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Scott P. Lawrence Nicholas A. Bright J. Paul Luzio Katherine Bowers 《Molecular biology of the cell》2010,21(20):3540-3551
The pH and lumenal environment of intracellular organelles is considered essential for protein sorting and trafficking through the cell. We provide the first evidence that a mammalian NHE sodium (potassium)/proton exchanger, NHE8, plays a key role in the control of protein trafficking and endosome morphology. At steady state, the majority of epitope-tagged NHE8 was found in the trans-Golgi network of HeLa M-cells, but a proportion was also localized to multivesicular bodies (MVBs). Depletion of NHE8 in HeLa M-cells with siRNA resulted in the perturbation of MVB protein sorting, as shown by an increase in epidermal growth factor degradation. Additionally, NHE8-depleted cells displayed striking perinuclear clustering of endosomes and lysosomes, and there was a ninefold increase in the cellular volume taken up by LAMP1/LBPA-positive, dense MVBs. Our data points to a role for the ion exchange activity of NHE8 being required to maintain endosome morphology, as overexpression of a nonfunctional point mutant protein (NHE8 E225Q) resulted in phenotypes similar to those seen after siRNA depletion of endogenous NHE8. Interestingly, we found that depletion of NHE8, despite its function as a sodium (potassium)/proton antiporter, did not affect the overall pH inside dense MVBs. 相似文献
3.
Bruce R. Dorrbecker Susan H. Mercik Paul A. Kramer 《Journal of chromatography. B, Analytical technologies in the biomedical and life sciences》1984,336(1)
A high-performance liquid chromatographic procedure is reported for reproducibly and sensitively quantitating caffeine and its N-demethylated metabolite paraxanthine in micro-samples. A 5-μm reversed-phase radial compression column and 214-nm fixed wavelength ultraviolet detector were used to attain a sensitivity sufficient to quantitate these compounds at concentratios as low as 80 ng/ml using only 25 μl of sample. The assay is applicable to microliter samples of whole blood, serum, plasma, saliva, amniotic, cerebro-spinal and gastric fluids such as might be obtained in studies involving small animals or neonates. The utility of the assay is illustrated with caffeine and paraxanthine levels measured in several maternal and fetal fluids following constant-rate intravenous infusion of caffeine into a rabbit throughout pregnancy. 相似文献
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John G. Neuhoff Katherine L. Long Rebecca C. Worthington 《Evolution and human behavior》2012,33(4):318-322
Listeners consistently perceive approaching sounds to be closer than they actually are and perceptually underestimate the time to arrival of looming sound sources. In a natural environment, this underestimation results in more time than expected to evade or engage the source and affords a “margin of safety” that may provide a selective advantage. However, a key component in the proposed evolutionary origins of the perceptual bias is the appropriate timing of anticipatory motor behaviors. Here we show that listeners with poorer physical fitness respond sooner to looming sounds and with a larger margin of safety than listeners with better physical fitness. The anticipatory perceptual bias for looming sounds is negatively correlated with physical strength and positively correlated with recovery heart rate (a measure of aerobic fitness). The results suggest that the auditory perception of looming sounds may be modulated by the response capacity of the motor system. 相似文献
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E. Charles Kerr E Bruce Eaton L. Thomas Netzel 《Nucleosides, nucleotides & nucleic acids》2013,32(5-6):851-866
Abstract Syntheses of two analogs of deoxyuridine with N,N-dialkylaniline chromophores are reported. 5-[3-(N-methylphenylamino)propanoyl]-2′-deoxyuridine (1) and 5-[2-(4-N,N-dimethylaminophenyl)ethyl)]-2′-deoxyuridine (2) are prepared by palladium-mediated coupling. Preparation of 2 was facilitated by in situ transient O4-trimethylsilyl protection during alkynylation which suppressed secondary cyclization of the coupling adduct. 相似文献
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Modified moving average analysis of T-wave alternans to predict ventricular fibrillation with high accuracy. 总被引:6,自引:0,他引:6
T-wave alternans is a marker of cardiac electrical instability with the potential for arrhythmia risk stratification. The modified moving average method was developed to measure alternans in settings with artifacts, noise, and nonstationary data. Algorithms were developed and performance characteristics were validated with simulated electrocardiograms (ECGs). Experimental laboratory ECGs with dynamically changing alternans values were analyzed. Alternans values estimated by modified moving average analysis correlated strongly with input alternans values (r(2) = 0.9999). Rapidly changing alternans levels and phase reversals did not perturb the measurement. When heart rate was increased from 60 to 180 beats/min, with T-wave alternans apex moving from 237 to 103 ms after the R wave, the measured alternans peak varied <5% from input value. Simulated 50- to 1,000-microV motion artifact spikes typical of treadmill ECGs produced inaccuracies <2%. Alternans values in experimental laboratory study using standard electrodes tracked vulnerability to myocardial ischemia-induced ventricular fibrillation with 100% sensitivity and specificity at a cut point of 0.75 mV. Modified moving average analysis is a robust method that precisely measures T-wave alternans in settings with artifacts, noise, and nonstationary data typical of clinical ECGs and yields an accurate estimate of risk for ventricular fibrillation. 相似文献
10.
Bruce M. Taylor Ronald W. Sarver Gregory Fici Roger A. Poorman Barry S. Lutzke Antonio Molinari Thomas Kawabe Karl Kappenman Allen E. Buhl Dennis E. Epps 《The protein journal》2003,22(1):31-40
The time dependency of the spontaneous aggregation of the fibrillogenic β-Amyloid peptide, Aβ1–40, was measured by turbidity, circular dichroism, HPLC, and fluorescence polarization. The results by all methods were comparable and they were most consistent with a kinetic model where the peptide first slowly forms an activated monomeric derivative (AM), which is the only species able to initiate, by tetramerization, the formation of linear aggregates. The anti-Aβ antibody 6E10, raised against residues 1–17, at concentrations of 200–300 nM delayed significantly the aggregation of 50 μM amyloid peptide. The anti–Aβ antibody 4G8, raised against residues 17–24, was much less active in that respect, while the antibody A162, raised against the C-terminal residues 39–43 of the full-length Aβ was totally inactive at those concentrations. Concomitant with the aggregation experiments, we also measured the time dependency of the Aβ1–40–induced toxicity toward SH-EP1 cells and hippocampal neurons, evaluated by SYTOX Green fluorescence, lactate dehydrogenase release, and activation of caspases. The extent of cell damage measured by all methods reached a maximum at the same time and this maximum coincided with that of the concentration of AM. According to the kinetic scheme, the latter is the only transient peptide species whose concentration passes through a maximum. Thus, it appears that the toxic species of Aβ1–40 is most likely the same transient activated monomer that is responsible for the nucleation of fibril formation. These conclusions should provide a structural basis for understanding the toxicity of Aβ1–40 in vitro and possibly in vivo. 相似文献