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161.
Holly Bean Florian Radu Elise De Catherine Schuler Robert E. Leggett Robert M. Levin 《Molecular and cellular biochemistry》2009,323(1-2):139-142
Partial urinary bladder outlet obstruction mediates cyclic ischemia and reperfusion resulting in the generation of both reactive oxygen species and reactive nitrogen species. It is theorized that with an increase in the level of free radicals, the level of protective antioxidants should decrease. To test this hypothesis, two electron transfer assays, the FRAP method and the CUPRAC method, were used to determine the level of antioxidant reactivity of obstructed and control bladder tissue. The results showed that the CUPRAC assay detected a significant decrease in the reactivity of antioxidants found within the obstructed bladder tissue as compared to the control bladder tissue in both the muscle and mucosa. The FRAP assay did not detect any difference between the muscle and mucosa of the obstructed and control bladder tissue. 相似文献
162.
163.
Knowledge of temporal change in ecological condition is important for the understanding and management of ecosystems. However, analyses of trends in biological condition have been rare, as there are usually too few data points at any single site to use many trend analysis techniques. We used a Bayesian hierarchical model to analyse temporal trends in stream ecological condition (as measured by the invertebrate-based index SIGNAL) across Melbourne, Australia. The Bayesian hierarchical approach assumes dependency amongst the sampling sites. Results for each site "borrow strength" from the other data because model parameter values are assumed to be drawn from a larger common distribution. This leads to robust inference despite the few data that exist at each site. Utilising the flexibility of the Bayesian approach, we also modelled change over time as a function of catchment urbanisation, allowed for potential temporal and spatial autocorrelation of the data and trend estimates, and used prior information to improve the estimate of data uncertainty. We found strong evidence of a widespread decline in SIGNAL scores for edge habitats (areas of little or no flow). The rate of decline was positively associated with catchment urbanisation. There was no evidence of such declines for riffle habitats (areas with rapid and turbulent flow). Melbourne has experienced a decline in rainfall, indicative of either drought and/or longer-term climate change. The results are consistent with the expected coupled effects of these rainfall changes and increasing urbanisation, but more research is needed to isolate a causal mechanism. More immediately, however, the Bayesian hierarchical approach has allowed us to identify a pattern in a biological monitoring data set that might otherwise have gone un-noticed, and to demonstrate a large-scale temporal decline in biological condition. 相似文献
164.
Christina Krabbe† Elise Courtois‡ Pia Jensen Jesper R. Jørgensen† Jens Zimmer Alberto Martínez-Serrano‡ Morten Meyer 《Journal of neurochemistry》2009,110(6):1908-1920
Neural stem cells constitute a promising source of cells for transplantation in Parkinson's disease, but a protocol for controlled dopaminergic differentiation is not yet available. Here we investigated the effect of the anti-apoptotic protein Bcl-xL and oxygen tension on dopaminergic differentiation and survival of a human ventral mesencephalic stem cell line (hVM1). hVM1 cells and a Bcl-xL over-expressing subline (hVMbcl-xL ) were differentiated by sequential treatment with fibroblast growth factor-8, forskolin, sonic hedgehog, and glial cell line-derived neurotrophic factor. After 10 days at 20% oxygen, hVMbcl-xL cultures contained proportionally more tyrosine hydroxylase(TH)-positive cells than hVM1 control cultures. This difference was significantly potentiated from 11 ± 0.8% to 17.2 ± 0.2% of total cells when the oxygen tension was lowered to 3%. Immunocytochemistry and Q-PCR-analysis revealed expression of several dopaminergic markers besides of TH just as dopamine was detected in the culture medium by HPLC analysis. Although Bcl-xL -over-expression reduced cell death in the cultures, it did not alter the relative content of GABAergic, neurons, while the content of astroglial cells was reduced in hVMbcl-xL cell cultures compared with control. We conclude that Bcl-xL and lowered oxygen tension act in concert to enhance dopaminergic differentiation and survival of human neural stem cells. 相似文献
165.
Raymond W. Bourdeau Enrico Malito Alexandre Chenal Brian L. Bishop Mark W. Musch Mitch L. Villereal Eugene B. Chang Elise M. Mosser Richard F. Rest Wei-Jen Tang 《The Journal of biological chemistry》2009,284(21):14645-14656
Anthrolysin O (ALO) is a pore-forming, cholesterol-dependent cytolysin
(CDC) secreted by Bacillus anthracis, the etiologic agent for
anthrax. Growing evidence suggests the involvement of ALO in anthrax
pathogenesis. Here, we show that the apical application of ALO decreases the
barrier function of human polarized epithelial cells as well as increases
intracellular calcium and the internalization of the tight junction protein
occludin. Using pharmacological agents, we also found that barrier function
disruption requires increased intracellular calcium and protein degradation.
We also report a crystal structure of the soluble state of ALO. Based on our
analytical ultracentrifugation and light scattering studies, ALO exists as a
monomer. Our ALO structure provides the molecular basis as to how ALO is
locked in a monomeric state, in contrast to other CDCs that undergo
antiparallel dimerization or higher order oligomerization in solution. ALO has
four domains and is globally similar to perfringolysin O (PFO) and
intermedilysin (ILY), yet the highly conserved undecapeptide region in domain
4 (D4) adopts a completely different conformation in all three CDCs.
Consistent with the differences within D4 and at the D2-D4 interface, we found
that ALO D4 plays a key role in affecting the barrier function of C2BBE cells,
whereas PFO domain 4 cannot substitute for this role. Novel structural
elements and unique cellular functions of ALO revealed by our studies provide
new insight into the molecular basis for the diverse nature of the CDC
family.Cholesterol-dependent cytolysins
(CDCs)4 are a family
of pore-forming toxins from many organisms, including but not limited to the
genera Archanobacterium, Bacillus, Clostridium, Listeria, and
Streptococcus. Recently, work in vertebrates has revealed that CDCs
and membrane attack complex/perforin superfamily domain-containing proteins
share a similar fold, suggesting that vertebrates use a similar mechanism for
defense against infection (1,
2). A common feature of the CDC
family is the requirement of cholesterol in the membrane to form pores
(3). In addition to
cholesterol, certain members of the family also require a cellular receptor,
such as CD59 for the toxin ILY from Streptococcus intermedius
(4). The specific mechanism by
which CDCs form pores is not completely resolved; however, what is generally
known is that ring-shaped oligomerization at the cellular membrane is followed
by large conformational changes in each unit of the oligomer, resulting in the
insertion of a β-barrel into the cellular membrane
(5). Pore formation results in
a variety of downstream signaling effects, including but not limited to the
influx of Ca2+ into the cell
(6).A good deal is known about structures of the prepore conformation of CDCs.
The crystal structures of prepore PFO, from Clostridium perfringens,
and ILY have previously been elucidated
(7,
8). Each structure shows a
characteristic four-domain architecture, in which domain 4 (D4) is involved in
membrane recognition, domain 3 (D3) is involved in β-sheet insertion, and
domain 2 (D2) is the hinge region that undergoes a large conformational change
(9-11).
Nevertheless, despite the similarities, structural differences in D4
orientation and the conformation of a highly conserved segment named the
undecapeptide region confer functional differences to PFO and ILY
(8). Noting these differences,
we decided to explore the structure and function of another member of the CDC
family, anthrolysin O (ALO).ALO is secreted by Bacillus anthracis, the etiologic agent for
anthrax. ALO is chromosomally encoded by a gene whose regulation is poorly
understood, and it is highly homologous to other members of the CDC family
(12). ALO has been shown to
have hemolytic and cytolytic activity
(13,
14). Although clinical studies
have shown that B. anthracis is weakly hemolytic
(15), anthrax bacteria do
produce biologically relevant amounts of hemolytic ALO, although the levels of
expression are under complex regulation and are dependent on the culture media
and growth conditions (12,
13,
16). At lower concentrations,
ALO can disrupt cell signaling
(13,
14). Search for a cellular
receptor of ALO has lead to the conclusion that it is a TLR4 agonist
(17). However, it is not known
that ALO binds to TLR4 directly and, if so, whether ALO also binds other
cellular receptors.In addition to ALO, B. anthracis secrete ∼400 proteins, termed
the anthrax secretome (18). Of
those, two exotoxins, edema toxin (ET) and lethal toxin (LT) have been
characterized in greatest detail. ET raises intracellular cAMP to pathologic
levels, whereas LT impairs mitogenic and stress responses by inactivating
mitogen-activating protein kinase kinase
(19,
20). The complex interplay
between these two toxins on various aspects of host cellular functions have
been demonstrated
(20-25).
ALO could also work in conjunction with other anthrax virulence factors to
modulate their cellular toxicity. For example, ALO and LF together induce
macrophage apoptosis, whereas ALO and PLC play a redundant role in a murine
inhalation anthrax model (17,
26). Interplay among anthrax
secreted factors on cells relevant to anthrax infection is just beginning to
be understood. This network of interactions is vital to the molecular basis of
how anthrax bacteria interact with the hosts during anthrax infection.Anthrax infection initiates when B. anthracis spores enter the
host through one of three routes: cutaneous, inhalational, or gastrointestinal
(GI) (27,
28). All three routes of
infection can lead to systemic infection and are ultimately lethal. Different
from inhalational anthrax, spores are ingested and germinate on or within the
epithelium of the GI tract in GI anthrax
(29). This is primarily based
on pathological observations that primary lesions of the GI tract are found in
GI anthrax, whereas no primary lesions of the lung are found in inhalational
anthrax (29). Inhalational
anthrax is a disease of choice for biological weapons because of its high
infectivity and mortality
(30). The initiation of GI
anthrax requires much higher doses of spores than inhalational anthrax, and
the molecular basis for the initiation of GI anthrax remains elusive
(31).Since the primary function of GI epithelia is to control the flux of
material into the body, disruption of this barrier can lead to movement of
bacteria into the surrounding tissue
(32). The barrier is produced
by a matrix of transmembrane and membrane-associated proteins. These cell to
cell contacts, or tight junctions, are sometimes altered during bacterial
infection to specifically disrupt the barrier function of epithelial cells.
Using a functional model for the gut epithelium, human gut epithelial Caco-2
brush border expressor (C2BBE) cells, we report that ALO decreases the barrier
function of C2BBE cells through disruption of tight junctions. We also show
that ALO disruption of barrier function is dependent on epithelial cell
polarity. We also present the crystal structure of the soluble state of ALO
and compare it with the known structures of other CDCs. In addition, we show
that ALO exists primarily as a monomer, in contrast to its closely related
homologue PFO, which exists as a dimer. Finally, we used domain swapping to
examine the structural components that confer specificity of ALO to gut
epithelial cells. 相似文献
166.
Although helping behavior is ubiquitous throughout the animal kingdom, actual rescue activity is particularly rare. Nonetheless, here we report the first experimental evidence that ants, Cataglyphis cursor, use precisely directed rescue behavior to free entrapped victims; equally important, they carefully discriminate between individuals in distress, offering aid only to nestmates. Our experiments simulate a natural situation, which we often observed in the field when collecting Catagyphis ants, causing sand to collapse in the process. Using a novel experimental technique that binds victims experimentally, we observed the behavior of separate, randomly chosen groups of 5 C. cursor nestmates under one of six conditions. In five of these conditions, a test stimulus (the “victim”) was ensnared with nylon thread and held partially beneath the sand. The test stimulus was either (1) an individual from the same colony; (2) an individual from a different colony of C cursor; (3) an ant from a different ant species; (4) a common prey item; or, (5) a motionless (chilled) nestmate. In the final condition, the test stimulus (6) consisted of the empty snare apparatus. Our results demonstrate that ants are able to recognize what, exactly, holds their relative in place and direct their behavior to that object, the snare, in particular. They begin by excavating sand, which exposes the nylon snare, transporting sand away from it, and then biting at the snare itself. Snare biting, a behavior never before reported in the literature, demonstrates that rescue behavior is far more sophisticated, exact and complexly organized than the simple forms of helping behavior already known, namely limb pulling and sand digging. That is, limb pulling and sand digging could be released directly by a chemical call for help and thus result from a very simple mechanism. However, it''s difficult to see how this same releasing mechanism could guide rescuers to the precise location of the nylon thread, and enable them to target their bites to the thread itself. 相似文献
167.
Ludovic S. Mure Pierre-Loic Cornut Camille Rieux Elise Drouyer Philippe Denis Claude Gronfier Howard M. Cooper 《PloS one》2009,4(6)
In addition to rods and cones, the human retina contains light-sensitive ganglion cells that express melanopsin, a photopigment with signal transduction mechanisms similar to that of invertebrate rhabdomeric photopigments (IRP). Like fly rhodopsins, melanopsin acts as a dual-state photosensitive flip-flop in which light drives both phototransduction responses and chromophore photoregeneration that bestows independence from the retinoid cycle required by rods and cones to regenerate photoresponsiveness following bleaching by light. To explore the hypothesis that melanopsin in humans expresses the properties of a bistable photopigment in vivo we used the pupillary light reflex (PLR) as a tool but with methods designed to study invertebrate photoreceptors. We show that the pupil only attains a fully stabilized state of constriction after several minutes of light exposure, a feature that is consistent with typical IRP photoequilibrium spectra. We further demonstrate that previous exposure to long wavelength light increases, while short wavelength light decreases the amplitude of pupil constriction, a fundamental property of IRP difference spectra. Modelling these responses to invertebrate photopigment templates yields two putative spectra for the underlying R and M photopigment states with peaks at 481 nm and 587 nm respectively. Furthermore, this bistable mechanism may confer a novel form of “photic memory” since information of prior light conditions is retained and shapes subsequent responses to light. These results suggest that the human retina exploits fly-like photoreceptive mechanisms that are potentially important for the modulation of non-visual responses to light and highlights the ubiquitous nature of photoswitchable photosensors across living organisms. 相似文献
168.
Cocklin S Jost M Robertson NM Weeks SD Weber HW Young E Seal S Zhang C Mosser E Loll PJ Saunders AJ Rest RF Chaiken IM 《Journal of molecular recognition : JMR》2006,19(4):354-362
Bacillus anthracis has recently been shown to secrete a potently hemolytic/cytolytic protein that has been designated anthrolysin O (ALO). In this work, we initiated a study of this potential anthrax virulence factor in an effort to understand the membrane-binding properties of this protein. Recombinant anthrolysin O (rALO35-512) and two N-terminally truncated versions of ALO (rALO390-512 and rALO403-512) from B. anthracis were overproduced in Escherichia coli and purified to homogeneity. The role of cholesterol in the cytolytic activity of ALO was probed in cellular cholesterol depletion assays using mouse and human macrophage-like lines, and also Drosophila Schneider 2 cells. Challenging the macrophage cells with rALO35-512, but not rALO390-512 or rALO403-512, resulted in cell death by lysis, with this cytolysis being abolished by depletion of the membrane cholesterol. Drosophila cells, which contain ergosterol as their major membrane sterol, were resistant to rALO-mediated cytolysis. In order to determine the molecular mechanism of this resistance, the interaction of rALO with model membranes comprised of POPC alone, or with a variety of structurally similar sterols including ergosterol, was probed using Biacore. Both rALO35-512 and rALO403-512 demonstrated robust binding to model membranes composed of POPC and cholesterol, with amount of protein bound proportional to the cholesterol content. Ergosterol supported greatly reduced binding of both rALO35-512 and rALO403-512, whereas other sterols tested did not support binding. The rALO403-512--membrane interaction demonstrated an equilibrium dissociation constant (KD) in the low nanomolar range, whereas rALO35-512 exhibited complex kinetics likely due to the multiple events involved in pore formation. These results establish the pivotal role of cholesterol in the action of rALO. The biosensor method developed to measure ALO recognition of cholesterol in a membrane environment could be extended to provide a platform for the screening of inhibitors of other membrane-binding proteins and peptides. 相似文献
169.
Jean-Marc Collombet Mikael Elias Guillaume Gotthard Elise Four Frédérique Renault Aurélie Joffre Dominique Baubichon Daniel Rochu Eric Chabrière 《PloS one》2010,5(2)
Background
DING proteins encompass an intriguing protein family first characterized by their conserved N-terminal sequences. Some of these proteins seem to have key roles in various human diseases, e.g., rheumatoid arthritis, atherosclerosis, HIV suppression. Although this protein family seems to be ubiquitous in eukaryotes, their genes are consistently lacking from genomic databases. Such a lack has considerably hampered functional studies and has fostered therefore the hypothesis that DING proteins isolated from eukaryotes were in fact prokaryotic contaminants.Principal Findings
In the framework of our study, we have performed a comprehensive immunological detection of DING proteins in mice. We demonstrate that DING proteins are present in all tissues tested as isoforms of various molecular weights (MWs). Their intracellular localization is tissue-dependant, being exclusively nuclear in neurons, but cytoplasmic and nuclear in other tissues. We also provide evidence that germ-free mouse plasma contains as much DING protein as wild-type.Significance
Hence, data herein provide a valuable basis for future investigations aimed at eukaryotic DING proteins, revealing that these proteins seem ubiquitous in mouse tissue. Our results strongly suggest that mouse DING proteins are endogenous. Moreover, the determination in this study of the precise cellular localization of DING proteins constitute a precious evidence to understand their molecular involvements in their related human diseases. 相似文献170.
Yolima Carrillo Elise Pendall Feike A. Dijkstra Jack A. Morgan Joanne M. Newcomb 《Plant and Soil》2011,347(1-2):339-350
Warming and elevated atmospheric CO2 (eCO2) can elicit contrasting responses on different SOM pools, thus to understand the effects of combined factors it is necessary to evaluate individual pools. Over two years, we assessed responses to eCO2 and warming of SOM pools, their susceptibility to decomposition, and whether these responses were mediated by plant inputs in a semi-arid grassland at the PHACE (Prairie Heating and CO2 Enrichment) experiment. We used long-term soil incubations and assessed relationships between plant inputs and the responses of the labile and resistant pools. We found strong and contrasting effects of eCO2 and warming on the labile C pool. In 2008 labile C was increased by eCO2 and was positively related to plant biomass. In contrast, in 2007 eCO2 and warming had interactive effects on the labile C, and the pool size was not related to plant biomass. Effects of warming and eCO2 in this year were consistent withtreatment effects on soil moisture and temperature and their effects on labile C decomposition. The decomposition rate of the resistant C was positively related to indicators of plant C inputs. Our approach demonstrated that SOM pools in this grassland can have early and contrasting responses to climate change factors. The labile C pool in the mixed-grass prairie was highly responsive to eCO2 and warming but the factors behind such responses were highly dynamic across years. Results suggest that in this grassland the resistant C pool could be negatively affected by increases in plant-production driven available soil C. 相似文献