Synthesis of cathepsin B by cells derived from the HL60 promyelocytic leukaemia cell line

Cells of the human promyelocytic HL60 line were induced to differentiate into granulocyte-like cells with dimethylsulphoxide (DMSO) or macrophage-like cells with 12-0-tetradecanoylphorbol-13-acetate (TPA). The synthesis of Cathepsin B by these cells was studied by immunoperoxidase staining and assay of cell lysates using the fluorimetric substrate benzoyloxycarbonyl-phenylanalyl-arginine-4-methyl-7-coumarylamide. Only 2–5% of the uninduced HL60 cells and DMSO-induced cells were immunohistochemically positive for Cathepsin B, compared with over 80% of the TPA-induced cells. Cathepsin B activity was lowest in the lysates of uninduced HL60s. DMSO-induced cells contained 1.5–2-fold the enzyme activity of HL60s and TPA-induced cell lysates demonstrated 5–14-fold the activity of uninduced HL60s. Induction of Cathepsin B synthesis was therefore associated with differentiation of the promyelocytes into cells of the monocyte/macrophage type, but not granulocyte-like cells. Cathepsin B was located immunohistochemically in human palatine tonsils. The enzyme was only demonstrated within macrophages in these tissues. Cathepsin B may therefore be a useful immunohistochemical marker for malignant and nonmalignant cells of the monocyte/macrophage lineage.     

Constitutively hyposialylated human T-lymphocyte clones in the Tn-syndrome: binding characteristics of plant and animal lectins

Previously, 1,3-galactosyltransferase-deficient (Tn+) and normal (TF+) T-lymphocyte clones have been established from a patient suffering from Tn-syndrome [Thurnheret al. (1992)Eur J Immunol 22: 1835–42], Tn+ T lymphocytes express only Tn antigen (GalNAc1-O-R) while other O-glycan structures such as sialosyl-Tn (Neu5Ac2,6GalNAc1-O-R) or TF (Gal1-3GalNAc1-O-R) antigens are absent from these cells as shown by flow cytometry using specific mABs for TF and sialosyl-Tn antigen, respectively. Normal T lymphocytes express the TF antigen and derivatives thereof. The surface glycans of Tn+ and TF+ cells were then analysed by flow cytometry using the following sialic acid-binding lectins:Amaranthus caudatus (ACA),Maackia amurensis (MAA),Limax flavus (LFA),Sambucus nigra (SNA) andTriticum vulgare (WGA). Equal and weak binding of MAA and SNA to both TF+ and Tn+ cells was found. WGA, LFA and ACA bound more strongly to TF+ cells than to Tn+ cells. Binding of ACA to TF+ cells was enhanced after sialidase treatment. To investigate the possible biological consequences of hyposialylation, binding of three sialic acid-dependent adhesion molecules to Tn+ and TF+ cells was estimated using radiolabelled Fc-chimeras of sialoadhesin (Sn), myelin-associated glycoprotein (MAG) and CD22. Equal and strong binding of human CD22 to both TF+ and Tn+ cells was found. Whereas binding of Sn and MAG to TF+ cells was strong (100%), binding to Tn+ cells amounted only to 33% (Sn) and 19% (MAG). These results indicate that thein vivo interactions of T lymphocytes in the Tn syndrome with CD22 are not likely to be affected, whereas adhesion mediated by Sn or MAG could be strongly reduced.     

Sediment-related growth limitation of Elodea nuttallii as indicated by a fertilization experiment

1. A fertilization experiment was performed to identify the limiting nutrient for the growth of submerged vegetation in ditches of a peat-grassland system in the Netherlands, in which restoration measures involved ceasing fertilization, exporting nutrients by removal of above-ground plant mass and large-scale introduction of calcium-rich, nutrient-poor artesian water.
2. Growth of Elodea was significantly enhanced by enrichment with nitrogen alone, and by fertilization with nitrogen in combination with phosphorus, and by nitrogen in combination with phosphorus and potassium.
3. Plant tissue nutrient concentrations increased significantly, for nitrogen by enrichment with nitrogen alone, and with nitrogen in combination with phosphorus and potassium; for phosphorus by enrichment with phosphorus alone and with phosphorus in combination with nitrogen and potassium; tissue concentrations of potassium were not enhanced by any treatment.
4. The elemental ratios of treated plants indicated that nitrogen, rather than phosphorus, was limiting in all treatments, except in those involving nitrogen and NK enrichment (when phosphorus was limiting).
5. The efficiency with which plants used nutrients declined with increased supply of nitrogen and phosphorus, but was unchanged when potassium was increased. Efficiencies were similar to those of other aquatic macrophytes.     

Sialoadhesin, a macrophage sialic acid binding receptor for haemopoietic cells with 17 immunoglobulin-like domains.

Sialoadhesin is a macrophage-restricted adhesion molecule of 185 kDa that mediates sialic acid-dependent binding to cells. It is expressed strongly by macrophages in lymphoid and haemopoietic tissues where it is likely to mediate cell-cell interactions. Here we report the molecular cloning of murine sialoadhesin and show that it is a new member of the immunoglobulin (Ig) superfamily with 17 Ig-like domains. COS cells transfected with a cDNA encoding full-length sialoadhesin bound mouse bone marrow cells in a sialic acid-dependent manner. Alternatively spliced cDNAs, predicting soluble forms of sialoadhesin containing the first three or 16 Ig-like domains of sialoadhesin, were expressed in COS cells and the respective proteins purified. When immobilized on plastic, the 16-domain form bound cells in a sialic acid-dependent manner, suggesting that sialoadhesin can function in both secreted and membrane-bound forms. The most similar proteins in the database were CD22, myelin-associated glycoprotein, Schwann cell myelin protein and CD33. Like sialoadhesin, CD22 mediates sialic acid-dependent cell adhesion. The sequence similarity of sialoadhesin to CD22 and related members of the Ig superfamily indicates the existence of a novel family of sialic acid binding proteins involved in cell-cell interactions.     

Sediment-related growth limitation of Elodea nuttallii as indicated by a fertilization experiment

1. A fertilization experiment was performed to identify the limiting nutrient for the growth of submerged vegetation in ditches of a peat-grassland system in the Netherlands, in which restoration measures involved ceasing fertilization, exporting nutrients by removal of above-ground plant mass and large-scale introduction of calcium-rich, nutrient-poor artesian water.
2. Growth of Elodea was significantly enhanced by enrichment with nitrogen alone, and by fertilization with nitrogen in combination with phosphorus, and by nitrogen in combination with phosphorus and potassium.
3. Plant tissue nutrient concentrations increased significantly, for nitrogen by enrichment with nitrogen alone, and with nitrogen in combination with phosphorus and potassium; for phosphorus by enrichment with phosphorus alone and with phosphorus in combination with nitrogen and potassium; tissue concentrations of potassium were not enhanced by any treatment.
4. The elemental ratios of treated plants indicated that nitrogen, rather than phosphorus, was limiting in all treatments, except in those involving nitrogen and NK enrichment (when phosphorus was limiting).
5. The efficiency with which plants used nutrients declined with increased supply of nitrogen and phosphorus, but was unchanged when potassium was increased. Efficiencies were similar to those of other aquatic macrophytes.     

Photolysis of 3-aryl-3-(trifluoromethyl)diazirines: a caveat regarding their use in photoaffinity probes.

The photolysis of 3-(4-tolyl)-3-(trifluoromethyl)diazirine in the presence of benzene, methanol, carbon tetrachloride, cyclohexane, triethylsilane, or diethylamine led to photoproducts consistent with the intermediacy of a singlet carbene. In the case of diethylamine, the photoinsertion into the N-H bond of diethylamine produced the expected adduct, 1-(diethylamino)-2,2,2-trifluoro-1-(4-tolyl)ethane. However, the base-catalyzed elimination of hydrogen fluoride from this adduct afforded an enamine, alpha-(diethylamino)-beta,beta-difluoro-4-methylstyrene, and the subsequent hydrolysis of this enamine furnished diethylamine and 2,2-difluoro-1-(4-tolyl)ethanone. This elimination and hydrolysis sequence effectively reversed the photoinsertion process. A similar photoinsertion and hydrolysis process using 3-(4-n-octylphenyl)-3-(trifluoromethyl)diazirine also produced 2,2-difluoro-1-(4-n-octylphenyl)ethanone in modest yield. These results suggest that the photoinsertion products from 3-aryl-3-(trifluoromethyl)-diazirines in biological systems may suffer similar fates limiting, in part, their utility in obtaining primary sequence data.     

The Effects of IL-1β on Astrocytes are Conveyed by Extracellular Vesicles and Influenced by Age

Neurochemical Research - The aging brain is associated with significant pathophysiological changes reflected in changes in astrocyte function. In this study, we hypothesized that the response of...     

Cellulose-bound Peptide Arrays: Preparation and Applications

Spatial organization of metabolic enzymes may represent a general cellular mechanism to regulate metabolic flux. One recent example of this type of cellular phenomenon is the purinosome, a newly discovered multi-enzyme metabolic assembly that includes all of the enzymes within the de novo purine biosynthetic pathway. Our understanding of the components and regulation of purinosomes has significantly grown in recent years. This paper reviews the purine de novo biosynthesis pathway and its regulation, and presents the evidence supporting the purinosome assembly and disassembly processes under the control of G-protein-coupled receptor (GPCR) signaling. This paper also discusses the implications of purinosome and GPCR regulation in drug discovery.