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1.
Int Immunopharmacol ; 81: 106257, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32044659

RESUMO

The activation of NLRP3 inflammasome and NF-κB pathway, associating with oxidativestress, have been implicated in the development of acute lung injury (ALI) and acute respiratory distress syndrome (ARDS). NecroX-5 has been reported to exhibit theeffectsofanti-oxidation and anti-stress in various diseases. However, the role of NecroX-5 in ALI has not been explicitly demonstrated. The aim of this study was to explore the therapeutic effects and potential mechanism action of NecroX-5 on ALI. Here, we found that NecroX-5 pretreatment dramatically diminished the levels of IL-1ß, IL-18 and ROS in in RAW264.7 cells challenged with LPS and ATP. Furthermore, NecroX-5 suppressed the activation of NLRP3 inflammasome and NF-κB signalpathway. In addition, NecroX-5 also inhibited the thioredoxin-interacting protein (TXNIP) expression. In vivo, NecroX-5 reduced the LPS-induced lung histopathological injury, the number of TUNEL-positive cells, lung wet/dry (W/D) ratio, levels of total protein and inflammatory cytokines in the bronchoalveolar lavage fluid (BALF) in mice. Additionally, LPS-induced upregulation of myeloperoxidase (MPO), ROS production and malondialdehyde (MDA) were inhibited by NecroX-5 administration. Thus, our results demonstrate that NecroX-5 protects against LPS-induced ALI by inhibiting TXNIP/NLRP3 and NF-κB.

2.
Int Immunopharmacol ; 77: 105973, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31677992

RESUMO

Sepsis is a syndrome of life-threatening organ dysfunction caused by dysregulated host responses to infection. Macrophage polarization is a key process involved in the pathogenesis of sepsis. Recent evidence has demonstrated that autophagy participates in the regulation of macrophage polarization in different phases of inflammation. Here, we investigated whether trichostatin A (TSA), a histone deacetylase (HDAC) inhibitor, promotes the macrophage M2 phenotype by enhancing autophagy to counteract excessive inflammation in a cecal ligation and puncture (CLP) mouse model. TSA stimulation increased the proportions of M2 marker (CD206, CD124 and CD23)-labeled RAW264.7 macrophages. Furthermore, with increasing TSA doses, autophagy was enhanced gradually. Interestingly, the autophagy activator rapamycin (Rap), also known as an mTOR inhibitor, unexpectedly decreased the proportions of M2 marker-labeled macrophages. However, TSA treatment reversed the Rap-induced decreases in CD206-labeled macrophages. Next, we stimulated different groups of RAW264.7 cells with the autophagy inhibitors MHY1485 or 3-methyladenine (3-MA). Inhibition of autophagy at any stage in the process suppressed TSA-induced macrophage M2 polarization, but the effect was not associated with mTOR activity. In vivo, TSA administration promoted peritoneal macrophage M2 polarization, increased LC3 II expression, attenuated sepsis-induced organ (lung, liver and kidney) injury, and altered systemic inflammatory cytokine secretion. However, 3-MA abolished the protective effects of TSA in CLP mice and decreased the number of M2 peritoneal macrophages. Therefore, TSA promotes the macrophage M2 phenotype by enhancing autophagy to reduce systemic inflammation and ultimately improves the survival of mice with polymicrobial sepsis.


Assuntos
Autofagia/efeitos dos fármacos , Ácidos Hidroxâmicos/farmacologia , Inflamação/tratamento farmacológico , Macrófagos Peritoneais/efeitos dos fármacos , Sepse/tratamento farmacológico , Animais , Biomarcadores/metabolismo , Linhagem Celular , Citocinas/metabolismo , Modelos Animais de Doenças , Inibidores de Histona Desacetilases/farmacologia , Inflamação/metabolismo , Ligadura/métodos , Pulmão/efeitos dos fármacos , Pulmão/metabolismo , Ativação de Macrófagos/efeitos dos fármacos , Macrófagos Peritoneais/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Fenótipo , Punções/métodos , Células RAW 264.7 , Sepse/metabolismo
3.
Biosens Bioelectron ; 145: 111700, 2019 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-31539651

RESUMO

DNA methyltransferase (MTase) and polynucleotide kinase (PNK) are both DNA-dependent enzymes that play important roles in DNA methylation and DNA repair processes, respectively. Dysregulation of their activities is associated with various human diseases. Herein, we present a specific and sensitive biosensing strategy, named terminal deoxynucleotidyl transferase (TdT)-activated nicking enzyme amplification reaction (TdT-NEAR), for their activity detection. As for MTase detection, an enclosed dumbbell-shaped oligonucleotide substrate, whose symmetric stem containing a recognition site of Dam MTase and an incomplete recognition sequence of nicking endonuclease Nt.BbvCI, was used. Typically, the substrate is methylated by Dam MTase and subsequently cleaved by Dpn I. In the presence of TdT and dGTP, poly(guanine, G) sequences are extended from the released 3'-OH ends, achieving the conversion of the incomplete Nt.BbvCI recognition sequence to an intact one. The extension products can then be used to trigger Nt.BbvCI-catalyzed cyclic cleavage of fluorophore/quencher-labelled oligonucleotide probe, giving a significantly enhanced fluorescence output. Such a sensing system can achieve sensitive and specific detection of Dam MTase with a detection limit of 0.002 U/mL. The unique working mechanism endows the sensing system with improved anti-interference capability and thus increased application potential in complex biological samples. Moreover, it was also demonstrated to work well for Dam MTase inhibitor screening and inhibitory activity evaluation, thus holding great potential in disease diagnosis and drug discovery. Using a simpler 3'-phosphorylated linear substrate and the same fluorescent probe, the TdT-NEAR strategy can be easily extended to the activity analysis of PNK, thus revealing wide application potential in bioanalysis.


Assuntos
Técnicas Biossensoriais , Metilases de Modificação do DNA/isolamento & purificação , DNA Nucleotidilexotransferase/química , Polinucleotídeo 5'-Hidroxiquinase/isolamento & purificação , Metilação de DNA/genética , Metilases de Modificação do DNA/química , Corantes Fluorescentes/química , Humanos , Polinucleotídeo 5'-Hidroxiquinase/química , DNA Metiltransferases Sítio Específica (Adenina-Específica) , Espectrometria de Fluorescência
4.
Anal Chem ; 91(20): 13165-13173, 2019 10 15.
Artigo em Inglês | MEDLINE | ID: mdl-31512479

RESUMO

The introduction of nanotechnology can overcome some inherent drawbacks of traditional DNA probes, thus promoting their applications in living cells. Herein, a three-dimensional DNA nanostructure, a DNA nanolantern, was prepared via simple nucleotide hybridization of four short-stranded oligonucleotides and successfully applied to the construction of a novel DNA probe and signal amplifier. Compared to most reported DNA nanostructures, a DNA nanolantern shows the distinct advantages of low cost, easy design and preparation, more and arbitrary adjusted probe numbers, and high fluorescence resonance energy transfer (FRET) signal readout. Compared to traditional DNA probes, the constructed nanolantern-based one has improved cell internalization efficiency, enhanced biostability, accelerated reaction kinetics, excellent biocompatibility, and greatly reduced false-positive output and was demonstrated to work well for probing the expression level of tumor-related mRNA and microRNA in living cells. The DNA nanolantern can also be easily integrated with some reported signal amplification strategies, e.g., isothermal hybridization chain reaction (HCR), and the obtained signal amplifier combines the advantages of the DNA nanolantern and the HCR, enabling sensitive imaging detection of ultralow abundance targets in living cells. This work demonstrated that this simple DNA nanostructure can not only improve the performance of traditional DNA probes but can also be easily integrated with reported DNA-based strategy and technology, thus showing a broad application prospect.

5.
BMC Infect Dis ; 19(1): 592, 2019 Jul 08.
Artigo em Inglês | MEDLINE | ID: mdl-31286917

RESUMO

BACKGROUND: Norovirus (NoV) is recognized as a leading cause of acute gastroenteritis (AGE) outbreaks in settings globally. Studies have shown that employees played an important role in the transmission mode during some NoV outbreaks. This study aimed to investigate the prevalence of NoV infection and duration of NoV shedding among employees during NoV outbreaks, as well as factors affecting shedding duration. METHODS: Specimens and epidemiological data were collected from employees who were suspected of being involved in the transmission or with AGE symptoms during NoV outbreaks in Xuhui District, Shanghai, from 2015 to 2017. Specimens were detected using real-time RT-PCR to determine whether or not employees had become infected with NoV. Specimens were collected every 3-7 days from NoV-infected employees until specimens became negative for NoV. RESULTS: A total of 421 employees were sampled from 49 NoV outbreaks, and nearly 90% of them (377/421) were asymptomatic. Symptomatic employees showed significantly higher prevalence of NoV infection than asymptomatic ones (70.5% vs. 17.0%, P < 0.01). The average duration of NoV shedding was 6.9 days (95% confidence interval: 6.1-7.7 days) among 88 NoV-infected individuals, and was significantly longer in symptomatic individuals than in asymptomatic ones (9.8 days vs. 5.6 days, P < 0.01). In Cox proportional-hazards model, after adjusting age and gender, symptoms was the only factor associated with duration of NoV shedding. CONCLUSIONS: Compared with asymptomatic employees, symptomatic employees had higher prevalence of NoV infection and longer durations of NoV shedding. Since NoV shedding duration among NoV-infected employees tends to be longer than their isolation time during outbreaks, reinforcement of hygiene practices among these employees is especially necessary to reduce the risk of virus secondary transmissions after their return to work.


Assuntos
Infecções por Caliciviridae , Surtos de Doenças/estatística & dados numéricos , Gastroenterite , Norovirus/genética , Adulto , Canal Anal/virologia , Infecções Assintomáticas/epidemiologia , Infecções por Caliciviridae/diagnóstico , Infecções por Caliciviridae/epidemiologia , Infecções por Caliciviridae/virologia , China/epidemiologia , Feminino , Gastroenterite/diagnóstico , Gastroenterite/epidemiologia , Gastroenterite/virologia , Humanos , Masculino , Pessoa de Meia-Idade , Prevalência , RNA Viral/análise , RNA Viral/genética , Reação em Cadeia da Polimerase em Tempo Real
6.
Lab Invest ; 99(8): 1143-1156, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-30911150

RESUMO

Acute respiratory distress syndrome (ARDS) is a uniform progression of overwhelming inflammation in lung tissue with extensive infiltration of inflammatory cells. Neutrophil apoptosis is thought to be a significant process in the control of the resolution phase of inflammation. It has been proved that 5-Aza-2'-deoxycytidine (Aza) can inhibit cancer by activating death-associated protein kinase 1 (DAPK1) to promote apoptosis. However, the effect of DAPK1 on neutrophil apoptosis is unclear, and research on the role of Aza in inflammation is lacking. Here, we investigated whether Aza can regulate DAPK1 expression to influence the fate of neutrophils in ARDS. In vitro, we stimulated neutrophil-like HL-60 (dHL-60) cells with different concentrations of Aza for different durations and used RNA interference to up- or downregulate DAPK1 expression. We observed that culturing dHL-60 cells with Aza increased apoptosis by inhibiting NF-κB activation to modulate the expression of Bcl-2 family proteins, which was closely related to the levels of DAPK1. In vivo, ARDS was evoked by intratracheal instillation of lipopolysaccharide (LPS; 3 mg/kg). One hour after LPS administration, mice were treated with Aza (1 mg/kg, i.p.). To inhibit DAPK1 expression, mice were intraperitoneally injected with a DAPK1 inhibitor. Aza treatment accelerated inflammatory resolution in LPS-induced ARDS by suppressing pulmonary edema, alleviating lung injury and decreasing the infiltration of inflammatory cells in bronchoalveolar lavage fluid (BALF). Moreover, Aza reduced the production of proinflammatory cytokines. However, administration of the DAPK1 inhibitor attenuated the protective effects of Aza. Similarly, the proapoptotic function of Aza was prevented when DAPK1 was inhibited either in vivo or in vitro. In summary, Aza promotes neutrophil apoptosis by activating DAPK1 to accelerate inflammatory resolution in LPS-induced ARDS. This study provides the first evidence that Aza prevents LPS-induced neutrophil survival by modulating DAPK1 expression.

7.
Chem Sci ; 10(8): 2290-2297, 2019 Feb 28.
Artigo em Inglês | MEDLINE | ID: mdl-30881654

RESUMO

DNA methylation is a significant epigenetic mechanism involving processes of transferring a methyl group onto cytosine or adenine. Such DNA modification catalyzed by methyltransferase (MTase) plays important roles in the modulation of gene expression and other cellular activities. Herein, we develop a simple and sensitive biosensing platform for the detection of DNA MTase activity by using only two oligonucleotides. The fluorophore labeled molecular beacon (MB) can be methylated by MTase and subsequently cleaved by endonuclease DpnI at the stem, giving a shortened MB. The shortened MB can then hybridize with a primer DNA, initiating a cycle of strand displacement amplification (SDA) reactions. The obtained SDA products can unfold new MB and initiate another cycle of SDA reaction. Therefore, continuous enlargement of SDA and exponential amplification of the fluorescence signal are achieved. Because the triple functions of substrate, template and probe are elegantly integrated in one oligonucleotide, only two oligonucleotides are necessary for multiple amplification cycles, which not only reduces the complexity of the system, but also overcomes the laborious and cumbersome operation that is always a challenge in conventional methods. This platform exhibits an extremely low limit of detection of 3.3 × 10-6 U mL-1, which is the lowest to our knowledge. The proposed MTase-sensing platform was also demonstrated to perform well in a real-time monitoring mode, which can achieve a further simplified and high-throughput detection. The sensing strategy might be extended to the activity detection of other enzymes, thus showing great application potential in bioanalysis and clinical diagnosis.

8.
Int Immunopharmacol ; 69: 289-298, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30753968

RESUMO

The timely resolution of pulmonary inflammation coordinated by endogenous pro-resolving mediators helps limit lung tissue injury, but few endogenous pro-resolving mediators that are normally operative during acute inflammation. The protective effects of BML-111 (5(S)-6(R)-7-trihydroxyheptanoic acid methyl ester), a potent commercially available anti-inflammatory and pro-resolving mediator, on ventilation-induced lung injury (VILI) have been extensively studied, but its characteristics as a pro-resolving mediator have not. Here, anesthetized Sprague-Dawley rats were ventilated with a high tidal volume (20 mL/kg, HVT) for 1 h and randomly allocated to recover for 6, 12, 24, 48, 72, 96 or 168 h; BML-111 was administered at the peak of inflammation to evaluate its pro-resolving effect on VILI. The one-hour HVT induced a maximal pulmonary inflammatory response at 12 h that was largely resolved by 72 h. BML-111 largely resolved the maximal inflammatory response at 48 h; the resolution interval (Ri) was shortened by 26 h. Similarly, HVT elicited a time course of changes in histopathology and pulmonary edema, and BML-111 alleviates these changes. Mechanistically, neutrophil apoptosis was significantly increased in BML-111-treated rats subjected to HVT. The apoptosis inhibitor z-VAD-fmk partially reversed the proapoptotic actions of BML-111 on neutrophil and the resolving effects of BML-111 on VILI but had no effect alone. Importantly, the HVT treatment activated the nuclear factor E2-related factor 2(Nrf2)/heme oxygenase-1(HO-1) and NF-κB signaling pathways in the lung tissue, and BML-111 further induced Nrf2 and HO-1 expression but inhibited the NF-κB pathway. Intriguingly, when we inhibited the Nrf2/HO-1 pathway with the HO-1 inhibitor zinc protoporphyrin IX (ZnPPIX), Nrf2 expression was further increased, but the inhibitory effects of BML-111 on the NF-κB pathway and on the subsequent inflammatory response, and the proapoptotic actions on neutrophil were reversed. The results suggest that BML-111 promotes the resolution of HVT-induced inflammation to mitigate VILI in rats, perhaps by modulating the Nrf2/HO-1 and NF-κB pathways and subsequently increasing neutrophil apoptosis.


Assuntos
Ácidos Heptanoicos/uso terapêutico , Inflamação/tratamento farmacológico , Pulmão/efeitos dos fármacos , Neutrófilos/patologia , Lesão Pulmonar Induzida por Ventilação Mecânica/tratamento farmacológico , Animais , Apoptose , Células Cultivadas , Modelos Animais de Doenças , Heme Oxigenase-1/metabolismo , Humanos , Pulmão/patologia , Masculino , Fator 2 Relacionado a NF-E2/metabolismo , NF-kappa B/metabolismo , Protoporfirinas/farmacologia , Ratos , Ratos Sprague-Dawley , Transdução de Sinais
9.
Int J Mol Med ; 42(6): 3083-3092, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30280199

RESUMO

Pulmonary fibrosis is an aggressive end­stage disease. Transforming growth factor­ß1 (TGF­ß1) mediates lung fibroblast activation and is essential for the progress of pulmonary fibrosis. BML­111, a lipoxinA4 (LXA4) receptor (ALX) agonist, has been reported to possess anti­ï¬brotic properties. The present study aimed to elucidate whether BML­111 inhibits TGF­ß1­induced mouse embryo lung fibroblast (NIH3T3 cell line) activation in vitro and bleomycin (BLM)­induced pulmonary fibrosis in vivo. In vitro experiments demonstrated that BML­111 treatment inhibits TGF­ß1­induced NIH3T3 cell viability and the expression of smooth muscle α actin (α­SMA), fibronectin and total collagen. Furthermore, this suppressive effect was associated with mothers against decapentaplegic homolog (Smad)2/3, extracellular signal­regulated kinase (ERK) and Akt phosphorylation interference. In vivo experiments revealed that BML­111 treatment markedly improved survival rate and ameliorated the destruction of lung tissue structure. It also reduced interleukin­1ß (IL­1ß), tumor necrosis factor­α (TNF­α) and TGF­ß1 expression in the BLM intratracheal mouse model. In addition, the expression ofα­SMA and extracellular matrix (ECM) deposition (total collagen, hydroxyproline and fibronectin) were also suppressed following BML­111 treatment. However, BOC­2, an antagonist of ALX, partially weakened the effects of BML­111. In conclusion, these results indicated that BML­111 inhibits TGF­ß1­induced fibroblasts activation and alleviates BLM­induced pulmonary fibrosis. Therefore, BML­111 may be used as a potential therapeutic agent for pulmonary fibrosis treatment.


Assuntos
Fibroblastos/metabolismo , Ácidos Heptanoicos/farmacologia , Fibrose Pulmonar/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , Animais , Linhagem Celular , Modelos Animais de Doenças , Fibroblastos/efeitos dos fármacos , Masculino , Camundongos , Células NIH 3T3 , Prognóstico , Fibrose Pulmonar/tratamento farmacológico , Fibrose Pulmonar/etiologia , Fibrose Pulmonar/patologia , Transdução de Sinais/efeitos dos fármacos , Proteínas Smad/metabolismo , Fator de Crescimento Transformador beta1/farmacologia , Resultado do Tratamento
10.
Chin Med J (Engl) ; 131(10): 1167-1173, 2018 May 20.
Artigo em Inglês | MEDLINE | ID: mdl-29553051

RESUMO

Background: Acute lung injury (ALI) is a severe disease with high mortality and poor prognosis. Protectin DX (PDX), a pro-resolving lipid mediator, exhibits protective effects in ALI. Our experiment aimed to explore the effects and related mechanisms of PDX in mice with ALI induced by lipopolysaccharide (LPS). Methods: BALB/c mice were randomly divided into five groups: sham, LPS, LPS plus 1 ng of PDX (LPS + PDX-1 ng), LPS plus 10 ng of PDX (LPS + PDX-10 ng), and LPS plus 100 ng of PDX (LPS + PDX-100 ng). Bronchoalveolar lavage fluids (BALFs) were collected after 24 h, and total cells, polymorphonuclear leukocytes, monocyte-macrophages, and lymphocytes in BALF were enumerated. The concentration of interleukin (IL)-1ß, IL-6, IL-10, tumor necrosis factor-alpha (TNF-α), macrophage inflammatory protein (MIP)-1α, and MIP-2 in BALF was determined, and histopathological changes of the lung were observed. The concentration of protein in BALF and lung wet/dry weight ratios were detected to evaluate pulmonary edema. After determining the optimal dose of PDX, neutrophil-platelet interactions in whole blood were evaluated by flow cytometry. Results: The highest dose of PDX (100 ng/mouse) failed to provide pulmonary protective effects, whereas lower doses of PDX (1 ng/mouse and 10 ng/mouse), especially 1 ng PDX, alleviated pulmonary histopathological changes, mitigated LPS-induced ALI and pulmonary edema, inhibited neutrophil infiltration, and reduced pro-inflammatory mediator (IL-1ß, IL-6, TNF-α, and MIP-1α) levels. Meanwhile, 1 ng PDX exhibited pro-resolving functions in ALI including upregulation of monocyte-macrophage numbers and anti-inflammatory mediator IL-10 levels. The flow cytometry results showed that PDX could inhibit neutrophil-platelet interactions in ALI. Conclusion: PDX exerts protective effects in LPS-induced ALI by mitigating pulmonary inflammation and abrogating neutrophil-platelet interactions.


Assuntos
Lesão Pulmonar Aguda/induzido quimicamente , Lesão Pulmonar Aguda/tratamento farmacológico , Ácidos Docosa-Hexaenoicos/uso terapêutico , Lipopolissacarídeos/toxicidade , Animais , Quimiocina CXCL2/metabolismo , Citometria de Fluxo , Interleucina-10/metabolismo , Interleucina-1beta/metabolismo , Interleucina-6/metabolismo , Pulmão/efeitos dos fármacos , Pulmão/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Fator de Necrose Tumoral alfa/metabolismo
11.
Cell Death Dis ; 9(2): 94, 2018 01 24.
Artigo em Inglês | MEDLINE | ID: mdl-29367697

RESUMO

Gestational trophoblastic disease (GTD) encompasses a range of trophoblast-derived disorders. The most common type of GTD is hydatidiform mole (HM). Some of HMs can further develop into malignant gestational trophoblastic neoplasia (GTN). Aberrant expression of microRNA (miRNA) is widely reported to be involved in the initiation and progression of cancers. MiRNA expression profile also has been proved to be the useful signature for diagnosis, staging, prognosis, and response to chemotherapy. Till now, the profile of miRNA in the progression of GTD has not been determined. In this study, a total of 34 GTN and 60 complete HMs (CHM) trophoblastic tissues were collected. By miRNA array screening and qRT-PCR validating, six miRNAs, including miR-370-3p, -371a-5p, -518a-3p, -519d-3p, -520a-3p, and -934, were identified to be differentially expressed in GTN vs. CHM. Functional analyses further proved that miR-371a-5p and miR-518a-3p promoted proliferation, migration, and invasion of choriocarcinoma cells. Moreover, we demonstrated that miR-371a-5p was negatively related to protein levels of its predictive target genes BCCIP, SOX2, and BNIP3L, while miR-518a-3p was negatively related to MST1 and EFNA4. For the first time, we proved that miR-371a-5p and miR-518a-3p directly targeted to 3'-UTR regions of BCCIP and MST1, respectively. Additionally, we found that miR-371a-5p and miR-518a-3p regulated diverse pathways related to tumorigenesis and metastasis in choriocarcinoma cells. The results presented here may offer new clues to the progression of GTD and may provide diagnostic biomarkers for GTN.


Assuntos
Progressão da Doença , Perfilação da Expressão Gênica , Doença Trofoblástica Gestacional/genética , Doença Trofoblástica Gestacional/patologia , MicroRNAs/genética , Sequência de Bases , Carcinogênese/genética , Carcinogênese/patologia , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células , Regulação para Baixo/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Mola Hidatiforme/genética , MicroRNAs/metabolismo , Invasividade Neoplásica , Metástase Neoplásica , Gravidez , Reprodutibilidade dos Testes , Fase S , Regulação para Cima/genética
12.
Int J Gynecol Cancer ; 27(2): 364-374, 2017 02.
Artigo em Inglês | MEDLINE | ID: mdl-27922982

RESUMO

OBJECTIVE: The aims of this study were to make clear whether miR-21 was dysregulated in hydatidiform mole (HM) tissues and choriocarcinoma (CCA) cells, to elucidate whether aberrant miR-21 expression would affect the function of CCA cells, and to find out whether there was a relationship between miR-21 and AKT, PDCD4, and PTEN in CCA cells. METHODS: Fresh and formalin-fixed, paraffin-embedded trophoblastic tissues (normal first trimester placentas and HMs) were retrieved from the biobank in the International Peace Maternity and Child Health Hospital, Shanghai Jiao Tong University. Choriocarcinoma JAR and JEG-3 cells were cultured. Expression of miR-21 in trophoblast cells and tissues was examined by quantitative real-time polymerase chain reaction. Location and distribution of miR-21 in trophoblast tissues were determinated by in situ hybridization and fluorescent in situ hybridization. The effect of miR-21 on JAR and JEG-3 cells was tested by miR-21 mimics and inhibitor transfection, followed by cell viability assay, flow cytometric analysis, and Transwell analysis. Interaction between miR-21 and its target genes in CCA cells was verified by quantitative real-time polymerase chain reaction, Western blot, and luciferase report system. RESULTS: We originally found miR-21 was markedly upregulated in HM tissues compared with normal first trimester placentas. The expression of miR-21 was exclusively confined in trophoblastic layers. Furthermore, we discovered miR-21 was significantly increased in JAR and JEG-3 cells compared with normal primary human trophoblastic cells. Moreover, we demonstrated miR-21 could promote proliferation, migration, and invasion of CCA cells. We furthermore proved miR-21 negatively regulated PDCD4 and PTEN in CCA cells and targeted to PDCD4 3'UTR directly. In addition, we confirmed that miR-21 could activate Akt pathway by phosphorylating Akt at Ser 473. CONCLUSIONS: Our results suggested miR-21 was responsible for aggressive phenotype of gestational trophoblastic disease and had the potential diagnostic and therapeutic values for gestational trophoblastic neoplasm.


Assuntos
Coriocarcinoma/genética , Coriocarcinoma/patologia , Mola Hidatiforme/genética , Mola Hidatiforme/patologia , MicroRNAs/biossíntese , Neoplasias Uterinas/genética , Neoplasias Uterinas/patologia , Proteínas Reguladoras de Apoptose/genética , Materiais Biomiméticos/farmacologia , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Coriocarcinoma/metabolismo , Feminino , Humanos , Mola Hidatiforme/metabolismo , Hibridização In Situ , Hibridização in Situ Fluorescente , MicroRNAs/antagonistas & inibidores , MicroRNAs/genética , Invasividade Neoplásica , PTEN Fosfo-Hidrolase/genética , Fosforilação , Gravidez , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas de Ligação a RNA/genética , Transfecção , Regulação para Cima , Neoplasias Uterinas/metabolismo
13.
Molecules ; 20(9): 16419-34, 2015 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-26378507

RESUMO

A series of 5H-thiazolo[3,2-a]pyrimidin-5-ones were synthesized by the cyclization reactions of S-alkylated derivatives in concentrated H2SO4. Upon treatment of S-alkylated derivatives at different temperatures, intramolecular cyclization to 7-(substituted phenylamino)-5H-thiazolo[3,2-a]pyrimidin-5-ones or sulfonation of cyclized products to sulfonic acid derivatives occurred. The structures of the target compounds were confirmed by IR, ¹H-NMR, (13)C-NMR and HRMS studies. The compounds were evaluated for their preliminary in vitro antibacterial activity against some Gram-positive and Gram-negative bacteria and screened for antitubercular activity against Mycobacterium tuberculosis by the broth dilution assay method. Some compounds showed good antibacterial and antitubercular activities.


Assuntos
Antibacterianos/síntese química , Antibacterianos/farmacologia , Antituberculosos/síntese química , Antituberculosos/farmacologia , Antibacterianos/química , Antituberculosos/química , Bactérias Gram-Negativas/efeitos dos fármacos , Espectroscopia de Ressonância Magnética , Espectrometria de Massas , Testes de Sensibilidade Microbiana , Mycobacterium tuberculosis/efeitos dos fármacos , Espectroscopia de Prótons por Ressonância Magnética , Espectrofotometria Infravermelho , Ácidos Sulfônicos/química
14.
Shock ; 44(4): 371-80, 2015 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-26196844

RESUMO

Acute lung injury (ALI) is characterized by lung inflammation and diffuse infiltration of neutrophils. Neutrophil apoptosis is recognized as an important control point in the resolution of inflammation. Maresin 1 (MaR1) is a new docosahexaenoic acid-derived proresolving agent that promotes the resolution of inflammation. However, its function in neutrophil apoptosis is unknown. In this study, isolated human neutrophils were incubated with MaR1, the pan-caspase inhibitor z-VAD-fmk, and lipopolysaccharide (LPS) to determine the mechanism of neutrophil apoptosis. Acute lung injury was induced by intratracheal instillation of LPS. In addition, mice were treated with MaR1 intravenously at the peak of inflammation and administered z-VAD-fmk intraperitoneally. We found that culture of isolated human neutrophils with LPS dramatically delayed neutrophil apoptosis through the phosphorylation of AKT, ERK, and p38 to upregulate the expression of the antiapoptotic proteins Mcl-1 and Bcl-2, which was blocked by pretreatment with MaR1 in vitro. In mice, MaR1 accelerated the resolution of inflammation in LPS-induced ALI through attenuation of neutrophil accumulation, pathohistological changes, and pulmonary edema. Maresin 1 promoted resolution of inflammation by accelerating caspase-dependent neutrophil apoptosis. Moreover, MaR1 also reduced the LPS-induced production of proinflammatory cytokines and upregulated the production of the anti-inflammatory cytokine interleukin-10. In contrast, treatment with z-VAD-fmk inhibited the proapoptotic action of MaR1 and attenuated the protective effects of MaR1 in LPS-induced ALI. Taken together, MaR1 promotes the resolution of LPS-induced ALI by overcoming LPS-mediated suppression of neutrophil apoptosis.


Assuntos
Lesão Pulmonar Aguda/tratamento farmacológico , Anti-Inflamatórios não Esteroides/uso terapêutico , Ácidos Docosa-Hexaenoicos/uso terapêutico , Neutrófilos/efeitos dos fármacos , Lesão Pulmonar Aguda/patologia , Clorometilcetonas de Aminoácidos/farmacologia , Animais , Anti-Inflamatórios não Esteroides/administração & dosagem , Anti-Inflamatórios não Esteroides/antagonistas & inibidores , Anti-Inflamatórios não Esteroides/farmacologia , Apoptose/efeitos dos fármacos , Líquido da Lavagem Broncoalveolar/citologia , Inibidores de Caspase/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Ácidos Docosa-Hexaenoicos/administração & dosagem , Ácidos Docosa-Hexaenoicos/antagonistas & inibidores , Ácidos Docosa-Hexaenoicos/farmacologia , Relação Dose-Resposta a Droga , Avaliação Pré-Clínica de Medicamentos/métodos , Humanos , Lipopolissacarídeos/administração & dosagem , Lipopolissacarídeos/antagonistas & inibidores , Masculino , Camundongos Endogâmicos BALB C , Neutrófilos/patologia , Transdução de Sinais/efeitos dos fármacos
15.
Gene ; 569(1): 60-5, 2015 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-25979673

RESUMO

In this study, we aimed to assess the neuroprotective effect of sevoflurane preconditioning in a cerebral focal ischemia-reperfusion rat model. Sixty Sprague Dawley rats were divided into six groups: sham operated group, cerebral focal ischemia-reperfusion (CIR) group, CIR+sevoflurane preconditioning (SP) (2%) group, CIR+sevoflurane preconditioning (2.5%) group, CIR+sevoflurane preconditioning (3%) group, and CIR+sevoflurane preconditioning (3.5%) group. All subjects were euthanized 2days post-surgery and their hippocampus tissues were removed. Tissue malondialdehyde (MDA), superoxide dismutase (SOD), glutathione (GSH) and glutathione peroxidase (GSH-Px) levels were measured and hippocampus tissue samples were examined histopathologically. Results showed that significant difference in antioxidant, immunity indexes, and apoptosis-related protein expression was detected in hippocampus tissue between sham-operated control and CIR groups. Sevoflurane preconditioning significantly dose-dependently reduced MDA, IL-1ß, IL-6, IL-10 and TNF-α levels and enhanced antioxidant enzyme activities in hippocampus tissue of CIR+SP groups compared to CIR group. In addition, sevoflurane preconditioning significantly dose-dependently upregulated PI3K, p-Akt and Bcl-2 levels and downregulated caspase-3 and Bax levels in hippocampus tissue of CIR+SP groups compared to CIR group. It can be concluded that sevoflurane preconditioning demonstrates a strong and ameliorative effect on cerebral I/R damage in rats. The neuroprotective mechanisms of sevoflurane preconditioning are associated with its properties of anti-apoptosis and anti-oxidation as well as regulation of PI3K and p-Akt signal activation.


Assuntos
Isquemia Encefálica/tratamento farmacológico , Éteres Metílicos/administração & dosagem , Traumatismo por Reperfusão/tratamento farmacológico , Animais , Antioxidantes/metabolismo , Apoptose/efeitos dos fármacos , Isquemia Encefálica/patologia , Regulação da Expressão Gênica/efeitos dos fármacos , Proteína Oncogênica v-akt/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Ratos , Traumatismo por Reperfusão/patologia , Sevoflurano , Transdução de Sinais/efeitos dos fármacos
16.
Dalton Trans ; 44(13): 6052-61, 2015 Apr 07.
Artigo em Inglês | MEDLINE | ID: mdl-25728056

RESUMO

This work reports the assembly, topological structure, supramolecular isomerism and luminescence of three solvent-controlled families of coordination compounds, [Zn(bpz)2(H2O)3·2Hpta] (1), [Zn(bpz)(pta)]n (2), [Zn(bpz)(tpa)(H2O)]n (3), [Zn(bpz)(tpa)·4H2O]n (4), and [Zn(bpz)(npa)·H2O]n (5 and 6) (bpz = 3,3',5,5'-tetramethyl-4,4'-bipyrazole, H2pta = phthalic acid, H2tpa = terephthalic acid, H2npa = 4-nitrophthalic acid). The six transition metal compounds reported in this study were definitely characterized by X-ray crystallography to reveal how networks with different topologies are constructed around the same four-connected metal centers. Compound 1 is a 0D discrete molecule, in which Zn(II) is in a trigonal bipyramidal coordination geometry, whereas the guest Hpta(-) as counteranion is hydrogen-bonded with the [Zn(bpz)2(H2O)3]. In compound 2, the Zn(II) center is linked by two bpz and two pta; thus, a 4-connected dia network with the point symbol {6(6)} is formed. In 3, the Zn(II) center is six-coordinated, but in fact it is also a 4-connected node in the whole network due to the terminal aqua ligand and bidentate chelating carboxylate group thus, the resultant network has a 4-connected cds topology with the point symbol {6(5)·8}. Compound 4 exhibits a chiral two-fold interpenetrated 4-connected qtz network with the point symbol {6(4)·8(2)}. Compounds 5 and 6 are a pair of genuine supramolecular isomers with identical 4-connected dia topology. The three families of compounds, namely, 1/2, 3/4, and 5/6, are structurally controlled by the solvent systems H2O/CH3OH-H2O, H2O/DMF-CH3OH, and CH3OH-H2O/CH3CN-H2O, respectively. Except for the discrete molecule 1, the other five compounds have the same 4-connected coordination networks, but with different topologies ranging from dia (2, 5, 6), cds (3) to qtz (4), suggesting the important influences that the linkage orientations of the ligand and different geometries of the 4-connected node exert in self-assembly. Interestingly, discrete 1 can be irreversibly transformed from a 0D discrete molecule to an infinite 3D structure (2) by heating it in CH3OH-H2O solvent, indicating a solvent-induced structural transformation. In addition, results about thermal stabilities and photoluminescence spectra are also discussed in detail.

17.
Gene ; 562(1): 76-82, 2015 May 10.
Artigo em Inglês | MEDLINE | ID: mdl-25701598

RESUMO

TGF-ß/Smad2/3 signal pathway is regarded as a central regulator in various tumors, but its roles in brain cancer therapy remain unknown. In this study, we identify that the TGF-ß/Smad2/3 signal pathway is activated in human brain glioma cells; inhibitor (SB203580) and siRNA against Smad2/3 quickly inhibited the phosphorylation of Smad2 and 3, expression of its major downstream gene, Ki-67, arrested cells in the G2/M phase and induced apoptosis of cells. The findings suggest that TGF-ß/Smad2/3 pathway plays a key role in U251 cell growth and metastasis, which suggests its potential role in the molecular therapy of brain cancer.


Assuntos
Regulação Neoplásica da Expressão Gênica , Transdução de Sinais/genética , Proteína Smad2/genética , Proteína Smad3/genética , Fator de Crescimento Transformador beta/genética , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Humanos , Imidazóis/farmacologia , Antígeno Ki-67/genética , Antígeno Ki-67/metabolismo , Neuroglia/efeitos dos fármacos , Neuroglia/metabolismo , Neuroglia/patologia , Fosforilação/efeitos dos fármacos , Piridinas/farmacologia , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Proteína Smad2/antagonistas & inibidores , Proteína Smad2/metabolismo , Proteína Smad3/antagonistas & inibidores , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta/metabolismo
18.
Br J Pharmacol ; 171(14): 3539-50, 2014 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-24697684

RESUMO

BACKGROUND AND PURPOSE: Acute lung injury (ALI) is a severe illness with a high rate of mortality. Maresin 1 (MaR1) was recently reported to regulate inflammatory responses. We used a LPS-induced ALI model to determine whether MaR1 can mitigate lung injury. EXPERIMENTAL APPROACH: Male BALB/c mice were injected, intratracheally, with either LPS (3 mg·kg(-1) ) or normal saline (1.5 mL·kg(-1) ). After this, normal saline, a low dose of MaR1 (0.1 ng per mouse) or a high dose of MaR1 (1 ng per mouse) was given i.v. Lung injury was evaluated by detecting arterial blood gas, pathohistological examination, pulmonary oedema, inflammatory cell infiltration, inflammatory cytokines in the bronchoalveolar lavage fluid and neutrophil-platelet interactions. KEY RESULTS: The high dose of MaR1 significantly inhibited LPS-induced ALI by restoring oxygenation, attenuating pulmonary oedema and mitigating pathohistological changes. A combination of elisa and immunohistochemistry showed that high-dose MaR1 attenuated LPS-induced increases in pro-inflammatory cytokines (TNF-α, IL-1ß and IL-6), chemokines [keratinocyte chemokine, monocyte chemoattractant protein-5, macrophage inflammatory protein (MIP)-1α and MIP-1γ], pulmonary myeloperoxidase activity and neutrophil infiltration in the lung tissues. Consistent with these observations, flow cytometry and Western blotting indicated that MaR1 down-regulated LPS-induced neutrophil adhesions and suppressed the expression of intercellular adhesion molecule (ICAM)-1, P-selection and CD24. CONCLUSIONS AND IMPLICATIONS: High-dose MaR1 mitigated LPS-induced lung injury in mice by inhibiting neutrophil adhesions and decreasing the levels of pro-inflammatory cytokines.


Assuntos
Lesão Pulmonar Aguda/tratamento farmacológico , Ácidos Docosa-Hexaenoicos/farmacologia , Lipopolissacarídeos/antagonistas & inibidores , Lesão Pulmonar Aguda/induzido quimicamente , Lesão Pulmonar Aguda/patologia , Administração por Inalação , Animais , Adesão Celular/efeitos dos fármacos , Ácidos Docosa-Hexaenoicos/administração & dosagem , Mediadores da Inflamação/metabolismo , Lipopolissacarídeos/administração & dosagem , Lipopolissacarídeos/toxicidade , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Neutrófilos/efeitos dos fármacos , Neutrófilos/patologia
19.
Gene ; 542(1): 46-51, 2014 May 25.
Artigo em Inglês | MEDLINE | ID: mdl-24630969

RESUMO

Tea polyphenols (TP) was investigated in rats for its protective effect on renal ischemia/reperfusion injury (RIRI). Rats were randomized into groups as follows: (I) sham group (n=10); (II) RIRI group (n=10); (III) RIRI+TP (100mg/kg) group (n=5); (IV) RIRI+TP (200mg/kg) group (n=5); (V) RIRI+TP+ Astragalus mongholicus aqueous extract (AMAE) (300 mg/kg+100mg/kg) group (n=5). For the IRI+TP groups, rats were orally given with tea polyphenols (100, 200 and 300 mg/kg body weight) once daily 10 days before induction of ischemia, followed by renal IRI. For the sham group and RIRI group, rats were orally given with equal volume of saline once daily 10 days before induction of ischemia, followed by renal IRI. Results showed that tea polyphenol pretreatment significantly suppressed ROS level and MDA release. On the other hand, in rats subjected to ischemia-reperfusion, the activities of endogenous antioxidant enzymes including superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GR) and glutathione peroxidase (GSH-Px) showed recovery, whereas the levels of urea nitrogen and serum creatinine were reduced by administration of tea polyphenols orally for 10 days prior to ischemia-reperfusion. Moreover, tea polyphenol pretreatment significantly decreased TLR4 and NF-κB p65 protein expression levels in RIRI rats. At the same time, tea polyphenol pretreatment attenuated the increased level of serum IL-1ß, IL-6, ICAM-1 and TNF-α, and enhanced IL-10 production in RIRI rats. Furthermore, tea polyphenol pretreatment significantly decreased renal epithelial tubular cell apoptosis induced by renal ischemia/reperfusion, alleviating renal ischemia/reperfusion injury. These results cumulatively indicate that tea polyphenol pretreatment could suppress the TLR4/NF-κB p65 signaling pathway, protecting renal tubular epithelial cells against ischemia/reperfusion-induced apoptosis, which implies that antioxidants may be a potential and effective agent for prevention of the ischemic/reperfusion injury through the suppression extrinsic apoptotic signal pathway induced by TLR4/NF-κB p65 signal pathway. Moreover, supplement of AMAE can increased renal protection effect of TP.


Assuntos
Apoptose/efeitos dos fármacos , Camellia sinensis/química , Rim/irrigação sanguínea , Extratos Vegetais/administração & dosagem , Polifenóis/administração & dosagem , Traumatismo por Reperfusão/tratamento farmacológico , Animais , Antioxidantes/uso terapêutico , Nitrogênio da Ureia Sanguínea , Catalase/metabolismo , Creatinina/sangue , Glutationa Peroxidase/metabolismo , Glutationa Redutase/metabolismo , Molécula 1 de Adesão Intercelular/sangue , Interleucina-10/sangue , Interleucina-1beta/sangue , Interleucina-6/sangue , Rim/efeitos dos fármacos , Túbulos Renais/irrigação sanguínea , Túbulos Renais/efeitos dos fármacos , Ativação de Macrófagos/imunologia , Macrófagos/imunologia , Masculino , Ativação de Neutrófilo/imunologia , Neutrófilos/imunologia , Estresse Oxidativo/efeitos dos fármacos , Ratos , Ratos Wistar , Espécies Reativas de Oxigênio/metabolismo , Traumatismo por Reperfusão/metabolismo , Traumatismo por Reperfusão/patologia , Transdução de Sinais/efeitos dos fármacos , Superóxido Dismutase/metabolismo , Receptor 4 Toll-Like/antagonistas & inibidores , Receptor 4 Toll-Like/biossíntese , Fator de Transcrição RelA/antagonistas & inibidores , Fator de Transcrição RelA/biossíntese , Fator de Necrose Tumoral alfa/sangue
20.
PLoS One ; 9(1): e85767, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24465692

RESUMO

The highly pathogenic porcine reproductive and respiratory syndrome virus (HP-PRRSV) is still a serious threat to the swine industry. However, the pathogenic mechanism of HP-PRRSV remains unclear. We infected host porcine alveolar macrophages (PAMs) with the virulent HuN4 strain and the attenuated HuN4-F112 strain and then utilized fluorescent two-dimensional difference gel electrophoresis (2D-DIGE) to screen for intracellular proteins that were differentially expressed in host cells infected with the two strains. There were 153 proteins with significant different expression (P<0.01) observed, 42 of which were subjected to mass spectrometry, and 24 proteins were identified. PAM cells infected with the virulent strain showed upregulated expression of pyruvate kinase M2 (PKM2), heat shock protein beta-1 (HSPB1), and proteasome subunit alpha type 6 (PSMA6), which were downregulated in cells infected with the attenuated strain. The upregulation of PKM2 provides sufficient energy for viral replication, and the upregulation of HSPB1 inhibits host cell apoptosis and therefore facilitates mass replication of the virulent strain, while the upregulation of PSMA6 facilitates the evasion of immune surveillance by the virus. Studying on those molecules mentioned above may be able to help us to understand some unrevealed details of HP-PRRSV infection, and then help us to decrease its threat to the swine industry in the future.


Assuntos
Macrófagos Alveolares/metabolismo , Macrófagos Alveolares/virologia , Síndrome Respiratória e Reprodutiva Suína/metabolismo , Síndrome Respiratória e Reprodutiva Suína/virologia , Vírus da Síndrome Respiratória e Reprodutiva Suína/patogenicidade , Proteômica , Animais , Animais Recém-Nascidos , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Ontologia Genética , Anotação de Sequência Molecular , Mapas de Interação de Proteínas , Proteoma/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Reprodutibilidade dos Testes , Software , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Estatística como Assunto , Suínos , Transcrição Genética , Eletroforese em Gel Diferencial Bidimensional , Virulência
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