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971.
Pang JJ Xu RK Xu XB Cao JM Ni C Zhu WL Asotra K Chen MC Chen C 《American journal of physiology. Heart and circulatory physiology》2004,286(3):H1063-H1069
Loss of cardiomyocytes by apoptosis is proposed to cause heart failure. Angiotensin II (ANG II), an important neurohormonal factor during heart failure, can induce cardiomyocyte apoptosis. Inasmuch as hexarelin has been reported to have protective effects in this process, we examined whether hexarelin can prevent cardiomyocytes from ANG II-induced cell death. Cultured cardiomyocytes from neonatal rats were stimulated with ANG II. Apoptosis was evaluated using fluorescence microscopy, TdT-mediated dUTP nick-end labeling (TUNEL) method, flow cytometry, DNA laddering, and analysis of cell viability by (3,4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT). It was found that incubation with 0.1 micromol/l ANG II for 48 h increased cardiomyocyte apoptosis. Administration of 0.1 micromol/l hexarelin significantly decreased this ANG II-induced apoptosis and DNA fragmentation and increased myocyte viability. To further investigate the underlying mechanisms, caspase-3 activity assay and mRNA expression of Bax, Bcl-2, and growth hormone secretagogue receptor (GHS-R; the supposed hexarelin binding site) were examined. GHS-R mRNA was abundantly expressed in cardiomyocytes and was upregulated after administration of hexarelin. These results suggest that hexarelin abates cardiomyocytes from ANG II-induced apoptosis possibly via inhibiting the increased caspase-3 activity and Bax expression induced by ANG II and by increasing the expression of Bcl-2, which is depressed by ANG II. Whether the upregulated expression of GHS-R induced by hexarelin is associated with this antiapoptotic effect deserves further investigation. 相似文献
972.
Interleukin-6-induced JAK2/STAT3 signaling pathway in endothelial cells is suppressed by hemodynamic flow 总被引:2,自引:0,他引:2
Ni CW Hsieh HJ Chao YJ Wang DL 《American journal of physiology. Cell physiology》2004,287(3):C771-C780
Endothelial cells (ECs) are constantly exposed to shear stress, the action of which triggers signaling pathways and cellular responses. During inflammation, cytokines such as IL-6 increase in plasma. In this study, we examined the effects of steady flow on IL-6-induced endothelial responses. ECs exposed to IL-6 exhibited STAT3 activation via phosphorylation of Tyr705. However, when ECs were subjected to shear stress, shear force-dependent suppression of IL-6-induced STAT3 phosphorylation was observed. IL-6 treatment increased the phosphorylation of JAK2, an upstream activator of STAT3. Consistently, shear stress significantly reduced IL-6-induced JAK2 activation. Pretreatment of ECs with an inhibitor of MEK1 did not alter this suppression by shear stress, indicating that extracellular signal-regulated kinase (ERK1/2) was not involved. However, pretreatment of ECs with an endothelial nitric oxide synthase inhibitor (nitro-L-arginine methyl ester) attenuated this inhibitory effect of shear stress on STAT3 phosphorylation. Shear stress-treated ECs displayed decreased nuclear transmigration of STAT3 and reduced STAT3 binding to DNA. Intriguingly, ECs exposed to IL-6 entered the cell cycle, as evidenced by increasing G2/M phase, and shear stress to these ECs significantly reduced IL-6-induced cell cycle progression. STAT3-mediated IL-6-induced cell cycle was confirmed by the inhibition of the cell cycle in ECs infected with adenovirus carrying the inactive mutant of STAT3. Our study clearly shows that shear stress exerts its inhibitory regulation by suppressing the IL-6-induced JAK2/STAT3 signaling pathway and thus inhibits IL-6-induced EC proliferation. This shear force-dependent inhibition of IL-6-induced JAK2/STAT3 activation provides new insights into the vasoprotective effects of steady flow on ECs against cytokine-induced responses. shear stress; nitric oxide; cell cycle 相似文献
973.
Neurotensin (NT) regulates a variety of biological processes primarily through interaction with neurotensin receptor-1 (NTR1), a heterotrimeric G-protein-coupled receptor (GPCR). Stimulation of NTR1 has been linked to activation of multiple signaling transduction pathways via specific coupling to G(q), G(i/o), or G(s), in various cell systems. However, the function of NT/NTR1 in the regulation of the Akt pathway remains unknown. Here, we report that activation of NTR1 by NT inhibits Akt activity as determined by the dephosphorylation of Akt at both Ser473 and Thr308 in AV12 cells constitutively expressing human NTR1 (NTR1/AV12). The inactivation of Akt by NT was rapid and dose-dependent. This effect of NT was completely blocked by the specific NTR1 antagonist, (S)-(+)-[1-(7-chloro-4-quinolinyl)-5-(2,6-dimethoxyphenyl)pyrazol-3-yl)-carbonylamino] cyclohexylacetic acid (SR 48527), but unaffected by the less active enantiomer ((R)-(-)-[1-(7-chloro-4-quinolinyl)-5-(2,6-dimethoxyphenyl)pyrazol-3-yl)-carbonylamino] cyclohexylacetic acid (SR 49711)), indicating the stereospecificity of NTR1 in the negative regulation of Akt. In addition, NT prevented insulin- and epidermal growth factor (EGF)-mediated Akt activation. Our results provide insight into the role of NT in the modulation of Akt signaling and the potential physiological significance of Akt regulation by NT. 相似文献
974.
Wenpeng?YangEmail author Younglian?Zheng Shen?Ni Jing?Wu 《Plant Molecular Biology Reporter》2004,22(4):361-374
Recessive allelic variations were investigated at 3 microsatellite (SSR) sites within theO2 gene by using 14 inbredo2 lines and a wild-type line in maize. Among the 15 lines, allelic variations were observed at umc1066, phi057, and phi112
sites. Two alleles were found at the umc1066 site—a recessive allele with 2 perfect GCCAGA repeats and a dominant allele with
3 perfect repeats. Three alleles were found at the phi057 site—2 recessive alleles with 3 and 5 perfect GCC repeats, respectively,
and another with 4 perfect repeats consistent with a dominant allele. At least 4 alleles exist at the phi112 site—among which
1 recessive allele has a 1-bp deletion, another has a 15-bp deletion, and other has no PCR products compared to the dominant
allele; all the alleles have unchanged AG repeats. The phi057 site in exon 6 was identified to be a hypervariable region in
the coding sequence of the02 gene, in addition to the 2 hypervariable regions in exon 1 previously reported. The primary mechanisms underlying the variations
in repeat numbers and regions flanking the SSR within theO2 gene appear to be unequal crossing over and replication slippage. Furthermore, base substitution of SSR motif can create
heteroalleles and modify the repeat number of SSR. The lysine content of kernel in theO2 ando2 lines correlates to a considerable extent with nucleotide variations at the umc1066, phi057, and phi112 sites. Our study
suggests that it is best to use the 3 markers together in molecular marker-assisted selection for high-lysine maize materials. 相似文献
975.
Urea Induced Inactivation and Unfolding of Arginine Kinase from the Sea Cucumber Stichopus japonicus
Urea titration was used to study the inactivation and unfolding equilibrium of arginine kinase (AK) from the sea cucumber Stichopus japonicus. Both fluorescence spectral and circular dichroism spectral data indicated that an unfolding intermediate of AK existed in the presence of 1.0 to 2.0 M urea. This was further supported by the results of size exclusion chromatography. The spectral data suggested that this unfolding intermediate shared many structural characteristics with the native form of AK including its secondary structure, tertiary structure, as well as its quaternary structure. Furthermore, according to the residual activity curve, this unfolding intermediate form still retained its catalytic function although its activity was lower than that of native AK. Taken together, the results of our study give direct evidence that an intermediate with partial activity exists in unfolding equilibrium states of AK during titration with urea. 相似文献
976.
Polyethylenimine (PEI) and other polycations are good vehicles for transferring genes into the cells. In earlier reports, poly-L-lysine and protamine have been shown to improve gene delivery with cationic liposomes. In this study, PEI, combined with different cationic liposomes, was studied to determine the optimal conditions for gene delivery. The reporter genes, luciferase and green fluorescent protein, were used to transfect human HeLa, HepG2 and hepatoma 2.2.15 cells with various combinations of PEIs (0.8 and 25 kDa), poly-L-lysine (15-30 kDa), protamine and cationic liposomes. The highest expression level was achieved by using the combination of PEI 25 kDa (0.65 microg/microg of DNA, nitrogen-to-DNA phosphate (N/P) ratio=4.5) with 10 nmol of DOTAP-cholesterol (DOTAP-Chol, 1:1 w/w). This DNA complex formulation dramatically increased the luciferase expression 10- to 100-fold, which was much higher than those of other polycations alone, cationic liposomes alone or the combination. In addition, PEI/DOTAP-Chol combination had little cytotoxicity than DOTAP-Chol or other cationic liposomes alone. The effect of oligonucleotide (ODN) delivery facilitated by PEI and cationic liposomes was also studied in the hepatoma cell lines. We demonstrated an antisense ODN of p53 delivered by PEI/DOTAP-Chol combination effectively inhibited the biosynthesis of p53 protein in HepG2 (68% inhibiton) and 2.2.15 cells (43% inhibition). Thus, the large PEI could synergistically increase the transfection efficiency when combined with the cationic liposomes. 相似文献
977.
HLA-E allelic variants. Correlating differential expression,peptide affinities,crystal structures,and thermal stabilities 总被引:6,自引:0,他引:6
Strong RK Holmes MA Li P Braun L Lee N Geraghty DE 《The Journal of biological chemistry》2003,278(7):5082-5090
Previous studies of HLA-E allelic polymorphism have indicated that balancing selection may be acting to maintain two major alleles in most populations, indicating that a functional difference may exist between the alleles. The alleles differ at only one amino acid position, where an arginine at position 107 in HLA-E*0101 (E(R)) is replaced by a glycine in HLA-E*0103 (E(G)). To investigate possible functional differences, we have undertaken a study of the physical and biochemical properties of these two proteins. By comparing expression levels, we found that whereas steady-state protein levels were similar, the two alleles did in fact differ with respect to cell surface levels. To help explain this difference, we undertook studies of the relative differences in peptide affinity, complex stability, and three-dimensional structure between the alleles. The crystal structures for HLA-E(G) complexed with two distinct peptides were determined, and both were compared with the HLA-E(R) structure. No significant differences in the structure of HLA-E were induced as a result of binding different peptides or by the allelic substitution at position 107. However, there were clear differences in the relative affinity for peptide of each heavy chain, which correlated with and may be explained by differences between their thermal stabilities. These differences were completely consistent with the relative levels of the HLA-E alleles on the cell surface and may indeed correlate with functional differences. This in turn may help explain the apparent balancing selection acting on this locus. 相似文献
978.
The large amount of repeats, especially high copy repeats, in the genomes of higher animals and plants makes whole genome assembly (WGA) quite difficult. In order to solve this problem, we tried to identify repeats and mask them prior to assembly even at the stage of genome survey. It is known that repeats of different copy number have different probabilities of appearance in shotgun data, so based on this principle, we constructed a statistical model and inferred criteria for mathematically defined repeats (MDRs) at different shotgun coverages. According to these criteria, we developed software MDRmasker to identify and mask MDRs in shotgun data. With repeats masked prior to assembly, the speed of assembly was increased with lower error probability. In addition, clone-insert size affects the accuracy of repeat assembly and scaffold construction. We also designed length distribution of clone-inserts using our model. In our simulated genomes of human and rice, the length distribution of repeats is differ 相似文献
979.
980.
Detection of mitochondrial DNA deletion by a modified PCR method in a 60Co radiation-exposed patient
A new PCR based method was developed to detect deleted mitochondrial DNA (mtDNA). Peripheral blood cell DNA was obtained from a victim who was accidently exposed to a 60Co radiation source in 1990. Using the DNA as template, first PCR was performed to generate multiple products including true deletions and artifacts. The full length product was recovered and used as template of secondary PCR. The suspicious deletion product of mtDNA could be confirmed only if it was yielded by first PCR. Using either original primers or their nested primers, the suspicious deletion product was amplified and authenticated as a true deletion product. The template was recovered and determined to be a deletion by sequencing directly. The results show that a new mtDNA deletion, which spans 889 bp from nt 11688 to nt 12576, was detected in the peripheral blood cells of the victim. It indicates that this new PCR-based method was more efficient at detecting small populations of mtDNA deletion than other routine methods. MtDNA deletion was found in the victim, suggesting the relationship between the deletion and phenotypes of the disease. 相似文献