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981.
Jiang J Xia XB Xu HZ Xiong Y Song WT Xiong SQ Li Y 《Journal of cellular physiology》2009,218(1):66-74
Retinal neovascularization (NV) occurs in various ocular disorders including proliferative diabetic retinopathy, retinopathy of prematurity and secondary neovascular glaucoma, which often result in blindness. Vascular endothelial growth factor (VEGF) is an essential growth factor for angiogenesis, and is particularly regulated by hypoxia inducible factor-1alpha (HIF-1alpha) under hypoxic conditions. Therefore, HIF-1alpha and VEGF could provide targets for therapeutic intervention on retinal NV. In this study, we investigated the inhibitory effects of small interfering RNA (siRNA) targeting HIF-1alpha and VEGF on the expression of HIF-1alpha and VEGF in human umbilical vein endothelial cells (HUVEC) in vitro and on retinal NV in vivo. siRNA-expressing plasmids targeting human HIF-1alpha (HIF-1alpha siRNA) and human VEGF(165) (VEGF siRNA) were constructed. They were transfected and co-transfected to HUVEC and C57BL/6J mice of ischemic retinopathy model. HIF-1alpha siRNA and VEGF siRNA specifically downregulated HIF-1alpha and VEGF at both mRNA and protein levels in vitro and in vivo. Neovascular tufts and neovascular nuclei were decreased in gene therapy group compared to control hypoxia group. Co-transfection of HIF-1alpha siRNA and VEGF siRNA resulted in maximal effects on VEGF suppression in vitro and in vivo. It also manifested the maximal inhibitory effect on retinal NV. These results indicate that the application of HIF-1alpha siRNA and VEGF siRNA technology holds great potential as a novel therapeutic for retinal NV. 相似文献
982.
Min Chen Qian Chen Xue-Wen Cheng Ting-Jia Lu Han-Xing Liu Jie-Min Jia Chi Zhang Li Xu Zhi-Qi Xiong 《Journal of neurochemistry》2009,111(5):1094-1103
Deposition of ubiquitinated protein aggregates is a hallmark of neurodegeneration in both acute neural injuries, such as stroke, and chronic conditions, such as Parkinson's disease, but the underlying mechanisms are poorly understood. In the present study, we examined the role of Zn2+ in ischemia-induced impairment of the ubiquitin-proteasome system in the CA1 region of rat hippocampus after transient global ischemia. We found that scavenging endogenous Zn2+ reduced ischemia-induced ubiquitin conjugation and free ubiquitin depletion. Furthermore, exposure to zinc chloride increased ubiquitination and inhibited proteasomal enzyme activity in cultured hippocampal neurons in a concentration- and time-dependent manner. Further studies of the underlying mechanisms showed that Zn2+ -induced ubiquitination required p38 activation. These findings indicate that alterations in Zn2+ homeostasis impair the protein degradation pathway. 相似文献
983.
984.
Ming-hon Yau Yu Wang Karen S. L. Lam Jialiang Zhang Donghai Wu Aimin Xu 《The Journal of biological chemistry》2009,284(18):11942-11952
Lipoprotein lipase (LPL) is a principal enzyme responsible for the
clearance of chylomicrons and very low density lipoproteins from the
bloodstream. Two members of the Angptl (angiopoietin-like protein) family,
namely Angptl3 and Angptl4, have been shown to inhibit LPL activity in
vitro and in vivo. Here, we further investigated the structural
basis underlying the LPL inhibition by Angptl3 and Angptl4. By multiple
sequence alignment analysis, we have identified a highly conserved 12-amino
acid consensus motif that is present within the coiled-coil domain (CCD) of
both Angptl3 and Angptl4, but not other members of the Angptl family.
Substitution of the three polar amino acid residues (His46,
Gln50, and Gln53) within this motif with alanine
abolishes the inhibitory effect of Angptl4 on LPL in vitro and also
abrogates the ability of Angptl4 to elevate plasma triglyceride levels in
mice. The CCD of Angptl4 interacts with LPL and converts the catalytically
active dimers of LPL to its inactive monomers, whereas the mutant protein with
the three polar amino acids being replaced by alanine loses such a property.
Furthermore, a synthetic peptide consisting of the 12-amino acid consensus
motif is sufficient to inhibit LPL activity, although the potency is
much lower than the recombinant CCD of Angptl4. In summary, our data suggest
that the 12-amino acid consensus motif within the CCD of Angptl4, especially
the three polar residues within this motif, is responsible for its interaction
with and inhibition of LPL by blocking the enzyme dimerization.Lipoprotein lipase
(LPL)3 is an
endothelium-bound enzyme that catalyzes the hydrolysis of plasma triglyceride
(TG) associated with chylomicrons and very low density lipoproteins
(1,
2). This enzyme plays a major
role in maintaining lipid homeostasis by promoting the clearance of TG-rich
lipoproteins from the bloodstream. Abnormality in LPL functions has been
associated with a number of pathological conditions, including
atherosclerosis, dyslipidemia associated with diabetes, and Alzheimer disease
(1).LPL is expressed in a wide variety of cell types, particularly in
adipocytes and myocytes (2). As
a rate-limiting enzyme for clearance of TG-rich lipoproteins, the activity of
LPL is tightly modulated by multiple mechanisms in a tissue-specific manner in
response to nutritional changes
(3,
4). The enzymatic activity of
LPL in adipose tissue is enhanced after feeding to facilitate the storage of
TG, whereas it is down-regulated during fasting to increase the utilization of
TG by other tissues (5). The
active form of LPL is a noncovalent homodimer with the subunits associated in
a head-to-tail manner, and the dissociation of its dimeric form leads to the
formation of a stable inactive monomeric conformation and irreversible enzyme
inactivation (6). At the
post-translational level, the LPL activity is regulated by numerous
apolipoprotein co-factors. For instance, apoCII, a small apolipoprotein
consisting of 79 amino acid residues in human, activates LPL by directly
binding to the enzyme (7,
8). By contrast, several other
apolipoproteins such as apoCI, apo-CIII, and apoE have been shown to inhibit
the LPL activity in vitro
(3).Angiopoietin-like proteins (Angptl) are a family of secreted proteins
consisting of seven members, Angptl1 to Angptl7
(9,
10). All the members of the
Angptl family share a similar domain organization to those of angiopoietins,
with an NH2-terminal coiled-coil domain (CCD) and a COOH-terminal
fibrinogen-like domain. Among the seven family members, only Angptl3 and
Angptl4 have been shown to be involved in regulating triglyceride metabolism
(10,
11). The biological functions
of Angptl3 in lipid metabolism were first discovered by Koishi et al.
(12) in their positional
cloning of the recessive mutation gene responsible for the hypolipidemia
phenotype in a strain of obese mouse KK/snk. Subsequent studies have
demonstrated that Angptl3 increases plasma TG levels by inhibiting the LPL
enzymatic activity
(13–15).
Angptl4, also known as fasting-induced adipocyte factor, hepatic
fibrinogen/angiopoietin-related protein, or peroxisome proliferator-activated
receptor-γ angiopoietin-related, is a secreted glycoprotein abundantly
expressed in adipocyte, liver, and placenta
(16–18).
In addition to its role in regulating angiogenesis, a growing body of evidence
demonstrated that Angptl4 is an important player of lipid metabolism
(10,
11). Elevation of circulating
Angptl4 by transgenic or adenoviral overexpression, or by direct
supplementation of recombinant protein, leads to a marked elevation in the
levels of plasma TG and low density lipoprotein cholesterol in mice
(19–22).
By contrast, Angptl4 knock-out mice exhibit much lower plasma TG and
cholesterol levels compared with the wild type littermates
(19,
20). Notably, treatment of
several mouse models (such as C57BL/6J, ApoE–/–,
LDLR–/–, and db/db obese/diabetic mice) with a
neutralizing antibody against Angptl4 recapitulate the lipid phenotype found
in Angptl4 knock-out mice
(19). The role of Angptl4 as a
physiological inhibitor of LPL is also supported by the finding that its
expression levels in adipose tissue change rapidly during the fed-to-fasting
transitions and correlate inversely with LPL activity
(23). In humans, a genetic
variant of the ANGPTL4 gene (E40K) has been found to be associated
with significantly lower plasma TG levels and higher high density lipoprotein
cholesterol concentrations in several ethnic groups
(24–26).Angptl3 and Angptl4 share many common biochemical and functional properties
(10). In both humans and
rodents, Angptl3 and Angptl4 are proteolytically cleaved at the linker region
and circulate in plasma as two truncated fragments, including
NH2-terminal CCD and COOH-terminal fibrinogen-like domain
(14,
27–29).
The effects of both Angptl3 and Angptl4 on elevating plasma TG levels are
mediated exclusively by their NH2-terminal CCDs
(15,
22,
23,
27,
30). The CCDs of Angptl3 and
Angptl4 have been shown to inhibit the LPL activity in vitro as well
as in mice
(23,30,31).
Angptl4 inhibits LPL by promoting the conversion of the catalytically active
LPL dimers into catalytically inactive LPL monomers, thereby leading to the
inactivation of LPL (23,
31). However, the detailed
structural and molecular basis underlying the LPL inhibition by Angptl3 and
Angptl4 remain poorly characterized at this stage.In this study, we analyzed all known amino acid sequences of Angptl3 and
Angptl4 from various species and found a short motif,
LAXGLLXLGXGL (where X represents polar
amino acid residues), which corresponds to amino acid residues 46–57 and
44–55 of human Angptl3 and Angptl4, respectively, is highly conserved
despite the low degree of their overall homology (∼30%). Using both in
vitro and in vivo approaches, we demonstrated that this 12-amino
acid sequence motif, in particular the three polar amino acid residue within
this motif, is essential for mediating the interactions between LPL and
Angpt4, which in turn disrupts the dimerization of the enzyme. 相似文献
985.
Xiaoyan Hui Weidong Zhu Yu Wang Karen S. L. Lam Jialiang Zhang Donghai Wu Edward W. Kraegen Yixue Li Aimin Xu 《The Journal of biological chemistry》2009,284(21):14050-14057
Major urinary protein-1 (MUP-1) is a low molecular weight secreted protein
produced predominantly from the liver. Structurally it belongs to the
lipocalin family, which carries small hydrophobic ligands such as pheromones.
However, the physiological functions of MUP-1 remain poorly understood. Here
we provide evidence demonstrating that MUP-1 is an important player in
regulating energy expenditure and metabolism in mice. Both microarray and
real-time PCR analysis demonstrated that the MUP-1 mRNA abundance in the liver
of db/db obese mice was reduced by ∼30-fold compared
with their lean littermates, whereas this change was partially reversed by
treatment with the insulin-sensitizing drug rosiglitazone. In both dietary and
genetic obese mice, the circulating concentrations of MUP-1 were markedly
decreased compared with the lean controls. Chronic elevation of circulating
MUP-1 in db/db mice, using an osmotic pump-based protein
delivery system, increased energy expenditure and locomotor activity, raised
core body temperature, and decreased glucose intolerance as well as insulin
resistance. At the molecular level, MUP-1-mediated improvement in metabolic
profiles was accompanied by increased expression of genes involved in
mitochondrial biogenesis, elevated mitochondrial oxidative capacity, decreased
triglyceride accumulation, and enhanced insulin-evoked Akt signaling in
skeletal muscle but not in liver. Altogether, these findings raise the
possibility that MUP-1 deficiency might contribute to the metabolic
dysregulation in obese/diabetic mice, and suggest that the beneficial
metabolic effects of MUP-1 are attributed in part to its ability in increasing
mitochondrial function in skeletal muscle.The liver is the primary organ for carbohydrate and lipid metabolism,
including gluconeogenesis, glycogenesis, cholesterol biosynthesis, and
lipogenesis (1,
2). These metabolic events in
the liver are tightly controlled by several pancreatic hormones including
insulin and glucagon. In addition, the liver itself is one of the largest
endocrine organs in the body, secreting numerous humoral factors involved in
the regulation of systemic glucose and lipid homeostasis. The importance of
the liver-derived humoral factors in maintaining glucose metabolism is
highlighted by the observation that glucose uptake by skeletal muscle is
severely impaired by surgical or pharmacological blockade of hepatic
parasympathetic nerves (3). In
the past several years, a number of liver-derived humoral metabolic factors,
including bone morphogenetic protein-9 (BMP-9)
(4), fibroblast growth factor
21 (FGF21)
(5–7),
retinol-binding protein 4 (RBP4)
(8,
9), adropin
(10), and angiopoietin-like
proteins (Angptl) 3, 4, and 6
(11–13),
have been identified, and their roles in glucose and lipid metabolism have
been characterized in great detail. Noticeably, BMP-9, FGF21, and Angptl6
exhibit potent insulin-sensitizing and glucose-lowering effects in animal
models, and they have been proposed as potential candidates for the treatment
of insulin resistance and type II diabetes
(4,
6,
7,
13).To search for novel liver-derived secretory factors involved in the
regulation of glucose homeostasis, we used microarray analysis as a global
screening for systematic identification of genes differentially expressed in
the liver of C57BLKS db/db mice (a genetically inherited
diabetic mouse model that is characterized by severe insulin resistance and
hyperglycemia) and their lean littermates. We found that the mRNA level of
mouse major urinary protein-1
(MUP-1)2 was markedly
down-regulated in db/db mice, and the change was largely
normalized upon treatment with the PPARγ agonist rosiglitazone. MUP-1 is
a small molecular weight secreted protein abundantly expressed in the liver
(14). Its expression in the
liver is enhanced by administration of the hepatotoxic agent
dimethylnitrosamine (15) but
is reduced by interleukin 6-induced acute phase response in mice
(16). Like other members of
the MUP family, MUP-1 has been proposed to act as a pheromone-binding protein
in urine (17), thereby
accelerating puberty and promoting aggressive behavior in male mice. However,
the precise functions of MUPs have yet to be determined.MUP-1 belongs to the lipocalin superfamily, the members of which share a
common tertiary structure with a cup-shaped hydrophobic ligand binding pocket
surrounded by an eight-stranded β-barrel
(18,
19). This structure confers
upon lipocalins the ability to bind and transport a wide variety of small
lipophilic substances, including fatty acids, cholesterols, prostaglandins,
and pheromones. Noticeably, several members of the lipocalin family, including
RBP4, lipocalin-2, and adipocyte fatty acid-binding protein (A-FABP), have
recently been shown to be important mediators of obesity-related insulin
resistance and glucose intolerance
(8,
20–22).
Unlike MUP-1, the expression of RBP4, lipocalin-2, and A-FABP are elevated in
obesity and diabetes (9,
20,
23).In this study, we investigated the metabolic role of MUP-1 in mice. Our
results demonstrated that MUP-1 was abundantly present in the circulation. In
genetic and dietary obese mouse models, the serum and urine concentrations of
MUP-1 were remarkably decreased. Replenishment of recombinant MUP-1 led to
improved glucose tolerance and insulin sensitivity, as well as increased
energy expenditure and locomotor activity in db/db diabetic
mice. Our data suggest that MUP-1 not only serves as a circulating biomarker,
negatively correlated with obesity-related metabolic disorders, but also plays
an active role in regulating energy homeostasis and insulin sensitivity in
mice. 相似文献
986.
987.
988.
Wenling Zhang ;Zhaoyang Zeng ;Yanhong Zhou ;Wei Xiong ;Songqing Fan ;Lan Xiao ;Donghai Huang ;Zheng Li ;Dan Li ;Minghua Wu ;Xiaoling Li ;Shourong Shen ;Rong Wang ;Li Cao ;Ke Tang ;Guiyuan Li 《Acta biochimica et biophysica Sinica》2009,(5):414-428
Previous studies have revealed that Epstein-Barr virus (EBV) was closely associated with nasopharyngeal carcinoma (NPC). This study aimed to characterize the global pathways affected in the EBV-associated NPC. Combined with microdissection, gene expression profries in 22 NPCs and 10 non-tumor nasopharyngeal epithelial (NPE) tissue samples were analyzed. All NPC specimens served in the microarray analysis were positive for EBV, as judged by identification of the expression of EBV nuclear antigen 1 (EBNA1). Through gene set enrichment analysis (GSEA), we found that cell cycle pathway was the most disregulated pathway in NPC (P = 0.000, false discovery rate q-value = 0.007), which included some aberrant expressed components. We first found that overexpression of CDK4, cyclin D1, and Rb proteins, and loss of expression of proteins p16, p27, and p19 were statistically significant in NPC tissues compared with non-cancerous NPE (P〈 0.05) by real-time RT- PCR and tissue microarray. EBV-encoded small RNA-1 (EBER-1) hybridization signals in the NPC showed significant associations with the overexpression of Rb (P = 0.000), cyclin D1 (P = 0.000), CDK4 (P = 0.000), and the loss of expression of p16 proteins (P = 0.039). In the final logistic regression analysis model, EBER-1 and abnormal expression of p16, Rb, cyclin D1, and E2F6 were inde- pendent contributions to nasopharyngeal carcinogenesis. Through survival analysis, only cyclin D1 could predict the prognosis of NPC patients. These results suggested that cell cycle pathway was the most disregulated pathway in the EBV-associated NPC, and EBER-1 was closely associated with p16, CDK4, cyclin D1, and Rb. cyclin D1 could be the prognosis biomarker for NPC. 相似文献
989.
目的:探讨一氧化氮供体药物硝酸甘油对对裸鼠胰腺癌皮下成瘤的抑制作用及对瘤体组织中低氧诱导因子1α(hypoxia induced factor 1α,HIF-1α),P糖蛋白(P-glucoprotein,P-gP)和谷胱甘肽S转移酶-π(Glutathione S-transferase pi,GST-π)的表达影响.方法:40只6-8周的18-22g的BALB/c裸鼠双侧腹股沟皮下注射胰腺癌PANC-1细胞成瘤,瘤体超过150mm3后随机分组,干预组用硝酸甘油帖剂干预,对照组贴安慰剂,并分别绘制生长曲线.采用免疫组织化学方法检测成瘤裸鼠中瘤体组织中HIF-1α、P-gP和GST-π的表达.结果:32例裸鼠成瘤,成瘤率80%,干预28天后,干预组和对照组的瘤体大小分别为1550.4±148.6mm3和750.6±71.2mm3.16只对照组瘤体组织中HIF-1α阳性表达12例(75%),P-gp阳性表达10例(62.5%),GsT-π阳性表达8例(50%).在16例干预组裸鼠瘤体组织中HIF-1α、P-gP、GST-π阳性表达分别是11例(68.8%),8例(50%),7例(43.8%).结论:特定剂量硝酸甘油对裸鼠皮下成瘤有明显抑制作用,且对HIF-1α及P-gp、GsT-π的表达也有一定的抑制作用. 相似文献
990.
1998年,Thomson等成功分离出了人类胚胎干细胞(embryonic stem cell,ES)系为研究再生医学和人类早期发育开辟了一个新领域.通过转染人类ES细胞进行遗传操作(genetic manipulation)可以帮助我们更好地理解人类Es细胞的生物学特性,同时为研究人类ES细胞在细胞及基因治疗中潜在的治疗用途提供了基础.本文综述了人类ES细胞外源基因转染方法及其应用的进展. 相似文献