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1.
Lab Invest ; 99(12): 1770-1783, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31278346

RESUMO

Early pulmonary fibrosis is the leading cause of poor prognosis in patients with acute respiratory distress syndrome (ARDS). However, whether the renin-angiotensin system (RAS) can serve as a therapeutic target is unknown. In this study, an animal model of early pulmonary fibrosis was established via the LPS three-hit regimen. Afterwards, the animals were treated with intraperitoneal injections of Ang-(1-7), AVE0991, or A779 once per day for 20 days. The plasma and BALF AngII levels of the animals were increased, while there were no significant changes in Ang-(1-7) levels in lung tissue after LPS treatment. Furthermore, the AT1R protein levels were significantly increased and the Mas levels were significantly decreased on days 14 and 21. Administration of Ang-(1-7) downregulated LPS-induced AT1R mRNA expression, which was upregulated by A779. The expression of Mas mRNA responded in the opposite direction relative to AT1R. Moreover, LPS caused decreased levels of Mas and E-cadherin and increased AT1R, Vimentin, and Src phosphorylation levels. Ang-(1-7) or AVE0991 blocked these effects but was counteracted by A779 treatment. Our findings suggested that AngII and AT1R levels exhibit opposite dynamic trends during LPS-induced early pulmonary fibrosis, as do Ang-(1-7) and Mas. Ang-(1-7) exerts protective effects against early pulmonary fibrosis, mainly by regulating the balance between AngII and AT1R and between Ang-(1-7) and Mas and by inhibiting Src kinase activation.

2.
J Cell Physiol ; 234(8): 12865-12875, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-30537127

RESUMO

The balance between Ang II/AT1R and Ang-(1-7)/Mas plays a pivotal role in the development of lipopolysaccharides (LPS)-induced acute respiratory distress syndrome. However, the mechanisms underlying the balancing process still remain unclear. Here we investigated the roles of nuclear factor (NF)-κB and p53 in regulating AT1R and Mas expression. The results demonstrated that Ang II pretreatment resulted in downregulation of Mas and upregulation of AT1R, phosphorylated p65, and apoptosis in LPS-treated Human pulmonary microvascular endothelial cells (HPMVECs), but had no effect on p53 expression. Lentiviral vector-mediated P65 knockdown, but not a P53 knockdown, reversed all these effects of Ang II. On the other hand, Ang-(1-7) pretreatment lead to an increased in Mas expression and a decrease in AT1R, p53, and phosphorylated p65 expressions with suppressed apoptosis in LPS-treated cells. P65 knockdown promoted the protein expression of both AT1R and Mas while inhibiting p53 expression. P53 knockdown, but not a p65 knockdown, reversed all these effects of Ang-(1-7). Interestingly, p65 overexpression upregulated p53 and AT1R but downregulated Mas. P53 knockdown activated p65. These results suggest that there is a two-way feedback regulation between AT1R and Mas receptor via the NF-kB p65/P53 pathway, which may play a key role in LPS-induced HPMVECs apoptosis.

3.
Lab Invest ; 98(3): 339-359, 2018 03.
Artigo em Inglês | MEDLINE | ID: mdl-29200203

RESUMO

Pulmonary fibrosis triggered during the early stage of acute respiratory distress syndrome (ARDS) contributes to poor prognosis in patients. However, whether microRNAs (miRNAs) can serve as therapeutic targets for early pulmonary fibrosis during ARDS is still largely unknown. In this study, we evaluated the effects and mechanisms of miR-200s and its targets ZEB1/2 in lung tissue. An early pulmonary fibrosis mouse model caused by ARDS was established via a lipopolysaccharide (LPS) three-hit regimen. Lentiviral packaged miR-200b/c cDNA or ZEB1/2 shRNA was intratracheally administered into the lungs of C57BL/6 mice 1 day before an LPS injection was administered. In vitro, following a 30-min pretreatment with miR-200b/c or SB203580/SIS3, RLE-6TN cells were stimulated by LPS or LPS + transforming growth factor-ß (TGF-ß) for 24 h. miR-200b/c and E-cadherin protein expression declined, whereas ZEB1/2 mRNA and protein and vimentin and α-smooth muscle actin (α-SMA) protein levels gradually increased during the development of pulmonary fibrosis. Furthermore, both the overexpression of miR-200b/c and the silencing of ZEB1/2 significantly alleviated pulmonary inflammation and fibrosis, reduced vimentin and α-SMA expression, and increased E-cadherin protein levels. In RLE-6TN cells, LPS combined with TGF-ß exerts synergistic effects of increasing vimentin and α-SMA protein levels, increasing p38 and smad3 phosphorylation and reducing E-cadherin protein levels, which were reversed by pretreatment with miR-200b/c or SB203580/SIS3. Our findings demonstrate that miR-200b/c was downregulated, whereas ZEB1/2 was upregulated in the development of LPS-induced early pulmonary fibrosis. miR-200b/c exerts a protective effect by targeting ZEB1/2, which may be associated with the inhibition of p38 MAPK and TGF-ß /smad3 signaling pathways.


Assuntos
Sistema de Sinalização das MAP Quinases , MicroRNAs/metabolismo , Fibrose Pulmonar/metabolismo , Homeobox 2 de Ligação a E-box com Dedos de Zinco/metabolismo , Homeobox 1 de Ligação a E-box em Dedo de Zinco/metabolismo , Actinas/metabolismo , Animais , Líquido da Lavagem Broncoalveolar/química , Caderinas/metabolismo , Linhagem Celular , Modelos Animais de Doenças , Transição Epitelial-Mesenquimal , Lipopolissacarídeos , Pulmão/patologia , Camundongos Endogâmicos C57BL , Fibrose Pulmonar/etiologia , Fibrose Pulmonar/patologia , RNA Interferente Pequeno , Distribuição Aleatória , Ratos , Síndrome do Desconforto Respiratório do Adulto/complicações , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Vimentina/metabolismo
4.
J Surg Res ; 219: 325-333, 2017 11.
Artigo em Inglês | MEDLINE | ID: mdl-29078900

RESUMO

BACKGROUND: Potential of liver regeneration after living-donor liver transplantation is closely associated with the recipient's prognosis, whereas exogenous gene might regulate the liver regeneration progress. NM23 is a multifunctional gene, which inhibits tumor metastasis and regulates cell proliferation, differentiation, development, and apoptosis; however, there is little research about NM23 in promoting liver cell proliferation. METHODS: To investigate the effect of NM23-E2 on the liver cell proliferation, the NM23-E2 overexpression vector or negative control vector was transfected into BRL-3A cells and donor liver, respectively. NM23-E2, Cyclin D1, and PCNA expression levels in BRL-3A cells and liver tissues were detected by quantitative real-time polymerase chain reaction and Western blot analysis. Cell Counting Kit-8 was used to detect cell proliferation and flow cytometry for investigating cell cycle. The liver regeneration rate was determined by calculating (regenerated-liver weight of recipient - liver weight of donor/liver weight of donor) × 100%. RESULTS: NM23-E2 overexpression increased the NM23-E2, Cyclin D1, and PCNA levels significantly in BRL-3A cells and liver tissues (P < 0.05). The number of S phase cells was more than that of negative control group, and cell proliferation rate was higher than that of the control group in BRL-3A cells markedly (P < 0.05). Moreover, the liver regeneration rate in the NM23-E2 overexpression group was also higher than that in negative control group on postoperative day 1, day 3, day 5, and day 7. CONCLUSIONS: Overexpression of NM23-E2 can increase Cyclin D1 and PCNA expression, shorten cell cycle, and thereby promoting the proliferation of liver cells and accelerating the regeneration of liver after 40% decreased-size rat liver transplantation.


Assuntos
Terapia Genética , Regeneração Hepática , Transplante de Fígado , Nucleosídeo NM23 Difosfato Quinases/genética , Animais , Ciclo Celular , Linhagem Celular , Proliferação de Células , Ciclina D1/metabolismo , Lentivirus , Fígado/metabolismo , Masculino , Nucleosídeo NM23 Difosfato Quinases/metabolismo , Antígeno Nuclear de Célula em Proliferação/metabolismo , Ratos Sprague-Dawley
5.
Front Microbiol ; 8: 965, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28611760

RESUMO

Human breast milk is widely recognized as the best source of nutrients for healthy growth and development of infants; it contains a diverse microbiota. Here, we characterized the diversity of the microbiota in the breast milk of East Asian women and assessed whether delivery mode influenced the microbiota in the milk of healthy breast-feeding mothers. We profiled the microbiota in breast milk samples collected from 133 healthy mothers in Taiwan and in six regions of mainland China (Central, East, North, Northeast, South, and Southwest China) by using 16S rRNA pyrosequencing. Lactation stage (months postpartum when the milk sample was collected) and maternal body mass index did not influence the breast milk microbiota. Bacterial composition at the family level differed significantly among samples from the seven geographical regions. The five most predominant bacterial families were Streptococcaceae (mean relative abundance: 24.4%), Pseudomonadaceae (14.0%), Staphylococcaceae (12.2%), Lactobacillaceae (6.2%), and Oxalobacteraceae (4.8%). The microbial profiles were classified into three clusters, driven by Staphylococcaceae (abundance in Cluster 1: 42.1%), Streptococcaceae (Cluster 2: 48.5%), or Pseudomonadaceae (Cluster 3: 26.5%). Microbial network analysis at the genus level revealed that the abundances of the Gram-positive Staphylococcus, Streptococcus, and Rothia were negatively correlated with those of the Gram-negative Acinetobacter, Bacteroides, Halomonas, Herbaspirillum, and Pseudomonas. Milk from mothers who had undergone Caesarian section (C-section group) had a significantly higher abundance of Lactobacillus (P < 0.05) and a higher number of unique unclassified operational taxonomic units (OTUs) (P < 0.001) than that from mothers who had undergone vaginal delivery (vaginal group). These findings revealed that (i) geographic differences in the microbial profiles were found in breast milk from mothers living in Taiwan and mainland China, (ii) the predominant bacterial families Streptococcaceae, Staphylococcaceae, and Pseudomonadaceae were key components for forming three respective clusters, and (iii) a significantly greater number of unique OTUs was found in the breast milk from mothers who had undergone C-section than from those who had delivered vaginally.

6.
Int J Clin Pharmacol Ther ; 54(12): 987-991, 2016 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-27668696

RESUMO

PURPOSE: To evaluate the corrective effect of norepinephrine on hypotension induced by dexmedetomidine through monitoring sedation status, hemodynamics as well as oxygen metabolism. METHODS: 100 patients administered standard-dose dexmedetomidine therapy with RASS between -2 and 0 in the intensive care unit (ICU) were included in the study. According to the application of norepinephrine to correct hypotension after dexmedetomidine therapy, the patients were divided into two groups: group A and group B. Group A (dexmedetomidine + norepinephrine group): those who had a systolic arterial pressure < 90 mmHg, a mean arterial pressure < 70 mmHg, or a decline in systolic arterial pressure > 40 mmHg or more than 30% of its base value after dexmedetomidine therapy and then received additional norepinephrine intravenously in order to maintain the arterial pressure at its base value. Group B (dexmedetomidine group): patients received the equivalent dose of dexmedetomidine to maintain the pressure at normal value without extra vasoconstrictor substance. Sedation (CPOT and RASS) and hemodynamic and oxygen metabolism indexes (heart rate, mean arterial pressure, respiratory rate, arterial oxygen saturation, central venous pressure, venous oxygen saturation, arteriovenous carbon dioxide difference, blood lactate level, blood lactate clearance rate, and average hourly urine output) were evaluated in the two groups at baseline, 6th hour, 12th hour, and 24th hour after the administration of intravenous dexmedetomidine. RESULTS: 39 cases were enrolled in group A and 61 cases in group B. Patients of both groups received adequate analgesia and sedation, and there was no significant statistical difference in analgesia and sedation at any point (both p > 0.05). Basal hemodynamic indexes and oxygen metabolism indexes also had no significant statistical difference (both p > 0.05). Central venous pressure (CVP) of group A was significantly higher than that of group B at the 6th hour and 12th hour after administration of intravenous dexmedetomidine (p = 0.005), and the heart rate (HR) of group A was markedly higher than that of group B at the 24th hour after dexmedetomidine therapy (p = 0.017), while the other indexes had no significant difference at any point (both p > 0.05). CONCLUSION: Dexmedetomidine plays an important role in ICU patients due to its pharmacological ability of sedation and analgesia. In our study, dexmedetomidine was successfully applied to ensure goal-directed sedation therapy (GDST). Norepinephrine can correct hypotension and bradycardia induced by intravenous dexmedetomidine. According to the hemodynamic indexes and oxygen metabolism indexes, the application of dexmedetomidine or the combination of dexmedetomidine with norepinephrine are both safe and appropriate to maintain the sedation status and hemodynamic situation in ICU patients.
.


Assuntos
Dexmedetomidina/efeitos adversos , Hipotensão/tratamento farmacológico , Norepinefrina/uso terapêutico , Idoso , Estado Terminal , Feminino , Hemodinâmica/efeitos dos fármacos , Humanos , Hipotensão/induzido quimicamente , Masculino , Pessoa de Meia-Idade
7.
Sci Rep ; 6: 27911, 2016 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-27302421

RESUMO

Acute respiratory distress syndrome (ARDS) caused by severe sepsis remains a major challenge in intensive care medicine. ACE2 has been shown to protect against lung injury. However, the mechanisms of its protective effects on ARDS are largely unknown. Here, we report that ACE2 prevents LPS-induced ARDS by inhibiting MAPKs and NF-κB signaling pathway. Lentiviral packaged Ace2 cDNA or Ace2 shRNA was intratracheally administrated into the lungs of male SD rats. Two weeks after gene transfer, animals received LPS (7.5 mg/Kg) injection alone or in combination with Mas receptor antagonist A779 (10 µg/Kg) or ACE2 inhibitor MLN-4760 (1 mg/Kg) pretreatment. LPS-induced lung injury and inflammatory response were significantly prevented by ACE2 overexpression and deteriorated by Ace2 shRNA. A779 or MLN-4760 pretreatment abolished the protective effects of ACE2. Moreover, overexpression of ACE2 significantly reduced the Ang II/Ang-(1-7) ratio in BALF and up-regulated Mas mRNA expression in lung, which was reversed by A779. Importantly, the blockade of ACE2 on LPS-induced phosphorylation of ERK1/2, p38 and p50/p65 was also abolished by A779. Whereas, only the ERK1/2 inhibitor significantly attenuated lung injury in ACE2 overexpressing rats pretreated with A779. Our observation suggests that AEC2 attenuates LPS-induced ARDS via the Ang-(1-7)/Mas pathway by inhibiting ERK/NF-κB activation.


Assuntos
Lesão Pulmonar Aguda/metabolismo , Lipopolissacarídeos/toxicidade , NF-kappa B/metabolismo , Peptidil Dipeptidase A/metabolismo , Lesão Pulmonar Aguda/induzido quimicamente , Lesão Pulmonar Aguda/patologia , Angiotensina II/metabolismo , Animais , Regulação Enzimológica da Expressão Gênica , Técnicas de Silenciamento de Genes , Imidazóis/farmacologia , Lentivirus/genética , Leucina/análogos & derivados , Leucina/farmacologia , Pulmão/efeitos dos fármacos , Pulmão/metabolismo , Pulmão/patologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/fisiologia , Masculino , Peptidil Dipeptidase A/genética , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Ratos Sprague-Dawley , Receptores Acoplados a Proteínas-G/genética , Receptores Acoplados a Proteínas-G/metabolismo , Transdução de Sinais
8.
Biomed Rep ; 4(5): 523-527, 2016 May.
Artigo em Inglês | MEDLINE | ID: mdl-27123242

RESUMO

Acute lung injury (ALI) and the more severe acute respiratory distress syndrome (ARDS) are common and complex inflammatory lung diseases. MicroRNAs (miRNAs), a type of non-coding RNA molecule that regulate gene expression at the post-transcriptional level, have emerged as a novel class of gene regulators, which have critical roles in a wide range of human disorders and diseases, including ALI. Certain types of miRNAs are abnormally expressed in response to lung injury. miRNAs can regulate inflammation pathways by targeting specific molecules and modulate immune response in the process of lung injury and repair. The regulation of miRNA can relieve injury response and promote the recovery of ALI/ARDS. Therefore, miRNAs may serve as novel therapeutic targets in ALI/ARDS.

9.
Apoptosis ; 21(1): 69-84, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26456506

RESUMO

Pulmonary arterial hypertension (PAH) is a life-threatening disorder that ultimately causes heart failure. While the underlying causes of this condition are not well understood, previous studies suggest that the anti-apoptotic nature of pulmonary microvascular endothelial cells (PMVECs) in hypoxic environments contributes to PAH pathogenesis. In this study, we focus on the contribution of Bcl-2 and hypoxia response element (HRE) to apoptosis-resistant endothelial cells and investigate the mechanism. PMVECs obtained from either normal rats or apoptosis-resistant PMVECs obtained from PAH rats were transduced with recombinant lentiviral vectors carrying either Bcl-2-shRNA or HRE combined Bcl-2-shRNA, and then cultured these cells for 24 h under hypoxic (5% O2) or normoxic (21% O2) conditions. In normal PMVECs, Bcl-2-shRNA or HRE combined with Bcl-2-shRNA transduction successfully decreased Bcl-2 expression, while increasing apoptosis as well as caspase-3 and P53 expression in a normoxic environment. In a hypoxic environment, the effects of Bcl-2-shRNA treatment on cell apoptosis, and on Bcl-2, caspase-3, P53 expression were significantly suppressed. Conversely, HRE activation combined with Bcl-2-shRNA transduction markedly enhanced cell apoptosis and upregulated caspase-3 and P53 expression, while decreasing Bcl-2 expression. Furthermore, in apoptosis-resistant PMVECs, HRE-mediated Bcl-2 silencing effectively enhanced cell apoptosis and caspase-3 activity. The apoptosis rate was significantly depressed when Lv-HRE-Bcl-2-shRNA was combined with Lv-P53-shRNA or Lv-caspase3-shRNA transduction in a hypoxic environment. These results suggest that HRE-mediated Bcl-2 inhibition can effectively attenuate hypoxia-induced apoptosis resistance in PMVECs by downregulating Bcl-2 expression and upregulating caspase-3 and P53 expression. This study therefore reveals critical insight into potential therapeutic targets for treating PAH.


Assuntos
Apoptose/genética , Células Endoteliais/metabolismo , Hipertensão/genética , Proteínas Proto-Oncogênicas c-bcl-2/genética , Elementos de Resposta , Animais , Caspase 3/genética , Caspase 3/metabolismo , Hipóxia Celular , Células Endoteliais/patologia , Regulação da Expressão Gênica , Vetores Genéticos/química , Vetores Genéticos/metabolismo , Hipertensão/induzido quimicamente , Hipertensão/metabolismo , Hipertensão/patologia , Lentivirus/genética , Lentivirus/metabolismo , Pulmão/irrigação sanguínea , Pulmão/patologia , Masculino , Monocrotalina , Cultura Primária de Células , Proteínas Proto-Oncogênicas c-bcl-2/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Artéria Pulmonar/metabolismo , Artéria Pulmonar/patologia , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Ratos , Ratos Sprague-Dawley , Transdução de Sinais , Transdução Genética , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo
10.
J Nanosci Nanotechnol ; 15(2): 1821-30, 2015 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-26353738

RESUMO

SWNHs induces accumulate with the levels of cytotoxic effects on different cell types and organs in animal models; it in a wide range may used for biomedical imaging. Conjunctival melanoma is a rare but potentially fatal ocular surface tumor. Recent studies of us have indicated the ability of SWNHs to inhibit proliferation of conjunctival melanoma cell line CRMM-1. But the role and molecular mechanisms of it was unclear. To address this question, the research about direct role of SWNHs on the growth, proliferation and apoptosis of CRMM-1 had been performed. Our results indicate that followed with the increasing concentrations of SWNHs, the number of cells decreased and apoptotic cells increased significantly. SWNHs delayed obviously mitotic entry of cells, and these effects followed the cultured time and the gradually increasing concentrations of SWNHs. SWNHs inhibited proliferation of cells at each time point in a time and dose-dependent manner, too. The TEM images showed that individual spherical SWNHs particles smaller than 100 nm in diameters were localized inside cytoplasma of cells. In summary, SWNHs inhibited mitotic entry, growth and proliferation of conjunctival melanoma cells and promoted its apoptosis, inhibited energy metabolism of cells in a dose-dependent manner. The roles of SWNHS on conjunctival melanoma cells were implicating energy metabolism. It may be the effective methods for treatment to conjunctival melanoma.


Assuntos
Carbono/administração & dosagem , Neoplasias da Túnica Conjuntiva/tratamento farmacológico , Neoplasias da Túnica Conjuntiva/metabolismo , Melanoma/tratamento farmacológico , Melanoma/metabolismo , Nanopartículas/administração & dosagem , Antineoplásicos/administração & dosagem , Antineoplásicos/química , Carbono/química , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Humanos , Teste de Materiais , Nanopartículas/química , Nanopartículas/ultraestrutura , Consumo de Oxigênio/efeitos dos fármacos , Tamanho da Partícula , Resultado do Tratamento
11.
Mol Med Rep ; 12(5): 6962-8, 2015 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-26329673

RESUMO

Injury to the optic nerve may lead to axonal degeneration, followed by the gradual death of retinal ganglion cells (RGCs), which results in irreversible vision loss. In the present study the mechanism of optic nerve injury, and the following regeneration and repair processes associated with sirtuin 1 (SIRT1)­regulated lipid metabolism were analyzed. In addition, the treatment of optic nerve injury using resveratrol was investigated. A rat model of optic nerve damage was prepared, and rats were divided into control, model and resveratrol groups. The model rats (with optic nerve damage) were treated with phosphate­buffered saline or resveratrol once a day for seven, 14 and 21 days. The rats were then sacrificed and the optic nerve was dissected. The expression levels of SIRT1, sterol regulatory element­binding protein 2 (SREBP­2) and 3­hydroxy­3­methylglutaryl­CoA reductase (HMGCR) mRNA in the optic nerve were measured by reverse transcription­quantitative polymerase chain reaction, and the protein expression of SIRT1, SREBP2 and HMGCR was evaluated by western blotting. In addition, the cholesterol level of the optic nerve was assessed. The retina was excised and the surviving RGCs were observed and counted. The morphology of the RGCs in the rat model of optic nerve injury changed; however, the degree of damage in rats treated with resveratrol was relatively small. The number of surviving RGCs and the cholesterol level in the RGCs from the model rats was observed to be restored by treatment with resveratrol following optic nerve damage. Additionally, the expression levels of SIRT1, SREBP­2 and HMGCR mRNA and protein were restored by resveratrol treatment in the rats with optic nerve damage. Thus, resveratrol reversed certain features of the optic nerve injury damage via SIRT1 and SREBP­2­associated signaling pathways and the downstream regulated gene, HMGCR. Furthermore, resveratrol promoted cholesterol synthesis in, and repair of, RGCs. Therefore, SIRT1 may serve as a promising novel target for the treatment of optic nerve damage.


Assuntos
Metabolismo dos Lipídeos , Regeneração Nervosa , Traumatismos do Nervo Óptico/fisiopatologia , Nervo Óptico/fisiopatologia , Sirtuína 1/metabolismo , Animais , Hidroximetilglutaril-CoA Redutases/genética , Hidroximetilglutaril-CoA Redutases/metabolismo , Masculino , Nervo Óptico/metabolismo , Nervo Óptico/fisiologia , Traumatismos do Nervo Óptico/genética , Traumatismos do Nervo Óptico/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos Sprague-Dawley , Sirtuína 1/genética , Proteína de Ligação a Elemento Regulador de Esterol 2/genética , Proteína de Ligação a Elemento Regulador de Esterol 2/metabolismo
12.
Sci Rep ; 5: 8209, 2015 Feb 03.
Artigo em Inglês | MEDLINE | ID: mdl-25644821

RESUMO

ACE2 and Ang-(1-7) have important roles in preventing acute lung injury. However, it is not clear whether upregulation of the ACE2/Ang-(1-7)/Mas axis prevents LPS-induced injury in pulmonary microvascular endothelial cells (PMVECs) by inhibiting the MAPKs/NF-κB pathways. Primary cultured rat PMVECs were transduced with lentiviral-borne Ace2 or shRNA-Ace2, and then treated or not with Mas receptor blocker (A779) before exposure to LPS. LPS stimulation resulted in the higher levels of AngII, Ang-(1-7), cytokine secretion, and apoptosis rates, and the lower ACE2/ACE ratio. Ace2 reversed the ACE2/ACE imbalance and increased Ang-(1-7) levels, thus reducing LPS-induced apoptosis and inflammation, while inhibition of Ace2 reversed all these effects. A779 abolished these protective effects of Ace2. LPS treatment was associated with activation of the ERK, p38, JNK, and NF-κB pathways, which were aggravated by A779. Pretreatment with A779 prevented the Ace2-induced blockade of p38, JNK, and NF-κB phosphorylation. However, only JNK inhibitor markedly reduced apoptosis and cytokine secretion in PMVECs with Ace2 deletion and A779 pretreatment. These results suggest that the ACE2/Ang-(1-7)/Mas axis has a crucial role in preventing LPS-induced apoptosis and inflammation of PMVECs, by inhibiting the JNK/NF-κB pathways.


Assuntos
Angiotensina I/metabolismo , Células Endoteliais/metabolismo , Fragmentos de Peptídeos/metabolismo , Peptidil Dipeptidase A/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Receptores Acoplados a Proteínas-G/metabolismo , Angiotensina II/análogos & derivados , Angiotensina II/farmacologia , Animais , Apoptose/efeitos dos fármacos , Células Cultivadas , Células Endoteliais/citologia , Células Endoteliais/efeitos dos fármacos , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Proteínas Quinases JNK Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Lipopolissacarídeos/toxicidade , Pulmão/citologia , Pulmão/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Microvasos/citologia , NF-kappa B/metabolismo , Fragmentos de Peptídeos/farmacologia , Peptidil Dipeptidase A/química , Peptidil Dipeptidase A/genética , Fosforilação/efeitos dos fármacos , Proteínas Proto-Oncogênicas/antagonistas & inibidores , RNA Interferente Pequeno/metabolismo , Ratos , Ratos Sprague-Dawley , Receptores Acoplados a Proteínas-G/antagonistas & inibidores , Transdução de Sinais/efeitos dos fármacos
13.
Hepat Mon ; 13(9): e12280, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-24282423

RESUMO

BACKGROUND: The mutations of LHBs in pre-S, especially in pre-S2, are definitive in hepatocellular carcinoma (HCC) associated with HBV. However, the mechanisms of the N-glycosylation modification in LHBs are unclear. The N-glycosylation modification of LHBs affects Endoplasmic Reticulum stress, cell proliferation and its secretion which was further studied. OBJECTIVES: The objectives of our studies was to indentified that modification of LHBs by N glycosylation modulate their secretion, affect ER stress or expression of cycling, cell cycle and proliferation. MATERIALS AND METHODS: The LHBs was mutated; then expression of proteins related to endoplasmic reticulum stress and EAED path of L02 cells affected by LHBs and its mutations was evaluated. LHBs proteins bound to multiubiquitin chains and its glycosylation motif were studied. The subcellular localization and secretion of LHBs and its mutations were identified. The effect on cell cycle and proliferation by LHBs and its mutations were detected. RESULTS: These data demonstrated that the N-glycosylation motifs of LHBs were associated with ER stress. The N15S, N123S, and N177S mutated LHBs proteins could induce overexpression of EDEM in L02 cells. LHBs and its mutated proteins contained p62-derived UBA domain, which could affect expression of cyclins. The subcellular localization of LHBs in endoplasmic reticulum was similar to its mutations. The secretion of LHBs was blocked by N320K mutation, which could induce an increase in G1 phase and inhibition of S phase, and inhibited mitotic entry. CONCLUSIONS: In conclusion, our studies powerfully demonstrated that modification of LHBs by N glycosylation could modulate their secretion, affect ER stress or expression of cycling, cell cycle and proliferation. The N320K may be the key sites N-linked glycosylation modification of LHBs. It may be a mechanism of HBV-induced HCC.

14.
Hepat Mon ; 13(4): e6065, 2013 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-23914225

RESUMO

BACKGROUND: HBV infection is a serious public health problem worldwide, which can contribute to the incidence of chronic hepatitis B (CHB), cirrhosis, and hepatocellular carcinoma (HCC). OBJECTIVES: In the present report, we assessed the association between adiponectin, its receptors and hepatic steatosis, fibrosis, and inflammation with hepatitis B virus. PATIENTS AND METHODS: Liver biopsies from 89 patients with untreated chronic hepatitis B (34 steatosis vs. 55 without steatosis) were analyzed; liver biopsies from 50 healthy adults were used as control. The liver biopsies were subjected to routine histological examination, and stained immunohistochemically for adiponectin and adiponectin receptor2 (adipoR2). RESULTS: The two groups were found to be comparable with respect to demographic, biochemical, metabolic, histological, and viral characteristics. BMI, γ-GT, FPG, insulin, and insulin sensitivity estimated by the HOMA index were significantly higher in patients with steatosis. The viral load of HBV and HBeAg positivity was higher in patients with steatosis than those without steatosis. High serum adiponectin levels were significantly correlated with abnormal serum ALT level (vs. normal ALT, P = 0.000), and HBV genotype C (vs. genotype B, P = 0.018). In patients with chronic HBV, the insulin sensitizing adipokine adiponectin, and its receptor AdipoR2were associated with steatosis. While adiponectin may becorrelated with inflammation, adiponectin, and its receptors were not associated with viral factors. CONCLUSIONS: Our results suggest that the role of adiponectin might be impaired in chronic hepatitis B with steatosis. Reduced hepatic expression of adiponectin and adipoR2 might be of pathophysiological relevance in CHB patients with steatosis. These findings indicated that reduced liver adiponectin expression may play an important role in the pathogenesis, and progression of CHB patients with steatosis. However, hepatic expression of adiponectin, and adipoR2 was not associated with various measures of HBV infection.

15.
Mol Med Rep ; 7(6): 1889-95, 2013 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-23625030

RESUMO

The development of acute lung injury (ALI) during sepsis almost doubles the mortality rate of patients. The efficacy of current treatment strategies is low as treatment is usually initiated following the onset of symptoms. Inflammation is one of the main mechanisms of autoimmune disorders and is a common feature of sepsis. The suppression of inflammation is therefore an important mechanism for the treatment of sepsis. Sirtuin 1 (Sirt1) has been demonstrated to play a role in the regulation of inflammation. Resveratrol, a potent Sirt1 activator, exhibits anti­inflammatory properties. However, the role of resveratrol for the treatment of ALI during sepsis is not fully understood. In the present study, the anti­inflammatory role of Sirt1 in the lipopolysaccharide (LPS)­induced TC­1 cell line and its therapeutic role in ALI was investigated in a mouse model of sepsis. The upregulation of matrix metalloproteinase-9, interleukin (IL)­1ß, IL­6 and inducible nitric oxide synthase was induced by LPS in the mouse model of sepsis and the TC­1 cell line, and resveratrol suppressed the overexpression of these proinflammatory molecules in a dose­dependent manner. Resveratrol decreased pulmonary edema in the mouse model of sepsis induced by LPS. In addition, resveratrol improved lung function and reduced pathological alterations in the mouse model of sepsis. Knockdown of Sirt1 by RNA interference resulted in an increased susceptibility of TC­1 cells to LPS stimulation and diminished the anti­inflammatory effect of resveratrol. These results demonstrated that resveratrol inhibits LPS­induced ALI and inflammation via Sirt1, and indicated that Sirt1 is an efficient target for the regulation of LPS­induced ALI and inflammation. The present study provides insights into the treatment of ALI during sepsis.


Assuntos
Lesão Pulmonar Aguda/prevenção & controle , Anti-Inflamatórios não Esteroides/uso terapêutico , Sirtuína 1/metabolismo , Estilbenos/uso terapêutico , Lesão Pulmonar Aguda/patologia , Animais , Anti-Inflamatórios não Esteroides/farmacologia , Linhagem Celular , Modelos Animais de Doenças , Inflamação/metabolismo , Inflamação/prevenção & controle , Interleucina-1beta/metabolismo , Interleucina-6/metabolismo , Lipopolissacarídeos/toxicidade , Masculino , Metaloproteinase 9 da Matriz/metabolismo , Camundongos , Óxido Nítrico Sintase Tipo II/metabolismo , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Resveratrol , Sepse/tratamento farmacológico , Sepse/etiologia , Sepse/metabolismo , Sirtuína 1/antagonistas & inibidores , Sirtuína 1/química , Estilbenos/farmacologia , Regulação para Cima/efeitos dos fármacos
16.
Nanoscale Res Lett ; 8(1): 100, 2013 Feb 22.
Artigo em Inglês | MEDLINE | ID: mdl-23432919

RESUMO

Single-wall carbon nanohorns (SWNHs) have been demonstrated to accumulate in cytotoxic levels within organs of various animal models and cell types, which emerge as a wide range of promising biomedical imaging. Septic encephalopathy (SE) is an early sign of sepsis and associated with an increased rate of morbidity and mortality. Microglia activation plays an important role in neuroinflammation, which contributes to neuronal damage. Inhibition of microglia activation may have therapeutic benefits, which can alleviate the progression of neurodegeneration. Therefore, we investigated the functional changes of mice microglia cell lines pre-treated with or without lipopolysaccharide (LPS) induced by SWNHs. To address this question, the research about direct role of SWNHs on the growth, proliferation, and apoptosis of microglia cell lines in mice (N9 and BV2) pre-treated with or without LPS had been performed. Our results indicate that the particle diameter of SWNHs in water is between 342 to 712 nm. The images in scanning electron microscope showed that SWNHs on polystyrene surface are individual particles. LPS induced activation of mice microglia, promoted its growth and proliferation, and inhibited its apoptosis. SWNHs inhibited proliferation, delayed mitotic entry, and promoted apoptosis of mice microglia cells. The effects followed gradually increasing cultured time and concentrations of SWNHs, especially in cells pre-treated with LPS. SWNHs induced a significantly increase in G1 phase and inhibition of S phase of mice microglia cells in a dose-manner dependent of SWNHs, especially in cells pre-treated with LPS. The transmission electron microscope images showed that individual spherical SWNH particles smaller than 100 nm in diameters were localized inside lysosomes of mice microglia cells. SWNHs inhibited mitotic entry, growth and proliferation of mice microglia cells, and promoted its apoptosis, especially in cells pre-treated with LPS. SWNHs inhibited expression of Sirt3 and energy metabolism related with Sirt3 in mice microglia cells in a dose-dependent manner, especially in cells pre-treated with LPS. The role of SWNHs on mice microglia was implicating Sirt3 and energy metabolism associated with it.

17.
Mol Med Rep ; 7(3): 805-10, 2013 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-23358574

RESUMO

A lymphocyte inhibition model was created using a co-culture of donor and host lymphocytes, resulting in apoptosis of the latter, and subsequently, inducing immune tolerance. This method may be used to resolve the immune rejection problem prior to organ transplantation. Using mixed lymphocyte culture (MLC) and/or addition of anti-anti-IL-2 neutralizing monoclonal antibody, we successfully developed a lymphocyte inhibition model in vitro. In this model, the apoptosis of recipient lymphocytes co-cultured with donor lymphocytes was observed by Wright-Giemsa stain, electron microscopy imaging and flow cytometry. The growth and proliferation of mixed lymphocytes were detected by XTT and BrdU assays. Cell viability was determined using CCK-8 cell viability assay. The activity of the recipient lymphocytes was very high when stimulated by antigens alone [PMLC+D2 (Bm) group] but markedly lowered by anti-anti-IL-2 neutralizing monoclonal antibody [PMLC+D2 (Bm)+anti­IL-2 group]. The suppression of recipient lymphocyte activity was due to apoptosis mediated by p53 and caspase-3, and the optimal ratio of donor and recipient lymphocytes for apoptosis was explored. With the exception of the control group, the ratio of apoptotic cells was highest in the PMLC+D2 (Bm)+anti-IL-2 group and lowest in the PMLC+D2 (Bm) group. Blockade of IL-2 with anti-IL-2 neutralizing antibody resulted in an increased number of apoptotic lymphocytes in our experiment, which suggested that IL-2 inhibits the apoptosis of lymphocytes. These data suggest that IL-2 is involved in MLC-induced apoptosis of recipient lymphocytes, and that apoptosis may be associated with p53 and caspase-3 pathways.


Assuntos
Apoptose , Imunossupressão , Linfócitos/citologia , Proteína Supressora de Tumor p53/metabolismo , Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/imunologia , Caspase 3/metabolismo , Células Cultivadas , Técnicas de Cocultura , Humanos , Interleucina-2/imunologia , Interleucina-2/metabolismo , Teste de Cultura Mista de Linfócitos , Linfócitos/metabolismo
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