altered expression of galectin- induces cortical thymocyte depletion and premature exit of immature thymocytes during trypanosoma cruzi infection　
antibodies
rabbit anti-galectin-1 immune serum was obtained as described.11 goat anti-human galectin-1 antibody was purchased from r&d systems. anti-cd8/cy-chrome, anti-cd4/phycoerithrin (pe), and mouse anti-human galectin-3 (clone b2c10) monoclonal antibodies (mab) were purchased from pharmingen/becton dickinson (san diego, ca). biotinylated rat anti-mouse galectin-3 mab (clone m3/38) was purchased from cedarlane laboratories (hornby, on, canada). fluorescein isothiocyanate-conjugated goat anti-rabbit igg was from biosys (compigne, france). anti-mouse laminin and fibronectin sera were obtained from novotec (st. martin-la-garenne, france). rabbit pan anti-cytokeratin was obtained from sigma. alexa 543-conjugated goat anti-mouse igg, alexa 488-conjugated goat anti-rabbit igg, and alexa 633-conjugated streptavidin were purchased from molecular probes (eugene, or). rhodamine-conjugated anti-goat igg was from santa cruz biotechnology (santa cruz, ca).
tissues and cell cultures
thymuses were obtained from either control or infected balb/c mice at the peak of parasitemia and immediately frozen in liquid nitrogen. the thymic epithelial cell line it-76m1 was provided by dr. t. itoh (tohoku university, sendai, japan) and cultured in rpmi 1640 medium supplemented with 10% heat-inactivated fetal bovine serum, 2 mmol/l l-glutamine, 50 &mol/l 2-mercaptoethanol, 100 u/ml penicillin, and 100 &g/ml streptomycin at 37??c in a humidified atmosphere containing 5% co2. for in vitro infections, it-76m1 cells were plated onto sterile glass slides (105 cells/slide) and cultured for 24 hours. parasites obtained from infected vero cultures were added to these cells, in a ratio of 50 parasites/cell, for an additional period of 48 hours. cultures were then fixed in 4% buffered paraformaldehyde for 5 minutes and processed for immunocytochemistry. tncs, isolated from thymuses of control or infected mice as previously described,25,26 were plated onto sterile glass slides and cultured for 35 to 60 hours. adhered tncs were then washed with phosphate-buffered saline (pbs), fixed as mentioned above and processed for immunocytochemistry.
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全刊杂志赏析网 2015altered expression of galectin- induces cortical thymocyte depletion and premature exit of immature thymocytes during trypanosoma cruzi infection　
the cortical population of thymic nurse cells (tncs), lymphoepithelial complexes in which immature thymocytes undergo differentiation and occasionally die by apoptosis, is altered after infection. specifically, in vitro analysis of tncs recovered from the thymus of infected mice showed that they are smaller in both size and number, have altered expression of ecm molecules,and release thymocytes faster than those from control animals.8 in this regard, alterations in cell-cell and cell-ecm interactions in the thymic microenvironment after t. cruzi infection are accompanied by high numbers of immature cd4+cd8+ t cells found in lymph nodes of infected mice. interestingly, some of these lymphocytes bear potentially autoreactive t-cell receptors (tcrs), probably representing cells that escaped from the normal processes of thymic selection.9 the presence of these immature t cells in the periphery might contribute to the induction of chagasic cardiopathy through an autoimmune response.
galectins are a family of &-galactoside-binding proteins highly conserved throughout animal evolution, which are present at different subcellular compartments.10 these proteins modulate several biological processes, such as cell adhesion, migration, proliferation, and apoptosis.11 galectin-1, -3, and -9 are expressed in thymus, among which galectin-1 and -3 are present throughout all of the thymic parenchyma, being mainly produced by thymic epithelial cells (tecs).12-15 interestingly, recent evidence indicates that galectins can interact with ecm glycoproteins and modulate cell-cell interactions within the thymic microenvironment.12,15 we have demonstrated that galectin-3 produced by thymic stromal cells can interfere with tec/thymocyte adhesion, by acting as an anti-adhesive molecule and modulating protein-carbohydrate interactions.15
it has become increasingly apparent that galectin-1 and -3 function as regulatory proteins, sometimes with opposite effects. galectin-1 can induce apoptosis in immature thymocytes displaying the cd3lowcd4ccd8c and cd3lowcd4lowcd8low phenotypes, an effect that parallels the processes of selection and thymocyte differentiation.12,16 on the other hand, in vitro studies revealed that intracellular galectin-3 inhibits apoptosis induced by a wide variety of stimuli in activated t lymphocytes and tumor cells,17,18 whereas extracellular galectin-3 promotes apoptosis when added exogenously to t cells.19,20 interestingly, recent data suggest that galectin-1 and -3 kill t cells by binding to them and engaging different cell surface glycoproteins.20 in addition, these carbohydrate-binding proteins can trigger different cell death pathways, with or without caspase activation, in different cell types.21-23
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全刊杂志赏析网 2015Structure and physicochemical properties of starches from kidney bean seeds at immature, premature and mature stages of development.
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2003 Feb 14;338(5):463-9.Structure and physicochemical properties of starches from kidney bean seeds at immature, premature and mature stages of development.1, , , , , , , .1Department of Biochemical Science and Technology, Faculty of Agriculture, Kagoshima University, 890-0065, Kagoshima, Japan.AbstractStarches from kidney bean (Phaseolus vulgaris L. cv. Toramame) seeds at the immature, premature, mature stages of development were examined. The starch content increased from 94, 219 to 265 mg per seed. Starches showed the C(a)-crystalline type composed of small (&5 micrometer) and large (10-35 micrometer) granules, with the large granules largely increasing with maturity. The amylose content increased from 21, 26 to 27%, and rapid viscograms and DSC thermograms suggested that the mature-stage starch was gelatinized with ease. The amylose increased in size from DPn 820, 1000 to 1080 and a number of chains per molecule (NC) from 3.3, 4.2 to 4.5. The branched amylose was a minor component (11-18% by mole) with NC 20-22. The amylopectin was similar in CL (23), beta-amylolysis limit (59%), and chain-length distribution, but reduced in size (DPn 17,100-5270) and increased in content of phosphorus (114-174 ppm) with an increase in the amount of phosphorus linked to C-6 of the glucose residue (8-66%).PMID:
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