Role of STAT3 in glucocorticoid-induced expression of the human IL-10 gene.

In the present report we have determined the molecular mechanisms, which govern the expression of the human IL-10 gene when induced by the glucocorticoid Methyl-Prednisolone (MP). Treatment of cells with MP at 10(-6) M will readily induce IL-10 in CD19+ primary B cells and in a human B cell line. Analysis of the IL-10 promoter showed a robust 18-fold induction and demonstrated that a potential GRE motif was not required, while mutation of the -120 STAT-motif strongly reduced MP-induced trans-activation. A strong induction was also seen with a trimeric STAT-motif and over-expression of dominant-negative STAT3 could block MP induction of IL-10 mRNA. Finally, MP treatment induced binding of STAT3 to the promoter as shown by gelshift, supershift and by chromatin-immunoprecipitation. These data show that glucocorticoid-induced expression of the IL-10 gene is mediated by the transcription factor STAT3.

Epidemiology of tick bites and borreliosis in children attending kindergarten or so-called "forest kindergarten" in southwest Germany.

The so-called "forest kindergartens" have been increasingly popular in Germany since the beginning of the 1990s. These are nurseries located in forested areas where children spend all-season full-time outdoors. Fifty-three kindergartens in the state of Baden-Württemberg, Germany participated in this study. In a prospective clinical cohort study, the child's personal data, history, protective parental habits concerning tick bites, number of tick bites, and cases of borreliosis were recorded monthly (March-October 2004) using a questionnaire. Altogether, 1,707 children of 25 "forest kindergartens" (506 children) and 28 conventional kindergartens (1,201 children) were included. The response rate was 75% in "forest kindergartens" and 65% in conventional kindergartens. In the "forest kindergartens", 1,503 tick bites especially on the trunk and on the head were found, whereas 502 tick bites were registered in conventional kindergartens. Sixteen cases of borreliosis were diagnosed (10 in "forest kindergartens", six in conventional kindergarten), most frequently manifesting as erythema migrans. Children attending a "forest kindergarten" have a 2.8 times increased risk of experiencing tick bites and a 4.6 times increased risk of suffering from borreliosis compared to conventional kindergarten in Germany, although protective parental behavior in "forest kindergarten" children was significantly better than that in conventional kindergarten.

Mechanism of interferon-gamma mediated down-regulation of interleukin-10 gene expression.

Expression of the anti-inflammatory cytokine IL-10 is suppressed by the pro-inflammatory interferon gamma but the mechanism of this action is unknown. We analysed activity of IL-10 promoter luciferase reporter constructs in transfected RPMI 8226.1 B cells that were treated at -2 h with IFNgamma (1000 U/ml) followed by stimulation with LPS (100 ng/ml) at 0 h. IFNgamma treatment suppressed LPS-induced IL-10 promoter activity in a construct carrying the -1044 promoter and also one containing the -195 promoter. The suppression was independent of the IRF-motif at -182 but involved the Stat-motif at -120. In gelshift analysis this Stat motif did bind LPS-induced Stat3 and with IFNgamma treatment it did, in addition, bind Stat1. ChIP analysis for detection of transcription factor binding to chromatin in intact cells demonstrated Stat3 binding to the proximal IL-10 promoter when cells are stimulated with LPS only. Treatment with IFNgamma only led to Stat1 binding in ChIP analysis and treatment with IFNgamma plus LPS led to reduced Stat3 binding while Stat1 binding remained high. Finally, LPS-induced activity of the trimeric Stat-motif in front of the luciferase reporter was suppressed by IFNgamma. These data demonstrate that IFNgamma down-regulates expression of the IL-10 gene by a novel mechanism that involves displacement of transactiving Stat3 by IFNgamma-induced Stat1.

IFN-alpha induces the human IL-10 gene by recruiting both IFN regulatory factor 1 and Stat3.

The anti-inflammatory cytokine IL-10 can be induced by type I IFNs, but the molecular mechanisms involved have remained elusive. With in silico analysis of the human IL-10 promoter we identified a module consisting of an IFN regulatory factor 1 (IRF-1) site and a Stat3 site. We demonstrate that IFN-alpha will induce the binding of IRF-1 and Stat3 to the respective motifs. Mutational analysis revealed that inactivation of the IRF-1 motif substantially reduces trans-activation from 5- to 2-fold and that inactivation of the Stat3 motif completely ablates trans-activation by IFN-alpha. The dominant role of Stat3 in this module was confirmed with the blockade of trans-activation by a dominant negative Stat3. By contrast, Stat1 contributes a minor proportion to the DNA binding to the Stat site, and overexpression will counteract Stat3-mediated trans-activation. The data show that IFN-alpha induces the IL-10 gene via a module consisting of interdependent IRF-1 and Stat3 motifs. Of note, LPS-induced trans-activation does not target this module, since it is independent of the IRF-1 motif but completely depends on Stat3.