Alcohol consumption decreases IL-2-induced NF-kappaB activity in enriched NK cells from C57BL/6 mice.
Previously we showed that ethanol (EtOH) consumption suppressed IL-2-induced cytolytic activity of murine splenic natural killer (NK) cells. Although IL-2 receptor signaling is involved in activation of NK cells, neither the mechanism for this activation nor the role of EtOH consumption in modulating activation is completely understood. In this study we show by electrophoretic mobility-shift assay (EMSA) that enriched splenic NK cells from EtOH-consuming C57BL/6 mice exhibit reduced NF-kappaB and AP-1 binding activity in response to IL-2 stimulation as compared to the water-drinking mice. Semiquantitative RT-PCR and real-time PCR analyses indicated that EtOH consumption inhibits the induction of perforin, granzyme A, and granzyme B in response to IL-2. Pyrrolidine dithiocarbamate (PDTC) and N-tosyl-L-phenylalanine chloromethyl ketone (TPCK) blocked NFkappaB and AP-1 binding activity in nuclear extracts of IL-2-stimulated NK cells in an EMSA and also inhibited the IL-2-induced expression of perforin, granzyme A, and granzyme B gene expression in enriched NK cells. These inhibitors dramatically suppressed IL-2-stimulated NK cytolytic activity against YAC-1 lymphoma target cells. Taken together, these results suggest that NFkappaB and AP-1 are important regulators of NK cell cytolytic function through regulation of perforin, granzyme A, and granzyme B gene expression. The findings further suggest that the decreased cytolytic activity of IL-2-stimulated NK cytolytic activity in EtOH-consuming mice is due at least in part to impaired transactivation of these and possibly other genes involved in control of NK-cell target lysis.
Zhou
J
J
Cancer Prevention Research Center and the Department of Pharmaceutical Sciences, College of Pharmacy, Washington State University, Pullman 99164-6510, USA.
Meadows
G G
GG
D000818:Animals; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D002492:Central Nervous System Depressants / Q000633:toxicity*; D024202:Electrophoretic Mobility Shift Assay; D000431:Ethanol / Q000633:toxicity*; D005260:Female; D053804:Granzymes; D007376:Interleukin-2 / Q000037:antagonists & inhibitors* / Q000494:pharmacology*; D007694:Killer Cells, Natural / Q000187:drug effects / Q000378:metabolism*; D008562:Membrane Glycoproteins / Q000037:antagonists & inhibitors; D051379:Mice; D008810:Mice, Inbred C57BL; D016328:NF-kappa B / Q000037:antagonists & inhibitors / Q000378:metabolism*; D054353:Perforin; D052899:Pore Forming Cytotoxic Proteins; D011392:Proline / Q000031:analogs & derivatives* / Q000494:pharmacology; D012333:RNA, Messenger / Q000096:biosynthesis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012697:Serine Endopeptidases / Q000096:biosynthesis; D013859:Thiocarbamates / Q000494:pharmacology; D014108:Tosylphenylalanyl Chloromethyl Ketone / Q000494:pharmacology; D018808:Transcription Factor AP-1 / Q000378:metabolism
D002492:Central Nervous System Depressants; D007376:Interleukin-2; D008562:Membrane Glycoproteins; D016328:NF-kappa B; D052899:Pore Forming Cytotoxic Proteins; D012333:RNA, Messenger; D013859:Thiocarbamates; D018808:Transcription Factor AP-1; D054353:Perforin; C066229:prolinedithiocarbamate; D000431:Ethanol; D014108:Tosylphenylalanyl Chloromethyl Ketone; D011392:Proline; D053804:Granzymes; C509214:Gzmb protein, mouse; D012697:Serine Endopeptidases