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31.
Endostatin inhibits microvessel formation in the ex vivo rat aortic ring angiogenesis assay 总被引:21,自引:0,他引:21
Kruger EA Duray PH Tsokos MG Venzon DJ Libutti SK Dixon SC Rudek MA Pluda J Allegra C Figg WD 《Biochemical and biophysical research communications》2000,268(1):183-191
Endostatin has demonstrated potent antiangiogenic and antitumor activity in mouse models. We have investigated the ex vivo rat aortic ring assay and a human vein model to assess the biological activity of murine and human endostatin. Rat aortic rings were exposed to recombinant murine endostatin (Spodoptera frugipera; Calbiochem, San Diego, CA) or recombinant human endostatin (Pichia pastoris; EntreMed, Rockville, MD). After 5 days, murine endostatin (500 microgram/ml) demonstrated inhibition of microvessel outgrowth with dose-dependent effects (down to 16 microgram/ml). No significant inhibition was observed with human endostatin in the rat assay. Human endostatin at 250 and 500 microgram/ml inhibited outgrowths from human saphenous vein rings after a 14-day incubation. Electron microscopy assessed the formation of basal lamina, confirming that the microvessels were progenitors of patent vessels. Immunostaining for Factor VIII or CD34 demonstrated that the microvessel cells were endothelial. BrdU incorporation assays supported the presence of proliferating endothelial cells, correlating with neovascularization from the aortic wall. We conclude that the rat aortic ring assay confirms the antiangiogenic activity of murine but not human endostatin, suggesting that the model may have species specificity. However, the human form shows biological activity against human vascular tissue. 相似文献
32.
Ohdan H Swenson KG Kruger Gray HS Yang YG Xu Y Thall AD Sykes M 《Journal of immunology (Baltimore, Md. : 1950)》2000,165(10):5518-5529
Human natural Abs against Galalpha1-3Galbeta1-4GlcNAc (Gal) epitopes are a major barrier to xenotransplantation. Studies in this report, which use combined multiparameter flow cytometric sorting and enzyme-linked immunospot assay, demonstrate that anti-Gal IgM-producing cells are found exclusively in a small B cell subpopulation (i.e., CD21(-/low) IgM(high) B220(low) CD5(-) Mac-1(-) 493(-) cells) in the spleens of alpha1, 3-galactosyltransferase-deficient mice. All IgM-producing cells were detected in a similar splenic subpopulation of alpha1, 3-galactosyltransferase-deficient and wild-type mice. A higher frequency of B cells with anti-Gal surface IgM receptors was observed in the peritoneal cavity than in the spleen, but these did not actively secrete Abs, and showed phenotypic properties of B-1b cells (CD21(-/low) IgM(high) CD5(-) CD43(+) Mac-1(+)). However, these became Mac-1(-) and developed anti-Gal Ab-producing activity after in vitro culture with LPS. The splenic B cells with anti-Gal receptors consisted of both Mac-1(+) B-1b cells and Mac-1(-) B-1b-like cells. The latter comprised most anti-Gal IgM-producing cells. Our studies indicate that anti-Gal natural IgM Abs are produced by a B1b-like, Mac-1(-) splenic B cell population and not by plasma cells or B-1a cells. They are consistent with a model whereby B-1b cells lose Mac-1 expression upon Ag exposure and that these, rather than plasma cells, become the major IgM Ab-producing cell population. 相似文献
33.
Osteoblasts in culture can differentiate into mature mineralizing osteoblasts when stimulated with osteogenic agents. Clinical trials and in vivo animal studies suggest that specific polyunsaturated fatty acids (PUFAs) may benefit bone health. The aim of this study was to investigate whether arachidonic acid (AA) and docosahexaenoic acid (DHA) affect osteogenesis in osteoblasts and the transdifferentiation into adipocytes. Results from this study show that long‐term exposure to AA inhibited alkaline phosphatase (ALP) activity in these cells, which might be prostaglandin E2 (PGE2)‐mediated. DHA exposure also inhibited ALP activity which was evident after both short‐ and long‐term exposures. The mechanism whereby DHA inhibits ALP activity is not clear and needs to be investigated. Although long‐term exposure to PUFAs inhibited ALP activity, the mineralizing properties of these cells were not compromised. Furthermore, PUFA exposure did not induce adipocyte‐like features in these cells as evidenced by the lack of cytoplasmic triacylglycerol accummulation. More research is required to elucidate the cellular mechanisms of action of PUFAs on bone. Copyright © 2008 John Wiley & Sons, Ltd. 相似文献
34.
Background
Whilst the use of the mannitol/lactulose test for intestinal permeability has been long established it is not known whether the doses of these sugars modify transit time Similarly it is not known whether substances such as aspirin that are known to increase intestinal permeability to lactulose and mannitol and those such as ascorbic acid which are stated to be beneficial to gastrointestinal health also influence intestinal transit time.Methods
Gastric and intestinal transit times were determined with a SmartPill following consumption of either a lactulose mannitol solution, a solution containing 600 mg aspirin, a solution containing 500 mg of ascorbic acid or an extract of blackcurrant, and compared by doubly repeated measures ANOVA with those following consumption of the same volume of a control in a cross-over study in six healthy female volunteers. The dominant frequencies of cyclic variations in gastric pressure recorded by the Smartpill were determined by fast Fourier transforms.Results
The gastric transit times of lactulose mannitol solutions, of aspirin solutions and of blackcurrant juice did not differ from those of the control. The gastric transit times of the ascorbic acid solutions were significantly shorter than those of the other solutions. There were no significant differences between the various solutions either in the total small intestinal or colonic transit times. The intraluminal pHs during the initial quartiles of the small intestinal transit times were lower than those in the succeeding quartiles. This pattern did not vary with the solution that was consumed. The power of the frequencies of cyclic variation in intragastric pressure recorded by the Smartpill declined exponentially with increase in frequency and did not peak at the reported physiological frequencies of gastric contractile activity.Conclusions
Whilst the segmental residence times were broadly similar to those using other methods, the high degree of variation between subjects generally precluded the identification of all but gross variation between treatments. The lack of any differences between treatments in either total small or large intestinal transit times indicates that the solutions administered in the lactulose mannitol test of permeability had no consistent influence on the temporal pattern of absorption. The negatively exponential profile and lack of any peaks in the frequency spectra of cyclic variation in gastric intraluminal pressure that were consistent with reported physiological frequencies of contractile activity profile suggests that the principal source of this variation is stochastic likely resulting from the effects of external events occasioned by normal daily activities on intra-abdominal pressure.Trial Registration
Australian New Zealand Clinical Trials Registry ACTRN12615000596505 相似文献35.
Laishram R. Singh Sapna Gupta Nicholaas H. Honig Jan P. Kraus Warren D. Kruger 《PLoS genetics》2010,6(1)
Missense mutant proteins, such as those produced in individuals with genetic diseases, are often misfolded and subject to processing by intracellular quality control systems. Previously, we have shown using a yeast system that enzymatic function could be restored to I278T cystathionine β-synthase (CBS), a cause of homocystinuria, by treatments that affect the intracellular chaperone environment. Here, we extend these studies and show that it is possible to restore significant levels of enzyme activity to 17 of 18 (94%) disease causing missense mutations in human cystathionine β-synthase (CBS) expressed in Saccharomyces cerevisiae by exposure to ethanol, proteasome inhibitors, or deletion of the Hsp26 small heat shock protein. All three of these treatments induce Hsp70, which is necessary but not sufficient for rescue. In addition to CBS, these same treatments can rescue disease-causing mutations in human p53 and the methylene tetrahydrofolate reductase gene. These findings do not appear restricted to S. cerevisiae, as proteasome inhibitors can restore significant CBS enzymatic activity to CBS alleles expressed in fibroblasts derived from homocystinuric patients and in a mouse model for homocystinuria that expresses human I278T CBS. These findings suggest that proteasome inhibitors and other Hsp70 inducing agents may be useful in the treatment of a variety of genetic diseases caused by missense mutations. 相似文献
36.
Lauren K. Pietersen Patrick Govender Hendrik G. Kruger Glenn E. M. Maguire James Wesley-Smith Thavendran Govender 《International journal of peptide research and therapeutics》2010,16(4):291-295
By designing and coupling two functional peptides, CKAFKRK and C(KAFKRK)3 in differing ratios to the surface of gold nanoparticles (GNPs), we evaluated the effect of loading on aggregation and proteolysis. Transmission electron microscopy images of the functionalised GNPs indicated a direct relationship between the degree of aggregation of the particles and the extent of peptide loading: The greater the percentage of the C(KAFKRK)3 peptide, the greater the dispersion (less aggregation) of the peptide-capped GNPs. The functionalised GNPs were subjected to trypsin digestion over increasing time periods and it was found that the peptides were cleaved at the site of Lys and Arg. The extent of cleavage was analysed by mass spectrometry. The results indicate that the rate of enzymatic degradation was directly proportional to the extent of loading, such that the greater percentage of the C(KAFKRK)3 peptide, the greater the rate and efficiency of the cleavage. These results could be attributed to the different peptide distribution of the particles and the entropy of the peptides with varying peptide ratios. 相似文献
37.
Bruce T Martinez IB Maia Neto O Vicente AC Kruger RH Thompson FL 《Microbial ecology》2010,60(4):840-849
The aim of this study was to characterize the bacterial community diversity of the Brazilian Atlantic forest soil by means of both cultivation and 16S rRNA clone libraries. A collection of 86 representative isolates, obtained from six samples of Atlantic forest soils from the National Park of Serra dos Órgãos (PARNASO), belonged to the genera Arthrobacter, Bacillus, Burkholderia, Leifsonia, Paenibacillus, Pseudomonas, Ralstonia, Serratia, and Streptomyces according to the 16S rRNA sequences. Representative isolates from the different genera degraded cellulose and lignin. The culture-independent analysis based on 894 partial 16S rRNA gene sequences revealed that the most frequently retrieved groups belonged to the phyla Acidobacteria (29–54%), Proteobacteria (16–38%), and Verrucomicrobia (0.6–14%). The majority of the sequences (82.6%) were unidentified singletons and doubletons, indicating a high diversity of rare unique sequences. Chao1 estimator disclosed a high number of phyla (41–152) and species (263–446). This is the first survey on the Atlantic Forest soils using a combination of cultivation and culture-independent approaches. We conclude that the Brazilian Atlantic Forest soil represents a vast source of novel bacteria. 相似文献
38.
Lack of fructose 2,6‐bisphosphate compromises photosynthesis and growth in Arabidopsis in fluctuating environments
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Alistair J. McCormick Nicholas J. Kruger 《The Plant journal : for cell and molecular biology》2015,81(5):670-683
The balance between carbon assimilation, storage and utilisation during photosynthesis is dependent on partitioning of photoassimilate between starch and sucrose, and varies in response to changes in the environment. However, the extent to which the capacity to modulate carbon partitioning rapidly through short‐term allosteric regulation may contribute to plant performance is unknown. Here we examine the physiological role of fructose 2,6‐bisphosphate (Fru‐2,6‐P2) during photosynthesis, growth and reproduction in Arabidopsis thaliana (L.). In leaves this signal metabolite contributes to coordination of carbon assimilation and partitioning during photosynthesis by allosterically modulating the activity of cytosolic fructose‐1,6‐bisphosphatase. Three independent T‐DNA insertional mutant lines deficient in 6‐phosphofructo‐2‐kinase/fructose‐2,6‐bisphosphatase (F2KP), the bifunctional enzyme responsible for both the synthesis and degradation of Fru‐2,6‐P2, lack Fru‐2,6‐P2. These plants have normal steady‐state rates of photosynthesis, but exhibit increased partitioning of photoassimilate into sucrose and have delayed photosynthetic induction kinetics. The F2KP‐deficient plants grow normally in constant environments, but show reduced growth and seed yields relative to wildtype plants in fluctuating light and/or temperature. We conclude that Fru‐2,6‐P2 is required for optimum regulation of photosynthetic carbon metabolism under variable growth conditions. These analyses suggest that the capacity of Fru‐2,6‐P2 to modulate partitioning of photoassimilate is an important determinant of growth and fitness in natural environments. 相似文献
39.
Bahareh Honarparvar Sachin A Pawar Cláudio Nahum Alves Jer?nimo Lameira Glenn EM Maguire José Rogério A Silva Thavendran Govender Hendrik G Kruger 《Journal of biomedical science》2015,22(1)
Background
Novel pentacycloundecane (PCU)-lactone-CO-EAIS peptide inhibitors were designed, synthesized, and evaluated against wild-type C-South African (C-SA) HIV-1 protease. Three compounds are reported herein, two of which displayed IC50 values of less than 1.00 μM. A comparative MM-PB(GB)SA binding free energy of solvation values of PCU-lactam and lactone models and their enantiomers as well as the PCU-lactam-NH-EAIS and lactone-CO-EAIS peptide inhibitors and their corresponding diastereomers complexed with South African HIV protease (C-SA) was performed. This will enable us to rationalize the considerable difference between inhibitory concentration (IC50) of PCU-lactam-NH-EAIS and PCU-lactone-CO-EAIS peptides.Results
The PCU-lactam model exhibited more negative calculated binding free energies of solvation than the PCU-lactone model. The same trend was observed for the PCU-peptide inhibitors, which correspond to the experimental activities for the PCU-lactam-NH-EAIS peptide (IC50 = 0.076 μM) and the PCU-lactone-CO-EAIS peptide inhibitors (IC50 = 0.850 μM). Furthermore, a density functional theory (DFT) study on the natural atomic charges of the nitrogen and oxygen atoms of the three PCU-lactam, PCU-lactim and PCU-lactone models were performed using natural bond orbital (NBO) analysis. Electrostatic potential maps were also used to visualize the electron density around electron-rich regions. The asymmetry parameter (η) and quadrupole coupling constant (χ) values of the nitrogen and oxygen nuclei of the model compounds were calculated at the same level of theory. Electronic molecular properties including polarizability and electric dipole moments were also calculated and compared. The Gibbs theoretical free solvation energies of solvation (∆Gsolv) were also considered.Conclusions
A general trend is observed that the lactam species appears to have a larger negative charge distribution around the heteroatoms, larger quadrupole constant, dipole moment and better solvation energy, in comparison to the PCU-lactone model. It can be argued that these characteristics will ensure better eletronic interaction between the lactam and the receptor, corresponding to the observed HIV protease activities in terms of experimental IC50 data.Electronic supplementary material
The online version of this article (doi:10.1186/s12929-015-0115-5) contains supplementary material, which is available to authorized users. 相似文献40.
Jason A Roberts Michael S Roberts Andrew Semark Andrew A Udy Carl MJ Kirkpatrick David L Paterson Matthew J Roberts Peter Kruger Jeffrey Lipman 《BMC anesthesiology》2011,11(1):1-7