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21.
Silva Vde A Cargnelutti MT Giesel GM Palmieri LC Monteiro RQ Verli H Lima LM 《PloS one》2011,6(9):e24735
Thrombin is a serine proteinase that plays a fundamental role in coagulation. In this study, we address the effects of ligand site recognition by alpha-thrombin on conformation and energetics in solution. Active site occupation induces large changes in secondary structure content in thrombin as shown by circular dichroism. Thrombin-D-Phe-Pro-Arg-chloromethyl ketone (PPACK) exhibits enhanced equilibrium and kinetic stability compared to free thrombin, whose difference is rooted in the unfolding step. Small-angle X-ray scattering (SAXS) measurements in solution reveal an overall similarity in the molecular envelope of thrombin and thrombin-PPACK, which differs from the crystal structure of thrombin. Molecular dynamics simulations performed with thrombin lead to different conformations than the one observed in the crystal structure. These data shed light on the diversity of thrombin conformers not previously observed in crystal structures with distinguished catalytic and conformational behaviors, which might have direct implications on novel strategies to design direct thrombin inhibitors. 相似文献
22.
Thereza de A. Garbelotto Luiz A. Campos Jocelia Grazia 《Zoological Journal of the Linnean Society》2013,168(3):452-472
Alitocoris Sailer, 1950, consists of four valid species described from Central America. In a recent cladistic analysis of Ochlerini, the genus was considered paraphyletic in the Herrichella Distant, 1911, group of taxa. The present study provides a cladistic analysis of the Herrichella clade, using 88 morphological characters and 40 taxa representing 21 genera of Ochlerini, including all known species of Alitocoris plus 16 new species. Outgroups included Eritrachys bituberculata Ruckes, 1959, Phereclus pluto Stål, 1862, and Adoxoplatys comis Breddin, 1903, with the last used for rooting. The cladistic analysis was conducted using TNT under heuristic searches and implied weighting of characters; 11 K‐values calculated for an average character fit ranged from 50 to 90% of a perfectly hierarchical character. The results corroborated the paraphyly of Alitocoris, calling for changes in the classification of the genus with the proposition of three new genera for two, three, and ten species, respectively, that will be described elsewhere. Alitocoris is redescribed and a key for the species is presented. Alitocoris brunneus, Alitocoris maculosus, and Alitocoris parvus are removed from the genus, and the new species Alitocoris grandis sp. nov. , Alitocoris lateralis sp. nov. , and Alitocoris ornatus sp. nov. are described. © 2013 The Linnean Society of London 相似文献
23.
Alfred L.M. Bothwell Michael Paskind Michael Reth Thereza Imanishi-Kari Klaus Rajewsky David Baltimore 《Cell》1981,24(3):625-637
To examine germ line genes of the heavy chain variable region (VH) that might contribute to formation of antibodies of the NPb family, we have derived cDNA clones from two hybridomas making NPb antibodies. One, B1–8, made an IgM protein and was derived during a primary response; the other, S43, made an IgG2a protein and was derived during a hyperimmune response. Sequence comparison of the two clones showed that they differed by only 10 bp in the VH region, had very different D segments and had identical J segments (J2). A set of closely related germ line VH genes was then cloned from a partial Eco RI library of C57BI/6 DNA. By comparing the germ line VH regions to the cDNA VH regions, we identified seven potential candidates for encoding the VH regions of NPb antibodies. The seven VH regions were sequenced, and one V(186-2) contained exactly the DNA sequence found in the clone derived from B1–8. None of the DNA sequence differences that distinguished the S43-derived clone from the B1–8 clone was found in any of the other six germ line genes. Because the S43 sequence was more closely related to the V(186-2) germ line sequence than to any of the other VH genes, we conclude that the differences between the genes resulted from somatic mutation and that the two hybridomas derived their VH regions from the same germ line gene. Certain of the sequenced VH genes contain crippling mutations; the repertoire of germ line VH genes that can contribute to the diversity of antibodies may therefore be less than the total number of genes detectable by hybridization. 相似文献
24.
To assess the significance of somatic point mutation in the hyperimmune response to the hapten NP, an in vivo enrichment procedure was followed. Mice that expressed high titers of B1-8 idiotopic determinants were selected as donors for serial transfer of small numbers of immune spleen cells into syngeneic irradiated recipient mice. Cells expressing B1-8 idiotopic determinants were chosen for enrichment because B1-8 cross-reactive determinants constitute a significant portion of the primary response. Furthermore, B1-8 is a monoclonal antibody derived from a primary response to NP, and its heavy and light chains are unmutated products of the germ-line genes VH186.2 and VL lambda 1, respectively. The germ-line sequence is thus available for comparison with the somatic mutants that arise during enrichment and hyperimmunization. The data show that serial transfer of spleen cells from mice with a high titer of idiotypic determinants results in a dramatic decrease in the titers of antibodies that bind antigen. Three lines of evidence indicate that progeny cells from the initial lambda-positive, idiotype-bearing, antigen-binding cells are successfully transferred and expanded during successive adoptive transfers. First, the proportion of lambda-bearing antibodies relative to NP-specific lambda-bearing antibodies increases with transfer, which is consistent with mutation away from antigen binding. Second, analysis of serum antibodies and hybridoma proteins derived from transfer-recipient mice confirm the presence of idiotype-positive antibodies that do not bind antigen. Third, RNA dot blot analysis of hybridomas constructed from a recipient mouse in the fourth transfer indicates a high frequency of expression of the VH gene predominantly used in the NP response. Many of the antibodies expressed by these hybridomas not only do not bind antigen, but have also lost the determinants recognized by the anti-idiotypic reagents. Most of these VH-positive hybridomas express lambda L chain. The most likely interpretation of the data is that somatic mutation is occurring during the hyperimmune response. Because we selected donor mice that expressed a high titer of idiotype-positive, antigen-specific antibody and immunized the recipient mice, we expected to observe a selective expansion of somatic variants that bound antigen. This was not the case. The observed loss of antigen binding suggests that the majority of mutations arising result in antibodies with lower affinity for the immunizing antigen. 相似文献
25.
Alexandre A. Hoeller Ana Paula R. Costa Maíra A. Bicca Filipe C. Matheus Gilliard Lach Francesca Spiga Stafford L. Lightman Roger Walz Graham L. Collingridge Zuner A. Bortolotto Thereza C. M. de Lima 《PloS one》2016,11(1)
Extensive evidence indicates the influence of the cholinergic system on emotional processing. Previous findings provided new insights into the underlying mechanisms of long-term anxiety, showing that rats injected with a single systemic dose of pilocarpine—a muscarinic receptor (mAChR) agonist—displayed persistent anxiogenic-like responses when evaluated in different behavioral tests and time-points (24 h up to 3 months later). Herein, we investigated whether the pilocarpine-induced long-term anxiogenesis modulates the HPA axis function and the putative involvement of NMDA receptors (NMDARs) following mAChRs activation. Accordingly, adult male Wistar rats presented anxiogenic-like behavior in the elevated plus-maze (EPM) after 24 h or 1 month of pilocarpine injection (150 mg/kg, i.p.). In these animals, mAChR activation disrupted HPA axis function inducing a long-term increase of corticosterone release associated with a reduced expression of hippocampal GRs, as well as consistently decreased NMDAR subunits expression. Furthermore, in another group of rats injected with memantine–an NMDARs antagonist (4 mg/kg, i.p.)–prior to pilocarpine, we found inhibition of anxiogenic-like behaviors in the EPM but no further alterations in the pilocarpine-induced NMDARs downregulation. Our data provide evidence that behavioral anxiogenesis induced by mAChR activation effectively yields short- and long-term alterations in hippocampal NMDARs expression associated with impairment of hippocampal inhibitory regulation of HPA axis activity. This is a novel mechanism associated with anxiety-like responses in rats, which comprise a putative target to future translational studies. 相似文献
26.
André M. Lopes Pérola O. Magalhães Priscila G. Mazzola Carlota O. Rangel‐Yagui João C. M. de Carvalho Thereza C.V. Penna Adalberto Pessoa Jr 《Biotechnology progress》2010,26(6):1644-1653
In biotechnology, endotoxin (LPS) removal from recombinant proteins is a critical and challenging step in the preparation of injectable therapeutics, as endotoxin is a natural component of bacterial expression systems widely used to manufacture therapeutic proteins. The viability of large‐scale industrial production of recombinant biomolecules of pharmaceutical interest significantly depends on the separation and purification techniques used. The aim of this work was to evaluate the use of aqueous two‐phase micellar system (ATPMS) for endotoxin removal from preparations containing recombinant proteins of pharmaceutical interest, such as green fluorescent protein (GFPuv). Partition assays were carried out initially using pure LPS, and afterwards in the presence of E. coli cell lysate. The ATPMS technology proved to be effective in GFPuv recovery, preferentially into the micelle‐poor phase (KGFPuv < 1.00), and LPS removal into the micelle‐rich phase (%REMLPS > 98.00%). Therefore, this system can be exploited as the first step for purification in biotechnology processes for removal of higher LPS concentrations. © 2010 American Institute of Chemical Engineers Biotechnol. Prog., 2010 相似文献
27.
Capillaria hepatica in rats: focal parasitic hepatic lesions and septal fibrosis run independent courses 总被引:1,自引:0,他引:1
Gomes AT Cunha LM Bastos CG Medrado BF Assis BC Andrade ZA 《Memórias do Instituto Oswaldo Cruz》2006,101(8):895-898
Capillaria hepatica causes two main lesions in the liver of rats: multifocal chronic inflammation, directly related to the presence of disintegrating parasites and their eggs, and a process of systematized septal fibrosis. The comparative behavior of these two lesions was investigated in rats experimentally infected with 600 embryonated eggs, following either corticosteroid treatment or specific antigenic stimulation, in an attempt to understand the relationship between these two lesions, and the pathogenesis of septal fibrosis. The two treatments differently modified the morphological aspects of the focal parasitic-related lesions, but did not interfere with the presentation of diffuse septal fibrosis, although a mild decrease in the degree of fibrosis occurred in corticoid-treated animals. These findings indicate that although the two lesions are C. hepatica induced, they are under different pathogenetic control, the induction of septal fibrosis being triggered during early infection to follow an independent pathway. 相似文献
28.
Verícimo MA França KM Arnholdt AC Kipnis TL 《Microbes and infection / Institut Pasteur》2006,8(12-13):2811-2820
Paracoccidiodomycosis (PCM) is a systemic mycosis that presents a wide spectrum of clinical manifestations caused by Paracoccidiodes brasiliensis. The experimental murine model has been used to approach the disease with susceptible and resistant mouse strains that reproduce most of the main human immunological features. We investigated whether the pattern of apoptosis of peritoneal cells from two polar strains of mice after infection with P. brasiliensis could be associated with the susceptibility or resistance to this pathogen. Apoptosis of A/J mouse cells (resistant), cultured in the presence or absence of LPS as stimuli, was observed as early as on the first day of infection. Cells from the infected susceptible strain BALB/c did not exhibit apoptosis in absence of LPS and persistently at a lesser degree than that observed in resistant mice. The apoptosis induced by the infection in resistant mice was not due to nitric oxide, since its blockage either in vitro or in vivo did not revert it. Analysis of additional strains of polar susceptibilities to PCM assured the dissociation of NO production and apoptosis. Interestingly, IL-6 and IL-10 were secreted in high amounts, by BALB/c cells and might be involved in shielding cells from apoptosis induced by P. brasiliensis. Furthermore, IFNgamma(-/-) mice did not show apoptosis of peritoneal cells while the Wt controls presented levels similar to those of A/J strain that secreted high amounts of IFNgamma and IL-1beta. The expression of Fas was increased in both strains and in Wt mice, whereas FasL was decreased in the susceptible strain and not significantly modulated in TNFRI and IFNgamma KO mice. These results suggest that apoptosis might be a mechanism of control of engagement of cells that could otherwise contribute to the susceptible phenotype observed in some strains of mice. 相似文献
29.
Thereza C. C. Medeiros Everardo V. S. B. Sampaio 《Wetlands Ecology and Management》2013,21(3):209-217
Leaves are major components of mangrove productivity, but data on leaf dynamics are scarce. We marked the shoot tips of three species in four sites of a riverine mangrove and monitored leaf formation, senescence and abscission and flower formation. The leaf area and biomass in the mangrove were estimated using phytosociological data. Leaf size and formation were similar among the four sites. The tips of Rhizophora mangle had more leaf scars (41), more leaves present (9.7), a faster leaf formation rate (one every 26 days) and a shorter life span (8.4 months) than those of Avicennia schaueriana (10, 8.1, 48 days and 13.1 months, respectively) and Laguncularia racemosa, except for the shorter life span (15, 6.6, 31 days and 6.8 months, respectively). The proportion of tips that flowered was higher in L. racemosa (13 %) and in R. mangle (11 %) than in A. schaueriana (2 %). The largest biomass of the average R. mangle leaf (0.75 vs. 0.53 and 0.37 g leaf?1, of L. racemosa and A. schaueriana, respectively) and the highest plant density of this species (2,590 vs. 694 and 202 plant ha?1, respectively) resulted in it having the greatest leaf productivity (10.6 Mg ha?1 year?1 compared to 2.4 Mg ha?1 year?1 for L. racemosa and 0.3 Mg ha?1 year?1 for A. schaueriana). The total leaf production is higher in this mangrove than most of those reported for other mangroves in the world. 相似文献
30.
Paulo Victor Pereira Baio Juliana Nunes Ramos Louisy Sanches dos Santos Morgana Fonseca Soriano Elisa Martins Ladeira M?nica Cristina Souza Thereza Cristina Ferreira Camello Marcio Garcia Ribeiro Raphael Hirata Junior Ver?nica Viana Vieira Ana Luíza Mattos-Guaraldi 《PLoS neglected tropical diseases》2013,7(12)