCR and Western blotting, and the cellular NO production was examined with Griess assay. RESULTS: The collateral vessels of rats with normal feeding expressed LOX-1, which was significantly increased in the collateral vessels of hypercholesterolemic rats; atorvastatin treatment significantly lowered LOX-1 expressions in the hypercholesterolemic rats. In normally fed rats, the growing collateral vessels exhibited strong eNOS expressions, which were lowered in hypercholesterolemic rats and enhanced after atorvastatin treatment. In the cell experiment, HUVECs with oxLDL treatment showed a high LOX-1 expression and a low eNOS expression, and atorvastatin treatment of the cells down-regulated LOX-1 and up-regulated eNOS expressions. Inhibition of LOX-1 mediated by a specific LOX-1 siRNA abolished the effect of oxLDL stimulation on eNOS expression in the cells. CONCLUSIONS: Both hypercholesterolemia and oxLDL can induce endothelial dysfunction and impair collateral vessel growth via the LOX-1/eNOS pathway in rats, and atorvastatin treatment can restore the LOX-1/eNOS pathway to promote the growth of the collateral vessels, suggesting the potential of atorvastatin as a therapeutic agent to promote repair of collateral vessel injuries in ischemic diseases.


Assuntos
Receptores Depuradores Classe E/metabolismo , Animais , Atorvastatina , Células Endoteliais da Veia Umbilical Humana , Humanos , Lipoproteínas LDL , Masculino , Óxido Nítrico Sintase Tipo III , Ratos , Ratos Sprague-Dawley
4.
Zhonghua Yi Xue Za Zhi ; 99(48): 3814-3818, 2019 Dec 24.
Artigo em Chinês | MEDLINE | ID: mdl-31874520

RESUMO

Objective: To investigate the effects of miR-106a-5p on the proliferation and migration of vascular endothelial cells, and the possible target gene of miR-106a-5p in endothelial cells. Methods: Human umbilical vein endothelial cells (HUVEC) were cultured in vitro, and the expression of mir-106a-5p in HUVECs was detected by real-time fluorescent quantitative polymerase chain reaction (RT-qPCR). Cell counting kit-8 (CCK-8) and wound healing assays were used to detected the proliferation and migration of HUVECs respectively.Dual luciferase repoter assay was used to identify the possible target gene of miR-106a-5p--STAT3. Results: mir-106a-5p inhibited the proliferation function of HUVECs (F=13.62, P<0.01). And mir-106a-5p expressed the migration of HUVEC,the closure rate in the mimic group was reduced after 12 h and 24 h of scratching (49.93/31.31) (χ(2)=8.240, P<0.05), (78.87/44.80) (χ(2)=10.50, P<0.01). The direct target gene of mir-106a-5p was STAT3, and miR-106a-5p regulated the expression of STAT3 through post-transcriptionally controlling. Conclusion: mir-106a-5p could inhibit proliferation and migration of HUVEC, and the possible target gene was STAT3.


Assuntos
Células Endoteliais da Veia Umbilical Humana , Humanos , MicroRNAs , Reação em Cadeia da Polimerase em Tempo Real , Fator de Transcrição STAT3 , Veias Umbilicais
5.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi ; 35(9): 806-811, 2019 Sep.
Artigo em Chinês | MEDLINE | ID: mdl-31750822

RESUMO

Objective To explore the effect of tanshinone IIA (TSA) on hydrogen peroxide (H2O2)-induced senescence of human umbilical vein endothelial cells (HUVECs) and the underlying mechanism. Methods HUVECs were cultured in vitro and divided into the control group, model group and TSA group. The cells in the TSA group were pre-treated with TSA for 24 hours. H2O2 was used to induce cell senescence in the model and TSA groups. Transfection with SIRT1 siRNA was used for the knockdown of SIRT1 in HUVECs. CCK-8 assay was performed to detect cell viability. The expression levels of senescence-related proteins (P21 and P26), SIRT1, phosphorylated endothelial nitric oxide synthase (p-eNOS), and eNOS were detected by Western blot analysis. Senescence-associated ß-galactosidase (SA-ß-gal) staining was performed to evaluate cell senescence. Results Pretreatment with TSA at low concentrations (10, 20 and 40 µg/mL) for 24 hours did not affect cell viability, while high concentrations (80, 160 and 320 µg/mL) decreased cell viability significantly. In addition, 10, 20 and 40 µg/mL of TSA promoted H2O2-mediated cell viability of HUVECs in a concentration-dependent manner. Compared with the control group, the positive rate of SA-ß-gal staining in the model group increased, while the positive rate in the TSA group was significantly lower than that in the model group. The expression levels of P21 and P16 protein in the model group were higher than those in the control group, while SIRT1 and p-eNOS/eNOS were lower than those in the control group. Conversely, the expression of P21 and P16 proteins in the TSA group were lower than those in the model group, and SIRT1 and p-eNOS/eNOS were higher in the TSA group than those in the model group. Transfected with SIRT1 siRNA significantly down-regulated the expression of SIRT1 in HUVECs and the positive rate of SA-ß-gal staining was notably raised when SIRT1 was silenced in TSA-treated HUVECs. Conclusion TSA attenuates H2O2-induced endothelial cell senescence by activating SIRT1/eNOS signaling pathway.


Assuntos
/farmacologia , Senescência Celular , Óxido Nítrico Sintase Tipo III/metabolismo , Sirtuína 1/metabolismo , Células Cultivadas , Células Endoteliais da Veia Umbilical Humana/citologia , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Humanos , Peróxido de Hidrogênio , Transdução de Sinais
6.
Cell Physiol Biochem ; 53(5): 865-886, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31724838

RESUMO

BACKGROUND/AIMS: Heart failure is characterized by chronic low-grade vascular inflammation, which in itself can lead to endothelial dysfunction. Clinical trials showed reductions in heart failure-related hospitalizations of type 2 diabetic patients using sodium glucose co-transporter 2 inhibitors (SGLT2i's). Whether and how SGLT2i's directly affect the endothelium under inflammatory conditions is not completely understood. The aim of the study was to investigate whether the SGLT2i Empagliflozin (EMPA) and Dapagliflozin (DAPA) reduce tumor necrosis factor α (TNFα) induced endothelial inflammation in vitro. METHODS: Human coronary arterial endothelial cells (HCAECs) and human umbilical vein endothelial cells (HUVECs) were (pre-)incubated with 1 µM EMPA or DAPA and subsequently exposed to 10 ng/ml TNFα. ROS and NO were measured using live cell imaging. Target proteins were either determined by infrared western blotting or fluorescence activated cell sorting (FACS). The connection between Cav-1 and eNOS was determined by co-immunoprecipitation. RESULTS: Nitric oxide (NO) bioavailability was reduced by TNFα and both EMPA and DAPA restored NO levels in TNFα-stimulated HCAECs. Intracellular ROS was increased by TNFα, and this increase was completely abolished by EMPA and DAPA in HCAECs by means of live cell imaging. eNOS signaling was significantly disturbed after 24 h when cells were exposed to TNFα for 24h, yet the presence of both SGLT2is did not prevent this disruption. TNFα-induced enhanced permeability at t=24h was unaffected in HUVECs by EMPA. Similarly, adhesion molecule expression (VCAM-1 and ICAM-1) was elevated after 4h TNFα (1.5-5.5 fold increase of VCAM-1 and 4-12 fold increase of ICAM-1) but were unaffected by EMPA and DAPA in both cell types. Although we detected expression of SGLT2 protein levels, the fact that we could not silence this expression by means of siRNA and the mRNA levels of SGLT2 were not detectable in HCAECs, suggests aspecificity or our SGLT2 antibody and absence of SGLT2 in our cells. CONCLUSION: These data suggest that EMPA and DAPA rather restore NO bioavailability by inhibiting ROS generation than by affecting eNOS expression or signaling, barrier function and adhesion molecules expression in TNFα-induced endothelial cells. Furthermore, the observed effects cannot be ascribed to the inhibition of SGLT2 in endothelial cells.


Assuntos
Compostos Benzidrílicos/farmacologia , Regulação para Baixo/efeitos dos fármacos , Glucosídeos/farmacologia , Óxido Nítrico/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Fator de Necrose Tumoral alfa/farmacologia , Vasos Coronários/citologia , Células Endoteliais/citologia , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Células Endoteliais da Veia Umbilical Humana , Humanos , Molécula 1 de Adesão Intercelular/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Permeabilidade/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Transportador 2 de Glucose-Sódio/genética , Transportador 2 de Glucose-Sódio/metabolismo , Molécula 1 de Adesão de Célula Vascular
7.
Chem Biol Interact ; 314: 108842, 2019 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-31586451

RESUMO

BACKGROUND AND AIMS: Chronic psychosocial stress is a risk factor for cardiovascular disease. In view of the important role of dipeptidyl peptidase-4 (DPP-4) in human pathophysiology, we studied the role of DPP-4 in stress-related vascular aging in mice, focusing on oxidative stress and the inflammatory response. METHODS AND RESULTS: Male mice were randomly divided into a non-stress group and an immobilization stress group treated for 2 weeks. Chronic stress accelerates aortic senescence and increases plasma DPP-4 levels. Stress increased the levels of gp91phox, p22phox, p47phox, p67phox, p53, p27, p21, p16INK4A, vascular cell adhesion molecule-1, intracellular adhesion molecule-1, monocyte chemoattractant protein-1, matrix metalloproteinase-2 (MMP-2), MMP-9, cathepsin S (Cat S), and Cat K mRNAs and/or protein in the aorta of the stressed mice and decreased their levels of endothelial nitric oxide synthase and SirTuin1 (SirT1). DPP-4 inhibitors can improve stress-induced targeting molecules and morphological changes. In vitro, the inhibition of DPP-4 also alleviated the changes in the oxidative and inflammatory molecules in response to hydrogen peroxide in human umbilical vein endothelial cells. CONCLUSIONS: DPP-4 inhibition can improve vascular aging in stressed mice, possibly by improving oxidative stress production and vascular inflammation. Our results suggest that DPP-4 may become a new therapeutic target for chronic stress-related vascular aging in metabolic cardiovascular diseases.


Assuntos
Dipeptidil Peptidase 4/metabolismo , Inibidores da Dipeptidil Peptidase IV/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Animais , Aorta/metabolismo , Aorta/patologia , Dipeptidil Peptidase 4/sangue , Dipeptidil Peptidase 4/química , Regulação para Baixo/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana , Humanos , Peróxido de Hidrogênio/metabolismo , Inflamação/patologia , Inflamação/prevenção & controle , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Óxido Nítrico Sintase/genética , Óxido Nítrico Sintase/metabolismo , Pirimidinas/farmacologia , Sirtuína 1/metabolismo , Estresse Fisiológico
8.
Zhongguo Zhong Yao Za Zhi ; 44(16): 3441-3447, 2019 Aug.
Artigo em Chinês | MEDLINE | ID: mdl-31602907

RESUMO

To observe the effect of Tripterygium Glycosides Tablets on angiogenesis of rats with type Ⅱ collagen-induced arthritis( CIA) and on the tube formation of human umbilical vein endothelial cells( HUVEC) in vitro. The HUVEC were induced by 20 µg·L-1 vascular endothelial growth factor( VEGF) in vitro,and were treated with 0. 1,1,10 mg·L-1 Tripterygium Glycosides Tablets continuously for 7 hours. The numbers of branches of tube formation were measured. SD rats were immunized to establish CIA. CIA rats were treated with 9,18,36 mg·kg-1·d-1 Tripterygium Glycosides Tablets for 42 days. Histopathological examination( HE) was performed to observe the vascular morphology and vascular density in the synovial membrane of the inflamed joints. Immunohistochemistry and immunofluorescence were performed to observe the expression of platelets-endothelial cell adhesion molecule( CD31) and αsmooth muscle actin( αSMA) in synovial membrane. Immunohistochemistry and Western blot were performed to observe the expression of hypoxia-inducible factors 1α( HIF1α) and angiotensin 1( Ang1) in the synovial tissue. The results showed that the numbers of branches of tube formation of HUVEC induced by VEGF were improved,and declined significantly after treated by Tripterygium Glycosides Tablets. Compared with the normal group,the vascular density,CD31 positive expression,CD31 +/αSMA-immature and total vascular positive expression in the synovial membrane of the model group were significantly increased,and so as HIF1α and Ang1 in the synovium. Tripterygium Glycosides Tablets reduced the synovial vascular density and inhibited the positive expression of CD31,CD31+/αSMA-immature blood vessels and total vascular,but has no effect on CD31+/αSMA+mature blood vessels. Tripterygium Glycosides Tablets also inhibited the expression of HIF1α and Ang1 in synovial membrane of inflammatory joints. Our results demonstrated that Tripterygium Glycosides Tablets could inhibit the angiogenesis of synovial tissue in CIA rats and the tube formation of HUVEC,which is related to the down-regulation of HIF1α/Ang1 signal axis.


Assuntos
Artrite Experimental/tratamento farmacológico , Medicamentos de Ervas Chinesas/farmacologia , Glicosídeos/farmacologia , Tripterygium/química , Inibidores da Angiogênese/farmacologia , Angiotensina I/metabolismo , Animais , Artrite Experimental/induzido quimicamente , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Ratos , Ratos Sprague-Dawley , Transdução de Sinais , Membrana Sinovial/efeitos dos fármacos , Comprimidos , Fator A de Crescimento do Endotélio Vascular
9.
Zhonghua Yi Xue Yi Chuan Xue Za Zhi ; 36(10): 975-979, 2019 Oct 10.
Artigo em Chinês | MEDLINE | ID: mdl-31598939

RESUMO

OBJECTIVE: To assess the effect of miR-137 on the proliferation and apoptosis of human umbilical vein endothelial cells (HUVECs) induced by high glucose and its mechanism. METHODS: HUVECs cells were divided into low-glucose group (5.5 mmol/L glucose-treated cells), high-glucose group (33.36 mmol/L glucose-treated cells), anti-NC group (cells treated with 33.36 mmol/L glucose after anti-NC transfection) and anti-miR-137 group (cells treated with 33.36 mmol/L glucose after anti-miR-137 transfection). After 48 hours, qRT-PCR was used to determine the expression of miR-137. CCK-8 assay and flow cytometry were used to detect cell proliferation and apoptosis rate, respectively. The targeting relationship between miR-137 and AKT2 was validated by dual fluorescence reporter gene detection system and AKT2 protein expression after overexpression or inhibition of miR-137. RESULTS: High glucose could significantly up-regulate the expression of miR-137 in HUVECs cells, and the expression of miR-137 in HUVECs cells transfected with miR-137 inhibitor was significantly decreased (P<0.05). High glucose can significantly inhibit HUVECs cell proliferation and induce apoptosis, while inhibition of miR-137 expression can weaken the effect of high glucose on HUVECs cell proliferation inhibition and apoptosis promotion (P<0.05). Inhibiting AKT2 expression could weaken the inhibitory effect of miR-137 inhibitor on HUVECs cell proliferation and apoptosis (P<0.05). CONCLUSION: Inhibiting the expression of miR-137 gene can attenuate the proliferation inhibition and apoptosis promotion of HUVECs induced by high glucose, and the mechanism is related to activating the expression of AKT2.


Assuntos
Apoptose , Proliferação de Células , Células Endoteliais da Veia Umbilical Humana/citologia , MicroRNAs/genética , Proteínas Proto-Oncogênicas c-akt/genética , Células Cultivadas , Glucose , Humanos
10.
Int J Nanomedicine ; 14: 7399-7417, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31571858

RESUMO

Purpose: We studied the effects of silver nanoparticles (AgNPs) on human blood platelet function. We hypothesized that AgNPs, a known antimicrobial agent, can be used as blood-compatible, "ideal material'' in medical devices or as a drug delivery system. Therefore, the aim of the current study was to investigate if functionalized AgNPs affect platelet function and platelets as well as endothelial cell viability in vitro. Methods: AgNPs, functionalized with reduced glutathione (GSH), polyethylene glycol (PEG) and lipoic acid (LA) were synthesized. Quartz crystal microbalance with dissipation was used to measure the effect of AgNPs on platelet aggregation. Platelet aggregation was measured by changes in frequency and dissipation, and the presence of platelets on the sensor surface was confirmed and imaged by phase contrast microscopy. Flow cytometry was used to detect surface abundance of platelet receptors. Lactate dehydrogenase test was used to assess the potential cytotoxicity of AgNPs on human blood platelets, endothelial cells, and fibroblasts. Commercially available ELISA tests were used to measure the levels of thromboxane B2 and metalloproteinases (MMP-1, MMP-2) released by platelets as markers of platelet activation. Results: 2 nm AgNPs-GSH, 3.7 nm AgNPs-PEG both at 50 and 100 µg/mL, and 2.5 nm AgNPs-LA at 100 µg/mL reduced platelet aggregation, inhibited collagen-mediated increase in total P-selectin and GPIIb/IIIa, TXB2 formation, MMP-1, and MMP-2 release. The tested AgNPs concentrations were not cytotoxic as they did not affect, platelet, endothelial cell, or fibroblast viability. Conclusion: All tested functionalized AgNPs inhibited platelet aggregation at nontoxic concentrations. Therefore, functionalized AgNPs can be used as an antiplatelet agent or in design and manufacturing of blood-facing medical devices, such as vascular grafts, stents, heart valves, and catheters.


Assuntos
Plaquetas/efeitos dos fármacos , Nanopartículas Metálicas/química , Agregação Plaquetária/efeitos dos fármacos , Prata/farmacologia , Colágeno/metabolismo , Fibroblastos/efeitos dos fármacos , Citometria de Fluxo , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , L-Lactato Desidrogenase/metabolismo , Ligantes , Metaloproteinases da Matriz/metabolismo , Nanopartículas Metálicas/ultraestrutura , Selectina-P/metabolismo , Tamanho da Partícula , Complexo Glicoproteico GPIIb-IIIa de Plaquetas/metabolismo , Polietilenoglicóis/química , Técnicas de Microbalança de Cristal de Quartzo , Espectroscopia de Infravermelho com Transformada de Fourier , Tromboxano B2/metabolismo
11.
Braz Oral Res ; 33: e059, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31664357

RESUMO

We recently demonstrated that a co-culture system of human umbilical vein endothelial cells (HUVECs) and human dental pulp stem cells (hDPSCs) could enhance angiogenesis ability in vitro. However, whether tumor necrosis factor α (TNF-α) could promote blood vessel formation during pulp regeneration remained unknown. The aim of this study was to investigate the effects of TNF-α on the formation of endothelial tubules and vascular networks in a co-culture system of hDPSCs and HUVECs. hDPSCs were co-cultured with HUVECs at a ratio of 1:5. The Matrigel assay was performed to detect the total tubule branching lengths and numbers of branches, and the Cell-Counting Kit 8 assay was performed to examine the effect of TNF-α on cell proliferation. Real-time polymerase chain reactions and western blot were used to detect vascular endothelial growth factor (VEGF) mRNA and protein expression. The Matrigel assay showed significantly greater total branching lengths and numbers of branches formed in the experimental groups treated with different concentrations of TNF-α compared with the control group. The decomposition times of the tubule structures were also significantly prolonged (P < 0.05). Treatment with 50 ng/ml TNF-α did not significantly change the proliferation of co-cultured cells, but it significantly increased the VEGF mRNA and protein expression levels (p < 0.05). In addition, the migration abilities of HUVECs and hDPSCs increased after co-culture with TNF-α (p < 0.05). TNF-α enhanced angiogenic ability in vitro in the co-culture system of hDPSCs and HUVECs.


Assuntos
Indutores da Angiogênese/farmacologia , Polpa Dentária/citologia , Polpa Dentária/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Neovascularização Fisiológica/efeitos dos fármacos , Fator de Necrose Tumoral alfa/farmacologia , Adolescente , Adulto , Western Blotting , Contagem de Células , Ensaios de Migração Celular , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/fisiologia , Células Cultivadas , Colágeno , Polpa Dentária/fisiologia , Combinação de Medicamentos , Células Endoteliais da Veia Umbilical Humana/fisiologia , Humanos , Laminina , Neovascularização Fisiológica/fisiologia , Proteoglicanas , Reação em Cadeia da Polimerase em Tempo Real , Valores de Referência , Reprodutibilidade dos Testes , Fatores de Tempo , Fator A de Crescimento do Endotélio Vascular/análise , Fator A de Crescimento do Endotélio Vascular/efeitos dos fármacos , Adulto Jovem
12.
Cell Physiol Biochem ; 53(3): 480-495, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31486323

RESUMO

BACKGROUND/AIMS: Hypoxia Inducible Factor-1α (HIF-1α) is involved in cancer progression and is stabilized by the chaperone HSP90 (Heat Shock Protein 90), preventing degradation. Previously identified HSP90 inhibitors bind to the N-terminal pocket of HSP90, which blocks binding to HIF-1α and induces HIF-1α degradation. N-terminal inhibitors have failed in the clinic as single therapy treatments partially because they induce a heat shock response. SM molecules are HSP90 inhibitors that bind to the C-terminus of HSP90 and do not induce a heat shock response. The effects of these C-terminal inhibitors on HIF-1α are unreported. METHODS: HCT116, MDA-MB-231, PC3, and HEK293T cells were treated with HSP90 inhibitors. qRT-PCR and western blotting was performed to assess mRNA and protein levels of HIF-1α, HSP- and RACK1-related genes. siRNA was used to knockdown RACK1, while MG262 was used to inhibit proteasome activity. Dimethyloxalylglycine (DMOG) was used to inhibit activity of the prolyl hydroxylases (PHDs). Anti-angiogenic activity of HSP90 inhibitors was assessed using a HUVEC tubule formation assay. RESULTS: We show that SM compounds decrease HIF-1α target expression at the mRNA and protein level under hypoxia in colorectal, breast and prostate cancer cells, leading to cell death, without inducing a heat shock response. Surprisingly, we found that when the C-terminal of HSP90 is inhibited, HIF-1α degradation occurs through the proteasome and prolyl hydroxylases in an oxygen-dependent manner even in very low levels of oxygen (tumor hypoxia levels). RACK1 was not required for proteasomal degradation of HIF-1α. CONCLUSION: Our results suggest that by targeting the C-terminus of HSP90 we can exploit the prolyl hydroxylase and proteasome pathway to induce HIF-1α degradation in hypoxic tumors.


Assuntos
Hipóxia Celular/fisiologia , Proteínas de Choque Térmico HSP90/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Aminoácidos Dicarboxílicos/metabolismo , Western Blotting , Hipóxia Celular/genética , Sobrevivência Celular/genética , Sobrevivência Celular/fisiologia , Células HCT116 , Células HEK293 , Proteínas de Choque Térmico HSP90/genética , Células Endoteliais da Veia Umbilical Humana , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Células PC-3 , Prolil Hidroxilases/genética , Prolil Hidroxilases/metabolismo , Complexo de Endopeptidases do Proteassoma/genética , Complexo de Endopeptidases do Proteassoma/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo
14.
Undersea Hyperb Med ; 46(4): 509-519, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31509907

RESUMO

Nitric oxide (NO) may protect against gas bubble formation and risk of decompression sickness. We have previously shown that the crucial co-factor tetrahydrobiopterin (BH4) is oxidized in a dose-dependent manner when exposed to hyperoxia similar to diving conditions but with minor effects on the NO production by nitric oxide synthase. By manipulating the intracellular redox state, we further investigated the relationship between BH4 levels and production of NO in human endothelial cells (HUVECs). HUVECs were cultured with and without ascorbic acid (AA) and the glutathione (GSH) synthesis inhibitor buthionine sulfoximine, prior to hyperoxic exposure. The levels of biopterins and GSH were determined in cell lysates while the production of NO was determined in intact cells. Omitting AA resulted in a 91% decrease in BH4 levels (0.49 ± 0.08 to 0.04 ± 0.01 pmol/106 cells, p⟨0.001) at 20 kPa oxygen (O2), and 88% decrease (0.24 ± 0.03 to 0.03 ± 0.01 pmol/106 cells, p=0.01) after exposure to 60 kPa O2. The NO generation was decreased by 23% (74.5 ± 2.2 to 57.3 ± 5.6 pmol/min/mg protein, p⟨0.001) at 20 kPa O2, but no significant change was observed at 60 kPa O2. GSH depletion had no effects on the NO generation. No correlation was found between NO generation and the corresponding intracellular BH4 concentration (p=0.675, r=-0.055) or the BH4 to BH2 ratio (p=0.983, r=0.003), determined across 18 in vitro experiments. Decreased BH4 in HUVECs, due to hyperoxia or lack of ascorbic acid, does not imply corresponding decreases in NO generation.


Assuntos
Ácido Ascórbico/administração & dosagem , Biopterina/análogos & derivados , Células Endoteliais/metabolismo , Hiperóxia/metabolismo , Óxido Nítrico/biossíntese , Antimetabólitos , Biopterina/análise , Biopterina/metabolismo , Butionina Sulfoximina , Doença da Descompressão/etiologia , Doença da Descompressão/prevenção & controle , Endotélio Vascular , Glutationa/análise , Glutationa/metabolismo , Células Endoteliais da Veia Umbilical Humana , Humanos , Óxido Nítrico Sintase/metabolismo , Oxirredução , Oxigênio , Pressão Parcial
15.
Nan Fang Yi Ke Da Xue Xue Bao ; 39(8): 898-903, 2019 Aug 30.
Artigo em Chinês | MEDLINE | ID: mdl-31511208

RESUMO

OBJECTIVE: To investigate the effect of miR-186 inhibition on the expression of hypoxia-inducible factor-1α (HIF-α) and mitochondrial function in hypoxic vascular endothelial cells. METHODS: Human umbilical vein endothelial cells (HUVECs) cultured in routine or hypoxic conditions for 6 h were examined for the expression of miR-186. A miR-186 inhibitor was transfected in the HUVECs, and the cells were subsequently cultured in hypoxic condition for 6 h to observe the changes in the mitochondrial structure under an electron microscope. The changes in the mRNA and protein expressions of HIF-1α in response to miR-186 interference were tested using real-time fluorescent quantitative PCR and Western blotting. RESULTS: The expression of miR-18 was mildly increased in HUVECs after hypoxic exposure for 6 h (P=0.0188). Interference of miR-186 expression obviously promoted the mRNA and protein expressions of HIF-1α in HUVECs. In hypoxic conditions, miR-186 interference significantly reduced mitochondrial damage in HUVECs as observed under electron microscope (P=0.0297). CONCLUSIONS: Inhibition of miR-186 protects vascular endothelial cells against hypoxic injuries by promoting HIF-α expression to lessen mitochondrial damage, suggesting the possibility of targeted miR-186 interference for treatment of hemorrhagic shock.


Assuntos
Veias Umbilicais , Hipóxia Celular , Células Endoteliais da Veia Umbilical Humana , Humanos , Hipóxia , Subunidade alfa do Fator 1 Induzível por Hipóxia , MicroRNAs , Mitocôndrias
16.
Int J Nanomedicine ; 14: 5875-5894, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31534329

RESUMO

Background: Theranostics based on multifunctional nanoparticles (NPs) is a promising field that combines therapeutic and diagnostic functionalities into a single nanoparticle system. However, the major challenges that lie ahead are how to achieve accurate early diagnosis and how to develop efficient and noninvasive treatment. Sonodynamic therapy (SDT) utilizing ultrasound combined with a sonosensitizer represents a novel noninvasive modality for cancer therapy. Different ultrasound frequencies have been used for SDT, nevertheless, whether the effect of SDT can enhance synergistic HIFU ablation remains to be investigated. Materials and methods: We prepared a nanosystem for codelivery of a sonosensitizer (methylene blue, MB) and a magnetic resonance contrast agent (gadodiamide, Gd-DTPA-BMA) based on hydrophilic biodegradable polymeric NPs composed of poly (lactic-co-glycolic acid) (PLGA). To enhance accumulation and penetration of the NPs at the tumor site, the surface of PLGA NPs was decorated with a tumor-homing and penetrating peptide-F3 and polyethylene glycol (PEG). The physicochemical, imaging and therapeutic properties of F3-PLGA@MB/Gd and drug safety were thoroughly evaluated both in vitro and in vivo. F3-PLGA@MB/Gd was evaluated by both photoacoustic and resonance imaging. Results: F3-PLGA@MB/Gd NPs exhibited higher cellular association than non-targeted NPs and showed a more preferential enrichment at the tumor site. Furthermore, with good drug safety, the apoptosis triggered by ultrasound in the F3-PLGA@MB/Gd group was greater than that in the contrast group. Conclusion: F3-PLGA@MB/Gd can work as a highly efficient theranostic agent, and the incorporation of targeted multimodal and combined therapy could be an encouraging strategy for cancer treatment.


Assuntos
Peptídeos Penetradores de Células/química , Tratamento por Ondas de Choque Extracorpóreas , Nanopartículas/química , Neoplasias/diagnóstico por imagem , Ultrassonografia , Animais , Bovinos , Morte Celular , Linhagem Celular Tumoral , Terapia Combinada , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Imagem por Ressonância Magnética , Camundongos Endogâmicos BALB C , Camundongos Nus , Nanopartículas/ultraestrutura , Espécies Reativas de Oxigênio/metabolismo
17.
Cell Physiol Biochem ; 53(4): 638-647, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31556253

RESUMO

BACKGROUND/AIMS: Prolonged hyperosmotic shrinkage evokes expression of osmoprotective genes via nuclear factor NFAT5-mediated pathway and activates Na+ influx via hypertonicity-induced cation channels (HICC). In human umbilical vein endothelial cells (HUVEC) elevation of intracellular sodium concentration ([Na+]i) triggers transcription of dozens of early response genes (ERG). This study examined the role of monovalent cations in the expression of Na+i-sensitive ERGs in iso- and hyperosmotically shrunken HUVEC. METHODS: Cell volume was measured by 3D reconstruction of cell shape and as 14C-urea available space. Intracellular Na+ and K+ content was measured by flame atomic absorption spectrometry. ERG transcription was estimated by RT-PCR. RESULTS: Elevation of medium osmolality by 150 mM mannitol or cell transfer from hypo- to isosmotic medium decreased cell volume by 40-50%. Hyperosmotic medium increased [Na+]i by 2-fold whereas isosmotic shrinkage had no impact on this parameter. Hyperosmotic but not isosmotic shrinkage increased up-to 5-fold the content of EGR1, FOS, ATF3, ZFP36 and JUN mRNAs. Expression of these ERGs triggered by hyperosmotic shrinkage and Na+,K+-ATPase inhibition by 0.1 µM ouabain exhibited positive correlation (R2=0.9383, p=0.0005). Isosmotic substitution of NaCl by N-methyl-D-glucamine abolished an increment of [Na+]i and ERG expression triggered by mannitol addition. CONCLUSION: Augmented expression of ERGs in hyperosmotically shrunken HUVEC is mediated by elevation of [Na+]i.


Assuntos
Tamanho Celular , Sódio/metabolismo , Tamanho Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Proteína 1 de Resposta de Crescimento Precoce/genética , Proteína 1 de Resposta de Crescimento Precoce/metabolismo , Células Endoteliais da Veia Umbilical Humana , Humanos , Meglumina/farmacologia , Ouabaína/farmacologia , Potássio/metabolismo , Cloreto de Sódio/farmacologia , ATPase Trocadora de Sódio-Potássio/antagonistas & inibidores , ATPase Trocadora de Sódio-Potássio/metabolismo , Tristetraprolina/genética , Tristetraprolina/metabolismo
18.
Int J Nanomedicine ; 14: 6451-6464, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31496697

RESUMO

Background: We recently reported on long-term comprehensive biocompatibility and biodistribution study of fluorescent nanodiamond particles (NV)-Z-average 800nm (FNDP-(NV)) in rats. FNDP-(NV) primary deposition was found in the liver, yet liver function tests remained normal. Purpose: The present study aimed to gain preliminary insights on discrete localization of FNDP-(NV) in liver cells of the hepatic lobule unit and venous micro-vasculature. Kinetics of FDNP-(NV) uptake into liver cells surrogates in culture was conducted along with cell cytokinesis as markers of cells' viability. Methods: Preserved liver specimens from a pilot consisting of two animals which were stained for cytoskeletal elements (fluorescein-isothiocyanate-phalloidin) were examined for distribution of FNDP-(NV) by fluorescent microscopy (FM) and Confocal-FM (CFM) using near infra-red fluorescence (NIR). Hepatocellular carcinoma cells (HepG-2) and human umbilical vein endothelial cells (HUVEC) were cultured with FNDP-(NV) and assayed for particle uptake and location using spectrophotometric technology and microscopy. Results: HepG-2 and HUVEC displayed rapid (<30 mins) onset and concentration-dependent FNDP-(NV) internalization and formation of peri-nuclear corona. FM/CFM of liver sections revealed FNDP-(NV) presence throughout the hepatic lobules structures marked by spatial distribution, venous microvascular spaces and parenchyma and non-parenchyma cells. Conclusion: The robust presence of FNDP-(NV) throughout the hepatic lobules including those internalized within parenchyma cells and agglomerates in the liver venous micro-circulation were not associated with macro or micro histopathological signs nor vascular lesions. Cells cultures indicated normal cytokinesis in cells containing FNDP-(NV) agglomerates. Liver parenchyma cells and the liver microcirculation remain agnostic to presence of FNDP-(NV) in the sinusoids or internalized in the hepatic cells.


Assuntos
Materiais Biocompatíveis/farmacologia , Fígado/metabolismo , Nanodiamantes/química , Animais , Células Hep G2 , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Imagem Tridimensional , Cinética , Fígado/efeitos dos fármacos , Microscopia de Fluorescência , Tamanho da Partícula , Ratos Sprague-Dawley , Distribuição Tecidual
19.
Adv Exp Med Biol ; 1155: 471-482, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31468424

RESUMO

Endothelial cell dysfunction (ECD) is a broad term, which implies dysregulation of endothelial cell functions. Several factors contribute to ECD including high blood pressure, high cholesterol levels, diabetes, obesity, hyperglycemia, and advanced glycation end products (AGEs). The highly reactive dicarbonyl methylglyoxal (MGO) is mainly formed as byproduct of glycolysis. Therefore, high blood glucose levels result in increased MGO accumulation. Taurine-rich foods are considered to protect against various diseases including vasculopathy and to exert anti-aging effects. Here, we investigated the protective effect of hot water extract of Octopus ocellatus meat (OOM), which contains high amounts of taurine, on MGO-induced cell damage in human umbilical vein endothelial cells and zebrafish embryos. Hot water extract of OOMinhibited MGO-induced cytotoxicity and DNA damage, as well as AGEs accumulation. In addition, hot water extract of OOM protected against vascular damage in zebrafish embryos. These results suggest that hot water extract of OOM possesses protective activity against MGO-induced cytotoxicity in both umbilical vein endothelial cells and zebrafish embryos. Therefore, it could be used as a dietary source of an agent for the prevention of vascular diseases.


Assuntos
Extratos Celulares/farmacologia , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Octopodiformes/química , Aldeído Pirúvico/toxicidade , Taurina/farmacologia , Animais , Células Cultivadas , Embrião não Mamífero/efeitos dos fármacos , Humanos , Carne , Peixe-Zebra
20.
Adv Exp Med Biol ; 1155: 705-715, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31468441

RESUMO

Blood vessels become less flexible with senescence; arteries narrow and become less flexible, disturbing blood circulation in aging and other vascular diseases. Mechanistically, vascular senescence plays an important role in the pathogenesis of normal aging and age-related vascular diseases. Vascular senescence also causes vascular dysfunction, resulting in damage to the vessel wall. Vascular aging involves the senescence of endothelial cells. Hydrogen peroxide is widely used to achieve oxidative stress-induced premature senescence. Here, we investigated the protective effects of a hot water extract of Loliolus beka meat (LBM) against H2O2-exposed HUVECs, a human umbilical vein endothelial cells line. The hot water extract of LBM protected cells against H2O2-induced cytotoxicity while reducing the expression of senescence markers, including ß-galactosidase, p53, and p21. In addition, the hot water extract of LBM protected against H2O2-induced DNA damage. These findings suggest that the hot water extract of LBM protects HUVECs from H2O2-induced senescence by preventing cellular damage. LBM serve as a supplement or natural food with benefits against vascular disease.


Assuntos
Extratos Celulares/farmacologia , Decapodiformes/química , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Estresse Oxidativo , Animais , Células Cultivadas , Senescência Celular , Humanos , Peróxido de Hidrogênio , Carne
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