RESUMO
OBJECTIVE: To analyze the expression of lncRNA-MALAT1 in peripheral blood of patients with acute myeloid leukemia (AML) sepsis and explore its clinical significance. METHODS: From March 2018 to March 2019, 95 confirmed AML patients including 43 sepsis infected cases and 52 uninfected cases were selected for treatment in the Department of Oncology and Hematology, The First People's Hospital of Longquanyi District. Their peripheral blood samples were taken as study samples, and the blood samples from 50 healthy people were used as control. RT-qPCR was used to detect lncRNA-MALAT1 expression level in samples from healthy group, uninfected group, and sepsis group. The correlation between lncRNA-MALAT1 expression level and clinical characteristics and prognosis of AML patients with sepsis were analyzed. RESULTS: The expression level of lncRNA-MALAT1 in the sepsis group was significantly up-regulated compared with the healthy group and uninfected group (P<0.05), while there was no significant difference between the healthy group and uninfected group (P>0.05). In AML patients with sepsis, the expression of lncRNA-MALAT1 was associated with clinical characteristics such as NCCN risk classification, white blood cell count, hemoglobin and so on. The overall survival rate of high lncRNA-MALAT1 expression group was significantly lower than that of low expression group (χ2=23.157, P=0.002). COX regression analysis showed that lncRNA-MALAT1 could be an independent prognostic factor for AML sepsis patients. CONCLUSION: The up-regulated expression of lncRNA-MALAT1 is closely related to the clinical characteristics and survival rate, and is an independent prognostic factor for AML sepsis patients. LncRNA-MALAT1 is expected to become a new diagnostic marker and therapeutic target for AML sepsis.
Assuntos
Leucemia Mieloide Aguda , RNA Longo não Codificante , Sepse , Humanos , Prognóstico , RNA Longo não Codificante/genética , Taxa de SobrevidaRESUMO
Tumor-associated macrophages (TAMs) and their alternative activation contribute greatly to the development of hepatocellular carcinoma (HCC). Receptor-interacting protein 140 (RIP140) is widely expressed in macrophages and regulates macrophage-mediated energy metabolism, the inflammatory response and tumorigenesis. However, whether RIP140 is involved in the activation of TAMs has not been reported. In the present study, we determined the expression of RIP140 in macrophages after treatment with HCC-conditioned medium (HCM) for 24 h. We also analyzed the effects of RIP140 overexpression on macrophage polarization, invasion and apoptosis of HepG2 and Huh7 cells. Transwell and apoptosis assays were used to estimated cell invasion and apoptosis. In addition, we investigated the effects of RIP140 overexpression in macrophages on the growth of H22 cells by subcutaneous injection of H22 cells along with macrophages in BALB/c nude mice. Western blotting and qRT-PCR were used to detect protein and mRNA expression associated with the NF-κB/IL-6 axis in TAMs. Immunohistochemical and immunofluorescence staining were used to evaluate the protein expression of RIP140 or F4/80 in human HCC samples. The protein expression of RIP140 in peripheral blood mononuclear cells was detected by western blotting. KaplanMeier survival curve estimation of overall survival for patients with HCC was based on RIP140 or F4/80 expression in HCC samples. We found that HCM inhibited RIP140 expression and fostered the alternative activation of macrophages. RIP140 overexpression in TAMs significantly inhibited the alternative activation of macrophages by inhibiting the NF-κB/IL-6 axis in TAMs, and suppressed HCC cell growth both in vitro and in vivo. In addition, the protein expression of RIP140 in peripheral blood monocytes was significantly lower in patients with HCC than in healthy people, and this result was consistent with the expression of RIP140 in HCC samples. Furthermore, low RIP140 expression and high F4/80 expression were found to be closely correlated with shorter survival time of the patients with HCC.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Carcinoma Hepatocelular/patologia , Neoplasias Hepáticas/metabolismo , Macrófagos/patologia , NF-kappa B/metabolismo , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Animais , Apoptose , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Linhagem Celular Tumoral , Polaridade Celular , Proliferação de Células , Meios de Cultivo Condicionados , Feminino , Regulação Neoplásica da Expressão Gênica , Células Hep G2 , Humanos , Neoplasias Hepáticas/genética , Macrófagos/metabolismo , Masculino , Camundongos , Transplante de Neoplasias , Proteína 1 de Interação com Receptor Nuclear , Prognóstico , Transdução de Sinais , Análise de SobrevidaRESUMO
Like ubiquitination, several studies have demonstrated that neddylation is implicated to be involved in the double strand break repair. BRCA1 is one of the key repair factors in the homologous recombination repair and may play a downstream role of the neddylation. BRCA1 is also a frequently mutated gene in cancers, which serve as the targets for PARP inhibitors. Here we further investigated the correlation between neddylation and BRCA1 complex using neddylation inhibitor MLN4924. MLN4924 efficiently inhibited the recruitment of components of BRCA1 complex to DNA damage sites. Thus MLN4924 may collaborate with PARP inhibitor to suppress tumor. Our results showed that combination MLN4924 and PARP inhibitor Olaparib impaired the DNA repair process in NSCLC cells. Furthermore, MLN4924 and Olaparib significantly inhibited the cancer cell growth. Kaplan-Meier survival analysis from lung cancer patients showed that high expression of NEDD8, BRCA1 and PARPs correlate with worse overall survival. Thus the combination of MLN4924 and PARP inhibitor may serve as a new strategy for NSCLC treatment.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Ciclopentanos/farmacologia , Neoplasias Pulmonares/tratamento farmacológico , Pirimidinas/farmacologia , Proteína BRCA1 , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Carcinoma Pulmonar de Células não Pequenas/mortalidade , Proteínas de Transporte/metabolismo , Linhagem Celular Tumoral , Ciclopentanos/administração & dosagem , Dano ao DNA/efeitos dos fármacos , Proteínas de Ligação a DNA , Chaperonas de Histonas , Humanos , Estimativa de Kaplan-Meier , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/mortalidade , Complexos Multiproteicos , Proteína NEDD8 , Proteínas Nucleares/metabolismo , Ftalazinas/administração & dosagem , Ftalazinas/farmacologia , Piperazinas/administração & dosagem , Piperazinas/farmacologia , Poli(ADP-Ribose) Polimerase-1/metabolismo , Inibidores de Poli(ADP-Ribose) Polimerases/administração & dosagem , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Pirimidinas/administração & dosagem , Proteínas Supressoras de Tumor/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitinas/metabolismoRESUMO
2',4'-Dihydroxy-6'-methoxy-3',5'-dimethylchalcone (DMC), one of the flavonoids isolated and purified from the dried flower buds of Cleistocalyx operculatus, was explored for its function in glucose uptake/glycogen synthesis in insulin-sensitive tissue cells and its effect and mechanism on 3T3-L1 preadipocyte differentiation. DMC (10 µM) treatment remarkably promoted glucose uptake in differentiated 3T3-L1 adipocytes (P < 0.05 vs control group), whereas the glucose uptake in L6 myoblasts and glycogen synthesis in HepG2 hepatocytes were not affected by the treatment. DMC had paradoxical effects on lipid accumulation in 3T3-L1 cells compared with differentiation control. High concentrations of DMC (10 and 20 µM) markedly diminished lipid accumulation; however, a low concentration of DMC (2.5 µM) enhanced lipid storage in 3T3-L1 cells (P < 0.01 vs differentiation control group), and 5 µM DMC did not impose a significant effect. It was demonstrated that the effect of DMC in lipid accumulation was controlled by the expression of PPAR-γ.
Assuntos
Adipócitos/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Chalconas/farmacologia , Glucose/metabolismo , Myrtaceae/química , Extratos Vegetais/farmacologia , Células 3T3-L1 , Adipócitos/citologia , Adipócitos/metabolismo , Animais , Transporte Biológico/efeitos dos fármacos , Chalconas/efeitos adversos , Transportador de Glucose Tipo 4/genética , Transportador de Glucose Tipo 4/metabolismo , Células Hep G2 , Humanos , Camundongos , PPAR gama/genética , PPAR gama/metabolismo , Regulação para CimaRESUMO
2',4'-Dihydroxy-6'-methoxy-3',5'-dimethylchalcone (DMC), which is isolated and purified from the dried flower buds of Cleistocalyx operculatus (Roxb.) Merr. et Perry (Myrtaceae), was investigated for its insulinotropic benefits against glucotoxicity using in vitro methods. When exposed to high glucose at the cytotoxicity level for 48 h, RIN-5F ß-cells experienced a significant viability loss and impaired insulin secretion function, whereas cotreating with DMC could protect ß-cells against glucotoxicity-induced decrease in glucose-stimulated insulin secretion in a dose-dependent manner without affecting basal insulin secretion. It was demonstrated that DMC increased insulin secretion against glucotoxicity by simulating the effect of GLP-1 and enhancing the expression of GLP-1R, followed by activating the signal pathway of PDX-1, PRE-INS, and GLUT2-GCK. Another mechanism was that DMC avoided the pancreatic islet dysfunction resulting from cellular damage by suppressing the production of nitric oxide (NO) by iNOS, and the expression of MCP-1. The results indicated the potential application of DMC in the intervention against glucotoxicity-induced hyperglycemia.
Assuntos
Chalconas/farmacologia , Glucose/toxicidade , Células Secretoras de Insulina/efeitos dos fármacos , Insulina/metabolismo , Myrtaceae/química , Extratos Vegetais/farmacologia , Substâncias Protetoras/farmacologia , Animais , Linhagem Celular Tumoral , Expressão Gênica/efeitos dos fármacos , Glucose/metabolismo , Transportador de Glucose Tipo 2/genética , Transportador de Glucose Tipo 2/metabolismo , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Secreção de Insulina , Células Secretoras de Insulina/metabolismo , Ratos , Transativadores/genética , Transativadores/metabolismoRESUMO
2',4'-Dihydroxy-6'-methoxy-3',5'-dimethylchalcone (DMC) is a chalcone isolated from the buds of Cleistocalyx operculatus (Roxb.) Merr. et Perry, and the hepatoprotective effects of DMC on Kunming mice have been studied in previous study. However, the effects of DMC on hepatocyte toxicity and corresponding mechanism remain unclear. The aim of this study was to evaluate the hepatoprotective mechanism of DMC in human hepatocytes (L02) treated with H2O2. The results demonstrated that pretreatment with DMC effectively protected H2O2-induced cell viability loss, cell membrane damage (lactate dehydrogenase, nitric oxide production and caspase-3 accumulation. Besides, DMC pretreatment increased the amount of glutathione, decreased malondialdehyde and the percentage of apoptotic L02 cells compared with only H2O2 treated group. Taken together, these results indicated that DMC had hepatoprotective effects against H2O2-induced liver injury by alleviating oxidative stress and apoptosis process in L02 cells, and DMC might be a potential candidate for the intervention of liver diseases.
Assuntos
Antioxidantes/farmacologia , Apoptose/efeitos dos fármacos , Chalconas/farmacologia , Hepatócitos/efeitos dos fármacos , Peroxidação de Lipídeos/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Caspase 3/química , Caspase 3/metabolismo , Linhagem Celular , Membrana Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Etnofarmacologia , Flores/química , Flores/crescimento & desenvolvimento , Glutationa/agonistas , Glutationa/metabolismo , Hepatócitos/enzimologia , Hepatócitos/metabolismo , Humanos , Peróxido de Hidrogênio/antagonistas & inibidores , Peróxido de Hidrogênio/toxicidade , Medicina Tradicional Chinesa , Óxido Nítrico/metabolismo , Concentração Osmolar , Oxidantes/antagonistas & inibidores , Oxidantes/toxicidade , Syzygium/química , Syzygium/crescimento & desenvolvimentoRESUMO
2',4'-Dihydroxy-6'-methoxy-3',5'-dimethylchalcone (DMC), a compound isolated and purified from the dried flower buds of Cleistocalyx operculatus (Roxb.) Merr. et Perry (Myrtaceae), was investigated for its glucose control benefits using in vitro methods. DMC showed strong noncompetitive (IC(50) of 43 µM) inhibition of pancreatic α-amylase; it was, however, ineffective against intestinal α-glucosidase. In addition, DMC exhibited remarkable glucose transport inhibition effects in both simulated fasting and fed states in Caco-2 cell monolayers (P < 0.05). Besides, exposure of MIN6 cells to 250 µM H(2)O(2) for 1 h caused a significant viability loss and insulin secretion reduction. Pretreatment of MIN6 cells with DMC for 2 h protected against the H(2)O(2)-induced decrease in glucose-stimulated insulin secretion in a dose-dependent manner and also enhanced the impaired basal insulin secretion. Such effects highlight the therapeutic potential of DMC in the management of hyperglycemia.
Assuntos
Glicemia/análise , Chalconas/farmacologia , Células CACO-2 , Humanos , Técnicas In VitroRESUMO
OBJECTIVE: To investigate factors and neonatal outcomes associated with histologic chorioamnionitis (HCA) in preterm premature rupture of membranes (PPROM). METHODS: From Jan. 2008 to Jun. 2011, 230 women with PPROM at 28 - 33(+6) weeks of gestation undergoing deliveries in the Second Affiliated Hospital of Wenzhou Medical College were studied retrospectively. According to placental histopathologic findings, those patients were categorized into two groups, including 138 cases in histologic chorioamnionitis (HCA group) and 65 cases in non-chorioamnionitis (control) group. Age, parity, gestational age of PPROM and delivery, latency period, oligohydramnios, white blood cell (WBC) count and serum C-reactive protein (CRP) level at admission and before delivery, the incidence of neonatal respiratory distress syndrome (NRDS), neonatal pneumonia, bronchopulmonary dysplasia, necrotizing enterocolitis, early-onset neonatal sepsis, abnormal brain sonography findings and mortality were compared between two groups. RESULTS: (1) The incidence of HCA was 68.0% (138/203) in all 203 cases with PPROM. (2) The occurring ruptured membrane gestation in HCA group was (31.1 ± 1.5) weeks, which were significantly earlier than (32.0 ± 1.3) weeks in control group (P < 0.05). The level of CRP of (8.2 ± 14.9) mg/L before delivery in HCA group was significantly higher than (5.5 ± 7.2) mg/L in control group (P < 0.05). The rate of oligohydramnios and cesearean sections were 55.1% (76/138) and 45.7% (63/138) in HCA group, which were significantly higher than 30.8% (20/65) and 29.2% (19/65) in control group (P < 0.05). There were no significant difference in patient's age, parity, WBC count and CRP at admission between two groups (P > 0.05). The latency period did not show significant difference between (140 ± 116) hours in HCA group and (129 ± 125) hours in control group (P > 0.05). (3) Using multivariable logistic regression models, oligohydramnios (OR = 2.937), gestational age of PPROM < 32 weeks (OR = 2.352), serum CRP level > 8 mg/L before delivery (OR = 4.923) and latency period > 48 - 168 hours (OR = 4.439) were significantly associated with HCA (P < 0.05). (4) The gestational age of delivery and birth weight of HCA group were significantly lower than those of control group [(32.0 ± 1.5) weeks vs. (32.7 ± 1.5) weeks, (1680 ± 379) g vs. (2017 ± 333) g, respectively, P < 0.05]. The incidence of Apgar < 7, abnormal brain sonograhy findings, neonatal pneumonia, bronchopulmonary dysplasia, early-onset neonatal sepsis and mortality in HCA group were significantly higher than those in control group [20.3% (28/138) vs. 7.7% (5/65), 14.5% (20/138) vs. 4.6% (3/65), 12.3% (17/138) vs. 3.1% (2/65), 5.8% (8/138) vs. 0, 6.5% (9/138) vs. 0, 12.3% (17/138) vs. 3.1% (2/65), respectively, P < 0.05]. The incidence of necrotizing enterocolitis (1.5%, 2/138) in HCA group was higher than that of control group (0) and the incidence of NRDS (18.8%, 26/138) in HCA group did not show statistical difference with 21.4% (14/65) in control group (P > 0.05). CONCLUSIONS: It was found that HCA was significantly correlated with lower gestational age of PPROM, higher serum CRP level before delivery, prolonged latency period and oligohydramnios in PPROM. HCA could increase the neonatal morbidity and mortality.
Assuntos
Proteína C-Reativa/análise , Corioamnionite/etiologia , Ruptura Prematura de Membranas Fetais , Oligo-Hidrâmnio/epidemiologia , Resultado da Gravidez , Adulto , Peso ao Nascer , Corioamnionite/epidemiologia , Feminino , Idade Gestacional , Humanos , Recém-Nascido , Doenças do Recém-Nascido/epidemiologia , Contagem de Leucócitos , Análise Multivariada , Gravidez , Estudos Retrospectivos , Fatores de RiscoRESUMO
Fusarium head blight, caused by members of the Fusarium graminearum species complex (FGSC), is among the most destructive and economically important diseases of small grain crops, including wheat. To determine the phylogenetic species and mycotoxin (trichothecene) chemotypes of the FGSC in the major winter-wheat-producing areas of China, 530 isolates were collected from diseased wheat during the years 2008, 2009, and 2010, and typed using a polymerase chain reaction-based trichothecene genotype assay. Virulence of isolates with different chemotypes was also compared. Of the 530 isolates typed, 348 were F. asiaticum and 182 were F. graminearum. Subdividing the 530 isolates by the trichothecene predicted to be expressed, 482 were of the deoxynivalenol (DON) chemotype and 48 were nivalenol (NIV). Acetylated derivatives of DON included 3-acetyldeoxynivalenol (3-AcDON; 300 isolates), and 15-acetyldeoxynivalenol (15-AcDON; 182 isolates). Chemotypes of the F. asiaticum isolates were either 3-AcDON or NIV, with 3-AcDON being predominant. F. graminearum isolates were all of the 15-AcDON chemotype. F. asiaticum was the predominant phylogenetic species in the Yangtze River Basin and F. graminearum was dominant in the north of China. Two areas of co-occurrence of trichothecene chemotypes were found. The 3-AcDON and 15-AcDON isolates had similar levels of virulence. The DON isolates were significantly more virulent than those of the NIV. The 3-AcDON and 15-AcDON chemotypes were predominant in the Yangtze River Basin and areas north of the Yangtze River Basin, respectively, and it is suggested that geographic distribution is associated with differences in temperature as well as crop rotation systems.
RESUMO
OBJECTIVE: To explore the characteristics of LN and type I, III collagen in pulmonary fibrosis induced by uranium ore dust in rats. METHODS: 60 adult Wistar rats were divided randomly into two groups, control group (30 rats) and uranium ore dust group (30 rats). Non-exposed intratracheal instillation method was used. Uranium ore dust group was exposed 20 mg/ml uranium ore dust suspension 1ml per rat, meanwhile control group was exposed normal saline 1ml per rat. Post-exposed the 7, 14, 21, 30 and 60 d, 6 rats in each group were killed randomly, lung tissue were collected. The pathological changes in lung tissue were observed by microscope using HE staining, the collagen I and III in lungs were observed by polarizing microscope using Biebrich scarlet staining. The expression of LN protein in lung tissue was observed by immunohistochemistry-SP. RESULTS: During lung fibrosis, a large amount of the proliferated I and III collagen in lungs were observed. Post-exposure to uranium ore dust, the characteristics in proliferated collagen in lungs were type I collagen deposited in lung interstitium mainly in the early stage. The area percentage of collagen I and III was increased significantly at 7, 14, 21, 30 and 60d in the experimental group as compared with that in the control group (P < 0.05 or P < 0.01). The over expression of LN in the lung tissue were observed. The expression of LN was distributed in the lung tissue as thickening of the linear or cluster. The integral optical density of LN was increased significantly at 21, 30 and 60 d in the experimental group as compared with that in the control group (P < 0.05 or P < 0.01). CONCLUSIONS: After exposure to uranium ore dust, the characteristics in proliferated collagen in lungs are the type of I collagen deposited in lung interstitium mainly in the early stage, while the type of III collagen increase significantly at the later period. The overexpression of LN exists in the process of pulmonary fibrosis. It suggests that LN has a role effect in the process of pulmonary fibrosis.
Assuntos
Colágeno Tipo III/metabolismo , Colágeno Tipo I/metabolismo , Laminina/metabolismo , Fibrose Pulmonar/metabolismo , Fibrose Pulmonar/patologia , Urânio/efeitos adversos , Animais , Poeira , Feminino , Masculino , Fibrose Pulmonar/induzido quimicamente , Ratos , Ratos WistarRESUMO
Despite the significance of cigarette smoke for carcinogenesis, the molecular mechanisms that lead to increased susceptibility of human cancers are not well-understood. In our present study, the oncogenic transforming effects of cigarette smoke condensate (CSC) were examined using papillomavirus-immortalized human bronchial epithelial cells (BEP2D). Growth kinetics, saturation density, resistance to serum-induced terminal differentiation, anchorage-independent growth and tumorigenicity in nude mice were used to investigate the various stages of transformation in BEP2D cells. Illumina microarray platforms were used to explore the CSC-induced alteration of global mRNA expression profiles of the earlier period and the advanced stage of CSC-treated BEP2D cells. We showed here that a series of sequential steps arose among CSC-treated immortalized human bronchial epithelial cells, including altered growth kinetics, resistance to serum-induced terminal differentiation, and anchorage-independence growth. In the earlier period of CSC treatment, 265 genes were down-regulated and 63 genes were up-regulated, respectively, and in the advanced stage of CSC treatment, 313 genes were down-regulated and 145 genes were up-regulated, respectively. Notably, among those genes, the expression of some of imprinted genes such as IGF2, NDN, H19 and MEG3 were all silenced or down-regulated in CSC-treated cells. These genes reactivated after 5 microM 5-aza-2-deoxycytidine (5-aza-dC) treatment. These results demonstrated that long-term treatment of human bronchial epithelial cells with CSC may adversely affect their genetic and epigenetic integrity and lead to further transformation.
Assuntos
Brônquios/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Nicotiana/toxicidade , Transcrição Gênica/efeitos dos fármacos , Animais , Azacitidina/análogos & derivados , Azacitidina/farmacologia , Northern Blotting , Brônquios/citologia , Brônquios/metabolismo , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Transformação Celular Neoplásica/induzido quimicamente , Transformação Celular Neoplásica/efeitos dos fármacos , Decitabina , Células Epiteliais/metabolismo , Citometria de Fluxo , Perfilação da Expressão Gênica , Humanos , Camundongos , Análise de Sequência com Séries de Oligonucleotídeos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Nicotiana/químicaRESUMO
OBJECTIVE: To investigate the pathogenesis role of aquaporin 3 and aquaporin 9 in idiopathic polyhydramnios by detecting their expression and distribution in fetal membranes and placenta. METHODS: Twenty-one of term pregnancy women with idiopathic polyhydramnios were enrolled as patient group matched with 30 women with normal term pregnancy as control group. The expression and localization of aquaporin 3 and aquaporin 9 in fetal membranes and placenta were detected by real-time polymerase chain reaction and streptavidin peroxidase immunohistochemiscal staining. RESULTS: (1) The mRNA expressions of aquaporin 3 and aquaporin 9 were detected in amnion, chorion and placental tissue in both patient group and control group. Both aquaporin 3 and aquaporin 9 were demonstrated positive staining in the amnion epithelia, chorion cytotrophoblasts and placental trophoblast. (2) The ratio of aquaporin 3 and aquaporin 9 mRNA expressions in amnion in patient group comparing to those in control group were 5.00 and 3.25, while in chorion they were 2.03 and 2.08. When compared with those in amnion and chorion of control group, there was a significant difference (P < 0.01). However, the relative change fold of aquaporin 3 and aquaporin 9 in placental trophoblast in patient group were decreased in comparison of those in control group, which also showed statistical difference (P < 0.01). (3) The expression of aquaporin 3 and aquaporin 9 protein in amnion were 7.5 +/- 2.0 and 11.1 +/- 1.8 in patient group, while they were 5.3 +/- 1.6 and 5.6 +/- 2.3 in control group. In chorion, the expression of aquaporin 3 and aquaporin 9 protein was 7.5 +/- 2.0 and 10.0 +/- 1.6 in patient group, respectively, while in control group, they were 5.4 +/- 2.2 and 5.6 +/- 2.1. When compared with those proteins in control group, it exhibited statistical difference (P < 0.05). However, in placental trophoblast of patient group, the expression of aquaporin 3 and aquaporin 9 protein were 3.5 +/- 1.4 and 4.0 +/- 2.5, respectively, which were significantly decreased than 5.6 +/- 1.3 and 7.1 +/- 2.9 in control group (P < 0.05). CONCLUSIONS: The alterations of aquaporin 3 and aquaporin 9 expressions in fetal membrane and placenta might be an adaptive response to idiopathic polyhydramnios. Further investigation should be needed to clarify the regulatory mechanism of aquaporin 3 and aquaporin 9 expressions.
Assuntos
Aquaporina 3 , Poli-Hidrâmnios , Âmnio/metabolismo , Aquaporina 1/metabolismo , Aquaporina 3/metabolismo , Membranas Extraembrionárias/metabolismo , Feminino , Humanos , Placenta/metabolismo , Poli-Hidrâmnios/metabolismo , Gravidez , RNA Mensageiro/metabolismoRESUMO
Despite the significance of oxidative damage in carcinogenesis, the molecular mechanisms that lead to increased susceptibility to oxidative stress are not well understood. We now report a link between loss of protection against oxidative damage and loss of function of PTEN, a highly mutated tumor suppressor gene in a variety of human tumors. Using two-dimensional gel electrophoresis, combined with Western and Northern blot analyses, we found that PTEN deficiency in mouse embryonic fibroblasts (MEFs) displays deregulated expression of several antioxidant enzymes, including peroxiredoxins 1, 2, 5, and 6 and Cu, Zn superoxide dismutase. In these Pten-deleted MEFs, the basal levels of reactive oxygen species (ROS) were increased, and both the basal level and the ROS-induced oxidative damage of DNA were increased, as evidenced by increased levels of hydrogen peroxide (H2O2), superoxide anion, 8-hydroxy-2'-deoxyguanosine, and DNA double-strand breaks. We further show that Pten deletion is correlated with resistance to H2O2-induced expression of several antioxidants. These findings suggest an essential role for PTEN in maintaining the normal redox state of mouse embryonic fibroblasts against oxidative damage. They also provide a molecular link between PTEN, whose inactivation is known to be involved in a variety of human tumors, and antioxidants, whose perturbation leads to oxidative damage of cells.
Assuntos
Antioxidantes/metabolismo , Fibroblastos/metabolismo , Deleção de Genes , Estresse Oxidativo/genética , PTEN Fosfo-Hidrolase/metabolismo , Animais , Células Cultivadas , Quebras de DNA de Cadeia Dupla , Embrião de Mamíferos , Regulação da Expressão Gênica , Peróxido de Hidrogênio/farmacologia , Camundongos , PTEN Fosfo-Hidrolase/genética , Peroxirredoxinas/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Superóxido Dismutase/metabolismoRESUMO
BACKGROUND & OBJECTIVE: Pleiotrophin (Ptn), a secretive growth/differentiation factor, has diverse functions involved in cell activities, including adhesion, migration, survival, growth, and differentiation. Ptn has been suggested to be a potential target for the treatment of several types of cancer. Studies have showed that rRibozyme targeting Ptn suppresses the growth, angiogenesis, and metastasis of melanoma and pancreatic cancer cells. This study was to produce a small interfering RNA (siRNA) to inhibit Ptn expression. METHODS: A group of double strand oligonucleotide fragments were synthesized and cloned into pSilencer 3.1-H1 hygro vector. siRNA-expressing vectors were transiently transfected into 3T3 cells to observe the inhibitory effects of different siRNAs on Ptn expression. Lipofectamine 2000 transfection and hygromycin B screening were used to establish PTEN-/- MEF241 cell line which could stably express silenced Ptn. The expression of Ptn was measured by Northern blot. Cell proliferation was measured. Tumorigenecity in nude mice was also measured to test if silencing the expression of Ptn can change the malignant phenotypes of PTEN-/- MEF241 cells. RESULTS: Three Ptn-specific siRNAs were designed and cloned into pSilencer 3.1-H1 hygro vector. One of them, PTEN siRNA-B, was proven to be able to effectively inhibit Ptn gene expression in PTEN-/- MEF241 cells; the inhibition rate was over 95%. The growth of PTEN-/- MEF241 cell clones was significantly slowed. Moreover, inhibiting the expression of Ptn by siRNA suppressed tumor growth and prolonged tumorigenesis duration in PTEN-/- MEF241 cell-grafted nude mice. CONCLUSION: Ptn-specific siRNA could inhibit the proliferation of PTEN-/- MEF241 cells and inhibit tumorigenesis, therefore, may be a potential target of antitumor gene therapy.
Assuntos
Proteínas de Transporte/metabolismo , Proliferação de Células , Citocinas/metabolismo , PTEN Fosfo-Hidrolase/genética , RNA Interferente Pequeno/farmacologia , Células 3T3 , Animais , Testes de Carcinogenicidade , Proteínas de Transporte/genética , Citocinas/genética , Deleção de Genes , Vetores Genéticos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Camundongos Nus , Fenótipo , TransfecçãoRESUMO
BACKGROUND & OBJECTIVE: Smad7 is an inhibitor of transforming growth factor-beta (TGF-beta) signal pathway. TGF-beta could induce the expression of several genes through activating SMAD and ras/MEK/ERK pathways. This study was to determine whether Smad7 is involved in regulating mitogen-activated protein kinase (MAPK) signal pathway with TGF-beta in malignant transformation of human bronchial epithelial BEP2D cells. METHODS: Immortalized BEP2D cells and malignant BERP35T2 cells were co-transfected with full-length Smad7 cDNA constructed pCISmad7.neo or Smad7 siRNA, transactivator vector pTet-Elk or pTet-Jun, and reporter vector pTRE-Luc, and stimulated with TGF-beta. The regulatory effect of Smad7 on MAPK signal pathway was investigated by standard luciferase assay. RESULTS: In BEP2D cells, when treated with TGF-beta1, phosphorylated activities of Elk and Jun were up-regulated (P(Elk)=0.033, P(Jun)=0.016); after co-transfection of Elk or Jun with pCISmad7.neo, phosphorylated activity of Elk was increased, and that of Jun was decreased (P(Elk)=0.017, P(Jun)=0.028); after co-transfection of Elk or Jun with Smad7 siRNA, phosphorylated activity of Elk was decreased, and that of Jun was increased (P(Elk)=0.018, P(Jun)=0.005). In BERP35T2 cells, when treated with TGF-beta1, phosphorylated activity of Elk was up-regulated (P=0.006); after co-transfection of Elk and Smad7 siRNA, phosphorylated activity of Elk was decreased (P=0.000); no activity of Jun was detected in BERP35T2 cells. CONCLUSIONS: In the process of malignant transformation of BEP2D cells, the intervention of Smad7 in MAPK signal pathway leads to the activity imbalance between extracellular signal-related protein kinase (ERK) and c-Jun NH2-terminal kinase (JNK), which in turn promotes cell proliferation. All these could contribute to further malignant transformation of these cells.
Assuntos
Transformação Celular Neoplásica , Células Epiteliais/citologia , Sistema de Sinalização das MAP Quinases , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Proteína Smad7/metabolismo , Brônquios/citologia , Células Cultivadas , Humanos , Fosforilação , Proteínas Proto-Oncogênicas c-jun/metabolismo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Proteína Smad7/genética , Transfecção , Fator de Crescimento Transformador beta/farmacologia , Proteínas Elk-1 do Domínio ets/metabolismoRESUMO
OBJECTIVE: To study the effect of overexpression of Smad7 gene on cell proliferation in human bronchial epithelial cell lines. METHODS: Human bronchial epithelial cell lines, BEP2D and BERP35T2 cells, were cotransfected with the mammalian expression vectors PCISmad7.neo and pMyc-SEAP, the latter was ac-myc cis-acting enhancer element fused with alkaline phosphatase (SEAP) reporter gene. Expression of c-myc, p15 and p21 mRNA was detected by RT-PCR before and after stable transfection of Smad7 into BEP2D and BERP35T2 cells in order to study the regulation of TGF-beta-mediated growth inhibition. RESULTS: After BEP2D and BERP35T2 cells transfected with Smad7, the transcriptional activity of c-myc was significantly increased. Smad7 overexpressing cells showed upregulation of c-myc expression and downregulation of p15 and p21 expression, which contributed to the loss of TGF-beta responses in these cells. CONCLUSION: Overexpression of Smad7 may facilitate cell proliferation by antagonizing TGF-beta-mediated antiproliferative gene responses.
Assuntos
Brônquios/citologia , Proliferação de Células , Células Epiteliais/citologia , Proteína Smad7/biossíntese , Fator de Crescimento Transformador beta/biossíntese , Transformação Celular Neoplásica , Células Cultivadas , Inibidor de Quinase Dependente de Ciclina p15/biossíntese , Inibidor de Quinase Dependente de Ciclina p15/genética , Inibidor de Quinase Dependente de Ciclina p21/biossíntese , Inibidor de Quinase Dependente de Ciclina p21/genética , Humanos , Proteínas Proto-Oncogênicas c-myc/biossíntese , Proteínas Proto-Oncogênicas c-myc/genética , Transdução de Sinais , Proteína Smad7/genética , Transfecção , Fator de Crescimento Transformador beta/genéticaRESUMO
BACKGROUND & OBJECTIVE: Escape from transforming growth factor-beta(TGF-beta)-induced inhibition of growth and proliferation may contribute to tumorigenesis. Smad7 is inhibitory Smads of TGF-beta s signal transduction pathway and prevents TGF-beta signaling. The disorder of Smad7 may lead to the perturbation of TGF-beta signal pathway. In this study, The authors analyzed the expression of Smad7 mRNA and the regulation of Smad7 gene by TGF-beta 1 in the process of malignant transformation of BEP2D cells to investigate the mechanism of cells malignant transformation. METHODS: Cells were cultured and stimulated with TGF-beta 1 followed by RNA extraction. Purified total RNA from TGF-beta 1 treated cells and untreated controls and performed an expression analysis with a human Smad7-specific probe applying Northern blot. As a loading control for the Northern experiment, the membrane was hybridized with a human glyceraldehyde-3-phosphate dehydrogenase(GAPDH) probe. Proteins were extracted from BEP2D and BERP35T-2 cells, then perform Western blot to examine the expression level of TGF-beta 1. RESULTS: Before stimulation with TGF-beta 1, the expression level of Smad7 in the BERP35T-2 cells were higher than that in the BEP2D cells. When stimulated with TGF-beta 1, Smad7 expression levels was upregulated evidently in BEP2D cells, but not significant in BERP35T-2 cells. The expression level of endogenetic TGF-beta 1, BERP35T-2 cells was a little higher than BEP2D cells. CONCLUSION: Over expression of Smad7 mRNA and down-regulation of the cells' responsiveness to TGF-beta 1 in human lung cancer cell line which induced by alpha-particles should be one of the mechanism of radiation induced lung cancer.