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31.
We previously demonstrated that laminar shear stress enhances human coronary artery endothelial cell (HCAEC) wound closure via the mechanisms of cell spreading and migration. Because cell-cell junctional proteins such as vascular endothelial cell cadherin (VE-cadherin) are critical to cell-cell adhesion and motility, we tested the hypothesis that modulation of VE-cadherin expression under shear stress may be linked to this enhancement in wound closure. HCAEC monolayers were preconditioned to attain cellular alignment by shearing at 12 dynes/cm(2) for 18 hr in a parallel-plate flow chamber. Subsequently, they were divided into the following three groups: (i) control; (ii) treated with anti-cadherin-5 antibody; or (iii) treated with the calcium chelating agent EGTA. Next, the monolayers were wounded with a metal spatula and resheared at 20 dynes/cm(2) or left static. Time-lapse imaging was performed during the first 3 hr after imposition of these conditions. Immunocytochemistry or Western blot analyses for VE-cadherin expression were performed on all wounded monolayers. Deconvolution microscopy, three-dimensional cell-cell junctional reconstruction images, and histogram analyses of interendothelial junction signal intensities were performed on cells at the wound edge of a monolayer. Under shear, HCAEC demonstrated increased VE-cadherin immunofluorescence and protein expression despite an enhancement in wound closure compared with static conditions. In separate experiments, application with anti-cadherin-5 antibody or treatment with EGTA attenuated VE-cadherin expression and further enhanced wound closure compared with control shear and all static conditions. In addition, the pattern of VE-cadherin localization with these treatments became more intracellular and nuclear in appearance. These findings of changes in this junctional adhesion protein expression and localization may further our understanding of laminar shear stress-induced endothelial repair in the coronary circulation. 相似文献
32.
Empadinhas N Albuquerque L Henne A Santos H da Costa MS 《Applied and environmental microbiology》2003,69(6):3272-3279
The biosynthetic pathway for the synthesis of the compatible solute alpha-mannosylglycerate (MG) in the thermophilic bacterium Thermus thermophilus HB27 was identified based on the activities of recombinant mannosyl-3-phosphoglycerate synthase (MPGS) (EC 2.4.1.217) and mannosyl-3-phosphoglycerate phosphatase (MPGP) (EC 3.1.3.70). The sequences of homologous genes from the archaeon Pyrococcus horikoshii were used to identify MPGS and MPGP genes in T. thermophilus HB27 genome. Both genes were separately cloned and overexpressed in Escherichia coli, yielding 3 to 4 mg of pure recombinant protein per liter of culture. The molecular masses were 43.6 and 28.1 kDa for MPGS and MPGP, respectively. The recombinant MPGS catalyzed the synthesis of alpha-mannosyl-3-phosphoglycerate (MPG) from GDP-mannose and D-3-phosphoglycerate, while the recombinant MPGP catalyzed the dephosphorylation of MPG to MG. The recombinant MPGS had optimal activity at 80 to 90 degrees C and a pH optimum near 7.0; MPGP had maximal activity between 90 and 95 degrees C and at pH 6.0. The activities of both enzymes were strictly dependent on divalent cations; Mn(2+) was most effective for MPGS, while Mn(2+), Co(2+), Mg(2+), and to a lesser extent Ni(2+) activated MPGP. The organization of MG biosynthetic genes in T. thermophilus HB27 is different from the P. horikoshii operon-like structure, since the genes involved in the conversion of fructose-6-phosphate to GDP-mannose are not found immediately downstream of the contiguous MPGS and MPGP genes. The biosynthesis of MG in the thermophilic bacterium T. thermophilus HB27, proceeding through a phosphorylated intermediate, is similar to the system found in hyperthermophilic archaea. 相似文献
33.
Da Silva JB De Albuquerque CM De Araújo EC Peixoto CA Hurd H 《Journal of invertebrate pathology》2000,76(4):257-262
Mosquitoes have an efficient defense system against infection. The cellular immune defense mechanism initiated by the mosquito Culex quinquefasciatus infected with the fungus Candida albicans was investigated in this study. Differences in the hemocyte counts in hemolymph perfused from uninoculated, saline-inoculated, and C. albicans-infected mosquitoes were compared using a light microscope. Phagocytosis was also investigated using electron microscopy. Four types of hemocytes were identified in control mosquitoes: prohemocytes (9.8%), plasmatocytes (38.8%), granular cells (44.2%), and oenocytoids (7.3%). Between 3 and 18 h postinoculation the total hemocyte count was significantly higher in infected, compared to uninfected, mosquitoes. Differential hemocyte counts from infected mosquitoes at 3, 6, and 18 h after inoculation showed that the relative proportion of plasmatocytes (48.6, 50.7, 45%) was higher and, concomitantly, the proportion of granular cells was lower (38, 36.8, 35%, respectively). Yeast cells were phagocytosed and limited growth was observed within the plasmatocytes. Melanized nodules were found attached to different insect tissues at 24 to 72 h following infection. These results suggest that phagocytosis, followed by nodule formation, was capable of clearing the hemolymph of yeast cells. 相似文献
34.
Characterization of terminal NeuNAcalpha2-3Galbeta1-4GlcNAc sequence in lipooligosaccharides of Neisseria meningitidis 总被引:1,自引:0,他引:1
Group B and C Neisseria meningitidis are the major cause of meningococcal
disease in the United States and in Europe. N . meningitidis
lipooligosaccharide (LOS), a major surface antigen, can be divided into 12
immunotypes of which L1 through L8 were found among Group B and C
organisms. Groups B and C but not Group A may sialylate their LOSs with
N-acetylneuraminic acid (NeuNAc) at the nonreducing end because they
synthesize CMP-NeuNAc. Using sialic acid-galactose binding lectins as
probes in an ELISA format, six of the eight LOS immunotypes (L2, L3, L4,
L5, L7, and L8) in Groups B and C bound specifically to Maackia amurensis
leukoagglutinin (MAL), which recognizes NeuNAcalpha2- 3Galbeta1-4GlcNAc/Glc
sequence, but not to Sambucus nigra agglutinin, which binds
NeuNAcalpha2-6Gal sequence. The combination of SDS-PAGE and MAL-blot
analyses revealed that these six LOSs contained only the
NeuNAcalpha2-3Galbeta1-4GlcNAc trisaccharide sequence in their 4.1 kDa LOS
components, which have a common terminal lacto-N-neotetraose (LNnT,
Galbeta1-4GlcNAcbeta1-3Galbeta1-4Glc) structure when nonsialylated as shown
by previous studies. The LOS-lectin binding was abolished when the LOSs
were treated with Newcastle disease viral neuraminidase which cleaves
alpha2-->3 linked sialic acid. Methylation analysis of a representative
LOS (L2) confirmed that NeuNAc is 2-->3 linked to Gal. Thus, these LOSs
structurally mimic certain glycolipids, i.e., paragloboside (LNnT-ceramide)
and sialylparagloboside and some glycoproteins in having LNnT and
N-acetyllactosamine sequences, respectively, with or without alpha2-->3
linked NeuNAc. The molecular mimicry of the LOSs may play a role in the
pathogenesis of N.meningitidis by assisting the organism to evade host
immune defenses in man.
相似文献
35.
Pasquier CM; Promponas VI; Varvayannis NJ; Hamodrakas SJ 《Bioinformatics (Oxford, England)》1998,14(8):749-750
Summary : FT is a tool written in C++, which implements the Fourier
analysis method to locate periodicities in aminoacid or DNA sequences. It
is provided for free public use on a WWW server with a Java interface.
Availability : The server address is http://o2.db. uoa.gr/FT Contact :
shamodr@atlas.uoa.gr
相似文献
36.
Karen CM Moraes Lívia F Diniz Maria Terezinha Bahia 《Memórias do Instituto Oswaldo Cruz》2015,110(2):181-191
Chagas disease, caused by the intracellular protozoan Trypanosoma
cruzi, is a serious health problem in Latin America. During this
parasitic infection, the heart is one of the major organs affected. The pathogenesis
of tissue remodelling, particularly regarding cardiomyocyte behaviour after parasite
infection and the molecular mechanisms that occur immediately following parasite
entry into host cells are not yet completely understood. When cells are infected with
T. cruzi, they develop an inflammatory response, in which
cyclooxygenase-2 (COX-2) catalyses rate-limiting steps in the arachidonic acid
pathway. However, how the parasite interaction modulates COX-2 activity is poorly
understood. In this study, the H9c2 cell line was used as our model and we
investigated cellular and biochemical aspects during the initial 48 h of parasitic
infection. Oscillatory activity of COX-2 was observed, which correlated with the
control of the pro-inflammatory environment in infected cells. Interestingly,
subcellular trafficking was also verified, correlated with the control of Cox-2 mRNA
or the activated COX-2 protein in cells, which is directly connected with the
assemble of stress granules structures. Our collective findings suggest that in the
very early stage of the T. cruzi-host cell interaction, the parasite
is able to modulate the cellular metabolism in order to survives. 相似文献
37.
CM van der Loos 《Biotechnic & histochemistry》2013,88(3):141-154
Three fixation issues related to immunostaining are discussed here: 1) Generally, a tissue block is fixed, then embedded and sectioned (pre-fixation). The type of fixative applied, crosslinking or coagulating, has an impact on selecting an epitope retrieval method. Individual antigens have a fixation–retrieval characteristic. 2) A long fixation time, especially with crosslinking fixatives, may compromise the result of immunostaining. This negative effect varies among different antigens and can be partially restored by applying a more sensitive/efficient detection system such as tyramide amplification. 3) Sections cut from a fresh frozen tissue block usually are acetone fixed (post-fixation). This was accepted as the “gold standard” for a long time. Post-fixation, however, may have serious consequences for preservation of small peptides leaking from the cut open cells, whereas this is not the case with pre-fixed intact cells. Consequently, the concept of an acetone post-fixed cryostat tissue section as “gold standard” no longer exists and a more appropriate use of the terms immunohistochemistry and immunocytochemistry therefore seems justified. For many antibodies, it is not known whether a formalin fixed, paraffin embedded tissue specimen is appropriate. Suggestions are made for creating a positive control cell block for testing such antibodies. 相似文献
38.
Background
Designing maximally selective ligands that act on individual targets is the dominant paradigm in drug discovery. Poor selectivity can underlie toxicity and side effects in the clinic, and for this reason compound selectivity is increasingly monitored from very early on in the drug discovery process. To make sense of large amounts of profiling data, and to determine when a compound is sufficiently selective, there is a need for a proper quantitative measure of selectivity. 相似文献39.
Background
Paulinella chromatophora is a freshwater filose amoeba with photosynthetic endosymbionts (chromatophores) of cyanobacterial origin that are closely related to free-living Prochlorococcus and Synechococcus species (PS-clade). Members of the PS-clade of cyanobacteria contain a proteobacterial form 1A RubisCO (ribulose-1,5-bisphosphate carboxylase/oxygenase) that was acquired by horizontal gene transfer (HGT) of a carboxysomal operon. In rDNA-phylogenies, the Paulinella chromatophore diverged basal to the PS-clade, raising the question whether the HGT occurred before or after the split of the chromatophore ancestor. 相似文献40.
Antonio Paulo Nassar Junior Alberto Queiroz Farias Luiz Augusto Carneiro d’ Albuquerque Flair José Carrilho Luiz Marcelo Sá Malbouisson 《PloS one》2014,9(9)