Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.21.4 (trypsin)
 42,187 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A granulomonopoietic enhancing factor (GM-EF) capable of promoting the effect of colony-stimulating factors (CSFs) on myeloid progenitor cells has been purified to homogeneity from serum-free medium conditioned by fully mature human macrophages. GM-EF was a glycoprotein with an apparent molecular weight of 74 kd and an isoelectric point of 5.2-5.3. The purified protein was heat stable (75 degrees C for 30 min) and was sensitive to treatment with trypsin, papain, and bacterial protease but not to neuraminidase. The activity of GM-EF could be effectively neutralized by GM-EF-specific antiserum, and no antigenic cross-reactivity was observed using antisera against interleukin (IL)-1, IL-4, and IL-6. These results suggest that GM-EF is a unique cytokine that is different biochemically and antigenically from other hematopoietic enhancing factors such as IL-1, IL-4, and IL-6.
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PMID:Purification and characterization of human macrophage-derived granulomonopoietic enhancing factor (GM-EF). 137 59

In the murine system, a number of cytokines (including interleukin-3 [IL-3], IL-4, and stem cell factor [SCF]) promote the growth of mast cells (MCs). However, so far little is known about factors controlling differentiation of human MCs. Recent data suggest that human MCs express receptors (R) for SCF. The aim of the present study was to investigate whether recombinant human (rh) SCF induces differentiation of human MCs from their precursor cells. For this purpose, bone marrow (BM; normal donors, n = 6) and peripheral blood (PB; normal donors, n = 11) mononuclear cells (MNC) were cultured in the presence of rhSCF, rhIL-3, rhIL-4, rhIL-9, recombinant human macrophage colony-stimulating factor (rhM-CSF), or control medium in long-term (8 weeks) suspension cultures. After 4 weeks, up to 5% of the MNC (BM and PB) cultured in the presence of rhSCF, but not in the presence of other cytokines, were found to exhibit the characteristics of MCs. These MCs expressed the YB5.B8-reactive domain of the SCF R as well as IgE R, as determined by combined toluidine blue/immunofluorescence staining. Myeloid antigens, likewise expressed on human basophils (ie, CD11b, CDw65, and Bsp-1), could not be detected on these cells. Furthermore, rhSCF, but not rhIL-3, rhIL-4, rhIL-9, or rhM-CSF, induced dose- and time-dependent increases in the formation of cellular tryptase (an MC-specific enzyme) (rhSCF [100 ng/mL], 1,308 +/- 679 ng/mL v control medium, 18 +/- 6 ng/mL tryptase on day 35 of PB cell cultures), as well as an increase in cellular histamine. After 6 to 8 weeks, when other mature hematopoietic cells decreased, MCs still could be detected in culture, with up to 40% of all cells being MCs. To test whether rhSCF also activates tissue MCs, we performed histamine release experiments (dispersed tissue; lung, n = 3; uterus, n = 3). SCF was found to enhance (by up to 3.4-fold) the capacity of the MCs to release histamine upon cross-linkage of IgE R with anti-IgE. Together, these observations suggest that rhSCF induces in vitro differentiation of human MCs from their BM and PB precursor cells in long-term culture and upregulates MC releasability.
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PMID:Induction of differentiation of human mast cells from bone marrow and peripheral blood mononuclear cells by recombinant human stem cell factor/kit-ligand in long-term culture. 138 99

Immunocompetent cells of the epidermis can interact by the elaboration and recognition of cytokines. Although much new information has been reported concerning the cytokines secreted by keratinocytes, little is known about cytokines derived from Langerhans cells (LC). To address this deficiency, we examined cytokine mRNA profiles in different epidermal preparations from BALB/c mice, taking advantage of the sensitive technique of polymerase chain reaction (PCR), after reverse transcription of mRNA. In assays of epidermal sheets separated from dermis by ammonium thiocyanate, mRNA for IL-1 alpha, IL-1 beta, IL-6, IL-7, tumor necrosis factor alpha (TNF alpha), TNF beta, granulocyte macrophage/colony-stimulating factor (GM-CSF), and macrophage inflammatory protein-1 alpha (MIP-1 alpha) were unequivocally present. By contrast, faint bands were detected for IL-4, IL-5, and interferon gamma (IFN gamma), and no PCR signal was detected for IL-2. Importantly, assays of epidermal cells (EC) dissociated with trypsin revealed similar mRNA profiles. To determine the effects of cell isolation, fluorescence-activated cell sorter (FACS)-purified Ia- EC were first analyzed; all of the previously cited cytokine mRNA were present except for IL-1 beta and MIP-1 alpha. EC depleted of LC by a second technique, lysis using anti-Ia monoclonal antibody and complement, revealed similar profiles, with substantially reduced PCR signals for IL-1 beta and MIP-1 alpha. Finally, FACS-purified LC (Ia+ EC) clearly expressed IL-1 beta and MIP-1 alpha mRNA, a finding that was verified by Southern blotting using internal oligo probes. We conclude that these cell-isolation procedures did not produce substantial alterations in basal mRNA profiles and that LC are the principal source of mRNA for IL-1 beta and MIP-1 alpha among unstimulated EC in mice.
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PMID:Langerhans cells are the major source of mRNA for IL-1 beta and MIP-1 alpha among unstimulated mouse epidermal cells. 138 44

A GALT-derived B lymphoma, T560, that bears IgAR is described. T560 is IgG2a kappa +, Ia+, B220+, J11d+, Thy-1-, CD3-, CD4-, CD5-, Mac 1-, Mac 2-, nonspecific esterase negative and binds bromelain-treated mouse RBC but not SRBC or ORBC. It presents antigen, secretes IL-1, IL-4 and IL-6 but not IL-2, IL-5 or TGF beta and appears to be related to the Lyt 1+(CD5) lineage of B cells though it lacks Lyt 1. T560 bears IgAR that, on the cell surface, are completely cross-inhibited by low concentrations of IgM and by high concentrations of IgG2a and IgG2b. They do not appear to represent a cell-surface form of galactosyl transferase. They are inducible by high concentrations of IgA, sensitive to trypsin and insensitive to neuraminidase. They are down-regulated by activation of PKC with PMA, but their recovery is not inhibited by cycloheximide, indicating that they are not degraded or shed. They may either lose their affinity for IgA or be internalized without degradation. Seventy percent of IgA receptor activity is lost when T560 is treated with PI-PLC; part of this loss of activity is due to activation of PKC and is inhibited by staurosporine, but approximately 30% of it is not protected by staurosporine indicating that some, or all, of the IgA receptor of T560 is connected to the cell membrane via a GPI linker. The T560 IgA receptor could be related to the poly-Ig or M cell receptor.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Sensitivity of receptors for IgA on T560, a murine B lymphoma, to phorbol myristate acetate and to phosphatidylinositol-specific phospholipase C. 165 5

Trypanosoma cruzi, which causes Chagas' disease, has been shown to cause polyclonal proliferation of lymphocytes after infection in vivo. This paper demonstrates that coculture of human PBMC with T. cruzi CL strain leads to proliferation of lymphocytes, which peaks on days 5 to 7 after infection. Approximately 15% of lymphocytes in culture undergo blast transformation. The proliferation of lymphoblasts can be measured by [3H]TdR incorporation, because the parasites incorporate little TdR. Parasites derived from autologous PBMC cultures or xenogeneic rat fibroblasts stimulate lymphocyte transformation similarly. By immunofluorescent cytometry, lymphoblasts from these cultures are 23 to 46% B cells (CD19+) and 39 to 64% T cells (CD3+), and approximately half of the T cells are CD4+ and half CD8+. A high percentage of lymphoblasts express MHC class II and IL-2R p55, suggesting both B and T lymphoblasts express these molecules. Anti-MHC class II and anti-IL-2R p55 mAb significantly inhibit the proliferative response of PBMC to T. cruzi. The mRNA for cytokines IL-1 beta, IL-2, IL-5, IL-6, IFN-gamma, and TNF-alpha are detected after T. cruzi coculture with PBMC, peaking on day 3. No IL-4 or IL-10 mRNA are detected. Large quantities of bioactive IL-1 and IL-6 are found in the supernatants of these PBMC. Monocytes, infected in the apparent absence of lymphocytes, assume activated morphology and accumulate mRNA for IL-1 beta, TNF-alpha, and IL-6. T cells require accessory cells to proliferate and produce cytokine mRNA. A trypsin-sensitive activity in lysates of T. cruzi stimulates lymphocyte proliferation. The data presented demonstrate that T. cruzi coculture with PBMC leads to lymphocyte proliferation, monocyte activation, and cytokine production.
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PMID:Coculture of human peripheral blood mononuclear cells with Trypanosoma cruzi leads to proliferation of lymphocytes and cytokine production. 172 69

The present studies were undertaken to determine whether the interleukin 4 binding proteins (IL-4BPs) previously identified in the biological fluids of mice are soluble forms of IL-4Rs. We also studied the binding properties of IL-4BPs in order to gain insight into their physiological role in vivo. Affinity-purified IL-4BPs and recombinant soluble IL-4Rs generated similar one-dimensional (Cleveland) peptide maps after digestion with either Staphylococcus aureus V8 protease or trypsin, indicating structural similarities. Furthermore, a rat mAb directed against the murine IL-4Rs immunoprecipitated the IL-4BPs and completely inhibited binding of 125I-IL-4 to a purified preparation of IL-4BPs. Taken together these data indicate that the IL-4BPs are soluble IL-4Rs. At 4 degrees C the IL-4BPs competitively inhibited the binding of IL-4 to membrane IL-4Rs but their ability to prevent binding of IL-4 to cells at 37 degrees C, at the same concentrations, was significantly reduced. Kinetic binding studies of soluble IL-4BPs vs. membrane IL-4Rs disclosed important differences in their rates of dissociation from IL-4. Whereas dissociation at 4 degrees C was slow for both, dissociation of IL-4 from IL-BPs at 37 degrees C was considerably faster (t 1/2 of 2 min) than dissociation of IL-4 from membrane IL-4Rs (t 1/2 of approximately 69 min). Temperature-dependent changes in dissociation kinetics were reversible, and could not be accounted for by either inactivation of the IL-4BPs at 37 degrees C or receptor internalization. Additional experiments also demonstrated that when IL-4BPs bind to IL-4 at 37 degrees C, the IL-4/IL-4BPs complex can rapidly dissociate, allowing IL-4 to bind to membrane IL-4Rs. In addition, binding of IL-4 by the IL-4BPs protects IL-4 from proteolytic degradation. Taken together, these results suggest that the IL-4BPs are naturally occurring forms of soluble IL-4Rs and that some of their properties (fast dissociation kinetics and protection of IL-4 from proteolysis) are consistent with a potential role as carrier proteins for IL-4 in the circulation.
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PMID:Evidence that natural murine soluble interleukin 4 receptors may act as transport proteins. 187 67

The proliferation of mucosal mast cells (MMC) depends on the presence of interleukin 3 (IL 3) and can be further enhanced by interleukin 4 (IL 4). The supernatant of a TH2 cell clone (ST2/K.9) stimulated by concanavalin A was found to contain a factor, provisionally termed mast cell costimulatory activity (MCA), that substantially enhances the proliferation of MMC promoted by a combination of IL 3 and IL 4. In comparison to other lymphokines MCA is rather resistant to tryptic digestion but is very sensitive to pH values lower than 6.0 and to organic solvents. Chromatographic fractionation of MCA revealed that activity is associated with protein(s) or glycoprotein(s) of 35 to 40 kDa. Partially purified MCA that was functionally free of other T-cell-derived lymphokines did not stimulate mast cell proliferation in the absence of a combination of IL 3 and IL 4. In addition, MCA did not affect the proliferation of mast cells when employed together with either IL 3 or IL 4 alone. Control experiments demonstrated that MCA is identical to neither the T-cell-derived lymphokines IL 2 to IL 6, IL 9, interferon gamma, tumor necrosis factor alpha or beta, or granulocyte-macrophage colony-stimulating factor (CSF), nor to IL 7, granulocyte CSF, macrophage CSF, erythropoietin, leukemia inhibitory factor, or epidermal growth factor (EGF). Finally, experiments using a panel of PPD-reactive TH1- and TH2-like cell lines revealed that MCA is preferentially produced by TH2 cells. These data, especially the relative resistance of MCA to trypsin and the high sensitivity to low pH values and organic solvents, indicate that MCA is distinct from known T-cell-derived lymphokines.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Characterization of a T-cell-derived mast cell costimulatory activity (MCA) that acts synergistically with interleukin 3 and interleukin 4 on the growth of murine mast cells. 210 34

A novel growth factor for bone marrow derived murine mucosal type mast cells has been isolated from the conditioned medium of the Mlsa-reactive mouse Th cell line MLS-4.2. In proliferation assays this growth factor synergizes, like IL-4, with IL-3 on established mast cell lines and was therefore termed MEA: mast cell growth enhancing activity. MEA was characterized as a glycoprotein with a Mr range between 37,000 and 43,000. Apparent homogeneity was obtained by using a four-step purification scheme including cation exchange chromatography, Procion red affinity chromatography, IEF, and gel filtration. Inasmuch as MEA was N-terminally blocked during automated Edman-degradation, peptide fragments after digestion with trypsin were used for partial amino acid sequence determination. All evaluable MEA peptide fragments showed complete sequence homology to a recently purified and cloned novel T cell growth factor (P40/TCGF III), the mouse homologue of human IL-9.
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PMID:Purification of MEA, a mast cell growth-enhancing activity, to apparent homogeneity and its partial amino acid sequencing. 214 Mar 90

We have identified a late, committed stage in the differentiation of the mast cell progenitor just before granulation. Mast cell committed progenitors (MCCP) are nongranulated cells with a density of 1.060 to 1.070 g/ml which can be harvested from the mesenteric lymph node of mice infected with Nippostrongylus brasiliensis. Mast cell-committed progenitors are able to proliferate and differentiate in the absence of IL-3 or IL-4 when cultured on a monolayer of embryonic skin or 3T3 fibroblasts and can form colonies in methylcellulose supplemented with fibroblast conditioned medium. Fibroblast conditioned medium appears to contain a soluble MCCP proliferation factor that maintains biologic activity when heated to 56 degrees C for 45 min but is destroyed by incubation with either trypsin or chymotrypsin. It can be selectively precipitated with 60 to 70% saturated ammonium sulfate. The factor is not absorbed by immobilized antibodies to nerve growth factor. The MCCP proliferation activity of the factor could not be mimicked by IL-1, IL-2, IL-4, granulocyte-macrophage-CSF, granulocyte-CSF, macrophage-CSF, IFN-alpha/beta, IFN-gamma, nerve growth factor, epidermal growth factor, serum fibronectin, heparin, or a number of glycosaminoglycans. At high salt concentrations, the factor passes through a 50-kDa membrane and can be concentrated above a 5-kDa membrane. MCCP acquire a connective tissue phenotype when cultured on a fibroblast monolayer and a mucosal phenotype when cloned in the presence of conditioned medium from PWM-stimulated spleen cells. When cultured in the absence of IL-3 on a monolayer of embryonic skin or 3T3 fibroblasts, mast cell-committed progenitors produce mast cells which stain with berberine sulfate suggesting a connective tissue phenotype; however, the mast cells that develop when mast cell-committed progenitors are cultured in the presence of IL-3 or conditioned media from PWM-stimulated spleen cells do not stain with berberine sulfate. MCCP intercalate into monolayers of embryonic skin or 3T3 fibroblasts, but T cells are not able to associate with the monolayer and can be completely washed away. Attempts to enrich mast cell-committed progenitors by intercalation and elution from embryonic skin monolayers proved unsuccessful, but some enrichment of mast cell-committed progenitors could be achieved by discontinuous Percoll gradients. Thus, we have identified a way to obtain late-stage, mast cell-committed progenitors in an environment that is virtually uncontaminated with other hematopoietic progenitors.
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PMID:The mast cell-committed progenitor. I. Description of a cell capable of IL-3-independent proliferation and differentiation without contact with fibroblasts. 278 62

A growth factor acting synergistically with IL-3 on thiol-sensitive "mucosal type" bone marrow-derived mast cell lines, and therefore termed mast cell growth enhancing activity, is present in PWM stimulated spleen cell conditioned medium. Mast cell growth enhancing activity can be partially purified and completely separated from IL-3, IL-4, and IL-5, and for the most part from IL-6 and GM-CSF using strong cation exchange and Procion red affinity chromatography. Mast cell growth enhancing activity binds to Con A-Sepharose and can be digested with trypsin and chymotrypsin. It shows a Mr ranging from 37 to 43 kDa under nonreducing SDS-PAGE and a main isoelectric point ranging from 6.2 to 7.3.
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PMID:Partial purification of a mast cell growth-enhancing activity and its separation from IL-3 and IL-4. 278 57


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