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21.
Koch M Wendorf M Dendorfer A Wolfrum S Schulze K Spillmann F Schultheiss HP Tschöpe C 《American journal of physiology. Heart and circulatory physiology》2003,285(1):H418-H423
Diabetes mellitus impairs the cardiac kallikrein-kinin system by reducing cardiac kallikrein (KLK) and kininogen levels, a mechanism that may contribute to the deleterious outcome of cardiac ischemia in this disease. We studied left ventricular (LV) function and bradykinin (BK) coronary outflow in buffer-perfused, isolated working hearts (n = 7) of controls and streptozotocin (STZ)-induced diabetic rats before and after global ischemia. With the use of selective kininase inhibitors, the activities of angiotensin I-converting enzyme, aminopeptidase P, and neutral endopeptidase were determined by analyzing the degradation kinetics of exogenously administered BK during sequential coronary passages. Basal LV function and coronary flow were impaired in STZ-induced diabetic rats. Neither basal nor postischemic coronary BK outflow differed between control and diabetic hearts. Reperfusion after 15 min of ischemia induced a peak in coronary BK outflow that was of the same extent and duration in both groups. In diabetic hearts, total cardiac kininase activity was reduced by 41.4% with an unchanged relative kininase contribution compared with controls. In conclusion, despite reduced cardiac KLK synthesis, STZ-induced diabetic hearts are able to maintain kinin liberation under basal and ischemic conditions because of a primary impairment or a secondary downregulation of kinin-degrading enzymes. 相似文献
22.
D M Kolpashchikov K Weisshart H P Nasheuer S N Khodyreva E Fanning A Favre O I Lavrik 《FEBS letters》1999,450(1-2):131-134
Human replication protein A is a heterotrimeric protein involved in various processes of DNA metabolism. To understand the contribution of replication protein A individual subunits to DNA binding, we have expressed them separately as soluble maltose binding protein fusion proteins. Using a DNA construct that had a photoreactive group incorporated at the 3'-end of the primer strand, we show that the p70 subunit on its own is efficiently cross-linked to the primer at physiological concentrations. In contrast, crosslinking of the p32 subunit required two orders of magnitude higher protein concentrations. In no case was the p14 subunit labelled above background. p70 seems to be the predominant subunit to bind single-stranded DNA and this interaction positions the p32 subunit to the 3'-end of the primer. 相似文献
23.
Axel Joachim Krafft Patrik Zamecnik Florian Maier André de Oliveira Peter Hallscheidt Heinz-Peter Schlemmer Michael Bock 《Physica medica : PM : an international journal devoted to the applications of physics to medicine and biology : official journal of the Italian Association of Biomedical Physics (AIFB)》2013,29(6):607-614
PurposeIn this work, a passive tracking sequence employing a phase-only cross correlation (POCC) algorithm was studied with a focus on the in vivo applicability of the technique. Therefore, MR-guided needle interventions were performed in a phantom and two animal experiments.MethodsThe targeting accuracy was quantified in an agarose phantom with 15 fiducials. For each fiducial, the distance between needle trajectory and target point was measured. In a first animal experiment at 3 T, the prostate of a pig was punctured under POCC guidance. Second, POCC-based tracking was performed during a laser-induced thermal therapy procedure in peripheral porcine muscle tissue at 1.5 T.ResultsIn the phantom experiment, the 15 fiducials were penetrated with a mean accuracy of 1.5 ± 0.9 mm (mean duration for one puncture about 2 min). In the first animal experiment, the center of the pig's right prostatic lobe was accurately punctured within 15 min. In the second, targeting and insertion of the needle could be performed within 5 min and a thermal lesion was successfully created.ConclusionOur initial experience with the POCC-based tracking sequence indicates that this technique has the potential as an accurate and versatile tool for in vivo MR-guided needle interventions. 相似文献
24.
Weisshart K Pestryakov P Smith RW Hartmann H Kremmer E Lavrik O Nasheuer HP 《The Journal of biological chemistry》2004,279(34):35368-35376
The heterotrimeric replication protein A (RPA) has multiple essential activities in eukaryotic DNA metabolism and in signaling pathways. Despite extensive analyses, the functions of the smallest RPA subunit p14 are still unknown. To solve this issue we produced and characterized a dimeric RPA complex lacking p14, RPADeltap14, consisting of p70 and p32. RPADeltap14 was able to bind single-stranded DNA, but its binding mode and affinity differed from those of the heterotrimeric complex. Moreover, in the RPADeltap14 complex p32 only minimally recognized the 3'-end of a primer in a primer-template junction. Partial proteolytic digests revealed that p14 and p32 together stabilize the C terminus of p70 against degradation. Although RPADeltap14 efficiently supported bidirectional unwinding of double-stranded DNA and interacted with both the simian virus 40 (SV40) large T antigen and cellular DNA polymerase alpha-primase, it did not support cell-free SV40 DNA replication. This inability manifested itself in a failure to support both the primer synthesis and primer elongation reactions. These data reveal that efficient binding and correct positioning of the RPA complex on single-stranded DNA requires all three subunits to support DNA replication. 相似文献
25.
Schub O Rohaly G Smith RW Schneider A Dehde S Dornreiter I Nasheuer HP 《The Journal of biological chemistry》2001,276(41):38076-38083
DNA polymerase alpha-primase (pol-prim) is the only enzyme that can start DNA replication de novo. The 180-kDa (p180) and 68-kDa (p68) subunits of the human four-subunit enzyme are phosphorylated by Cyclin-dependent kinases (Cdks) in a cell cycle-dependent manner. Cyclin A-Cdk2 physically interacts with pol-prim and phosphorylates N-terminal amino acids of the p180 and the p68 subunits, leading to an inhibition of pol-prim in initiating cell-free SV40 DNA replication. Mutation of conserved putative Cdk phosphorylation sites in the N terminus of human p180 and p68 reduced their phosphorylation by Cyclin A-Cdk2 in vitro. In contrast to wild-type pol-prim these mutants were no longer inhibited by Cyclin A-Cdk2 in the initiation of viral DNA replication. Importantly, rather than inhibiting it, Cyclin A-Cdk2 stimulated the initiation activity of pol-prim containing a triple N-terminal alanine mutant of the p180 subunit. Together these results suggest that Cyclin A-Cdk2 executes both stimulatory and inhibitory effects on the activity of pol-prim in initiating DNA replication. 相似文献
26.
Miteva K Haag M Peng J Savvatis K Becher PM Seifert M Warstat K Westermann D Ringe J Sittinger M Schultheiss HP Tschöpe C Van Linthout S 《PloS one》2011,6(12):e28513
Background
Under conventional heart failure therapy, inflammatory cardiomyopathy typically has a progressive course, indicating a need for alternative therapeutic strategies to improve long-term outcomes. We recently isolated and identified novel cardiac-derived cells from human cardiac biopsies: cardiac-derived adherent proliferating cells (CAPs). They have similarities with mesenchymal stromal cells, which are known for their anti-apoptotic and immunomodulatory properties. We explored whether CAPs application could be a novel strategy to improve acute Coxsackievirus B3 (CVB3)-induced myocarditis.Methodology/Principal Findings
To evaluate the safety of our approach, we first analyzed the expression of the coxsackie- and adenovirus receptor (CAR) and the co-receptor CD55 on CAPs, which are both required for effective CVB3 infectivity. We could demonstrate that CAPs only minimally express both receptors, which translates to minimal CVB3 copy numbers, and without viral particle release after CVB3 infection. Co-culture of CAPs with CVB3-infected HL-1 cardiomyocytes resulted in a reduction of CVB3-induced HL-1 apoptosis and viral progeny release. In addition, CAPs reduced CD4 and CD8 T cell proliferation. All CAPs-mediated protective effects were nitric oxide- and interleukin-10-dependent and required interferon-γ. In an acute murine model of CVB3-induced myocarditis, application of CAPs led to a decrease of cardiac apoptosis, cardiac CVB3 viral load and improved left ventricular contractility parameters. This was associated with a decline in cardiac mononuclear cell activity, an increase in T regulatory cells and T cell apoptosis, and an increase in left ventricular interleukin-10 and interferon-γ mRNA expression.Conclusions
We conclude that CAPs are a unique type of cardiac-derived cells and promising tools to improve acute CVB3-induced myocarditis. 相似文献27.
RPA subunit arrangement near the 3'-end of the primer is modulated by the length of the template strand and cooperative protein interactions. 下载免费PDF全文
O I Lavrik D M Kolpashchikov K Weisshart H P Nasheuer S N Khodyreva A Favre 《Nucleic acids research》1999,27(21):4235-4240
To analyze the interaction of human replication protein A (RPA) and its subunits with the DNA template-primer junction in the DNA replication fork, we designed several template-primer systems differing in the size of the single-stranded template tail (4, 9, 13, 14, 19 and 31 nt). Base substituted photoreactive dNTP analogs-5-[ N -(2-nitro-5-azidobenzoyl)- trans -3-amino-propenyl-1]-2'-deoxyuridine-5'-triphosphate (NAB-4-dUTP) and 5-[ N -[ N -(2-nitro-5-azidobenzoyl)glycyl]- trans -3-aminopropenyl-1]-2'-deoxyuridine-5'-triphosphate (NAB-7-dUTP)-were used as substrates for elongation of radiolabeled primer-template by DNA polymerases in the presence or absence of RPA. Subsequent UV crosslinking showed that the pattern of p32 and p70 RPA subunit labeling, and consequently their interaction with the template-primer junction, is strongly dependent on the template extension length at a particular RPA concentration, as well as on the ratio of RPA to template concentration. Our results suggest a model of changes in the RPA configuration modulating by the length of the template extension in the course of nascent DNA synthesis. 相似文献
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29.
Different regions of primase subunit p48 control mouse polyomavirus and simian virus 40 DNA replication in vitro 下载免费PDF全文
DNA polymerase alpha-primase (pol-prim), a complex consisting of four subunits, is the major species-specific factor for mouse polyomavirus (PyV) and simian virus 40 (SV40) DNA replication. Although p48 is the most conserved subunit of pol-prim, it is required for in vitro PyV DNA replication but can inhibit cell-free SV40 DNA replication. Production of chimeric human-mouse p48 revealed that different regions of p48 are involved in supporting PyV DNA replication and inhibiting SV40 DNA replication. The N and C-terminal parts of p48 do not have species-specific functions in cell-free PyV DNA replication, but the central part (amino acids [aa] 129 to 320) controls PyV DNA replication in vitro. However, PyV T antigen physically binds to mouse, human, and chimeric pol-prim complexes independently, whether they support PyV DNA replication or not. In contrast to the PyV system, the inhibitory effects of mouse p48 on SV40 DNA replication are mediated by N- and C-terminal regions of p48. Thus, a chimeric p48 containing human aa 1 to 128, mouse aa 129 to 320, and human aa 321 to 418 is active in both PyV and SV40 DNA replication in vitro. 相似文献
30.
Troponin elevation in acute ischemic stroke (TRELAS) - protocol of a prospective observational trial
Jan F Scheitz Hans-Christian Mochmann Christian H Nolte Karl G Haeusler Heinrich J Audebert Peter U Heuschmann Ulrich Laufs Bernhard Witzenbichler Heinz-Peter Schultheiss Matthias Endres 《BMC neurology》2011,11(1):98