Su Wol Chung

Oxidized Low Density Lipoprotein Inhibits Interleukin-12 Production in Lipopolysaccharide-activated Mouse Macrophages via Direct Interactions between Peroxisome Proliferator-activated Receptor-γ and Nuclear Factor-κB

Lipopolysaccharide (LPS) increases the production of interleukin-12 (IL-12) from mouse macrophages via a κB site within the IL-12 p40 promoter. In this study, we found that oxidized low density lipoprotein (oxLDL) inhibited this LPS-stimulated production of IL-12 in a dose-dependent manner while native LDL did not. OxLDL inhibited p40 promoter activation in monocytic RAW264.7 cells transiently transfected with p40 promoter/reporter constructs, and the repressive effect mapped to a region in the p40 promoter containing a binding site for nuclear factor-κB (NF-κB) (p40-κB). Activation of macrophages by LPS in the presence of oxLDL resulted in markedly reduced binding to the κB site, as demonstrated by the electrophoretic mobility shift assays. In contrast, native LDL did not inhibit the IL-12 p40 promoter activation and NF-κB binding to the κB sites, suggesting that oxidative modification of LDL was crucial for the inhibition of NF-κB-mediated IL-12 production. 9-Hydroxyoctadecadienoic acid, a major oxidized lipid component of oxLDL, significantly inhibited IL-12 production in LPS-stimulated mouse macrophages and also suppressed NF-κB-mediated activation in IL-12 p40 promoter. The NF-κB components p50 and p65 directly bound peroxisome proliferator-activated receptor-γ (PPAR-γ) in vitro. In cotransfections of CV-1 and HeLa cells, PPAR-γ inhibited the NF-κB transactivation in an oxLDL-dependent manner. From these results, we propose that oxLDL-mediated suppression of the IL-12 production from LPS-activated mouse macrophages may, at least in part, involve both inhibition of the NF-κB-DNA interactions and physical interactions between NF-κB and PPAR-γ.

Retinoids Inhibit Interleukin-12 Production in Macrophages through Physical Associations of Retinoid X Receptor and NFκB

Lipopolysaccharide (LPS) increases the production of interleukin-12 (IL-12) from mouse macrophages via a κB site within the IL-12 p40 promoter. In this study, we found that retinoids inhibit this LPS-stimulated production of IL-12 in a dose-dependent manner. The NFκB components p50 and p65 bound retinoid X receptor (RXR) in a ligand-independent manner in vitro, and the interaction interfaces involved the p50 residues 1–245, the p65 residues 194–441, and the N-terminal A/B/C domains of RXR. Activation of macrophages by LPS resulted in markedly enhanced binding activities to the κB site, which significantly decreased upon addition of retinoids, as demonstrated by the electrophoretic mobility shift assays. In cotransfections of CV-1 and HeLa cells, RXR also inhibited the NFκB transactivation in a ligand-dependent manner, whereas a mutant RXR lacking the AF2 transactivation domain, which serves as ligand-dependent binding sites for transcription integrators SRC-1 and p300, was without any effect. In addition, coexpression of increasing amounts of SRC-1 or p300 relieved the retinoid-mediated inhibition of the NFκB transactivation. From these results, we propose that retinoid-mediated suppression of the IL-12 production from LPS-activated macrophages may involve both inhibition of the NFκB-DNA interactions and competitive recruitment of transcription integrators between NFκB and RXR.

Inhibition of interleukin-12 and interferon-γ production in immune cells by tanshinones from Salvia miltiorrhiza

Pharmacological control of interleukin-12 (IL-12) and interferon-gamma (IFN-gamma) production may be a key therapeutic strategy for modulating immunological diseases dominated by Th1-derived cytokine responses. In this study, we investigated the effects of three different tanshinone pigments from Salvia miltiorrhiza (tanshinone I, dihydrotanshinone, and cryptotanshinone) on IL-12 production in mouse macrophages and on IFN-gamma production in lymph node cells. All tested tanshinones significantly inhibited IL-12 production in lipopolysaccharide (LPS)-activated macrophages and also IFN-gamma production in keyhole limpet hemocyanin (KLH)-primed lymph node cells in a dose-dependent manner. Dihydrotanshinone was more effective than tanshinone I or cryptotanshinone. Tanshinones significantly inhibited the expression of IL-12 p40 gene at the mRNA level. Furthermore, tanshinones potently inhibited the promoter activation of IL-12 p40 gene and nuclear factor (NF)-kappaB binding to the kappaB site, suggesting that tanshinones may negatively regulate IL-12 production at the transcription level. These results may explain some known biological activities of tanshinones including their anti-inflammatory effect, and suggest a possible use of tanshinones in the treatment of immunological diseases dominated by Th1-derived cytokine responses.

Inhibition of interleukin-12 production in lipopolysaccharide-activated macrophages by curcumin.

Pharmacological control of interleukin-12 production may be a key therapeutic strategy for modulating immunological diseases dominated by type-1 cytokine responses. In this study we investigated the effects of curcumin (1,7-bis[4-hydroxy-3-methoxyphenyl]-1, 6-heptadiene-3,5-dione) on the production of interleukin-12 from mouse macrophages stimulated with lipopolysaccharide. Curcumin potently inhibited the production of interleukin-12 in a dose-dependent manner. The effect of curcumin on interleukin-12 p40 promoter activation was analyzed by transfecting RAW264.7 monocytic cells with p40 promoter/reporter constructs. The repressive effect mapped to a region in the p40 promoter containing a binding site for nuclear factor kappaB (p40-kappaB). Furthermore, activation of macrophages by lipopolysaccharide resulted in markedly enhanced binding activity to the kappaB site, which significantly decreased upon addition of curcumin. These results suggest that curcumin-induced inhibition of interleukin-12 production in macrophages may explain some of the biological effects of curcumin including its anti-inflammatory activity.

Enhancement of interleukin-4 production in activated CD4+ T cells by diphthalate plasticizers via increased NF-AT binding activity.

Background: Diethylhexyl phthalate (DEHP) and diisononyl phthalate (DINP), two commonly used plasticizers in flexible polyvinylchloride formulations, have potentially adverse effects on human health. However, the influence of these diphthalates on allergic responses remains unclear. In this study we examined the effects of DEHP and DINP on IL-4 production in CD4+ T cells and the level of IgE in sera, critical hallmarks associated with allergic diseases. Methods: Mouse T cells were exposed to two diphthalates in vitro and in vivo. The levels of IL-4 and IgE were determined by ELISA, and the degree of NF-AT activation was determined by IL-4 gene promoter assay and electrophoretic mobility shift assay. Results and Discussion: Both DEHP and DINP significantly enhanced IL-4 production in activated CD4+ T cells in a concentration-dependent manner. Treatment with DEHP or DINP in vivo resulted in a significant increase of IL-4 production in CD4+ T cells and of IgE levels in sera. Furthermore, DEHP and DINP enhanced the activation of IL-4 gene promoter in EL4 T cells and the enhancing effect mapped to a region in the IL-4 promoter containing binding sites for a transcription factor, NF-AT. The activation of T cells resulted in markedly enhanced binding activities to the NF-AT site, which significantly increased upon addition of DEHP or DINP, indicating that NF-AT was involved in the enhancing effect of DEHP and DINP on IL-4 production. These findings suggest that both DEHP and DINP enhance allergic responses by enhancement of IL-4 production in CD4+ T cells via stimulation of NF-AT-binding activity.

Retinoid-mediated inhibition of interleukin-12 production in mouse macrophages suppresses Th1 cytokine profile in CD4(+) T cells.

Interleukin‐12 (IL‐12) plays a central role in the immune system by driving the immune response towards T helper 1 (Th1) type responses characterized by high IFN‐γ and low IL‐4 production. In this study we investigated whether retinoid‐mediated inhibition of interleukin‐12 production in mouse macrophages could regulate cytokine profile of antigen (Ag)‐primed CD4+ Th cells. Pretreatment with retinoids (9‐cis‐RA, all‐trans‐RA, TTNPB) significantly inhibited IL‐12 production by mouse macrophages stimulated with lipopolysaccharide (LPS) or heated‐killed Listeria monocytogenes (HKL). Retinoid‐pretreated macrophages reduced their ability to induce IFN‐γ and increased the ability to induce IL‐4 in Ag‐primed CD4+ T cells. Addition of recombinant IL‐12 to cultures of retinoid‐pretreated macrophages and CD4+ T cells restored IFN‐γ production in CD4+ T cells. The in vivo administration of 9‐cis‐RA resulted in the inhibition of IL‐12 production by macrophages stimulated in vitro with either LPS or HKL, leading to the inhibition of Th1 cytokine profile (decreased IFN‐γ and increased IL‐4 production) in CD4+ T cells. These findings may explain some known effects of retinoids including the inhibition of encephalitogenicity, and point to a possible therapeutic use of retinoids in the Th1‐mediated immune diseases such as autoimmune diseases.

Involvement of p38 mitogen-activated protein kinase in the induction of interleukin-12 p40 production in mouse macrophages by berberine, a benzodioxoloquinolizine alkaloid.

Interleukin (IL)-12 plays a pivotal role in the development of T helper type 1 (Th1)-immune response, which may have therapeutic effects on diseases associated with pathologic Th2 responses such as allergic disorders and asthma. In this study, we investigated the effects of berberine, a benzodioxoloquinolizine alkaloid with anti-microbial and anti-tumor activities, on the production of IL-12 p40, an inducible subunit of IL-12, in mouse macrophages. Berberine-induced IL-12 p40 production and activation of p38 mitogen-activated protein kinase (MAPK) in dose-dependent manners, which were significantly inhibited by p38 MAPK inhibitors and yohimbine, indicating that p38 MAPK and alpha(2)-adrenergic receptor were involved in the induction of IL-12 p40 production in mouse macrophages by berberine. Furthermore, berberine significantly enhanced IL-12 p40 production in mouse macrophages when combined with lipopolysaccharide, a well-known inducer of IL-12 production. These findings may explain some of the known biological effects of berberine and suggests berberine as an immunotherapeutic compound for induction of IL-12, which is potentially applicable for tumors, infectious disease, and airway inflammation.

Inhibition of interleukin-12 production in lipopolysaccharide-activated mouse macrophages by parthenolide, a predominant sesquiterpene lactone in Tanacetum parthenium: involvement of nuclear factor-κB

Pharmacological control of interleukin-12 (IL-12) production may be a key therapeutic strategy for modulating immunological diseases dominated by type-1 cytokine responses. In this study, we investigated the effects of parthenolide, an anti-inflammatory sesquiterpene, on the production of IL-12 from mouse macrophages stimulated with lipopolysaccharide (LPS). Parthenolide potently inhibited the LPS-induced IL-12 production in a dose-dependent manner. The effect of parthenolide on IL-12 p40 promoter activation was analyzed by transfecting RAW264.7 monocytic cells with p40 promoter/luciferase constructs. The repressive effect mapped to a region in the p40 promoter containing a binding site for nuclear factor-kappaB (p40-kappaB). Furthermore, activation of macrophages by LPS resulted in markedly enhanced binding activity to the kappaB site, which significantly decreased upon addition of parthenolide. These results suggest that parthenolide-induced inhibition of IL-12 production in macrophages may explain some of the biological effects of parthenolide including its anti-inflammatory activity.

Hydroquinone, a reactive metabolite of benzene, enhances interleukin-4 production in CD4+ T cells and increases immunoglobulin E levels in antigen-primed mice

Exposure to cigarette smoke is known to increase the risk of the development of allergic disease. The mechanism is not well understood. In this study, we determined the effect of hydroquinone (HQ), a major metabolite of benzene present in large quantities in cigarette tar, on interleukin‐4 (IL‐4) production by CD4+ T cells. HQ significantly enhanced IL‐4 production by keyhole limpet haemocyanin (KLH)‐primed CD4+ T cells in a dose‐dependent manner. The enhancing effect of HQ on IL‐4 production was maximal at a concentration of 50 µm. It increased the level of IL‐4 production approximately 10‐fold. HQ enhanced IL‐4 mRNA expression and also IL‐4 gene promoter activity, suggesting that the enhancing effect of HQ on IL‐4 production may occur at the transcriptional level. Furthermore, the injection of KLH‐primed mice with HQ resulted in a significant increase in the levels of IL‐4 and immunoglobulin E. These findings provide evidence that HQ, a major component of cigarette tar, may enhance allergic immune responses by inducing the production of IL‐4 in CD4+ T cells.

Sulfasalazine prevents T-helper 1 immune response by suppressing interleukin-12 production in macrophages

Interleukin‐12 (IL‐12) plays a pivotal role in the development of T‐helper 1 (Th1) immune response, which may be involved in the pathogenesis of chronic inflammatory autoimmune disorders. In this study we investigated the effects of sulfasalazine, a drug for treating inflammatory bowel disease and rheumatoid arthritis, on the production of IL‐12 from mouse macrophages stimulated with lipopolysaccharide (LPS). Sulfasalazine potently inhibited the production of IL‐12 in a dose‐dependent manner, in part through the down‐regulation of nuclear factor κB (NFκB) activation in IL‐12 p40 gene. Activation of macrophages by LPS resulted in markedly enhanced binding activities to the κB site, which significantly decreased upon addition of sulfasalazine as demonstrated by an electrophoretic gel shift assay. Importantly, macrophages pretreated with sulfasalazine either in vitro or in vivo reduced their ability to induce interferon‐γ (IFN‐γ) and increased the ability to induce IL‐4 in antigen‐primed CD4+ T cells. From these results, sulfasalazine may induce the Th2 cytokine profile in CD4+ T cells by suppressing IL‐12 production in macrophages, and sulfasalazine‐induced inhibition of IL‐12 production in macrophages may explain some of the known biological effects of sulfasalazine.