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排序方式: 共有205条查询结果,搜索用时 31 毫秒
61.
Boorsma M Hoenke S Marrero A Fischer R Bailey JE Renner WA Bachmann MF 《Biotechnology and bioengineering》2002,79(6):602-609
The production and study of toxic proteins requires inducible expression systems with low basal level expression and high inducibility. Here, we describe bioprocess applications of the pCytTS temperature-regulatable Sindbis virus replicon-based expression system. We used green fluorescent protein as a marker protein to optimize the selection of stable transfected clones with increased expression levels. Using the optimized protocol, clones were constructed that produced the growth-inhibiting, anti-viral protein interferon beta (beta-IFN). Selected clones were analyzed for temperature-dependent beta-IFN production in adherent and suspension cultures in serum free medium. Specific expression levels were around 1.0 x 10(5) IU/10(6) cells/day (0.5 microg/10(6) cells/day) in suspension cultures and over 1.5 x 10(6) IU/mL/day (7.5 microg/mL/day) in hollow fiber reactors using adherent cells. Hexahistidine-tagged beta-IFN purified from T-flask cultures was highly glycosylated and showed high specific activity. beta-IFN mRNA amplified by the viral replicase for 10 days did not show an accumulation of mutations. These data suggest the applicability of the pCytTS-inducible expression system for the production of high-quality glycoproteins in different reactors. 相似文献
62.
Díaz-Vázquez LM García O Velázquez Z Marrero I Rosario O 《Journal of chromatography. B, Analytical technologies in the biomedical and life sciences》2005,825(1):11-20
Methodology using MAE/SPME/GC-MS is being pursued for the analysis of organic pollutants in sebum. The microwave-assisted extraction (MAE) of standards of semi volatile organic pollutants from sebum was optimized. All compounds were extracted from sebum with recoveries analyzed by GC/MS ranging from 94% to 100% under the optimum MAE conditions: 10mL acetone-hexane (2:1), 60 degrees C, and 10 min microwave heating. To improve the detection limits a SPME procedure was optimized. Linearity ranged from 0.70 ppb to 25 ppb. R.S.D. were in the range of 1-23% for the SPME step. Preliminary real samples were analyzed and a range of compounds was detected. The optimized MAE/SPME/GC-MS methodology promises to be useful for different applications. 相似文献
63.
Vascular endothelial growth factor activates STAT proteins in aortic endothelial cells 总被引:7,自引:0,他引:7
Bartoli M Gu X Tsai NT Venema RC Brooks SE Marrero MB Caldwell RB 《The Journal of biological chemistry》2000,275(43):33189-33192
64.
Presenilin-dependent gamma-secretase processing regulates multiple ERBB4/HER4 activities 总被引:6,自引:0,他引:6
Transmembrane receptors typically transmit cellular signals following growth factor stimulation by coupling to and activating downstream signaling cascades. Reports of proteolytic processing of cell surface receptors to release an intracellular domain (ICD) has raised the possibility of novel signaling mechanisms directly mediated by the receptor ICD. The receptor tyrosine kinase ERBB4/HER4 (referred to here as ERBB4) undergoes sequential processing by tumor necrosis factor-alpha converting enzyme and presenilin-dependent gamma-secretase to release the ERBB4 ICD (4ICD). Our recent data suggests that regulation of gene expression by the ERBB4 nuclear protein and the proapoptotic activity of ERBB4 involves the gamma-secretase release of 4ICD. To determine the role gamma-secretase processing plays in ERBB4 signaling, we generated an ERBB4 allele with the transmembrane residue substitution V673I (ERBB4-V673I). We demonstrate that ERBB4-V673I fails to undergo processing by gamma-secretase but retains normal cell surface signaling activity. In contrast to wild-type ERBB4, however, ERBB4-V673I was excluded from the nuclei of transfected cells and failed to activate STAT5A stimulation of the beta-casein promoter. These results support the contention that gamma-secretase processing of ERBB4 is necessary to release a functional 4ICD nuclear protein which directly regulates gene expression. We also demonstrate that 4ICD failed to accumulate within mitochondria of ERBB4-V673I transfected cells and the potent proapoptotic activity of ERBB4 was completely abolished in cells expressing ERBB4-V673I. Our results provide the first formal demonstration that proteolytic processing of ERBB4 is a critical event regulating multiple receptor signaling activities. 相似文献
65.
66.
M Marrero R A Prough R S Putnam M Bennett L Milewich 《The Journal of steroid biochemistry and molecular biology》1991,38(5):599-609
Dehydroepiandrosterone (DHEA), administered per os, serves to prevent or retard the development of a variety of genetic and induced disorders in mice and rats. This treatment also results in the development of hepatomegaly, a change of liver color from pink to mahogany, peroxisome proliferation in hepatocytes and alterations in hepatocyte mitochondria morphology and respiration. We used one- and two-dimensional polyacrylamide gel electrophoresis (PAGE) to identify changes in the relative levels of liver proteins produced by DHEA treatment of rodents. In mouse liver, there were apparent increases in the levels of 26 proteins and decreases in the levels of 7 proteins. Of the induced proteins the most prominent had Mr approximately 72 K; this protein was identified in a previous study as enoyl-CoA hydratase/3-hydroxyacyl-CoA dehydrogenase. Another protein of Mr approximately 28 K, of unknown nature, also was induced markedly by DHEA treatment of mice and rats. A protein of Mr approximately 160 K, which was identified as carbamoyl phosphate synthetase-I (CPS-I), was decreased markedly by DHEA action. This enzyme, which comprises approx. 15-20% of mitochondrial matrix protein, is involved in the entry and rate-limiting step of the urea cycle. The specific activity of CPS-I also was significantly decreased by DHEA, but serum urea levels were normal. To determine whether steroids other than DHEA also induced similar changes, mice were treated with various steroids for 14 days and, thereafter, liver proteins were evaluated by SDS-PAGE: estradiol-17 beta and isoandrosterone induced both the approximately 72 and approximately 28 kDa proteins, testosterone and androsterone induced the 28 kDa protein only, but etiocholanolone, pregnenolone and progesterone were without effect. The findings of this study serve to demonstrate that: (i) hepatic protein levels are affected by DHEA treatment of mice and rats; (ii) liver CPS-I activity is decreased significantly by DHEA treatment, but serum urea levels remain within the normal range; and (iii) sex steroids and some of their precursors, when administered per os, also alter liver protein levels. 相似文献
67.
Benigno?PadrónEmail author Manuel?Nogales Anna?Traveset Montserrat?Vilà Alejandro?Martínez-Abraín David?P.?Padilla Patricia?Marrero 《Biological invasions》2011,13(4):831-844
The success of many alien plant species depends on mutualistic relationships with other species. We describe the assemblage
of seed dispersers on three species of alien Opuntia invading Mediterranean and Macaronesian habitats, and examine the quality of such plant-animal interactions. We identified
vertebrates consuming O. maxima, O. dillenii and O. stricta fruits by direct observation and collecting droppings and pellets. Phenology of the alien species, as well as that of coexisting
native species, was monitored for an entire year. Germination tests of ingested and non-ingested seeds were performed both
in the greenhouse and in the field. Seed coat thickness and viability were also measured for all treatments. A great variety
of taxa, including reptiles, birds and mammals actively participate in the seed dispersal of Opuntia. Phenology of Opuntia fruits in Menorca and Tenerife overlaps with only a few native fleshy-fruited plants present in the study areas, which suggests
an advantage for the invader. Most seeds germinated during the second year of the experiment, independently of the effect
produced by the dispersers’ guts. We found great variation in the germination percentage of Opuntia after gut passage and in the effects of ingestion on seed coat thickness. Seed viability was somewhat reduced after gut passage
compared to manually depulped seeds. Our results show how different Opuntia species are integrated into native communities by means of mutualistic interactions, with both native and alien dispersers.
Although with heterogeneous effects, either type of disperser potentially contributes to the spread of these alien cacti in
the recipient areas. 相似文献
68.
Zhang Q Cigan AD Marrero L Lopreore C Liu S Ge D Savoie FH You Z 《Genesis (New York, N.Y. : 2000)》2011,49(2):75-82
The doublecortin (Dcx) gene encodes a microtubule-binding protein that was originally found in immature neurons. In this study, we used two mouse strains that express reporter genes (LacZ and enhanced green fluorescence protein, respectively) driven by the endogenous Dcx promoter. We found that Dcx was expressed in the mesenchymal cells in the mouse embryonic limb buds. A population of the mesenchymal cells continued Dcx expression after they differentiated into joint interzone cells and then articular chondrocytes. In contrast, the endochondral chondrocytes lost Dcx expression when the mesenchymal cells differentiated into endochondral chondrocytes. These data support a concept that the articular and endochondral chondrocytes originate from the same mesenchymal cells that express Dcx. In contrast to the notion that articular chondrocytes are derived from de-differentiated endochondral chondrocytes, our findings demonstrate that the lineages of articular and endochondral chondrocytes bifurcate at the stage of endochondral chondrogenesis. 相似文献
69.
MicroRNAs miR-122, miR-34a, miR-16 and miR-21 are commonly deregulated in liver fibrosis and hepatocellular carcinoma. This study examined whether circulating levels of these miRNAs correlate with hepatic histological disease severity in patients with chronic hepatitis C infection (CHC) or non-alcoholic fatty-liver disease (NAFLD) and can potentially serve as circulating markers for disease stage assessment. We first used an in vitro model of hepatitis C virus (HCV) infection to measure the extracellular levels of these four miRNAs. Whereas miR-21 extracellular levels were unchanged, extracellular levels of miR-122, miR-34a and to a lesser extent miR-16, steadily increased during the course of HCV infection, independently of viral replication and production. Similarly, in CHC patients, serum levels of miR-122, miR-34a and miR-16 were significantly higher than in control individuals, while miR-21 levels were unchanged. There was no correlation between the serum levels of any of these microRNAs and HCV viral loads. In contrast, miR-122 and miR-34a levels positively correlated with disease severity. Identical results were obtained in an independent cohort of CHC patients. We extended the study to patients with NAFLD. As observed in CHC patients, serum levels of miR-122, miR-34a and miR-16 were significantly higher in NAFLD patients than in controls, while miR-21 levels were unchanged. Again, miR-122 and miR-34a levels positively correlated with disease severity from simple steatosis to steatohepatitis. In both CHC and NAFLD patient groups, serum levels of miR-122 and miR-34a correlated with liver enzymes levels, fibrosis stage and inflammation activity. miR-122 levels also correlated with serum lipids in NAFLD patients. CONCLUSION: Serum levels of miR-34a and miR-122 may represent novel, noninvasive biomarkers of diagnosis and histological disease severity in patients with CHC or NAFLD. 相似文献
70.
Anne Lamontagne Ronald E. Long Mary Ann Comunale Julie Hafner Lucy Rodemich-Betesh Mengjun Wang Jorge Marrero Adrian M. Di Bisceglie Timothy Block Anand Mehta 《PloS one》2013,8(6)