首页 | 本学科首页   官方微博 | 高级检索  
文章检索
  按 检索   检索词:      
出版年份:   被引次数:   他引次数: 提示:输入*表示无穷大
  收费全文   926篇
  免费   51篇
  国内免费   1篇
  2023年   3篇
  2022年   4篇
  2021年   18篇
  2020年   9篇
  2019年   17篇
  2018年   22篇
  2017年   19篇
  2016年   29篇
  2015年   42篇
  2014年   37篇
  2013年   45篇
  2012年   80篇
  2011年   71篇
  2010年   46篇
  2009年   35篇
  2008年   70篇
  2007年   68篇
  2006年   49篇
  2005年   57篇
  2004年   66篇
  2003年   49篇
  2002年   62篇
  2001年   3篇
  2000年   5篇
  1999年   4篇
  1998年   9篇
  1997年   11篇
  1996年   7篇
  1995年   8篇
  1994年   11篇
  1993年   6篇
  1992年   6篇
  1991年   1篇
  1990年   2篇
  1989年   2篇
  1988年   1篇
  1986年   1篇
  1984年   1篇
  1982年   2篇
排序方式: 共有978条查询结果,搜索用时 31 毫秒
71.
Evidence is presented that RSK1 (ribosomal S6 kinase 1), a downstream target of MAPK (mitogen-activated protein kinase), directly phosphorylates nNOS (neuronal nitric oxide synthase) on Ser847 in response to mitogens. The phosphorylation thus increases greatly following EGF (epidermal growth factor) treatment of rat pituitary tumour GH3 cells and is reduced by exposure to the MEK (MAPK/extracellular-signal-regulated kinase kinase) inhibitor PD98059. Furthermore, it is significantly enhanced by expression of wild-type RSK1 and antagonized by kinase-inactive RSK1 or specific reduction of endogenous RSK1. EGF treatment of HEK-293 (human embryonic kidney) cells, expressing RSK1 and nNOS, led to inhibition of NOS enzyme activity, associated with an increase in phosphorylation of nNOS at Ser847, as is also the case in an in vitro assay. In addition, these phenomena were significantly blocked by treatment with the RSK inhibitor Ro31-8220. Cells expressing mutant nNOS (S847A) proved resistant to phosphorylation and decrease of NOS activity. Within minutes of adding EGF to transfected cells, RSK1 associated with nNOS and subsequently dissociated following more prolonged agonist stimulation. EGF-induced formation of the nNOS-RSK1 complex was significantly decreased by PD98059 treatment. Treatment with EGF further revealed phosphorylation of nNOS on Ser847 in rat hippocampal neurons and cerebellar granule cells. This EGF-induced phosphorylation was partially blocked by PD98059 and Ro31-8220. Together, these data provide substantial evidence that RSK1 associates with and phosphorylates nNOS on Ser847 following mitogen stimulation and suggest a novel role for RSK1 in the regulation of nitric oxide function in brain.  相似文献   
72.
Aoki J  Inoue A  Makide K  Saiki N  Arai H 《Biochimie》2007,89(2):197-204
Phospholipase A1 (PLA1) is an enzyme that hydrolyzes phospholipids and produces 2-acyl-lysophospholipids and fatty acids and is conserved in a wide range of organisms. Mammals have several enzymes that exhibit PLA1 activity in vitro. The extracellular PLA1s include phosphatidylserine (PS)-specific PLA1 (PS-PLA1), membrane-associated phosphatidic acid (PA)-selective PLA1s (mPA-PLA1alpha and mPA-PLA1beta), hepatic lipase (HL), endothelial lipase (EL) and pancreatic lipase-related protein 2 (PLRP2), all of which belong to the pancreatic lipase gene family. The former three PLA1s differ from other members in their substrate specificities, structural features and gene organizations, and form a subfamily in the pancreatic lipase gene family. PS-PLA1, mPA-PLA1alpha and mPA-PLA1beta exhibit only PLA1 activity, while HL, EL and PLRP2 show triacylglycerol-hydrolyzing activity in addition to PLA1 activity. The tertiary structures of lipases have two surface loops, the lid and the beta9 loop. The lid and the beta9 loop cover the active site in its closed conformation. An alignment of amino acid sequences of the pancreatic lipase gene family members revealed two molecular characteristics of PLA1s in the two surface loops. First, lipase members exhibiting PLA1 activity (PS-PLA1, mPA-PLA1alpha and mPA-PLA1beta, EL, guinea pig PLRP2 and PLA1 from hornet venom (DolmI)) have short lids. Second, PS-PLA1, mPA-PLA1alpha, mPA-PLA1beta and DolmI, which exhibit only PLA(1) activity, have short beta9 loops. Thus, the two surface loops appear to be involved in the ligand recognition. PS-PLA1 and mPA-PLA1s specifically hydrolyze PS and PA, respectively, producing their corresponding lysophospholipids. Lysophosphatidylserine and lysophosphatidic acid have been defined as lipid mediators with multiple biological functions. Thus, these PLA1s have a role in the production of these lysophospholipid mediators.  相似文献   
73.
Bone marrow-derived mesenchymal stem cells (MSCs) have been reported to prevent the development of liver fibrosis in a number of pre-clinical studies. Marked changes in liver histopathology and serological markers of liver function have been observed without a clear understanding of the therapeutic mechanism by which stem cells act. We sought to determine if MSCs could modulate the activity of resident liver cells, specifically hepatic stellate cells (SCs) by paracrine mechanisms using indirect cocultures. Indirect coculture of MSCs and activated SCs led to a significant decrease in collagen deposition and proliferation, while inducing apoptosis of activated SCs. The molecular mechanisms underlying the modulation of SC activity by MSCs were examined. IL-6 secretion from activated SCs induced IL-10 secretion from MSCs, suggesting a dynamic response of MSCs to the SCs in the microenvironment. Blockade of MSC-derived IL-10 and TNF-alpha abolished the inhibitory effects of MSCs on SC proliferation and collagen synthesis. In addition, release of HGF by MSCs was responsible for the marked induction of apoptosis in SCs as determined by antibody-neutralization studies. These findings demonstrate that MSCs can modulate the function of activated SCs via paracrine mechanisms provide a plausible explanation for the protective role of MSCs in liver inflammation and fibrosis, which may also be relevant to other models of tissue fibrosis.  相似文献   
74.
Natural antibiotic polyene amides such as myxalamides are potent inhibitors of mitochondrial complex I. Because of the significant instability of this series of compounds due to an extended pi-conjugation skeleton, a detailed characterization of their inhibitory action has not been performed. To elucidate the action mechanism as well as binding manner of polyene amides with complex I, identification of the roles of each functional group in the inhibitory action is needed. We here synthesized a series of amide analogues and carried out structure-activity studies with bovine heart mitochondrial complex I. With respect to the left-hand portion, the natural pi-conjugation skeleton common to many natural products is not required for the inhibition and can be substituted with a simpler substructure such as a conjugated diene. The geometry and shape of the left-hand portion were shown to be important for the inhibition, suggesting that this portion may bind to a narrow hydrophobic pocket in the enzyme rather than merely partitioning into the lipid membrane phase. Concerning the right-hand portion of the inhibitor, the presence of the 2-methyl, amide NH, and (S)-1'-methyl groups was crucial for the activity, suggesting that both methyl groups neighboring the amide group finely adjust the hydrogen-bonding ability of the amide group. In contrast, modifications of the 2'-OH group did not significantly influence the activity, suggesting that the role of this functional group is not to serve as a hydrogen bond donor to the enzyme but to act as a hydrophilic anchor directing the right-hand portion at or near the membrane surface. Detailed characterization of the action mechanism indicated that the polyene amides share a common binding domain with other complex I inhibitors, though their binding position (or manner) within the domain may differ considerably from that of other inhibitors.  相似文献   
75.
A flow-based immunoassay system utilizing secondary-antibody coated microbeads and Cy5-secondary antibody for signal production was successfully developed to quantitate target bacteria with a kinetic exclusion assay (KinExA 3000 Instrument). It directly measured the concentration of unliganded antibody separated from the equilibrated mixture of antibody and bacteria through a 0.2 microm polyethersulfone membrane, enabling it to quantify the concentration of bacteria. The novel method demonstrated the qualities of rapidness, sensitivity, high accuracy and reproducibility, and ease to perform. Detection of Pseudomonas aeruginosa and Staphylococcus aureus was accomplished with low detection limits of 4.10 x 10(6) and 5.20 x l0(4)cells/mL, respectively, with an assay time of less than 15 min. The working ranges for quantification were 4.10 x l0(6) to 1.64 x l0(10)cells/mL for P. aeruginosa, and 5.20 x l0(4) to 1.04 x l0(9)cells/mL for S. aureus. It yielded an assay with at least 10-fold greater sensitivity than ELISA and could correctly assess the concentration of predominant bacterium spiked in the mixture of P. aeruginosa and S. aureus. With this reliable platform, the average amount of antibody bound by one cell in the maximum capability could be further provided: (1.6-2.5) x l0(5) antibodies for one P. aeruginosa cell and (2.2-2.7) x l0(8) antibodies for one S. aureus cell. The KinExA system is flexible to determine different kinds of bacteria conveniently by using anti-mouse IgG as the same immobilizing agent. However, a higher specificity of the antibodies to the target bacteria will be required for the use of this system with higher detection sensitivity.  相似文献   
76.
Several reports have documented a better prognosis for HIV‐1‐infected patients co‐infected with GBV‐C, while other reports have contradicted such findings with the result that this issue remains controversial. We attempted to clarify the complicated status of the effect of GBV‐C co‐infection on HIV‐1‐infected patients. GBV‐C RNA was detected in 37 samples in 182 HIV‐1‐infected patients (20.3%) using RT/nested PCR. Of these, 3 were determined to be GBV‐C genotype 1, 12 were genotype 2, and the remaining 22 were genotype 3. The GBV‐C viral load quantified by real‐time PCR ranged from 7.8 × 103 to 3.3 × 106 copies/ml. Weakly negative correlation was observed between GBV‐C viral load and HIV‐1 viral load in 19 HAART‐naïve patients, indicating that a higher GBV‐C viral load is associated with a greater suppression of HIV‐1 replication. A previously published in vitro study suggested that GBV‐C infection would induce up‐regulation of RANTES, leading to suppression of HIV‐1 replication. However, in our present study, the blood RANTES level was significantly lower in the GBV‐C co‐infected group than in the uninfected group (190–9,959 vs. 264–31,038 pg/ml, P=0.004). Our results suggested that a suppression of HIV‐1 replication by GBV‐C co‐infection is not mediated by up‐regulated RANTES, and thus call for another as yet unknown factor.  相似文献   
77.
Focusing on the final step of osteoclastogenesis, we studied cell fusion from tartrate-resistant acid phosphatase (TRAP)-positive mononuclear cells into multinuclear cells. TRAP-positive mononuclear cells before generation of multinuclear cells by cell fusion were differentiated from RAW264.7 cells by treatment with receptor activator of nuclear factor kappa B ligand (RANKL), and then the cells were treated with lipopolysaccharide (LPS), followed by culturing for further 12 h. LPS-induced cell fusion even in the absence of RANKL. Similarly, tumor necrosis factor (TNF)-alpha and peptidoglycan (PGN) induced cell fusion, but M-CSF did not. The cell fusion induced by RANKL, TNF-alpha, and LPS was specifically blocked by osteoprotegerin (OPG), anti-TNF-alpha antibody, and polymyxin B, respectively. LPS- and PGN-induced cell fusion was partly inhibited by anti-TNF-alpha antibody but not by OPG. When TRAP-positive mononuclear cells fused to yield multinuclear cells, phosphorylation of Akt, Src, extracellular signal-regulated kinase (ERK), p38MAPK (p38), and c-Jun NH2-terminal kinase (JNK) was observed. The specific chemical inhibitors LY294002 (PI3K), PP2 (Src), U0126 (MAPK-ERK kinase (MEK)/ERK), and SP600125 (JNK) effectively suppressed cell fusion, although SB203580 (p38) did not. mRNA of nuclear factor of activated T-cells c1 (NFATc1) and dendritic cell-specific transmembrane protein (DC-STAMP) during the cell fusion was quantified, however, there was no obvious difference among the TRAP-positive mononuclear cells treated with or without M-CSF, RANKL, TNF-alpha, LPS, or PGN. Collectively, RANKL, TNF-alpha, LPS, and PGN induced cell fusion of osteoclasts through their own receptors. Subsequent activation of signaling pathways involving PI3K, Src, ERK, and JNK molecules was required for the cell fusion. Although DC-STAMP is considered to be a requisite for cell fusion of osteoclasts, cell fusion-inducing factors other than DC-STAMP might be necessary for the cell fusion.  相似文献   
78.
The chemokine receptor genes of the CCR cluster on human chromosome 3p21 play important roles in humoral and cellular immune responses. Several of these receptors have been shown to influence human immunodeficiency virus infection and progression to AIDS, and their homologues may play a role in feline immunodeficiency virus infection. We report the isolation and sequencing of a 150-kb domestic cat BAC clone containing the feline CCR genes CCR1, CCR2, CCR3, and CCR5 to further analyze these four receptor genes within the family Felidae. Comparative and phylogenetic analyses reveal evidence for historic gene conversion between the adjacent CCR2 and CCR5 genes in the Felidae and in three independent mammalian orders (Primates, Cetartiodactyla, and Rodentia), resulting in higher than expected levels of sequence similarity between the two paralogous genes within each order. The gene conversion was restricted to the structural (transmembrane) domains of the CCR2 and CCR5 genes. We also discovered a recent gene conversion event between the third extracellular loop of CCR2 and CCR5 genes that was fixed in Asian lions and found at low frequency in African lions (Panthera leo), suggesting that this domain may have an important functional role. Our results suggest that ongoing parallel gene conversion between CCR2 and CCR5 promotes receptor heterodimerization in independent evolutionary lineages and offers an effective adaptive strategy for gene editing and coevolution among interactive immune response genes in mammals.  相似文献   
79.
2,5-Diketo-d-gluconate (2,5DKG) is a compound that can be the intermediate for d-tartrate and also vitamin C production. Although Gluconobacter oxydans NBRC3293 produces 2,5DKG from d-glucose via d-gluconate and 2-keto-d-gluconate (2KG), with accumulation of the product in the culture medium, the efficiency of 2,5DKG production is unsatisfactory because there is a large amount of residual d-gluconate at the end of the biotransformation process. Oxidation of 2KG to 2,5DKG is catalyzed by a membrane-bound flavoprotein-cytochrome c complex: 2-keto-gluconate dehydrogenase (2KGDH). Here, we studied the kgdSLC genes encoding 2KGDH in G. oxydans NBRC3293 to improve 2,5DKG production by Gluconobacter spp. The kgdS, kgdL, and kgdC genes correspond to the small, large, and cytochrome subunits of 2KGDH, respectively. The kgdSLC genes were cloned into a broad-host-range vector carrying a DNA fragment of the putative promoter region of the membrane-bound alcohol dehydrogenase gene of G. oxydans for expression in Gluconobacter spp. According to our results, 2KGDH that was purified from the recombinant Gluconobacter cells showed characteristics nearly the same as those reported previously. We also expressed the kgdSLC genes in a mutant strain of Gluconobacter japonicus NBRC3271 (formerly Gluconobacter dioxyacetonicus IFO3271) engineered to produce 2KG efficiently from a mixture of d-glucose and d-gluconate. This mutant strain consumed almost all of the starting materials (d-glucose and d-gluconate) to produce 2,5DKG quantitatively as a seemingly unique metabolite. To our knowledge, this is the first report of a Gluconobacter strain that produces 2,5DKG efficiently and homogeneously.  相似文献   
80.
Geographical variation of elytra color pattern in two sibling ladybird species, Harmonia yedoensis and H. axyridis (Coleoptera: Coccinellidae), was examined. The two species are distributed sympatrically in central Japan; however, only H. yedoensis and H. axyridis occur in the Ryukyu Islands (southern Japan) and Hokkaido island (northern Japan), respectively. The frequency of elytra color patterns was significantly different between the two species in all sympatric locations and our results were inconsistent with the classical theory on Müllerian mimicry. The most dominant pattern of H. axyridis was the least dominant of H. yedoensis in all sympatric populations. Furthermore, the frequency of the non‐melanic form (red ground color with or without black spots) increased towards the south in H. yedoensis. This tendency was in contrast to the known geographical cline in H. axyridis in which the melanic form (black ground color with red spots) was gradually displaced with the non‐melanic form northwards in the Japanese archipelago. We discuss possible selective factors including predator avoidance, thermal adaptation and reproductive character displacement, all of which might contribute to the maintenance of the color polymorphism in the two Harmonia species.  相似文献   
设为首页 | 免责声明 | 关于勤云 | 加入收藏

Copyright©北京勤云科技发展有限公司  京ICP备09084417号