101 research outputs found
Evaluation of 4,6-Diamidino-2-phenylindole as a Fluorescent Probe Substrate for Rapid Assays of the Functionality of Human Multidrug and Toxin Extrusion Proteins
ABSTRACT: Multidrug and toxin extrusion protein 1 (MATE1) and MATE2-K are organic cation/H ؉ antiporters that have recently been identified and suggested to be responsible for the brush border secretory transport of many cationic drugs in renal tubules. We here report our finding that 4,6-diamidino-2-phenylindole (DAPI) can be used as a probe substrate for rapid assays of the functionality of the human MATEs, hMATE1, and hMATE2-K, by taking advantage of its fluorescent nature. The specific cellular uptakes of DAPI by cloned hMATE1 and hMATE2-K, which were assessed by fluorescence intensity, were found to be rapid and saturable with the Michaelis constants of 1.13 and 3.16 M, respectively, indicating that DAPI is a good substrate of both hMATEs. It was found that many organic cations inhibit the specific uptake of DAPI by hMATE1 and hMATE2-K, and the extents of inhibition are in good correlation with those of inhibition of the specific uptake of [ 3 H]cimetidine as a typical substrate, indicating comparable performances of both substrates as probes in identifying inhibitors. Thus, DAPI can be an alternative probe substrate that enables fluorometric rapid assays of the functionality of both hMATEs. It was also found that the other major renal organic cation transporters, human organic cation transporter 2 (hOCT2), hOCT3, human novel organic cation transporter 1 (hOCTN1), and hOCTN2, cannot transport DAPI, although hOCT1, which is mainly expressed in the liver, can. Therefore, the DAPI uptake assay can be a method specific to the hMATEs among organic cation transporters in the human kidney
Type II cGMP-dependent protein kinase negatively regulates fibroblast growth factor signaling by phosphorylating Raf-1 at serine 43 in rat chondrosarcoma cells
Although type II cGMP-dependent protein kinase (PKGII) is a major downstream effector of cGMP in chondrocytes and attenuates the FGF receptor 3/ERK signaling pathway, its direct target proteins have not been fully explored. In the present study, we attempted to identify PKGII-targeted proteins, which are associated with the inhibition of FGF-induced MAPK activation. Although FGF2 stimulation induced the phosphorylation of ERK1/2, MEK1/2, and Raf-1 at Ser-338 in rat chondrosarcoma cells, pretreatment with a cell-permeable cGMP analog strongly inhibited their phosphorylation. On the other hand, Ser-43 of Raf-1 was phosphorylated by cGMP in a dose-dependent manner. Therefore, we examined the direct phosphorylation of Raf-1 by PKGII. Wild-type PKGII phosphorylated Raf-1 at Ser-43 in a cGMP-dependent manner, but a PKGII D412A/R415A mutant, which has a low affinity for cGMP, did not. Finally, we found that a phospho-mimic mutant, Raf-1 S43D, suppressed FGF2-induced MAPK pathway. These results suggest that PKGII counters FGF-induced MEK/ERK activation through the phosphorylation of Raf-1 at Ser-43 in chondrocytes
The effects of the herbal medicine Daikenchuto (TJ-100) after esophageal cancer resection, open-label, randomized controlled trial
Background
Daikenchuto (TJ-100), a traditional Japanese herbal medicine, is widely used in Japan. Its effects on gastrointestinal motility and microcirculation and its anti-inflammatory effect are known. The purpose of this prospective randomized controlled trial was to investigate the effect of TJ-100 after esophagectomy in esophageal cancer patients.
Methods
Forty patients for whom subtotal esophageal resection for esophageal cancer was planned at our institute from March 2011 to August 2013 were enrolled and divided into two groups at the point of determination of the operation schedule after informed consent was obtained: a TJ-100 (15 g/day)-treated group (n = 20) and a control group (n = 20). The primary efficacy end-points were maintenance of the nutrition condition and the recovery of gastrointestinal function. The secondary efficacy end-points were the serum C-reactive protein (CRP) level and adrenomedullin level during the postoperative course, the incidence of postoperative complications, and the length of hospital stay after surgery.
Results
We examined 39 patients because one patient in the TJ-100 group was judged as having unresectable cancer after surgery. The mean age of the TJ-100 group patients was significantly older than that of the control group patients.The rate of body weight decrease at postoperative day 21 was significantly suppressed in the TJ-100 group (3.6% vs. the control group: 7.0%, p = 0.014), but the serum albumin level was not significantly different between the groups. The recovery of gastrointestinal function regarding flatus, defecation, and oral intake showed no significant between-group differences, but postoperative bowel symptoms tended to be rare in the TJ-100 group. There was no significant between-group difference in the length of hospital stay after surgery. The serum CRP level at postoperative day 3 was 4.9 mg/dl in the TJ-100 group and 6.9 mg/dl in the control group, showing a tendency of a suppressed serum CRP level in the TJ-100 group (p = 0.126). The rate of increase in adrenomedullin tended to be high postoperatively, but there was no significant difference between the two groups.
Conclusions
TJ-100 treatment after esophageal cancer resection has the effects of prompting the recovery of gastrointestinal motility and minimizing body weight loss, and it might suppress the excess inflammatory reaction related to surgery
Pathobiological implications of mucin (MUC) expression in the outcome of small bowel cancer.
Mucins have been associated with survival in various cancer patients, but there have been no studies of mucins in small bowel carcinoma (SBC). In this study, we investigated the relationships between mucin expression and clinicopathologic factors in 60 SBC cases, in which expression profiles of MUC1, MUC2, MUC3, MUC4, MUC5AC, MUC6 and MUC16 in cancer and normal tissues were examined by immunohistochemistry. MUC1, MUC5AC and MUC16 expression was increased in SBC lesions compared to the normal epithelium, and expression of these mucins was related to clinicopathologic factors, as follows: MUC1 [tumor location (p = 0.019), depth (p = 0.017) and curability (p = 0.007)], MUC5AC [tumor location (p = 0.063) and lymph node metastasis (p = 0.059)], and MUC16 [venous invasion (p = 0.016) and curability (p = 0.016)]. Analysis of 58 cases with survival data revealed five factors associated with a poor prognosis: poorly-differentiated or neuroendocrine histological type (
The ASTRO-H X-ray Observatory
The joint JAXA/NASA ASTRO-H mission is the sixth in a series of highly
successful X-ray missions initiated by the Institute of Space and Astronautical
Science (ISAS). ASTRO-H will investigate the physics of the high-energy
universe via a suite of four instruments, covering a very wide energy range,
from 0.3 keV to 600 keV. These instruments include a high-resolution,
high-throughput spectrometer sensitive over 0.3-2 keV with high spectral
resolution of Delta E < 7 eV, enabled by a micro-calorimeter array located in
the focal plane of thin-foil X-ray optics; hard X-ray imaging spectrometers
covering 5-80 keV, located in the focal plane of multilayer-coated, focusing
hard X-ray mirrors; a wide-field imaging spectrometer sensitive over 0.4-12
keV, with an X-ray CCD camera in the focal plane of a soft X-ray telescope; and
a non-focusing Compton-camera type soft gamma-ray detector, sensitive in the
40-600 keV band. The simultaneous broad bandpass, coupled with high spectral
resolution, will enable the pursuit of a wide variety of important science
themes.Comment: 22 pages, 17 figures, Proceedings of the SPIE Astronomical
Instrumentation "Space Telescopes and Instrumentation 2012: Ultraviolet to
Gamma Ray
The Quiescent Intracluster Medium in the Core of the Perseus Cluster
Clusters of galaxies are the most massive gravitationally-bound objects in
the Universe and are still forming. They are thus important probes of
cosmological parameters and a host of astrophysical processes. Knowledge of the
dynamics of the pervasive hot gas, which dominates in mass over stars in a
cluster, is a crucial missing ingredient. It can enable new insights into
mechanical energy injection by the central supermassive black hole and the use
of hydrostatic equilibrium for the determination of cluster masses. X-rays from
the core of the Perseus cluster are emitted by the 50 million K diffuse hot
plasma filling its gravitational potential well. The Active Galactic Nucleus of
the central galaxy NGC1275 is pumping jetted energy into the surrounding
intracluster medium, creating buoyant bubbles filled with relativistic plasma.
These likely induce motions in the intracluster medium and heat the inner gas
preventing runaway radiative cooling; a process known as Active Galactic
Nucleus Feedback. Here we report on Hitomi X-ray observations of the Perseus
cluster core, which reveal a remarkably quiescent atmosphere where the gas has
a line-of-sight velocity dispersion of 164+/-10 km/s in a region 30-60 kpc from
the central nucleus. A gradient in the line-of-sight velocity of 150+/-70 km/s
is found across the 60 kpc image of the cluster core. Turbulent pressure
support in the gas is 4% or less of the thermodynamic pressure, with large
scale shear at most doubling that estimate. We infer that total cluster masses
determined from hydrostatic equilibrium in the central regions need little
correction for turbulent pressure.Comment: 31 pages, 11 Figs, published in Nature July
Hitomi (ASTRO-H) X-ray Astronomy Satellite
The Hitomi (ASTRO-H) mission is the sixth Japanese x-ray astronomy satellite developed by a large international collaboration, including Japan, USA, Canada, and Europe. The mission aimed to provide the highest energy resolution ever achieved at E > 2 keV, using a microcalorimeter instrument, and to cover a wide energy range spanning four decades in energy from soft x-rays to gamma rays. After a successful launch on February 17, 2016, the spacecraft lost its function on March 26, 2016, but the commissioning phase for about a month provided valuable information on the onboard instruments and the spacecraft system, including astrophysical results obtained from first light observations. The paper describes the Hitomi (ASTRO-H) mission, its capabilities, the initial operation, and the instruments/spacecraft performances confirmed during the commissioning operations for about a month
Peptide Carrier-Mediated Transport in Intestinal Brush Border Membrane Vesicles of Rats and Rabbits: Cephradine Uptake and Inhibition
The uptake kinetics of cephradine, an amino-β-lactam antibiotic, were studied in rat and rabbit intestinal brush border membrane vesicles preparations using both the Ca 2+ and the Mg 2+ methods of preparation, in the presence of an inward proton gradient. The Ca 2+ method demonstrated greater uptake of cephradine in intestinal brush border vesicles prepared from both rat and rabbit and was used for these studies. The transport was observed to be of Michaelis–Menten carrier-mediated type with a passive transport component. The kinetic parameters obtained were as follows: for rat and rabbit, respectively, K m , 1.6 and 1.9 m M ; J max ′, 1.7 and 20.7 nmol/mg/min; P c ′ (= J max ′/ K m ), 1.1 and 10.9 µL/mg/min; and P m ′, 0.4 and 0.8 µL/mg/min. The kinetic parameters for the rat vesicles are consistent with those from our previous perfusion study using a conversion factor of 0.71 cm 2 /mg protein. The rabbit vesicles exhibited a similar Michaelis constant and a 10-fold larger maximal transport velocity, suggesting a quantitative advantage for the study of carrier-mediated transport in the rabbit compared to rat vesicles from the intestine. Cephradine uptake was inhibited by phenylpropionylproline, a proline derivative, and enalapril, an ACE inhibitor, which do not have an α-amino group, as well as dipeptides, tripeptides, and amino-β-lactam antibiotics in both rat and rabbit vesicles. These results support the suggestion that they share the same peptide carrier pathway for oral absorption and that the vesicles may be a useful tool in developing orally effective peptide-type drugs.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/41430/1/11095_2004_Article_304789.pd
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