Expression of protease and protease-inhibitory activity in human mononuclear leukocyte cultures

Hatcher, Victor B.; Norin, Allen J.

doi:10.1016/0014-4827(82)90392-5

Cited by 5 publications

(6 citation statements)

References 21 publications

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“…Cell lines used in this study were obtained from the American Type Culture Collection (RockviUe, MD) and were maintained in media according to their instructions. Human erythrocytes and human PBL (HPBL) were isolated as described previously (23,24). Briefly, blood obtained from healthy volunteers was diluted 1:1 with HBSS, layered on Ficoll-Hypaque (Pharmacia, Piscataway, NJ), and centrifuged at 200 g for 20 min.…”

Section: Methodsmentioning

confidence: 99%

“…Lymphocyte-mediated Cytotoxicity. Cytolysis of tumor cells by HPBL was measured as described previously (23,24). In brief, 100 /A of SlCr-labeled target cells (2 x 104 cells/ml) was mixed with 100/xl of HPBL (2 x 106 cells/ml) to yield serial twofold dilutions of E/T cell ratios of 100:1 to 6:1.…”

Section: Methodsmentioning

confidence: 99%

“…Cytolysis of tumor cells by HPBL was measured as described previously (23,24 Staphylococcus aureus and human endothelial cell plasma membrane proteins (25). Initial studies were performed with the NK-sensitive K562 tumor cell line.…”

Section: Binding Of Tumor Plasma Membrane Proteins To Viable Lymfho-mentioning

confidence: 99%

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A novel ligand in lymphocyte-mediated cytotoxicity: expression of the beta subunit of H+ transporting ATP synthase on the surface of tumor cell lines.

Das

Mondragon

Sadeghian

et al. 1994

The Journal of Experimental Medicine

126

109

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SummaryExtracellular adenosine triphosphate (eATP) has been suggested to play a role in lymphocyteinduced tumor destruction. We now provide evidence that a protein responsible for ATP synthesis in mitochondria may also play a physiologic role in major histocompatibility complex-independent, lymphocyte-mediated cytotoxicity. A 51.5-kD protein (p51.5) beating structural and immunologic characteristics of the B subunit of H + transporting ATP synthase (E.C. 3.6.1.34, B-H +ATPase, published molecular mass of 51.6 kD) was detected on the plasma membrane of three different human tumor cell lines studied. NH2-terminal amino acid sequence analysis of purified p51.5 from K562 tumor cells revealed 100% homology of 16 residues identified in the first 21 positions to the known sequence of human mitochondrial B-H § ATPase. Antibody directed against a 2l-iner peptide in the ATP binding region of B-H+ATPase (anti-B) reacted with only one band on Western blots of whole tumor extracts and tumor membrane extracts suggesting that the antiserum reacts with a single species of protein. Anti-B reacted with the cell membranes of tumor ceils as determined by fluorescence-activated flow cytometry and immunoprecipitated a 51.5-kD protein from surface-labeled neoplastic cells (but not human erythrocytes and lymphocytes). Purified p51.5 bound to human lymphocytes and inhibited natural killer (NK) cell-mediated cytotoxicity. Furthermore, anti-B treatment of the K562 and A549 tumor cell lines inhibited NK (by >95%) and interleukin 2-activated killer (LAK) cell (by 75%) cytotoxicity, respectively. Soluble p51.5 upon binding to lymphocytes retained its reactivity to anti-B suggesting that the ATP binding domain and the lymphocyte-receptor binding domain reside in distinct regions of the ligand. These results suggest that B-H +ATPase or a nearly identical molecule is an important ligand in the effector phase (rather than the recognition phase) of a cytolytic pathway used by naive NK and LAK cells.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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A novel ligand in lymphocyte-mediated cytotoxicity: expression of the beta subunit of H+ transporting ATP synthase on the surface of tumor cell lines.

Das

Mondragon

Sadeghian

et al. 1994

The Journal of Experimental Medicine

126

109

View full text Add to dashboard Cite

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“…In contrast, Loskutoff and Edgington described inhibitors of PA in the cytosol of endothelial cells that could be irreversibly inactivated by acidification [29]. Similarly, Hatcher and Norin have shown that human T-lymphocytes, after days of activation, secrete trypsin-inhibitory activity as well as activated protease activity [41].…”

Section: Discussionmentioning

confidence: 93%

Plasminogen‐dependent fibrinolytic activity in normal human lymphocytes: Diminished lymphocyte plasminogen activator in chronic lymphocytic leukemia

1985

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Discrepancies in correlations between fibrinolytic activity and metastatic potential of malignant cells has resulted in speculation on the putative role of plasminogen activators (PA) in cancer. In this report we have compared lymphocyte PA from 40 patients with chronic lymphocytic leukemia (CLL) to normal human B- and T-lymphocytes. Lymphocytes were isolated from peripheral blood by Ficoll-Hypaque centrifugation. The B- and T-cells were further separated on nylon wool columns. Cell PA activity and cell membrane PA were determined using 3H-fibrin-coated plates with added human plasminogen. Lymphocytes did not lyse 3H-fibrin in the absence of plasminogen. Plasminogen-dependent fibrinolytic activities of normal B- and T-lymphocytes were comparable. The addition of protease inhibitors with trypsin or plasmin specificity to lymphocytes significantly inhibited normal PA, thus substantiating the serine protease spectrum of lymphocyte PA. Examination of lymphocytes from greater than 95% of patients with chronic lymphocytic leukemia revealed a marked decrease in lymphocyte and cell membrane PA as compared to normals. No correlation between Stage of CLL and lymphocyte PA was observed. Likewise, an inhibitor of PA in CLL lymphocytes was not detected. The function of PA in normal B-lymphocyte physiology and the potential pathogenetic role of diminished PA in CLL lymphocytes remain to be explored.

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“…It has been shown in the past that human peripheral blood mononuclear cells (PBMC) contain neutral proteolytic activity(ies) either surface bound [21] or secretable by lymphocytes [22] and that PBMC express a serine-type pro- [23]. Since in all previous studies heterogeneous cell populations have been used, the cell source of the particular enzyme(s) has not yet been clearly identified.…”

Section: Introductionmentioning

confidence: 99%

A novel serine proteinase (HuTSP) isolated from a cloned human CD8⁺ cytolytic T cell line is expressed and secreted by activated CD4⁺ and CD8⁺ lymphocytes

et al. 1987

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A novel proteinase, termed human T cell-associated serine proteinase (HuTSP), has been highly purified from a human CD8+ T lymphocyte clone. By using a panel of chromogenic model peptide substrates the enzyme was found to specifically recognize L-arginine and to cleave Tos-Gly-Pro-Arg-nitroanilide with high efficiency at a pH optimum of 10.5-11. Exposure to class-specific proteinase inhibitors revealed that HuTSP is a serine endopeptidase. The enzyme has a mol. mass of approximately 50 kDa (non-reduced) and of approximately 25-30 kDa (reduced) when analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis suggesting HuTSP to be a disulfide-linked dimer. The enzyme is shown to be inducible by lectin in both CD4+ and CD8+ lymphocytes. Moreover, HuTSP was detected in a number of independent CD4+ and CD8+ T cell clones and was found to be released from effector cells upon ligand binding to the CD3-T cell receptor complex.

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Expression of protease and protease-inhibitory activity in human mononuclear leukocyte cultures

Cited by 5 publications

References 21 publications

A novel ligand in lymphocyte-mediated cytotoxicity: expression of the beta subunit of H+ transporting ATP synthase on the surface of tumor cell lines.

A novel ligand in lymphocyte-mediated cytotoxicity: expression of the beta subunit of H+ transporting ATP synthase on the surface of tumor cell lines.

Plasminogen‐dependent fibrinolytic activity in normal human lymphocytes: Diminished lymphocyte plasminogen activator in chronic lymphocytic leukemia

A novel serine proteinase (HuTSP) isolated from a cloned human CD8⁺ cytolytic T cell line is expressed and secreted by activated CD4⁺ and CD8⁺ lymphocytes

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Expression of protease and protease-inhibitory activity in human mononuclear leukocyte cultures

Cited by 5 publications

References 21 publications

A novel ligand in lymphocyte-mediated cytotoxicity: expression of the beta subunit of H+ transporting ATP synthase on the surface of tumor cell lines.

A novel ligand in lymphocyte-mediated cytotoxicity: expression of the beta subunit of H+ transporting ATP synthase on the surface of tumor cell lines.

Plasminogen‐dependent fibrinolytic activity in normal human lymphocytes: Diminished lymphocyte plasminogen activator in chronic lymphocytic leukemia

A novel serine proteinase (HuTSP) isolated from a cloned human CD8+ cytolytic T cell line is expressed and secreted by activated CD4+ and CD8+ lymphocytes

Contact Info

Product

Resources

About

A novel serine proteinase (HuTSP) isolated from a cloned human CD8⁺ cytolytic T cell line is expressed and secreted by activated CD4⁺ and CD8⁺ lymphocytes