title: TNF-alpha secretion and apoptosis of lymphocytes mediated by gene transfer.
abstract: Efficient gene transfer of lymphocytes is extremely difficult. Apoptosis may play a role in this gene transfer resistance of lymphocytes. Here we show that transfection of lymphocytes via non-viral vectors leads to induction of apoptosis in a significant proportion of cells. Since apoptosis may be mediated via tumor necrosis factor d (TNF-alpha) and the TNF-alpha receptor pathway, we studied the amount of TNF-alpha secreted by lymphocytes transfected without gene insert. TNF-alpha secretion was dependent on the gene transfer method used. High amounts were detected using receptor-mediated gene transfer and lipofection. In contrast, only low amounts of TNF-alpha were detected after electroporation and retroviral gene transfer. In receptor-mediated gene transfer, TNF-alpha secretion was due to the use of anti-CD3 antibody. Transfection of lymphocytes led to selective decrease in CD120b/TNF-alpha receptor II (TNFR-2)-positive cells. Induction of apoptosis and necrosis mediated by TNF-alpha via TNFR-2 (p80) was partially blocked using a neutralizing anti-TNF-alpha antibody. Blockage of apoptosis and necrosis could be further increased by adding anti-Fas-ligand (FasL) antibody, suggesting that induction of apoptosis via FasL and Fas receptor (Apo-1/CD95) may also play a role. This blockage led to a significant increase in the proliferation rate of lymphocytes transfected with cytokine genes. In conclusion, various gene transfer techniques led to TNF-alpha secretion, apoptosis and necrosis of lymphocytes. Apoptosis and necrosis could be partially blocked using a neutralizing anti-TNF-alpha antibody.
authors.affiliation: Abteilung für Innere Medizin und Poliklinik m. S. Hämatologie und Onkologie, Humboldt-Universität zu Berlin, Germany.
authors.forename: O
authors.initials: O
authors.lastname: Ebert
authors.forename: G
authors.initials: G
authors.lastname: Röpke
authors.forename: A
authors.initials: A
authors.lastname: Märten
authors.forename: P
authors.initials: P
authors.lastname: Lefterova
authors.forename: B
authors.initials: B
authors.lastname: Micka
authors.forename: P
authors.initials: P
authors.lastname: Buttgereit
authors.forename: S
authors.initials: S
authors.lastname: Niemitz
authors.forename: B
authors.initials: B
authors.lastname: Trojaneck
authors.forename: G
authors.initials: G
authors.lastname: Schmidt-Wolf
authors.forename: D
authors.initials: D
authors.lastname: Huhn
authors.forename: B
authors.initials: B
authors.lastname: Wittig
authors.forename: I G
authors.initials: IG
authors.lastname: Schmidt-Wolf
mesh_terms: D000911:Antibodies, Monoclonal / Q000494:pharmacology; D017209:Apoptosis; D016207:Cytokines / Q000494:pharmacology*; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D018014:Gene Transfer Techniques; D006801:Humans; D066298:In Vitro Techniques; D008214:Lymphocytes / Q000166:cytology* / Q000378:metabolism; D009336:Necrosis; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000276:immunology / Q000378:metabolism*; D019014:fas Receptor / Q000276:immunology
chemical_list: D000911:Antibodies, Monoclonal; D016207:Cytokines; D014409:Tumor Necrosis Factor-alpha; D019014:fas Receptor