The superoxide generating system of B cell lines. Structural homology with the phagocytic oxidase and triggering via surface Ig.
EBV-transformed B lymphocyte cell lines (EBV-BLCL) produce superoxide after stimulation with phorbol ester, a capacity unique among nonmyeloid cells. The superoxide producing system of EBV-BLCL (B cell oxidase) was compared with the phagocytic NADPH-oxidase and the relationship of the capacity to produce superoxide to the presence of the EBV-genome was analyzed. The two EBV-transformed B cell lines F1 and HELL generated superoxide in response to PMA (2.3 nmol/10(6) F1 cells x 1 h and 6.27 nmol/10(6) HELL cells x 1 h with 1 microgram/ml of PMA), whereas no superoxide release was detected with the EBV-positive Burkitt lymphoma line WIL-2 and the EBV-negative plasmocytoma line U-266. Also, F1 and HELL showed lucigenin-dependent chemiluminescence (CL) after PMA-treatment, whereas no CL responses were detected from WIL-2 or U-266. Further, F1 and HELL cells contained a low potential cytochrome b-245 (10.9 and 61.0 pmol/mg protein, respectively) and also a 45 kDa diphenylene-iodonium (DPI)-binding peptide, both components of the phagocytic NADPH-oxidase. In contrast, neither the cytochrome b-245 nor the 45 kDa DPI-binding peptide were detected in WIL-2 and U-266. In addition, DPI inhibited O2- production by PMA-stimulated EBV-BLCL and polymorphonuclear granulocytes. Further, F1 line cells showed superoxide dismutase-inhibitable lucigenin-dependent CL when triggered by protein A-bearing staphylococci (Cowan strain I) or by a mAb directed against human IgG in the presence of solid-phase goat anti-mouse-Ig antibody. From a panel of eight EBV-BLCL, only five responded with CL when exposed to protein A-bearing staphylococci, whereas all showed CL when treated with phorbol ester. Inasmuch as all eight EBV-BLCL possessed surface Ig and a "functional" oxidase, their differential response to cross-linking of surface Ig may be determined by differences in signal transduction. Superoxide production by EBV-BLCL appears thus related to expression of an electron transport chain structurally homologous, if not identical, with the "phagocytic" NADPH-oxidase. Apparently, the presence of EBV-genome in B cell lines does not per se lead to expression of this oxidase. This suggests that nontransformed B cells may, at a certain differentiation stage, also express a superoxide-generating chain. From the finding of stimulation of superoxide production of EBV-BLCL via surface Ig it appears possible that also Ag may be able to trigger such B cells to production of superoxide which might have an important role in the physiology of B cells.
Maly
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FE
Institute of Clinical Immunology, Inselspital, Bern, Switzerland.
Cross
A R
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Jones
O T
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Wolf-Vorbeck
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G
Walker
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C
Dahinden
C A
CA
De Weck
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D001402:B-Lymphocytes / Q000201:enzymology / Q000378:metabolism* / Q000502:physiology; D002460:Cell Line; D002461:Cell Line, Transformed; D002472:Cell Transformation, Viral; D004579:Electron Transport; D004789:Enzyme Activation; D004854:Herpesvirus 4, Human; D006801:Humans; D008213:Lymphocyte Activation / Q000187:drug effects; D009247:NADH, NADPH Oxidoreductases / Q000378:metabolism / Q000502:physiology*; D019255:NADPH Oxidases; D010586:Phagocytes / Q000201:enzymology*; D011947:Receptors, Antigen, B-Cell / Q000502:physiology*; D013481:Superoxides / Q000096:biosynthesis*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D011947:Receptors, Antigen, B-Cell; D013481:Superoxides; D009247:NADH, NADPH Oxidoreductases; D019255:NADPH Oxidases; D013755:Tetradecanoylphorbol Acetate