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Luciferase Assay to Screen Tumour-specific Promoters in Lung Cancer
Xu, Rong,Guo, Long-Jiang,Xin, Jun,Li, Wen-Mao,Gao, Yan,Zheng, You-Xian,Guo, You-Hong,Lin, Yang-Jun,Xie, Yong-Hua,Wu, Ya-Qing,Xu, Rui-An Asian Pacific Journal of Cancer Prevention 2013 Asian Pacific journal of cancer prevention Vol.14 No.11
Objective: Specific promoters could improve efficiency and ensure the safety of gene therapy. The aim of our study was to screen examples for lung cancer. Methods: The firefly luciferase gene was used as a reporter, and promoters based on serum markers of lung cancer were cloned. The activity and specificity of seven promoters, comprising CEACAM5 (carcinoembryonic antigen, CEA), GRP (Gastrin-Releasing Peptide), KRT19 (cytokeratin 19, KRT), SFTPB (surfactant protein B, SP-B), SERPINB3 (Squamous Cell Carcinoma Antigen, SCCA), SELP (Selectin P, Granule Membrane Protein 140kDa, Antigen CD62, GMP) and DKK1 (Dickkopf-1) promoters were compared in lung cancer cells to obtain cancer-specific examples with strong activity. Results: The CEACAM5, DKK1, GRP, SELP, KRT19, SERPINB3 and SFTPB promoters were cloned. Furthermore, we successfully constructed recombinant vector pGL-CEACAM5 (DKK1, GRP, SELP, KRT19, SERPINB3 and SFTPB) contained the target gene. After cells were transfectedwith recombinant plasmids, we found that the order of promoter activity from high to low was SERPINB3, DKK1, SFTPB, KRT19, CEACAM5, SELP and GRP and the order for promoters regarding specificity and high potential were SERPINB3, DKK1, SELP, SFTPB, CEACAM5, KRT19 and GRP. Conclusion: The approach adopted is feasible to screen for new tumour specific promoters with biomarkers. In addition, the screened lung-specific promoters might have potential for use in lung cancer targeted gene therapy research.
Yin, Yan,Wang, Rong-Rong,Wang, Yan,Wang, Jian-Jun,Xu, Gen-Xing Asian Pacific Journal of Cancer Prevention 2014 Asian Pacific journal of cancer prevention Vol.15 No.8
In this study, we demonstrated selenium (Se) accumulation in Bifidobacterium longum strain (B. longum) and evaluated the effect of Se-enriched B. longum (Se-B. longum) on tumor growth and immune function in tumor-bearing mice. Analysis using high-performance liquid chromatography-inductively coupled plasma mass spectrometry (HPLC-ICP-MS) revealed that more than 99% of Se in Se-B. longum was organic, the main component of which was selenomethionine (SeMet). In the in vivo experiments, tumor-bearing mice (n=8) were orally administrated with different doses of Se-B. longum alone or combined with cyclophosphamide (CTX). The results showed that the middle and high dose of Se-B. longum significantly inhibited tumor growth. When Se-B. longum and CTX were combined, the antitumor effect was significantly enhanced and the survival time of tumor-bearing mice (n=12) was prolonged. Furthermore, compared with CTX alone, the combination of Se-B. longum and CTX stimulated the activity of natural killer (NK) cells and T lymphocytes, increasing the levels of interleukin-2 (IL-2) and tumor necrosis factor-${\alpha}$ (TNF-${\alpha}$), and the leukocyte count of H22 tumor-bearing mice (n=12).
Chen, Peng,Wang, Xiu-Li,Ma, Zhong-Sen,Xu, Zhong,Jia, Bo,Ren, Jin,Hu, Yu-Xin,Zhang, Qing-Hua,Ma, Tian-Gang,Yan, Bing-Di,Yan, Qing-Zhu,Li, Yan-Lei,Li, Zhen,Yu, Jin-Yan,Gao, Rong,Fan, Na,Li, Bo,Yang, Jun Asian Pacific Journal of Cancer Prevention 2012 Asian Pacific journal of cancer prevention Vol.13 No.7
HMGN5 is a typical member of the HMGN (high mobility group nucleosome-binding protein) family which may function as a nucleosomal binding and transcriptional activating protein. Overexpression of HMGN5 has been observed in several human tumors but its role in tumorigenesis has not been fully clarified. To investigate its significance for human lung cancer progression, we successfully constructed a shRNA expression lentiviral vector in which sense and antisense sequences targeting the human HMGN5 were linked with a 9-nucleotide loop. Inhibitory effects of siRNA on endogenous HMGN5 gene expression and protein synthesis were demonstrated via real-time RT-PCR and western blotting. We found HMGN5 silencing to significantly inhibit A549 and H1299 cell proliferation assessed by MTT, BrdU incorporation and colony formation assays. Furthermore, flow cytometry analysis showed that specific knockdown of HMGN5 slowed down the cell cycle at the G0/G1 phase and decreased the populations of A549 and H1299 cells at the S and G2/M phases. Taken together, these results suggest that HMGN5 is directly involved in regulation cell proliferation in A549 and H1299 cells by influencing signaling pathways involved in cell cycle progression. Thus, our finding suggests that targeting HMGN5 may be an effective strategy for human lung cancer treatment.
Rong-hua Pan,Feng-yan Xie,Hui-mei Chen,Li-zhi Xu,Xiao-chun Wu,Ling-ling Xu,Gang Yao 대한약학회 2011 Archives of Pharmacal Research Vol.34 No.3
Salvianolic acid B (Sal B) is the most abundant bioactive molecule from Radix Salviae Miltiorrhizae, and has recently been used for treating renal fibrosis in traditional Chinese medicine. Here we investigated the ability reversal of Sal B to reverse the transdifferentiation of human kidney proximal tubular epithelial cells that was induced by transforming growth factor-beta 1 (TGF-β1). The effects of Sal B on HK-2 cell morphology were observed by phase contrast microscopy, while alpha smooth muscle actin and E-cadherin were studied by immunocytochemistry and real-time reverse transcription polymerase chain reaction, respectively. Exposure of HK-2 cells to TGF-β1 for 72 h induced a complete conversion of the epithelial cells to myofibroblasts. When HK-2 cells were co-incubated with Sal B and TGF-β1 for a further 72 h, the morphology of myofibroblasts returned to that of proximal tubular epithelial cells,whereas the myofibroblast phenotype was maintained after exposure of cells to TGF-β1 for 144 h. Sal B reduced alpha smooth muscle actin levels and increased E-cadherin levels compared with their epithelial-to-mesenchymal transition controls. The reversal effect of Sal B was dose-dependent. That Sal B reverses the epithelial-to-mesenchymal transition in vitro suggests that it could possibly facilitate the repair of tubular epithelial structures and the regression of renal fibrosis in injured kidneys.
Yue-Rong Liang,Shi-Cheng Ma,Xin-Qiang Zheng,Jing-Yi Xu,Ming-Yan Wu,Yi-Wen Luo,Xian-Yang Luo,Jian-Liang Lu 한국식품과학회 2011 Food Science and Biotechnology Vol.20 No.1
Three groups of spontaneously hypertensive rats were administered by gavage with distilled water (control group), low-dose green tea (LGT, 0.2 g/kg BW·day) and high-dose green tea (HGT, 1.0 g/kg BW·day) for 4 weeks,during which systolic blood pressure (SBP) was measured weekly by non-invasive tail-cuff method. At the end of experiment, left ventricular hypertrophy index (LHVI) and plasma biochemical indicators were determined, and ultrastructures of myocardium and aortic vascular smooth muscle cells were observed by transmission electron microscopy (TEM). The results show that green tea gavage suppressed the increase in SBP, along with decline in levels of plasma nitric oxide, aldosterone, malondialdehyde, and LHVI, but increased levels of plasma creatinine, calcitonin gene-related peptide, and glutathione peroxidase in a dose dependant manner. Green tea had no effect on plasma total cholesterol. TEM shows that green tea gavage protected mitochondria of left ventricular myocardium and aortic vascular smooth muscle cells from damage.
Yinglin Yan,Haichao Qin,Yiqi Wei,Rong Yang,Yunhua Xu,Liping Chen,Qiaole Li,Mangmang Shi 대한화학회 2018 Bulletin of the Korean Chemical Society Vol.39 No.5
Recently, three dimensional graphene aerogel (3DGA) supported sulfur microparticles was used as a cathode material for lithium?sulfur batteries, which was considered as one of the most promising next generation rechargeable batteries due to its ultra?high theoretical specific capacity (1675 mAh/g). However, the mechanical strength of 3DGA remains an issue for further application. Herein, a strengthened 3DGA (S3DGA) was achieved by soaking in a low concentration ammonia solution at a relative low temperature. Then the S3DGA loaded sulfur (S3DGA?S) was cut into a round piece and directly used as a cathode without additional binders or conductive additives in Li?S batteries. The mechanical strength, microstructure, and electrochemical properties were investigated by compare with a 3DGA prepared without strengthen. The S3DGA?S presented good mechanical strength, excellent capacity retention, and lower electrochemical impedance.
Feng Xu,Hua Cheng,Rong Cai,Lin Ling Li,Jie Chang,Jun Zhu,Feng Xia Zhang,Liu Ji Chen,Yan Wang,Shu Han Cheng,Shui Yuan Cheng 한국분자세포생물학회 2008 Molecules and cells Vol.26 No.6
Anthocyanidin synthase (ANS, leucoanthocyanidin oxygenase), a 2-oxoglutarate iron-dependent oxygenase, catalyzed the penultimate step in the biosynthesis of the anthocyanin class of flavonoids, from the colorless leucoanthocyanidins to the colored anthocyanidins. The full-length cDNA and genomic DNA sequences of ANS gene (designated as GbANS) were isolated from Ginkgo biloba for the first time. The full-length cDNA of GbANS contained a 1062-bp open reading frame (ORF) encoding a 354-amino-acid protein. The genomic DNA analysis showed that GbANS gene had three exons and two introns. The deduced GbANS protein showed high identities to other plant ANSs. The conserved amino acids (H-X-D) ligating ferrous iron and residues (R-X-S) participating in 2-oxoglutarate binding were found in GbANS at the similar positions like other ANSs. Southern blot analysis indicated that GbANS belonged to a multi-gene family. The expression analysis by real-time PCR showed that GbANS expressed in a tissue-specific manner in G. biloba. GbANS was also found to be up-regulated by all of the six tested abiotic stresses, UV-B, abscisic acid, sucrose, salicylic acid, cold and ethylene, consistent with the promoter region analysis of GbANS. The recombinant protein was successfully expressed in E. coli strain with pET-28a vector. The in vitro enzyme activity assay by HPLC indicated that recombinant GbANS protein could catalyze the formation the cyanidin from leucocyanidin and conversion of dihydroquercetin to quercetin, suggesting GbANS is a bifunctional enzyme within the anthocyanidin and flavonol biosynthetic pathway.