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981.
The reaction between [Fe2Ir2(CO)12]2− and diphenylacetylene in refluxing CH3CN yields the substituted cluster [Fe2Ir2(CO)10(PhC2Ph)]2− (1). In the crystals, the four metal atoms define a butterfly arrangement whose Ir-Ir hinge is parallel to the acetylenic C2 unit. The neutral triangular cluster [FeIr2(CO)9(PhC2Ph)] (2) is obtained by the treatment of 1 with acids at room temperature; in this 48 valence electrons species, the C-C and the Ir-Ir bonds are also parallel, in the coordination mode.The cluster [Fe2Rh(CO)10] reacts with diphenylacetylene in refluxing THF yielding [Fe2Rh(CO)8(PhC2Ph)] (3). In this 46 C.V.E.’s cluster, the C2 unit is perpendicular to the Fe-Fe edge, exemplifying the bonding mode. According to 13C NMR spectra, the structure of the three clusters is maintained in solution. Electrochemical investigations show that the one-electron oxidation of [Fe2Ir2(CO)10(L)]2− (L = 2CO, PhC2Ph) as well as the one-electron reduction of [Fe2Rh(CO)8(PhC2Ph)] only generates the respective short lived products.  相似文献   
982.
Inhibitor of apoptosis proteins (IAPs) such as XIAP, cIAP1, and cIAP2 are upregulated in many cancer cells. Several compounds targeting IAPs and inducing cell death in cancer cells have been developed. Some of these are synthesized mimicking the N-terminal tetrapeptide sequence of Smac/DIABLO, the natural endogenous IAPs inhibitor. Starting from such conceptual design, we generated a library of 4-substituted azabicyclo[5.3.0]alkane Smac-mimetics. Here we report the crystal structure of the BIR3 domain from XIAP in complex with Smac037, a compound designed according to structural principles emerging from our previously analyzed XIAP BIR3/Smac-mimetic complexes. In parallel, we present an in silico docking analysis of three Smac-mimetics to the BIR3 domain of cIAP1, providing general considerations for the development of high affinity lead compounds targeting three members of the IAP family.  相似文献   
983.
The studies and the researches carried out in the last years on the Palaeolithic site of Isernia La Pineta have brought to consider in new way the activities realized by the human group that lived the basin of Isernia during the Middle Pleistocene offering an important key of interpretation of the behavioural strategies of the prehistoric man. The analysis of the exploitation of the raw material has confirmed the presence on the site of two different lithotypes: flint and limestone; the lithological dichotomy is related to the functional dichotomy of the raw material that seems to have conditioned the activities of the human group in different areas of the site. The necessity to deepen the study on the limestone has derived from the evidence brought to light in the last excavation campaigns of a remarkable concentration of the flaked limestone pebbles and the flake scars in some areas of the explored archeosurfaces, particularly on the 3a and on the overlooking layers. The present study has the purpose to explain the characteristics of the limestone finds both in reference to the raw material and to its state of preservation both to the technotypological evidences and its spatial distribution with the purpose to better understand the modalities of the exploitation of the raw material. The information collected until today have permitted to obtain a precise knowledge of the environmental context and the territorial resources exploited by the human group showing an opportunistic capability to find the most advantageous behavioural solution for the necessities of subsistence.  相似文献   
984.
Pseudomonas pseudoalcaligenes KF707 is naturally resistant to the toxic metalloid tellurite, but the mechanisms of resistance are not known. In this study we report the isolation of a KF707 mutant (T5) with hyperresistance to tellurite. In order to characterize the bacterial response and the pathways leading to tolerance, we utilized Phenotype MicroArray technology (Biolog) and a metabolomic technique based on nuclear magnetic resonance spectroscopy. The physiological states of KF707 wild-type and T5 cells exposed to tellurite were also compared in terms of viability and reduced thiol content. Our analyses showed an extensive change in metabolism upon the addition of tellurite to KF707 cultures as well as different responses when the wild-type and T5 strains were compared. Even in the absence of tellurite, T5 cells displayed a “poised” physiological status, primed for tellurite exposure and characterized by altered intracellular levels of glutathione, branched-chain amino acids, and betaine, along with increased resistance to other toxic metals and metabolic inhibitors. We conclude that hyperresistance to tellurite in P. pseudoalcaligenes KF707 is correlated with the induction of the oxidative stress response, resistance to membrane perturbation, and reconfiguration of cellular metabolism.  相似文献   
985.
Sphingolipids such as ceramides (Cers) play important roles in cell proliferation, apoptosis, and cell cycle regulation. An increased Cer level is linked to the cytotoxic effects of several chemotherapeutics. Various selective cyclooxygenase-2 (COX-2) inhibitors induce anti-proliferative effects in tumor cells. We addressed the possible interaction of the selective COX-2 inhibitors, coxibs, with the sphingolipid pathway as an explanation of their anti-proliferative effects. Sphingolipids were measured using liquid chromatography tandem mass spectrometry. Treatment of various cancer cell lines with celecoxib significantly increased sphinganine, C(16:0)-, C(24:0)-, C(24:1)-dihydroceramide (dhCer) and led to a depletion of C(24:0)-, C(24:1)-Cer in a time- and concentration-dependent manner, whereas other coxibs had no effect. Using (13)C,(15)N-labeled l-serine, we demonstrated that the augmented dhCers after celecoxib treatment originate from de novo synthesis. Celecoxib inhibited the dihydroceramide desaturase (DEGS) in vivo with an IC(50) of 78.9 +/- 1.5 muM and increased total Cer level about 2-fold, indicating an activation of sphingolipid biosynthesis. Interestingly, inhibition of the sphingolipid biosynthesis by specific inhibitors of l-serine palmitoyltransferase diminished the anti-proliferative potency of celecoxib. In conclusion, induction of de novo synthesis of sphingolipids and inhibition of DEGS contribute to the anti-proliferative effects of celecoxib.  相似文献   
986.
We have previously shown that Fhit tumor suppressor protein interacts with Hsp60 chaperone machinery and ferredoxin reductase (Fdxr) protein. Fhit-effector interactions are associated with a Fhit-dependent increase in Fdxr stability, followed by generation of reactive oxygen species and apoptosis induction under conditions of oxidative stress. To define Fhit structural features that affect interactions, downstream signaling, and biological outcomes, we used cancer cells expressing Fhit mutants with amino acid substitutions that alter enzymatic activity, enzyme substrate binding, or phosphorylation at tyrosine 114. Gastric cancer cell clones stably expressing mutants that do not bind substrate or cannot be phosphorylated showed decreased binding to Hsp60 and Fdxr and reduced mitochondrial localization. Expression of Fhit or mutants that bind interactor proteins results in oxidative damage and accumulation of cells in G2/M or sub-G1 fractions after peroxide treatment; noninteracting mutants are defective in these biological effects. Gastric cancer clones expressing noncomplexing Fhit mutants show reduction of Fhit tumor suppressor activity, confirming that substrate binding, interaction with heat shock proteins, mitochondrial localization, and interaction with Fdxr are important for Fhit tumor suppressor function.Fhit protein is a powerful tumor suppressor that is frequently lost or reduced in cancer cells because of rearrangement of the exquisitely DNA damage-sensitive fragile FHIT gene. Restoration of Fhit expression suppresses tumorigenicity of cancer cells of various types, and the ability to induce apoptosis in cancer cells in vitro is reduced by specific Fhit mutations (1, 2).Through studies of signal pathways affected by Fhit expression, by searches for Fhit protein effectors, and by in vitro analyses of Fhit activity, we and others have defined Fhit enzymatic activity in vitro (3), apoptotic activity in cells and tumors (46), and most recently identification of a Fhit protein complex that affects Fhit stability, mitochondrial localization, and interaction with ferredoxin reductase (Fdxr)5 (7). The complex includes Hsp60 and Hsp10 that mediate Fhit stability and may affect import into mitochondria, where Fhit interacts with Fdxr, which is responsible for transferring electrons from NADPH to cytochrome P450 via ferredoxin. Virally mediated Fhit restoration in Fhit-deficient cancer cells increases production of intracellular reactive oxygen species (ROS), followed by increased apoptosis of cancer cells under oxidative stress conditions; conversely, Fhit-negative cells escape apoptosis, likely carrying oxidative DNA damage that contributes to accumulation of mutations.The Fhit protein sequence, showing high homology to the histidine triad (HIT) family of proteins, suggested that the protein product would hydrolyze diadenosine tetraphosphate or diadenosine triphosphate (Ap3A) (8), and in vitro studies showed that Ap3A was cleaved into ADP and AMP by Fhit. The catalytic histidine triad within Fhit was essential for catalytic activity (3), and a Fhit mutant that substituted Asn for His at the central histidine (H96N mutant) was catalytically inactive, although it bound substrate well (3). Early tumor suppression studies showed that cancer cells stably transfected with wild type (WT) or H96N mutant Fhit were suppressed for tumor growth in nude mice. This suggested the hypothesis that the Fhit-substrate complex sends the tumor suppression signal (9, 10). To test this hypothesis, a series of FHIT alleles was designed to reduce substrate-binding and/or hydrolytic rates and was characterized by quantitative cell-death assays on cancer cells virally infected with each allele. The allele series covered defects as great as 100,000-fold in kcat and increases as large as 30-fold in Km. Mutants with 2–7-fold increases in Km had significantly reduced apoptotic indices and the mutant with a 30-fold increase in Km retained little apoptotic function. Thus, the proapoptotic function of Fhit, which is likely associated with tumor suppressor function, is limited by substrate binding and is unrelated to substrate hydrolysis (11).Fhit, a homodimeric protein of 147 amino acids, is a target of tyrosine phosphorylation by the Src family protein kinases, which can phosphorylate Tyr-114 of Fhit in vitro and in vivo (12). After co-expression of Fhit with the Elk tyrosine kinase in Escherichia coli to generate phosphorylated forms of Fhit, unphosphorylated, mono-, and diphosphorylated Fhit were purified, and enzyme kinetics studies showed that monophosphorylated Fhit exhibited monophasic kinetics with Km and kcat values ∼2- and ∼7-fold lower, respectively, than for unphosphorylated Fhit. Diphosphorylated Fhit exhibited biphasic kinetics; one site had Km and kcat values ∼2- and ∼140-fold lower, respectively, than for unphosphorylated Fhit; the second site had a Km ∼60-fold higher and a kcat ∼6-fold lower than for unphosphorylated Fhit (13). Thus, it was possible that the alterations in Km and kcat values for phosphorylated forms of Fhit might favor formation and lifetime of the Fhit-Ap3A complex and enhance tumor suppressor activity (see
Fhit forms
Kinetic parameters
% Sub-G1
Direct binding
Subcellular location
Co-IP in vivo
8-OHdG
Apoptosis
Tumor suppressor
Km (mm)kcat (s–1)A549MKN74Hsp60FdxrHsp60Fdxr
Fhit WT 1.6 +/– 0.19 2.7 +/– 0.95 43 24 Yes Yes Cyt & mito Yes Yes Yes Yes Yes
Catalyt mutants
   H96D Up 2-fold Down >2 × 104 29 NT NT NT Cyt & mito Yes Yes NT Yes NT
   H96N Up 2-fold Down >5 × 105 31 14.4 NT NT Cyt & mito Yes Yes Yes Yes Yes
Loop mutants
   Y114A Up 23-fold Down 2-fold 3.7 NT NT NT Cyt +/– +/– +/– No No
   Y114D NT NT 2.9 6 NT NT Cyt +/– +/– No –/+
   Y114E NT NT NT NT NT NT Cyt & mito –/+ –/+ No NT
   Y114F Up 5-fold Up 1.1-fold 11.5 3 NT NT Cyt & mito –/+ –/+ No No
   Y114W Up 5-fold Up 1.4-fold NT NT NT NT Cyt & mito –/+ NT NT
   del113–117 Up 10-fold Down 38-fold 5 NT NT NT NT NT NT No NT
Other mutants
   L25W Up 7-fold Down 4-fold 15 NT NT NT Cyt NT –/+
   I10W,L25W Up 32-fold Down 6-fold 11 NT NT NT NT NT NT NT NT NT
   F5W Up 3.3 fold NT NT 5 NT NT NT NT NT +/– No NT
Purified pFhit
   pFhit Down 0.4-fold Down 7-fold NA NA –/+ Yes NA NA NA NA NA NA
   ppFhit Down 0.4-fold Down > 100-fold NA NA –/+ Yes NA NA NA NA NA NA
Up 60-fold Down 6-fold
Open in a separate windowTo explore the in vivo importance of the Tyr-114 phosphorylation site and define Fhit-mediated signaling events, Semba et al. (14) compared the differential biological effects of Ad-FHIT WT and Ad-FHIT Tyr-114 mutant expression in human lung cancer cells. Caspase-dependent apoptosis was effectively induced only by WT Fhit protein. However, the biological significance of phosphorylation at Tyr-114 has been difficult to study because the endogenous phosphorylated forms have very short half-lives; activation of epidermal growth facto receptor family members induces Fhit phosphorylation by Src and proteasome degradation of phosphorylated Fhit (15).Although there are possible connections among the various pathways known to be altered in Fhit-deficient cells, apoptosis, DNA damage-response checkpoint activation, ROS production, and related biological effects of Fhit loss or overexpression, details of the pathway(s) leading from Fhit overexpression to cell death and tumor suppression have not been delineated. Now that a Fhit signaling complex has been identified, we set out to examine which structural features of Fhit protein might participate in individual steps of the pathway leading from Fhit overexpression through complex formation, subcellular localization, interaction with mitochondrial Fdxr, DNA damage induction, cell cycle changes, apoptosis, and ultimately tumor suppression. The underlying hypotheses were as follows: substrate-binding mutants would behave similarly to WT; nonsubstrate-binding mutants would be defective in some step of the pathway, perhaps complexing with heat shock proteins or Fdxr or perhaps induction of DNA damage; and Tyr-114 mutants, which also affect formation or stability of the enzyme-substrate complex, would also be defective in executing some step of the Fhit overexpression pathway to cell death. One goal was to identify specific mutants that exhibited deficiency in specific steps of the pathway, so that such mutants could be used to dissect each step in more detail. Using in vitro Fhit and Fhit-effector protein interactions, we aimed to determine the following: 1) which proteins of the complex interact directly with Fhit, and 2) the biological role of these interactions in vivo. Using cancer cells expressing exogenous WT and mutant Fhit proteins, we were able to examine the structural features of Fhit that affect the direct interaction with its effectors, participate in ROS production, and are necessary for tumor suppression activity.  相似文献   
987.
A food safety control low mass-range proteomics platform for the detection of illicit treatments in veal calves by MALDI-TOF-MS serum profiling     
Lorenza Della Donna  Maurizio Ronci  Paolo Sacchetta  Carmine Di Ilio  Bartolomeo Biolatti  Giorgio Federici  Carlo Nebbia  Andrea Urbani Professor 《Biotechnology journal》2009,4(11):1596-1609
Performance enhancing agents (PEAs) are illegally used in cattle and other meat producing species to increase food conversion and lean meat production. Due to the very short breeding cycle, veal calves represent the meat producing bovine category mostly subjected to illicit treatments. These chemical agents are difficult to detect by conventional analytical approaches due to the employment of synergistic formulations at very low dosage and given the use of uncharacterized novel compounds. Such a scenario has fostered a strong interest in the discovery of functional molecular biomarkers for the detection of growth promoting agents in meat producing species. A multivariate MALDI-TOF-MS proteomics platform has been developed using bovine serum samples. Analytical performances have been thoroughly evaluated in order to enable reproducible profiles from 10 μL sera samples. We propose univariate and multivariate discrimination models capable to identify calves undergoing illicit treatments. In particular, we found a strong discrimination power associated with a polypeptide fragment from β2-glycoprotein-I. We provide a fundamental proof of concept in the potential application of MALDI-TOF-MS proteomics profiling in the food safety control.  相似文献   
988.
Computing parametric beta diversity with unequal plot weights: a solution based on resampling methods     
Carlo Ricotta 《Theoretical Ecology》2009,2(1):13-17
Jost (Ecology, 88:2427–2439, 2007) recently showed that the Shannon diversity is the only standard diversity measure that can be partitioned into meaningful independent alpha and beta components when plot weights are unequal. This conclusion is very disappointing if one wants to calculate the beta diversity of unequal weighted plots using a parametric measure with varying sensitivities to the occurrence of rare and abundant species. To overcome this impasse, at least partially, in this paper, I propose a parametric measure of beta diversity that is based on the combination of Shannon’s entropy with Hurlbert’s ‘expected species diversity’. Unlike most parametric measures of diversity, the proposed index has a clear probabilistic interpretation, allowing at the same time a multiplicative partition of diversity into independent alpha and beta components for unequally weighted plots.  相似文献   
989.
Homeodomain Interacting Protein Kinase 2 Activation Compromises Endothelial Cell Response to Laminar Flow: Protective Role of p21waf1,cip1,sdi1     
Stefania Mattiussi  Chiara Lazzari  Silvia Truffa  Annalisa Antonini  Silvia Soddu  Maurizio C. Capogrossi  Carlo Gaetano 《PloS one》2009,4(8)

Background

In the cardiovascular system, laminar shear stress (SS) is one of the most important source of endothelial protecting signals. Physical and chemical agents, however, including ionising radiations and anticancer drugs, may injure endothelial cells determining an increase in oxidative stress and genotoxic damage. Whether the SS protective function remains intact in the presence of strong oxidants or DNA damage is currently unclear.

Methods and Results

To investigate this aspect a series of experiments were performed in which HUVEC were exposed to sub-lethal doses of the radio-mimetic compound Bleomycin (Bleo; 10 µg/ml) which generated free radicals (ROS) without significantly compromising cell survival. Remarkably, the application of a SS of 12 dyne/cm2 did not protect endothelial cells but markedly accelerated apoptosis compared to controls kept in static culture and in the presence of Bleo. Experiments with the inducible nitric oxide synthase (iNOS) inhibitor GW274150 significantly reduced the SS-dependent apoptosis indicating that the production of NO was relevant for this effect. At molecular level, the ataxia-telangectasia-mutated (ATM) kinase, the homeodomain-interacting protein kinase-2 (HIPK2) and p53 were found activated along a pro-apoptotic signalling pathway while p21waf1,cip1,sdi1 was prevented from its protective action. RNA interference experiments revealed that HIPK2 and p53 were both important for this process, however, only the forced expression p21waf1,cip1,sdi1 fully restored the SS-dependent pro-survival function.

Conclusions

This study provides the first evidence that, in the presence of genotoxic damage, laminar flow contributes to endothelial toxicity and death and identifies molecular targets potentially relevant in endothelial dysfunction and cardiovascular disease pathogenesis.  相似文献   
990.
Nitric Oxide Administration Using an Oxygen Hood: A Pilot Trial     
Namasivayam Ambalavanan  George T. El-Ferzli  Claire Roane  Robert Johnson  Waldemar A. Carlo 《PloS one》2009,4(2)

Background

We have shown earlier that inhaled nitric oxide (iNO) administered by oxygen hood reduces pulmonary hypertension in an animal model (J Perinatol 2002; 22:50-6). Our objective in this study was to determine feasibility of iNO by oxygen hood in neonates with elevated alveolar-arterial oxygen gradients (A-aDO2).

Methods/Principal Findings

Masked randomized controlled pilot trial. Inclusion criteria were: gestation≥34 weeks, age<7 days, with post-ductal arterial line, and A-aDO2 400–600. Infants were randomized to study gas (iNO 20 ppm or equivalent O2 flow) for 1 hr which was then weaned over the next 4 hours. Primary outcome was PaO2 one hour post-randomization. Four infants each were randomized to iNO or O2 (controls). Two of the four infants given iNO had an increase in PaO2 of >100 torr, while oxygenation was unchanged in the controls. Methemoglobinemia and other adverse effects were not noted in any infant. Environmental levels of NO and NO2 were minimal (<1 ppm) at >0.3 m from the hood.

Conclusions

Administration of iNO by oxygen hood is feasible. Larger randomized controlled trials are required to measure the efficacy and determine an appropriate target population for this technique.

Trial Registration

ClinicalTrials.gov NCT00041548  相似文献   
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