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991.
Protective mechanisms during ischemic tolerance in skeletal muscle   总被引:3,自引:0,他引:3  
The purpose of this study was to test specific mechanisms of protection afforded the rat extensor digitorum longus (EDL) muscle during ischemic tolerance. Two days following five cycles of 10 min ischemia and 10 min reperfusion, heme oxygenase (HO) and calcium-dependent nitric oxide synthase (cNOS) activities were increased 2- and 2.5-fold (p <.05), respectively. Interestingly, calcium-independent NOS (iNOS) activity was completely downregulated (p <.05). The levels of superoxide dismutase (SOD) and catalase were increased 2-fold (p <.05), while glutathione peroxidase activity remained unchanged from non-preconditioned controls. Using intravital microscopy combined with chromium mesoporphyrin (CrMP), a selective HO inhibitor, and l-NAME, a NOS inhibitor, the roles of HO and cNOS were evaluated. Ischemic tolerance in the EDL muscle, 48 h after the preconditioning stimulus, was characterized by complete protection from both microvascular perfusion deficits and tissue injury after a 2-h period of ischemia. Removal of NOS activity completely removed the benefit afforded microvascular perfusion, while inhibition of HO activity prevented the parenchymal protection. These data suggest that ischemic tolerance within skeletal muscle is associated with the upregulation of specific cytoprotective proteins and that the benefits afforded by cNOS and HO activity are spatially discrete to the microvasculature and parenchyma, respectively.  相似文献   
992.
A molecular beacon-based real-time NASBA assay for detection and identification of Mycobacterium avium subsp. paratuberculosis has been developed. It targets and amplifies sequences from the dnaA gene which are specific for this bacterium. The assay includes an internal amplification control, to allow identification of inhibited reactions. The assay was tested against 18 isolates of M. avium subsp. paratuberculosis, 17 other mycobacterial strains and 25 non-mycobacterial strains, and was fully selective in that it detected all the targets but none of the non-targets. The lowest number of cells which the assay can detect with 99% probability is 150-200 cells per reaction (as determined using pure culture suspensions). Using centrifugation and nucleic acid extraction as sample treatment, the assay was able to consistently detect 10(3) M. avium subsp. paratuberculosis cells in 20 ml artificially contaminated drinking water. With a simple detergent and enzymatic sample pretreatment before centrifugation and nucleic acid extraction, the assay was able to consistently detect 10(4) M. avium subsp. paratuberculosis cells in 20 ml artificially contaminated semi-skimmed milk. The assay will be a useful addition to the range of diagnostic tools available for the study of M. avium subsp. paratuberculosis.  相似文献   
993.
Colonies of co-operatively breeding African mole-rats have traditionally been thought to be composed of a single breeding female, one or two breeding males, and their offspring. In the naked mole-rat (Heterocephalus glaber), the occurrence of facultative inbreeding means incest avoidance cannot prevent reproduction in subordinate group members, and physiological suppression of reproductive function by the breeding female occurs in both sexes. In contrast, previous studies of captive colonies of the Damaraland mole-rat (Cryptomys damarensis) suggest that breeding within a colony is restricted to a single breeding pair, simply because all other colony members are highly related (first- or second-order relatives) and this species is an obligate outbreeder. Using microsatellite markers, we investigated parentage and colony composition in 18 wild Damaraland mole-rat colonies to determine whether inbreeding avoidance alone can explain the high levels of reproductive skew in this species. Multiple and unidentified paternity was widespread within colonies and immigrants of both sexes were regularly identified. Unrelated, opposite-sex nonbreeders were found coexisting in two colonies. These results suggest that, in the wild, conditions exist where nonreproductive females can come into contact with unrelated males, even when they do not disperse from their natal colony. Inbreeding avoidance alone is therefore insufficient to maintain the high levels of reproductive skew identified in this species suggesting that the breeding female somehow suppresses the reproductive function in nonbreeding females.  相似文献   
994.
TMADH (trimethylamine dehydrogenase) is a complex iron-sulphur flavoprotein that forms a soluble electron-transfer complex with ETF (electron-transferring flavoprotein). The mechanism of electron transfer between TMADH and ETF has been studied using stopped-flow kinetic and mutagenesis methods, and more recently by X-ray crystallography. Potentiometric methods have also been used to identify key residues involved in the stabilization of the flavin radical semiquinone species in ETF. These studies have demonstrated a key role for 'conformational sampling' in the electron-transfer complex, facilitated by two-site contact of ETF with TMADH. Exploration of three-dimensional space in the complex allows the FAD of ETF to find conformations compatible with enhanced electronic coupling with the 4Fe-4S centre of TMADH. This mechanism of electron transfer provides for a more robust and accessible design principle for interprotein electron transfer compared with simpler models that invoke the collision of redox partners followed by electron transfer. The structure of the TMADH-ETF complex confirms the role of key residues in electron transfer and molecular assembly, originally suggested from detailed kinetic studies in wild-type and mutant complexes, and from molecular modelling.  相似文献   
995.
To study canine herpesvirus (CHV) reactivation from red foxes (Vulpes vulpes), 29 foxes with varying CHV antibody and CHV carrier status were treated with methylprednisolone acetate, a glucocorticosteroid drug with prolonged immunosuppressive effect in dogs. In the first experiment, 17 foxes with unknown CHV carrier status were treated once with methylprednisolone: in the second experiment, five foxes were treated twice, 4 mo after being intravenously CHV infected; and in the third experiment, six foxes were treated five times, 11 mo after peroral CHV infection. Infectious CHV was not isolated after treatment from either naturally or experimentally CHV-infected foxes or from untreated, CHV-seronegative in-contact foxes. Canine herpesvirus DNA was not detectable in mucosal secretions or white blood cells of any of the foxes, whereas all trigeminal ganglia of experimentally CHV-infected foxes were polymerase chain reaction-positive. In CHV-seropositive foxes, anti-CHV antibody titers did not change with time after treatment, and CHV-seronegative in-contact controls did not seroconvert. Hematologic parameters remained mostly unchanged. We conclude that CHV is not as easily reactivated in foxes following corticosteroid treatment as in dogs, although there was no obvious sign of immunosuppression. Canine herpesvirus was not spread from virus carriers to naive in-contact foxes, which may be among possible explanations for the reported low CHV prevalence in wild foxes.  相似文献   
996.
We studied whether the peptides of nine amino acids (9-mers) that are typically used in MHC class I presentation are sufficiently unique for self:nonself discrimination. The human proteome contains 28,783 proteins, comprising 107 distinct 9-mers. Enumerating distinct 9-mers for a variety of microorganisms we found that the average overlap, i.e., the probability that a foreign peptide also occurs in the human self, is about 0.2%. This self:nonself overlap increased when shorter peptides were used, e.g., was 30% for 6-mers and 3% for 7-mers. Predicting all 9-mers that are expected to be cleaved by the immunoproteasome and to be translocated by TAP, we find that about 25% of the self and the nonself 9-mers are processed successfully. For the HLA-A*0201 and HLA-A*0204 alleles, we predicted which of the processed 9-mers from each proteome are expected to be presented on the MHC. Both alleles prefer to present processed 9-mers to nonprocessed 9-mers, and both have small preference to present foreign peptides. Because a number of amino acids from each 9-mer bind the MHC, and are therefore not exposed to the TCR, antigen presentation seems to involve a significant loss of information. Our results show that this is not the case because the HLA molecules are fairly specific. Removing the two anchor residues from each presented peptide, we find that the self:nonself overlap of these exposed 7-mers resembles that of 9-mers. Summarizing, the 9-mers used in MHC class I presentation tend to carry sufficient information to detect nonself peptides amongst self peptides.  相似文献   
997.
Several independent studies of bacterial degradation of nitrate ester explosives have demonstrated the involvement of flavin-dependent oxidoreductases related to the old yellow enzyme (OYE) of yeast. Some of these enzymes also transform the nitroaromatic explosive 2,4,6-trinitrotoluene (TNT). In this work, catalytic capabilities of five members of the OYE family were compared, with a view to correlating structure and function. The activity profiles of the five enzymes differed substantially; no one compound proved to be a good substrate for all five enzymes. TNT is reduced, albeit slowly, by all five enzymes. The nature of the transformation products differed, with three of the five enzymes yielding products indicative of reduction of the aromatic ring. Our findings suggest two distinct pathways of TNT transformation, with the initial reduction of TNT being the key point of difference between the enzymes. Characterization of an active site mutant of one of the enzymes suggests a structural basis for this difference.  相似文献   
998.
BACKGROUND: Experiments on the fatigue of tendons have shown that cyclic loading induces failure at stresses lower than the ultimate tensile strength (UTS) of the tendons. The number of cycles to failure (Nf) has been shown to be dependent upon the magnitude of the applied cyclic stress. METHOD OF APPROACH: Utilizing data collected by Schechtman (1995), we demonstrate that the principles of Linear Elastic Fracture Mechanics (LEFM) can be used to predict the fatigue behavior of tendons under cyclic loading for maximum stress levels that are higher than 10% of the ultimate tensile strength (UTS) of the tendon (the experimental results at 10% UTS did not fit with our equations). CONCLUSIONS: LEFM and other FM approaches may prove to be very valuable in advancing our understanding of damage accumulation in soft connective tissues.  相似文献   
999.
Starting from 6-(4'-methoxyphenyl)fulvene (1a), 6-(2',4',6'-trimethoxyphenyl)fulvene (1b), or 6-(3',5'-dimethoxyphenyl)fulvene (1c), [1,2-di(cyclopentadienyl)-1,2-di(4'-methoxyphenyl)-ethanediyl] titanium dichloride (2a), [1,2-di(cyclopentadienyl)-1,2-bis(2',4',6'-trimethoxyphenyl)-ethanediyl] titanium dichloride (2b), and [1,2-di(cyclopentadienyl)-1,2-bis(3',5'-dimethoxyphenyl)-ethanediyl] titanium dichloride (2c) were synthesised. When titanocenes 2a-c were tested against pig kidney carcinoma cells (LLC-PK) inhibitory concentrations (IC50) of 2.8 x 10(-4), 3.6 x 10(-4) and 2.1 x 10(-4) M, respectively, were observed.  相似文献   
1000.
Horseradish peroxidase: a modern view of a classic enzyme   总被引:18,自引:0,他引:18  
Veitch NC 《Phytochemistry》2004,65(3):249-259
Horseradish peroxidase is an important heme-containing enzyme that has been studied for more than a century. In recent years new information has become available on the three-dimensional structure of the enzyme and its catalytic intermediates, mechanisms of catalysis and the function of specific amino acid residues. Site-directed mutagenesis and directed evolution techniques are now used routinely to investigate the structure and function of horseradish peroxidase and offer the opportunity to develop engineered enzymes for practical applications in natural product and fine chemicals synthesis, medical diagnostics and bioremediation. A combination of horseradish peroxidase and indole-3-acetic acid or its derivatives is currently being evaluated as an agent for use in targeted cancer therapies. Physiological roles traditionally associated with the enzyme that include indole-3-acetic acid metabolism, cross-linking of biological polymers and lignification are becoming better understood at the molecular level, but the involvement of specific horseradish peroxidase isoenzymes in these processes is not yet clearly defined. Progress in this area should result from the identification of the entire peroxidase gene family of Arabidopsis thaliana, which has now been completed.  相似文献   
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