Activation and attenuation of transcription factor NF-kB in mouse glomerular mesangial cells in response to tumor necrosis factor-alpha, immunoglobulin G, and adenosine 3':5'-cyclic monophosphate. Evidence for involvement of reactive oxygen species.
The transcription factor NF-kB may play an important role in the response to tissue injury and activation of cytokines. We therefore examined the regulation of NF-kB in mesangial cells. Treatment of mesangial cells with TNF-alpha increased nuclear proteins that bound to an NF-kB-specific DNA oligonucleotide. IgG aggregates also increased nuclear NF-kB demonstrating Fc-tau receptor-mediated activation of NF-kB. Treatment of a cytosolic preparation with the detergent deoxycholate also activated NF-kB. The binding characteristics were typical for NF-kB transcription factors as determined by competition experiments with NF-kB-binding wild type kB DNA oligonucleotides or mutated oligonucleotides. Furthermore, a monoclonal antibody against the p65 subunit of NF-kB prevented the binding of NF-kB to the kB oligonucleotide. To evaluate the potential role of reactive oxygen intermediates in the activation of NF-kB, we used PDTC as a scavenger and HMAP as an inhibitor of NADPH-dependent oxidase. Both PDTC and HMAP attenuated the increase in nuclear NF-kB in response to either TNF-alpha or IgG complexes. Finally, generation of superoxide anion by xanthine oxidase activated NF-kB, an effect also mitigated by PDTC. In contrast, exogenous H2O2 did not activate NF-kB. Preincubation of cells with 8 br-cAMP, forskolin, or PGE2 attenuated the increase in nuclear NF-kB in response to TNF-alpha, aggregated IgG, or superoxide anion. Our results provide support for a role of reactive oxygen intermediates as mediators for activation of NF-kB in MC after stimulation with TNF-alpha or IgG aggregates. As an unexpected novel finding we report that cAMP can inhibit activation of NF-kB in MC. These observations may help to explain effects of TNF-alpha, IgG aggregates and cAMP on generation of cytokines by mesangial cells and the resulting glomerular pathophysiology.
Satriano
J
J
Department of Medicine, Albert Einstein College of Medicine, Bronx, New York 10461.
Schlondorff
D
D
D000098:Acetophenones / Q000494:pharmacology; D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000494:pharmacology*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000494:pharmacology; D004247:DNA / Q000378:metabolism; D015232:Dinoprostone / Q000494:pharmacology; D016166:Free Radical Scavengers / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects; D005920:Glomerular Mesangium / Q000166:cytology / Q000378:metabolism*; D007074:Immunoglobulin G / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D016328:NF-kappa B / Q000235:genetics* / Q000378:metabolism; D017382:Reactive Oxygen Species; D017452:Receptors, IgG / Q000502:physiology; D013481:Superoxides; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*; D014969:Xanthine Oxidase
D000098:Acetophenones; D016166:Free Radical Scavengers; D007074:Immunoglobulin G; D016328:NF-kappa B; D017382:Reactive Oxygen Species; D017452:Receptors, IgG; D014409:Tumor Necrosis Factor-alpha; D013481:Superoxides; D005576:Colforsin; D004247:DNA; C056165:acetovanillone; D000242:Cyclic AMP; D014969:Xanthine Oxidase; D017868:Cyclic AMP-Dependent Protein Kinases; D015232:Dinoprostone