Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.21.4 (trypsin)
42,187 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

UVB irradiation can cause considerable changes in the composition of cells in the skin and in cutaneous cytokine levels. We found that a single exposure of normal human skin to UVB induced an infiltration of numerous IL-4(+) cells. This recruitment was detectable in the papillary dermis already 5 h after irradiation, reaching a peak at 24 h and declining gradually thereafter. The IL-4(+) cells appeared in the epidermis at 24 h postradiation and reached a plateau at days 2 and 3. The number of IL-4(+) cells was markedly decreased in both dermis and epidermis at day 4, and at later time points, the IL-4 expression was absent. The IL-4(+) cells did not coexpress CD3 (T cells), tryptase (mast cells), CD56 (NK cells), and CD36 (macrophages). They did coexpress CD15 and CD11b, showed a clear association with elastase, and had a multilobed nucleus, indicating that UVB-induced infiltrating IL-4(+) cells are neutrophils. Blister fluid from irradiated skin, but not from control skin, contained IL-4 protein as well as increased levels of IL-6, IL-8, and TNF-alpha. In contrast to control cultures derived from nonirradiated skin, a predominant type 2 T cell response was detected in T cells present in primary dermal cell cultures derived from UVB-exposed skin. This type 2 shift was abolished when CD15(+) cells (i.e., neutrophils) were depleted from the dermal cell suspension before culturing, suggesting that neutrophils favor type 2 T cell responses in UVB-exposed skin.
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PMID:Ultraviolet B radiation induces a transient appearance of IL-4+ neutrophils, which support the development of Th2 responses. 1193 23

The alpha thalassaemias are the commonest known human genetic disorders. Although they have almost certainly risen to their current frequencies through natural selection by malaria, the precise mechanism of malaria protection remains unknown. We have investigated the characteristics of red blood cells (RBCs) from individuals heterozygous for alpha(0)thalassaemia (-/alphaalpha) from a range of perspectives. On the basis of the hypothesis that defects in membrane transport could be relevant to the mechanism of malaria protection, we investigated sodium and potassium transport and the activity of the Plamodium falciparum-induced choline channel but found no significant differences in -/alphaalpha RBCs. Using flow cytometry, we found that thalassaemic P. falciparum-infected RBCs (IRBCs) bound 44% more antibody from immune plasma than control IRBCs. This excess binding was abrogated by predigestion of IRBCs with trypsin but was not directed at the variant surface molecule PfEMP1. Furthermore, we found no evidence for altered cytoadhesion of alpha-thalassaemic IRBCs to the endothelial receptors intercellular adhesion molecule-1 (ICAM-1), CD36 or thrombospondin. We hypothesize that altered red-cell membrane band 3 protein may be a target for enhanced antibody binding to alpha-thalassaemic IRBCs and could be involved in the mechanism of malaria protection.
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PMID:The membrane characteristics of Plasmodium falciparum-infected and -uninfected heterozygous alpha(0)thalassaemic erythrocytes. 1213 62

Gametocytes, the sexual stages of malaria parasites (Plasmodium spp.) that are transmissible to mosquitoes, have been the focus of much recent research as potential targets for novel drug and vaccine therapies. However, little is known about the host clearance of gametocyte-infected erythrocytes (GEs). Using a number of experimental strategies, we found that the scavenger receptor CD36 mediates the uptake of nonopsonized erythrocytes infected with stage I and IIA gametocytes of Plasmodium falciparum by monocytes and culture-derived macrophages (Mphis). Light microscopy and immunofluorescence assays revealed that stage I and IIA gametocytes were readily internalized by monocytes and Mphis. Pretreating monocytes and Mphis with a monoclonal antibody that blocked CD36 resulted in a significant reduction in phagocytosis, as did treating GEs with low concentrations of trypsin to remove P. falciparum erythrocyte membrane protein 1 (PfEMP-1), a parasite ligand for CD36. Pretreating monocytes and Mphis with peroxisome proliferator-activated receptor gamma-retinoid X receptor agonists, which specifically upregulate CD36, resulted in a significant increase in the phagocytosis of GEs. Murine CD36 on mouse Mphis also mediated the phagocytosis of P. falciparum stage I and IIA gametocytes, as determined by receptor blockade with anti-murine CD36 monoclonal antibodies and the lack of uptake by CD36-null Mphis. These results indicate that phagocytosis of stage I and IIA gametocytes by monocytes and Mphis appears to be mediated to a large extent by the interaction of PfEMP-1 and CD36, suggesting that CD36 may play a role in innate clearance of these early sexual stages.
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PMID:CD36-mediated nonopsonic phagocytosis of erythrocytes infected with stage I and IIA gametocytes of Plasmodium falciparum. 1249 89

Transforming growth factor (TGF)-beta1 exerts autocrine and paracrine effects on hematopoiesis. Here, we have attempted to evaluate the effect of endogenous TGF-beta1 on early erythroid development from primitive human hematopoietic stem cells (HSCs) and to assess the effects of TGF-beta1 on different phases of erythropoiesis. Cord blood CD34(+)CD38(-) lineage-marker-negative (Lin(-)) cells were cultured in serum-free conditions using various combinations of stem cell factor (SCF), erythropoietin (Epo), and TGF-beta-neutralizing antibody. Generation of erythroid progenitors was assessed using colony assay and flow cytometry. Terminal erythroid differentiation was examined when SCF/Epo-stimulated cells were recultured in the presence of Epo with and without TGF-beta1. Anti-TGF-beta augmented the proliferation of CD34(+)CD38(-)Lin(-) cells (day 21) in SCF-stimulated (6.4-fold +/- 1.5-fold) and SCF/Epo-stimulated (2.9-fold +/- 1.2-fold) cultures. Cells stimulated by SCF/Epo underwent similar levels of erythroid differentiation with and without anti-TGF-beta. While SCF alone stimulated the production of tryptase-positive mast cells, cells stimulated by SCF/anti-TGF-beta were predominantly erythroid (CD36(+)CD14(-) and glycophorin A positive). A distinct expansion of erythroid progenitors (CD34(+)CD36(+)CD14(-)) with the potential to form erythroid colonies was seen, revealing early Epo-independent erythroid development. In contrast, the kinetics of erythroid progenitor generation from primitive HSCs indicate that TGF-beta1 is not inhibitory in late erythropoiesis, but it accelerated the conversion of large BFU-E into colony-forming units-erythroid. Finally, TGF-beta1 accelerated Epo-induced terminal erythroid differentiation and resulted in a greater level of enucleation (22% +/- 6% versus 7% +/- 3%) in serum-free conditions. Serum addition stimulated enucleation (54% +/- 18%), which was lower (26% +/- 14%) with anti-TGF-beta, suggesting that optimal erythroid enucleation is Epo dependent, requiring serum factors including TGF-beta1.
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PMID:Neutralization of autocrine transforming growth factor-beta in human cord blood CD34(+)CD38(-)Lin(-) cells promotes stem-cell-factor-mediated erythropoietin-independent early erythroid progenitor development and reduces terminal differentiation. 1296 10

Hyaluronic acid (HA) and chondroitin sulfate A (CSA) have been identified as receptors for adhesion of Plasmodium falciparum-infected erythrocytes (IEs) and appear to be involved in mediating parasite accumulation in the placenta. We demonstrate here that some, but not all, parasite populations can adhere to both receptors, and we identify distinguishing features of adhesion. Adhesion to HA and CSA was greatest among pigmented trophozoite-infected erythrocytes and at physiologic pH and was associated with a lack of rosette formation and little adhesion to CD36 and intercellular adhesion molecule-1. Adhesion to HA was sensitive to trypsin cleavage of the IE surface, whereas trypsin-resistant and trypsin-sensitive CSA adhesion were both observed. Soluble HA, but not CSA, could cause aggregation or clumping of IEs. Different HA types varied in adhesion-inhibitory activity, which was altered by physical treatment, suggesting that structural features of HA influence IE interactions. These findings have important implications for understanding the pathogenesis of malaria, particularly in pregnancy.
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PMID:Plasmodium falciparum-infected erythrocytes demonstrate dual specificity for adhesion to hyaluronic acid and chondroitin sulfate A and have distinct adhesive properties. 1472 79

Erythrocytes (RBCs) opsonized by IgG and complement are prevalently recognized and phagocytosed by complement receptor CR1. This mechanism, effective in senescent and damaged RBCs seems to be operative in ring-parasitized RBCs, since infection by Plasmodium falciparum induces stage-dependent binding of auto-antibodies and activated C3 to the RBC membrane. Later, parasite forms are also recognized by non-opsonic receptors, such as scavenger receptor CD36. Malaria parasites induce the oxidative formation of hemichromes which are the trigger for the auto-antigen development. Band 3 protein is the most plausible candidate of the RBC auto-antigen, induced by hemichromes. Auto-antigens isolated from trophozoites were found only in a high-molecular-weight protein aggregates not present in the normal RBC. The immunocomplex was purified by protein-A affinity chromatography, purified proteins digested by trypsin and analyzed by MALDI-TOF. Peptide mapping showed that the main antigen consisted of band 3 protein aggregates that also contained hemichromes, IgGs, complement factor 3 (C3), and traces of spectrin and glycophorin but no parasite proteins. Two cysteines located in the band 3 cytoplasmic domain were found to be particularly reactive to oxidants and mediated band 3 covalent dimerization via disulfide bonds. Thus, parasites promote oxidative alterations in the membrane of the host which lead to exposure of antigenic sites recognized by anti-band 3 auto-antibodies. Formation of band 3 clusters appears to be mediated by cytoplasmic binding of hemichromes and also by direct band 3 oxidation, whereby clustered, oxidized and antigenic band 3 was underglycosylated.
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PMID:Mechanisms of band 3 oxidation and clustering in the phagocytosis of Plasmodium falciparum-infected erythrocytes. 1496 70

Villous trophoblast cells (TC) obtained from first trimester and term human placentae after trypsin/Percoll gradient isolation were immunodepleted of contaminant cells. The level of purity was assessed by the intracellular expression of the pan trophoblast marker cytokeratin-7 (CK7) and comparisons were made with the GB25 trophoblast-specific (cytotrophoblast+syncytiotrophoblast) cell surface marker. The presence of contaminating cells was traced with intracellular vimentin, or cell surface CD2, CD36, and CD163 markers and evaluated by flow cytometric analysis. The pattern of CK7 expression by trophoblast cells was also analyzed by immunofluorescence microscopy. Most batches of TC from first trimester or term placentae (92+/-3% and 96+/-2%, respectively) showed a high percentage of CK7 expressing cells, with less than 2% contaminating vimentin positive cells. In some batches of TC with a lower percentage (65+/-4%) of CK7-expressing cells, no vimentin was found, but a low percentage of CD36-expressing cells was evidenced, with no presence of CD2, and/or CD163-expressing cells. The intracellular CK7 signal correlated significantly with that of GB25 (p<0.05) cell surface expression in TC of term placentae. The choriocarcinoma BeWo and Jar cell lines also showed high levels (>92%) of CK7-expressing cells. Conversely, the control U87 astrocytoma cell line showed a high percentage (>90%) of vimentin but no CK7-expressing cells. These results provide evidence that the mutually exclusive pattern of intracellular CK7/vimentin expression of human TC can be used for evaluation by flow cytometry of the purity of primary human trophoblast cells.
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PMID:Evaluation of Cytokeratin 7 as an accurate intracellular marker with which to assess the purity of human placental villous trophoblast cells by flow cytometry. 1508 19

Caveolae, the specialized invaginations of plasma membranes, formed sealed vesicles with outwards-orientated cytosolic surface after isolation from primary human adipocytes. This morphology allowed differential, vectorial identification of proteins at the opposite membrane surfaces by proteolysis and MS. Extracellular-exposed caveolae-specific proteins CD36 and copper-containing amine oxidase were concealed inside the vesicles and resisted trypsin treatment. The cytosol-orientated caveolins were efficiently digested by trypsin, producing peptides amenable to direct MS sequencing. Isolation of peripheral proteins associated with the cytosolic surface of caveolae revealed a set of proteins that contained nuclear localization signals, leucine-zipper domains and PEST (amino acid sequence enriched in proline, glutamic acid, serine and threonine) domains implicated in regulation by proteolysis. In particular, PTRF (polymerase I and transcript release factor) was found as a major caveolae-associated protein and its co-localization with caveolin was confirmed by immunofluorescence confocal microscopy. PTRF was present at the surface of caveolae in the intact form and in five different truncated forms. Peptides (44 and 45 amino acids long) comprising both the PEST domains were sequenced by nanospray-quadrupole-time-of-flight MS from the full-length PTRF, but were not found in the truncated forms of the protein. Two endogenous cleavage sites corresponding to calpain specificity were identified in PTRF; one of them was in a PEST domain. Both cleavage sites were flanked by mono- or diphosphorylated sequences. The phosphorylation sites were localized to Ser-36, Ser-40, Ser-365 and Ser-366 in PTRF. Caveolae of human adipocytes are proposed to function in targeting, relocation and proteolytic control of PTRF and other PEST-domain-containing signalling proteins.
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PMID:Vectorial proteomics reveal targeting, phosphorylation and specific fragmentation of polymerase I and transcript release factor (PTRF) at the surface of caveolae in human adipocytes. 1524 32

Introduction to bioorganic chemistry by Prof. Kanaoka at the entrance of my research works affects greatly throughout the life afterward. Chemical modification studies of enzyme proteins taught me quality of chemical reactions. For example, triethyloxonium fluoroborate (Et3O+ BF4(-)), a Meerwein reagent, selectively reacted with a particular carboxyl group (Asp-177) in the substrate binding site of trypsin, even though the reaction was performed in aqueous solution. A series of ion channel studies intoxicate me how exciting the science works are. Purification of sodium channel protein from electric eels initiated the collaboration work to reveal total primary structure of the molecule, as an inaugurating work of ion channel molecules. Photoaffinity labeling proved to be an efficient method to elucidate ligand binding sites, such as TTX binding site within the sodium channel and the sites for calcium anatagonists in L-type calcium channels. Encounter with CD36 molecule expands our works to more pathobiochemical field. We revealed CD36, a class B scavenger receptor, is related to development of atherosclerosis by phagocytosis of ox-LDL in macrophages and even matured adipocytes. In microglia, however, CD36 plays clearance role of oligomeric beta-amyloid peptides in IL-4 activated type-2 microglia, suggesting the activation of type-2 microglia may be useful for developing a new method to treat or prevent from Alzheimer's disease.
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PMID:[Seeking "Etwas Neues"--from bioorganic chemistry to Alzheimer's disease]. 1898 99

Cerebral malaria (CM) is characterized by accumulation of circulating cells within brain microvessels, among which platelets play an important role. In vitro, platelets modulate the cytoadherence of Plasmodium falciparum-parasitized red blood cells (PRBCs) to brain endothelial cells. Here we show for the first time that platelet microparticles (PMPs) are able to bind to PRBCs, thereby transferring platelet antigens to the PRBC surface. This binding is largely specific to PRBCs, because PMPs show little adherence to normal red blood cells. PMP adherence is also dependent on the P. falciparum erythrocyte membrane protein 1 variant expressed by PRBCs. PMP binding to PRBCs decreases after neutralization of PRBC surface proteins by trypsin or after treatment of PMPs with a mAb to platelet-endothelial cell adhesion molecule-1 (CD31) and glycoprotein IV (CD36). Furthermore, PMP uptake is a dynamic process that can be achieved by human brain endothelial cells (HBECs), inducing changes in the endothelial phenotype. Lastly, PMPs dramatically increase PRBC cytoadherence to HBECs. In conclusion, our study identifies several mechanisms by which PMPs may participate in CM pathogenesis while interacting with both PRBCs and HBECs. PMPs thereby provide a novel target for antagonizing interactions between vascular cells that promote microvascular sludging and blood brain barrier alteration during CM.
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PMID:Platelet microparticles: a new player in malaria parasite cytoadherence to human brain endothelium. 1953 85


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