Zhang X.,Liaocheng Peoples Hospital |
Yu B.,Liaocheng Peoples Hospital |
Hu R.,Liaocheng Infection Hospital |
Hao L.,Yancheng City No1 Peoples Hospital
Biochimie | Year: 2017
Purpose We examined the epigenetic regulation of microRNA-137 (miR-137) on lysine-specific demethylase 1 (KDM1A, or LSD1) induced oncogenic effects in NSCLC. Methods NSCLC cell lines, A549 and H460 cells were transfected with a mammalian LSD1 overexpression plasmid. It's effects on endogenous KDM1A gene and LSD1 protein expressions were examined by qRT-PCR and western blot assays. NSCLC proliferation and migration were also examined by MTT proliferation and wound-scratch assays, respectively. In LSD1-overexpeseed NSCLC cells, lentiviral transfection was conducted to upregulated miR-137 expression. The subsequent effects of miR-137 upregulation on LSD1-mediated cancer regulations were also examined. In addition, key components of histone deacetylases-associated signaling pathways, including EZH2, HDAC1 and HDAC2 were also examined by western blot in LSD1-and miR-137-mediated NSCLC cells. Results Mammalian LSD1 overexpression plasmid was efficient in upregulating KDM1A gene and LSD1 protein in A549 and H460 cells. It also exerted oncogenic effects in NSCLC by promoting cancer proliferation and migration. MiR-137 was inversely correlated with LSD1 in NSCLC, as lentivirus-mediated miR-137 upregulation suppressed KDM1A/LSD1 productions and inhibited proliferation or migration in LSD1-overexpressed A549 and H460 cells. Further western blot analysis demonstrated EZH2, HDAC1 and HDAC2 were activated by LSD1, but inhibited by miR-137 in NSCLC. Conclusion Oncogenic effects of LSD1 were reversely regulated by its upstream epigenetic modulator miR-137 in NSCLC. The interaction between LSD1 and miR-137 may very well involve the regulation on histone deacetylases-associated signaling pathways. © 2017
Meng Q.,Yancheng City No1 Peoples Hospital |
Zhao J.,Shanghai JiaoTong University |
Liu H.,Yancheng City No1 Peoples Hospital |
Zhou G.,Yancheng City No1 Peoples Hospital |
And 3 more authors.
Tumor Biology | Year: 2014
Osteosarcoma is the most common primary malignant bone tumor in children and adolescents. Unfortunately, treatment failures are common due to the metastasis and chemoresistance, but the underlying molecular mechanism remains unclear. Accumulating evidence indicated that the deregulation of DNA-binding protein high-mobility group box 1 (HMGB1) was associated with the development of cancer. This study aimed to explore the expression of HMGB1 in osteosarcoma tissues and its correlation to the clinical pathology of osteosarcoma and to discuss the role of HMGB1 in the development of osteosarcoma. The results from RT-PCR and Western blot showed that the expression rate of HMGB1 messenger RNA (mRNA) and the expression of HMGB1 in the osteosarcoma tissues were significantly higher than those in normal bone tissue (p < 0.05), the expression rate of HMGB1 mRNA and the expression of HMGB1 in the carcinoma tissues with positive lung metastasis were significantly higher than those without lung metastasis (p < 0.05), and with increasing Enneking stage, the expression rate of HMGB1 mRNA and the expression of HMGB1 also increased (p < 0.05). In order to explore the role of HMGB1 in osteosarcoma, the expression of HMGB1 in the human osteosarcoma MG-63 cell line was downregulated by the technique of RNA interference. Western blot results showed that the protein expression of HMGB1 was significantly decreased in the MG-63 cells from HMGB1-siRNA transfection group (p < 0.05), which suggested that HMGB1 was successfully downregulated in the MG-63 cells. Then the changes in proliferation, apoptosis, and invasion of MG-63 cells were examined by MTT test, PI staining, annexin V staining, and transwell chamber assay. Results showed that the abilities of proliferation and invasion were suppressed in HMGB1 knockdown MG-63 cells, and the abilities of apoptosis were enhanced in HMGB1 knockdown MG-63 cells. The expression of cyclin D1, MMP-9 was downregulated in HMGB1 knockdown MG-63 cells, and the expression of caspase-3 was upregulated in HMGB1 knockdown MG-63 cells. Taken together, the overexpression of HMGB1 in osteosarcoma might be related to the tumorigenesis, invasion, and metastasis of osteosarcoma, which might be a potential target for the treatment of osteosarcoma. © 2014, International Society of Oncology and BioMarkers (ISOBM).
WIN55, 212-2 promotes differentiation of oligodendrocyte precursor cells and improve remyelination through regulation of the phosphorylation level of the ERK 1/2 via cannabinoid receptor 1 after stroke-induced demyelination
Sun J.,China Pharmaceutical University |
Fang Y.,China Pharmaceutical University |
Chen T.,China Pharmaceutical University |
Guo J.,China Pharmaceutical University |
And 4 more authors.
Brain Research | Year: 2013
In stroke, a common cause of neurological disability in adults is that the myelin sheaths are lost through the injury or death of mature oligodendrocytes, and the failure of remyelination may be often due to insufficient proliferation and differentiation of oligodendroglial progenitors. In the current study, we used middle cerebral artery occlusion (MCAO) to induced transient focal cerebral ischemia, and found that WIN55, 212-2 augmented actively proliferating oligodendrocytes measured by CC1 immunoreactive cells within the peri-infarct areas. To establish whether these effects were associated with changes in myelin formation, we analyzed the expression of myelin basic protein (MBP) and myelin ultrastructure. We found that WIN55, 212-2 showed more extensive remyelination than vehicle at 14 days post injection (dpi). The extracellular signal-regulated kinase/mitogen-activated protein kinase (ERK/MAPK) signaling pathway may be involved in OPCs differentiation. To determine the regulatory effect of WIN55, 212-2 post-treatment on phospho-ERK 1/2 (p-ERK 1/2) after ischemia/reperfusion, Western blot analysis was performed. We found that WIN55, 212-2 regulated the phosphorylation level of the ERK 1/2 to promote OPCs survival and differentiation. Notably, cannabinoid receptor 1 is coupled to the activation of the ERK cascade. Following rimonabant combined treatment, the effect of WIN55, 212-2 on regulating the phosphorylation level of the ERK 1/2 was reversed, and the effect of accelerated myelin formation was partially inhibited. Together, we first found that WIN55, 212-2 promoted OPCs differentiation and remyelination through regulation of the level of the p-ERK 1/2 via cannabinoid receptor 1. © 2012 Elsevier B.V. All rights reserved.
Shan X.,Yancheng City No1 Peoples Hospital |
Miao Y.,Soochow University of China |
Miao Y.,Yancheng City No1 Peoples Hospital |
Fan R.,Yancheng City No1 Peoples Hospital |
And 6 more authors.
International Journal of Molecular Sciences | Year: 2013
Hepatocellular carcinoma is one of the most common and lethal cancers worldwide, especially in developing countries. In the present study, we found that the expression of a microRNA, miR-590-5P, was down-regulated and S100A10 was up-regulated in six hepatocellular carcinoma cell lines. The reporter gene assay showed that overexpression of miR-590-5P effectively reduced the activity of luciferase expressed by a vector bearing the 3' untranslated region of S100A10 mRNA. Ectopic miR-590-5P overexpression mediated by lentiviral infection decreased expression of S100A10. Infection of Lv-miR-590-5P inhibited cell growth and induced cell cycle G1 arrest in HepG2 cells. In addition, miR-590-5P expression suppressed the expression of Wnt5a, cMyc and cyclin D1, and increased the phosphorylation of β-catenin and expression of Caspase 3, which may contribute to the inhibitory effect of miR-590-5P on cell growth. Taken together, our data suggest that down-regulation of miR-590-5P is involved in hepatocellular carcinoma and the restoration of miR-590-5P can impair the growth of cancer cells, suggesting that miR-590-5P may be a potential target molecule for the therapy of hepatocellular carcinoma. © 2013 by the authors; licensee MDPI, Basel, Switzerland.
Xu Y.,Xuzhou Medical College |
Gao A.-M.,Fudan University |
Ji L.-J.,Xuzhou Medical College |
Li X.,Yancheng City No1 Peoples Hospital |
And 3 more authors.
Cellular Physiology and Biochemistry | Year: 2016
Background/Aims: Hypoxia has recently been proposed as one of the most important factors in progressive renal injury. Hypoxia-induced vascular endothelial growth factor (VEGF) expression may play a critical role in maintaining peritubular capillary endothelium in renal disease. This study was designed to investigate the effect and underlying mechanism of all-trans retinoic acid (ATRA) on hypoxia-induced injury in NRK52E cells. Methods: For mimicking hypoxia, cells were treated with 100 μM of cobalt chloride (CoCl2). The cell viability, expression of VEGF, p65, transforming growth factor-β2 (TGF-β2) and serine carboxypeptidase 1 (Scpep1), and nuclear factor of kappaB (NF-κB) activities after ATRA treatment were determined by MTT, western blot and electrophoretic mobility shift assay. Co-immunoprecipitation analysis was performed to demonstrate whether Scpep1 interacted with TGF-β2. Results: It was found that CoCl2 triggered hypoxia injury and significantly reduced cell viability. ATRA pretreatment increased the cell survival rate. Under hypoxic conditions, the expression of VEGF, p65 and TGF-β2 increased. Addition of ATRA significantly attenuated the expression of VEGF, p65 and TGF-β2. There was a corresponding variation of NF-κB/DNA binding activities. In addition, ATRA stimulated Scpep1 expression under normoxic and hypoxia condition. Furthermore, TGF-β2 interacted with Scpep1. Conclusions: This study indicated that ATRA may attenuate hypoxia-induced injury in NRK52E cells via inhibiting NF-κB/VEGF and TGF-β2/VEGF pathway. © 2016 The Author(s) Published by S. Karger AG, Basel.
Shao K.,Yancheng City No1 Peoples Hospital |
Shao K.,Nantong University |
Ding W.,Nantong University |
Wang F.,Nantong University |
And 3 more authors.
PLoS ONE | Year: 2011
Aptamers are short RNA or DNA oligonucleotides which can bind with different targets. Typically, they are selected from a large number of random DNA sequence libraries. The main strategy to obtain aptamers is systematic evolution of ligands by exponential enrichment (SELEX). Low efficiency is one of the limitations for conventional PCR amplification of random DNA sequence library in aptamer selection because of relative low products and high by-products formation efficiency. Here, we developed emulsion PCR for aptamer selection. With this method, the by-products formation decreased tremendously to an undetectable level, while the products formation increased significantly. Our results indicated that by-products in conventional PCR amplification were from primer-product and product-product hybridization. In emulsion PCR, we can completely avoid the product-product hybridization and avoid the most of primer-product hybridization if the conditions were optimized. In addition, it also showed that the molecule ratio of template to compartment was crucial to by-product formation efficiency in emulsion PCR amplification. Furthermore, the concentration of the Taq DNA polymerase in the emulsion PCR mixture had a significant impact on product formation efficiency. So, the results of our study indicated that emulsion PCR could improve the efficiency of SELEX. © 2011 Shao et al.
Zhang X.,Yancheng City No1 Peoples Hospital |
Chen Y.,Yancheng City No1 Peoples Hospital |
Zhang T.,Yancheng City No1 Peoples Hospital |
Zhang Y.,Yancheng City No1 Peoples Hospital
African Health Sciences | Year: 2015
Background: Da Huang (Radix et Rhizoma Rhei) is the dried root or rhizome of Rheum palmatum L., Rheum tanguticum Maxim ex Balf. or Rheum officinale Braill of family Polygonaceae. It has heat clearing, damp drying, fire purging and toxin removing effects. Because of its definite curative efficacy, it has been widely applied in clinical settings. Objective: To study the inhibitory effect of emodin on human hepatoma cell line SMMC-7721 and its mechanism. Methods: MTT assay, flow cytometry and electron microscopy were used to investigate the inhibitory effect of different concentrations of emodin on human hepatoma cell line SMMC-7721. Results: 12 h, 24 h and 48 h after the action of 20, 40 and 80 umol/L emodin on SMMC-7721 cells, the proliferation of human hepatoma SMMC-7721 cells was inhibited; the inhibitory effects showed time-and concentration-dependence. 48 h after the action of different concentrations of emodin on SMMC-7721 cells, cells in G2/M phase increased significantly, while the proportion of S phase cells gradually declined. Conclusion: Emodin can inhibit human hepatoma cell line SMMC-7721. © Makerere University, Medical School. All rights reserved.
Wan X.,Nanjing Medical University |
Li X.,Yancheng City No1 Peoples Hospital |
Bo H.,Nanjing Medical University |
Zhao Y.,Nanjing Medical University |
And 4 more authors.
Transplantation Proceedings | Year: 2013
Background: It has been reported that the all-trans retinoic acid (atRA)-mediated protective effects in various cells are related to the inhibition of nuclear factor (NF)-κB activities. There exists some evidence that an increase in vascular endothelial growth factor (VEGF), which is expressed by proximal tubular epithelial cells and regulated by NFκB, may play a critical role in maintaining peritubular capillary endothelium in renal disease. By stimulating the production of VEGF, hypoxia is involved in tubulointerstitial fibrosis processes in various renal diseases. Methods: NRK52E cells survival rate was proportional to absorbance in dimethyl-thiazol- diphenyltetrazoliumbromide tests. Quantitative real-time polymerase chain reaction and Western blot were performed to assay the expression of VEGF, p65, and Scpep1. The activation of NFκB was determined by electrophoretic mobility shift assay. Co-immunoprecipitation analysis demonstrates that whether the Scpep1 and NFκB protein interacted. Results: We demonstrated that the hypoxia-mimicking agent CoCl2 triggered hypoxia injury of rat proximal tubular epithelial cells and significantly reduced cell viability. Addition of atRA increased the cell survival rate. Under CoCl 2-mimicking hypoxic conditions, the expression of VEGF and p65 increased. The addition of atRA significantly attenuated the expression of VEGF and p65. There was a similar variation of NFκB/DNA binding activities. atRA not only activated distinct pathways to stimulate the expression of Scpep1, a retinoid-inducible gene, under normoxic conditions, but also acted as a CoCl2-mimicking hypoxia. Conclusion: The protective effects of atRA against hypoxia-induced injury might be involved in suppression of VEGF expression via stimulating Scpep1 distinct pathways and inhibiting the NFκB pathway. © 2013 Elsevier Inc.
PubMed | Yancheng City No1 Peoples Hospital and Soochow University of China
Type: Journal Article | Journal: Annals of clinical and laboratory science | Year: 2016
Long non-coding RNAs (lncRNAs) have been proven to serve a critical role in cancer development and progression. The aim of this study was to elucidate clinical significance of lncRNA MALAT1 expression in breast cancer (BC). A total of 78 BC patients treated with radical resection were enrolled in this study. Quantitative reverse transcription-polymerase chain reaction was used to detect MALAT1 expression in tissues and serum samples. The receiver operating characteristics (ROC) curve was constructed to describe diagnostic specificity and sensitivity. Lentivirus-mediated RNA interference was used to knockdown MALAT1 in the MDA-MB-231 cell line, and then cell proliferation and invasion were explored. Results showed that MALAT1 expression was significantly up-regulated in 85.9% (67/78) of cancerous tissues compared with normal counterparts (P<0.01). Further, an elevated MALAT1 expression in BC tissue was significantly associated with lymph metastasis (P=0.037) and adverse 5-year disease-free survival (mean 48.5 months vs 62.7 months, P=0.012). Suppression of lncRNA MALAT1 significantly inhibited BC cells proliferation, migration and invasion, induced apoptosis and cell cycle G1 arrest. In addition, serum MALAT1 levels in BC patients were much higher than levels in patients with benign breast disease (P<0.001), its diagnostic efficacy was satisfactory, area under the curve (AUC) was 0.833. In conclusion, MALAT1 upregulation plays an important rolein BC development, and serum MALAT1 level may be a potential tumor marker for BC diagnosis.
PubMed | Yancheng City No1 Peoples Hospital
Type: Journal Article | Journal: International journal of molecular sciences | Year: 2013
Hepatocellular carcinoma is one of the most common and lethal cancers worldwide, especially in developing countries. In the present study, we found that the expression of a microRNA, miR-590-5P, was down-regulated and S100A10 was up-regulated in six hepatocellular carcinoma cell lines. The reporter gene assay showed that overexpression of miR-590-5P effectively reduced the activity of luciferase expressed by a vector bearing the 3 untranslated region of S100A10 mRNA. Ectopic miR-590-5P overexpression mediated by lentiviral infection decreased expression of S100A10. Infection of Lv-miR-590-5P inhibited cell growth and induced cell cycle G1 arrest in HepG2 cells. In addition, miR-590-5P expression suppressed the expression of Wnt5a, cMyc and cyclin D1, and increased the phosphorylation of -catenin and expression of Caspase 3, which may contribute to the inhibitory effect of miR-590-5P on cell growth. Taken together, our data suggest that down-regulation of miR-590-5P is involved in hepatocellular carcinoma and the restoration of miR-590-5P can impair the growth of cancer cells, suggesting that miR-590-5P may be a potential target molecule for the therapy of hepatocellular carcinoma.