Interleukin (IL)-10 inhibits IL-6 production in microglia by preventing activation of NF-kappaB.
The purpose of this study was to determine if interleukin (IL)-10 inhibits lipopolysaccharide (LPS)-induced IL-6 production in microglia by inhibiting activation of nuclear factor-kappaB (NF-kappaB). N13 microglia (a murine microglial cell line) and primary microglia from neonatal mice were cultured in the presence or absence of LPS and increasing amounts of murine IL-10 for 24 h. As predicted, LPS treatment increased supernatant IL-6 concentration in both N13 and primary microglia cultures. Pretreatment with IL-10, however, decreased LPS-induced IL-6 secretion in a dose-dependent manner in both culture systems. Likewise, ribonuclease protection assays showed that LPS increased steady-state IL-6 mRNA levels, but that pretreatment with IL-10 blocked the LPS-induced increase in IL-6 mRNA. Because NF-kappaB is the predominant transcription factor responsible for IL-6 transcription in response to inflammatory stimuli, it was hypothesized that IL-10 inhibited IL-6 production by preventing nuclear translocation of NF-kappaB. Consistent with this idea, LPS increased nuclear translocation of NF-kappaB as assessed by gel mobility shift assay. Supershift assays and immunocytochemical staining showed that both the p50 and p65 subunits of NF-kappaB translocated from the cytoplasm to the nucleus upon LPS stimulation. Pretreatment with IL-10, however, inhibited LPS-induced activation of NF-kappaB. Furthermore, inhibition of NF-kappaB activity with tosyl-Phe-chloromethlyketone (a serine protease inhibitor that prevents degradation of the NF-kappaB-IkappaB complex), completely blocked LPS-induced IL-6 production. These data suggest that IL-10 inhibited IL-6 production in microglia by decreasing the activity of NF-kappaB and, therefore, extend what little is known of the intricate relationship between anti-inflammatory and inflammatory cytokines in the central nervous system.
Heyen
J R
JR
Laboratory of Integrative Biology, Department of Animal Sciences, University of Illinois at Urbana-Champaign, Urbana, IL, USA.
Ye
S
S
Finck
B N
BN
Johnson
R W
RW
D000818:Animals; D000831:Animals, Newborn; D001921:Brain / Q000276:immunology*; D002460:Cell Line; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D002478:Cells, Cultured; D003593:Cytoplasm / Q000187:drug effects / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects / Q000276:immunology*; D016753:Interleukin-10 / Q000494:pharmacology*; D015850:Interleukin-6 / Q000235:genetics*; D008070:Lipopolysaccharides / Q000494:pharmacology; D051379:Mice; D008807:Mice, Inbred BALB C; D017628:Microglia / Q000187:drug effects / Q000276:immunology*; D016328:NF-kappa B / Q000037:antagonists & inhibitors*; D014107:Tosyllysine Chloromethyl Ketone / Q000494:pharmacology; D014108:Tosylphenylalanyl Chloromethyl Ketone / Q000494:pharmacology
D015850:Interleukin-6; D008070:Lipopolysaccharides; D016328:NF-kappa B; D016753:Interleukin-10; D014107:Tosyllysine Chloromethyl Ketone; D014108:Tosylphenylalanyl Chloromethyl Ketone