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941.
Gao Y Li HX Xu LT Wang P Xu LY Cohen L Yang PY Gu K Meng ZQ 《Molecular biology reports》2012,39(2):1683-1689
The purpose of this study was to investigate the effect of bufalin on the anti-proliferative activity of sorafenib in the
human hepatocellular carcinoma (HCC) cell lines PLC/PRF/5 and Hep G-2 and to determine the relevant molecular mechanism. Concurrent
treatment with sorafenib and bufalin at a fixed ratio (25:1) for 48 h resulted in synergistic growth inhibition in HCC cell
lines as determined by CCK-8 cell viability assays. Exposure of both PLC/PRF/5 and Hep G-2 cells to this combination of sorafenib
(6.25 μM) and bufalin (50 nM) resulted in noticeable increases in apoptotic cell death, as evidenced by the disruption of
mitochondria, compared to treatment with either agent alone. Although both sorafenib (6.25 μM) and bufalin (250 nM) alone
inhibited the phosphorylation of ERK, the reduction in pERK was more pronounced in the cells treated with a combination of
bufalin (50 nM) and sorafenib (250 nM). Furthermore, the inhibitory effect of bufalin on pERK was blocked by the PI3kinase
inhibitor LY294002, suggesting that the reduction in pERK induced by bufalin might be mediated by AKT in these two HCC cell
lines. Taken together, the results of our study suggest that bufalin enhances the anti-cancer effects of sorafenib on PLC/PRF/5
and Hep G-2 by contributing to the downregulation of ERK. 相似文献
942.
Obtaining random homozygous mutants in mammalian cells for forward genetic studies has always been problematic due to the diploid genome. With one mutation per cell, only one allele of an autosomal gene can be disrupted, and the resulting heterozygous mutant is unlikely to display a phenotype. In cells with a genetic background deficient for the Bloom's syndrome helicase, such heterozygous mutants segregate homozygous daughter cells at a low frequency due to an elevated rate of crossover following mitotic recombination between homologous chromosomes. We constructed DNA vectors that are selectable based on their copy number and used these to isolate these rare homozygous mutant cells independent of their phenotype. We use the piggyBac transposon to limit the initial mutagenesis to one copy per cell, and select for cells that have increased the transposon copy number to two or more. This yields homozygous mutants with two allelic mutations, but also cells that have duplicated the mutant chromosome and become aneuploid during culture. On average, 26% of the copy number gain events occur by the mitotic recombination pathway. We obtained homozygous cells from 40% of the heterozygous mutants tested. This method can provide homozygous mammalian loss-of-function mutants for forward genetic applications. 相似文献
943.
944.
Xuelian Meng Aijiang Guo Wei Gong Wanzhong JiaXuenong Luo Junjun ZhaiYongxi Dou Xuepeng Cai 《Gene》2012
Ovine β2 subunit of the interleukin (IL)-12 receptor (IL-12Rβ2) was cloned from mRNA preparation of mitogen-activated peripheral blood mononuclear cells (PBMCs). The complete coding sequence for ovine IL-12 Rβ2 was found to be 2586 nucleotides in length encoding 862-amino-acid residue protein. It showed 96.4% homology at the nucleotide level and 94.1% homology at the amino acid level with bovine IL-12 Rβ2. The ovine IL-12 Rβ2 subunit shares common structural and functional elements with their counterparts from the other species. Phylogenetic tree showed that ovine IL-12Rβ2 was clustered into the Artiodactyla group, together with those of cattle and pig, which was distinct from the other groups. Real-time RT-PCR was used to investigate expression of the IL-12Rβ2 in different tissues of sheep in order to determine the characterization of this receptor in tissue. Expression analysis showed that IL-12Rβ2 mRNA expression was detected at all the detected tissues with the exception of thymus. 相似文献
945.
Bacteria are present extensively in the environment. Investigation of their antioxidant properties will be useful for further study on atrazine stress tolerance of bacteria and the defense mechanism of antioxidant enzymes against atrazine or other triazine herbicides. Superoxide dismutase (SOD), catalase (CAT), glutathione S-transferase (GST) and total antioxidant capacity (T-AOC) from one Gram-negative representative strain Escherichia coli K12 and one Gram-positive representative strain Bacillus subtilis B19, respectively, were tested for response to atrazine stress. The results indicated that SOD, CAT, GST and T-AOC were induced upon exposure to atrazine. The growth of two bacteria was better in the absence than in the presence of atrazine, indicating that atrazine can decrease bacterial growth. The changes of enzyme activities indicate the presence of oxidative stress. Oxidative stress induced by atrazine may be due to imbalance of redox potential in bacterial cells, which leads to bacterial metabolic disorder. 相似文献
946.
Assembly and maintenance of nodes of ranvier rely on distinct sources of proteins and targeting mechanisms 总被引:1,自引:0,他引:1
Zhang Y Bekku Y Dzhashiashvili Y Armenti S Meng X Sasaki Y Milbrandt J Salzer JL 《Neuron》2012,73(1):92-107
We have investigated the source(s) and targeting of components to PNS nodes of Ranvier. We show adhesion molecules are freely diffusible within the axon membrane and accumulate at forming nodes from local sources, whereas ion channels and cytoskeletal components are largely immobile and require transport to the node. We further characterize targeting of NF186, an adhesion molecule that pioneers node formation. NF186 redistributes to nascent nodes from a mobile, surface pool. Its initial accumulation and clearance from the internode require extracellular interactions, whereas targeting to mature nodes, i.e., those flanked by paranodal junctions, requires intracellular interactions. After incorporation into the node, NF186 is immobile, stable, and promotes node integrity. Thus, nodes assemble from two sources: adhesion molecules, which initiate assembly, accumulate by diffusion trapping via interactions with Schwann cells, whereas ion channels and cytoskeletal components accumulate via subsequent transport. In mature nodes, components turnover slowly and are replenished via transport. VIDEO ABSTRACT: 相似文献
947.
D Ma GF Dargush SK Natarajan EI Levy AH Siddiqui H Meng 《Journal of biomechanics》2012,45(13):2256-2263
Flow diverter (FD) is an emerging neurovascular device based on self-expandable braided stent for treating intracranial aneurysms. Variability in FD outcome has underscored a need for investigating the hemodynamic effect of fully deployed FD in patient-specific aneurysms. Image-based computational fluid dynamics, which can provide important hemodynamic insight, requires accurate representation of FD in deployed states. We developed a finite element analysis (FEA) based workflow for simulating mechanical deployment of FD in patient-specific aneurysms. We constructed FD models of interlaced wires emulating the Pipeline Embolization Device, using 3D finite beam elements to account for interactions between stent strands, and between the stent and other components. The FEA analysis encompasses all steps that affect the final deployed configuration including stent crimping, delivery and expansion. Besides the stent, modeling also includes key components of the FD delivery system such as microcatheter, pusher, and distal coil. Coordinated maneuver of these components allowed the workflow to mimic clinical operation of FD deployment and to explore clinical strategies. The workflow was applied to two patient-specific aneurysms. Parametric study indicated consistency of the deployment result against different friction conditions, but excessive intra-stent friction should be avoided. This study demonstrates for the first time mechanical modeling of braided FD stent deployment in cerebral vasculature to produce realistic deployed configuration, thus paving the way for accurate CFD analysis of flow diverters for reliable prediction and optimization of treatment outcome. 相似文献
948.
949.
Bian F Mao G Guo M Mao G Wang J Li J Han Y Chen X Zhang M Xia G 《Journal of cellular physiology》2012,227(5):2230-2239
Selective spermatozoa movement from storage of the oviduct to fertilization site is suggested to be a result of chemotaxis. In the present study, Natriuretic peptide precursor A (NPPA) induced sperm chemotaxis in capillaries and enhanced intracellular Ca(2+) level, both of which could be blocked by the Natriuretic Peptide Receptor 1 (NPR1) inhibitor anantin and the cGMP-dependent protein kinase (PKG) inhibitors, KT5823 and Rp-8-Br-PET-cGMPS. NPPA also increased spermatozoa kinetic parameters of VAP, VSL, LIN, STR, and BCF. Only 2.0% of positive staining for NPR1 was detected in fresh spermatozoa. The positive rate was increased in capacitated spermatozoa (20.5%), and further increased in spermatozoa of NPPA treatment (70.2%). Nppa mRNA level in the ampullae was significantly higher compared with that in isthmus and uterotubal junction, and NPPA protein had an ascending gradient (AG) from the uterotubal junction to ampullae in gonadotropin-treated mice. NPPA induced sperm chemotaxis in diestrus oviducts without a NPPA gradient, and sperm chemotaxis occurred in the oviducts of gonadotropin-treated mice. These effects were inhibited by anantin. Meanwhile, sperm chemotaxis also occurred in unilateral ovariectomized oviducts of gonadotropin-treated mice, in which the possible effect of follicular fluid and oocyte-cumulus mass were eliminated when ovulation occurs. Furthermore, anantin significantly decreased the rate of fertilization in a dose-dependent manner (0.1 μM, 57.1%; 1 μM, 33.8%) compared with control (78.5%). These results suggest that a NPPA gradient originating in the oviduct induces sperm chemotaxis by binding to its receptor NPR1 and then activating PKG pathway, and plays a physiological role in fertilization. 相似文献
950.
Q. Meng C. Garcia-Rodriguez G. Manzanarez M.A. Silberg F. Conrad J. Bettencourt X. Pan T. Breece R. To M. Li D. Lee L. Thorner M.T. Tomic J.D. Marks 《Analytical biochemistry》2012,430(2):141-150
Quantitation of individual monoclonal antibodies (mAbs) within a combined antibody drug product is required for preclinical and clinical drug development. We have developed two antitoxins, XOMA 3B and XOMA 3E, each consisting of three mAbs that neutralize type B and type E botulinum neurotoxin (BoNT/B and BoNT/E) to treat serotype B and E botulism. To develop mAb-specific binding assays for each antitoxin, we mapped the epitopes of the six mAbs. Each mAb bound an epitope on either the BoNT light chain (LC) or translocation domain (HN). Epitope mapping data were used to design LC-HN domains with orthogonal mutations to make them specific for only one mAb in either XOMA 3B or XOMA 3E. Mutant LC-HN domains were cloned, expressed, and purified from Escherichia coli. Each mAb bound only to its specific domain with affinity comparable to the binding to holotoxin. Further engineering of domains allowed construction of enzyme-linked immunosorbent assays (ELISAs) that could characterize the integrity, binding affinity, and identity of each of the six mAbs in XOMA 3B and 3E without interference from the three BoNT/A mAbs in XOMA 3AB. Such antigen engineering is a general method allowing quantitation and characterization of individual mAbs in a mAb cocktail that bind the same protein. 相似文献