Propolis has documented anti-inflammatory properties, although its mechanisms of action are poorly understood. In this study, the anti-inflammatory effects of polyphenol-rich propolis extracts (PPE) from China (CPPE) and Brazil (BPPE) were examined. Oral administration of PPE to lipopolysaccharide (LPS)-challenged mice decreased serum proinflammatory cytokine concentrations and inhibited pulmonary nuclear factor (NF)-κB activation. Both PPE types modulated LPS-induced key inflammatory mediators production in RAW 264.7 macrophages. They also suppressed NF-κB activation in HEK 293T cells, correlating well with their inhibitory effects on IκB phosphorylation and p65 nuclear translocation in LPS-activated macrophages. We found PPE suppressed NF-κB activation through delaying the ubiquitination of TRAF6 in HeLa-T6RZC stable cells and by directly disrupting the polyubiquitin synthesis in an in vitro kinase assay system. Overall, analysis showed substantial compositional differences between CPPE and BPPE; nevertheless, they both displayed similar anti-inflammatory properties through NF-κB-responsive inflammatory gene expressions by inhibiting TRAF6 dependent canonical NF-κB pathway. © 2015 Elsevier Ltd.19 464478Baker, R.G., Hayden, M.S., Ghosh, S., NF-κB, inflammation, and metabolic disease (2011) Cell Metabolism, 13 (1), pp. 11-22Bankova, V., Chemical diversity of propolis and the problem of standardization (2005) Journal of Ethnopharmacology, 100 (1-2), pp. 114-117Berbaum, K., Shanmugam, K., Stuchbury, G., Wiede, F., Korner, H., Munch, G., Induction of novel cytokines and chemokines by advanced glycation endproducts determined with a cytometric bead array (2008) Cytokine, 41 (3), pp. 198-203Bueno-Silva, B., Alencar, S.M., Koo, H., Ikegaki, M., Silva, G.V., Napimoga, M.H., Rosalen, P.L., Anti-inflammatory and antimicrobial evaluation of neovestitol and vestitol isolated from Brazilian red propolis (2013) Journal of Agricultural and Food Chemistry, 61 (19), pp. 4546-4550Bufalo, M.C., Ferreira, I., Costa, G., Francisco, V., Liberal, J., Cruz, M.T., Lopes, M.C., Sforcin, J.M., Propolis and its constituent caffeic acid suppress LPS-stimulated pro-inflammatory response by blocking NF-kappaB and MAPK activation in macrophages (2013) Journal of Ethnopharmacology, 149 (1), pp. 84-92Chan, G.C.-F., Cheung, K.-W., Sze, D.M.-Y., The immunomodulatory and anticancer properties of propolis (2013) Clinical Reviews in Allergy & Immunology, 44 (3), pp. 262-273Chawla, A., Nguyen, K.D., Goh, Y.S., Macrophage-mediated inflammation in metabolic disease (2011) Nature Reviews. Immunology, 11 (11), pp. 738-749Cuevas, A., Saavedra, N., Cavalcante, M.F., Salazar, L.A., Abdalla, D.S.P., Identification of microRNAs involved in the modulation of pro-angiogenic factors in atherosclerosis by a polyphenol-rich extract from propolis (2014) Archives of Biochemistry and Biophysics, 557 (1), pp. 28-35Daleprane, J.B., Freitas Vda, S., Pacheco, A., Rudnicki, M., Faine, L.A., Dorr, F.A., Ikegaki, M., Abdalla, D.S., Anti-atherogenic and anti-angiogenic activities of polyphenols from propolis (2012) Journal of Nutritional Biochemistry, 23 (6), pp. 557-566Deng, L., Wang, C., Spencer, E., Yang, L., Braun, A., You, J., Slaughter, C., Chen, Z.J., Activation of the IκB kinase complex by TRAF6 requires a dimeric ubiquitin-conjugating enzyme complex and a unique polyubiquitin chain (2000) Cell, 103 (2), pp. 351-361dos Santos, J.S., Monte-Alto-Costa, A., Caffeic acid phenethyl ester improves burn healing in rats through anti-inflammatory and antioxidant effects (2013) Journal of Burn Care & Research, 34 (6), pp. 682-688Haque, A., Koide, N., Noman, A.S.M., Odkhuu, E., Badamtseren, B., Naiki, Y., Komatsu, T., Yokochi, T., Flavopiridol inhibits lipopolysaccharide-induced TNF-α production through inactivation of nuclear factor-κB and mitogen-activated protein kinases in the MyD88-dependent pathway (2011) Microbiology and Immunology, 55 (3), pp. 160-167Hämäläinen, M., Nieminen, R., Vuorela, P., Heinonen, M., Moilanen, E., Anti-inflammatory effects of flavonoids: Genistein, kaempferol, quercetin, and daidzein inhibit STAT-1 and NF-κ B activations, whereas flavone, isorhamnetin, naringenin, and pelargonidin inhibit only NF-κ B activation along with their inhibitory effect on iNOS expression and NO production in activated macrophages (2007) Mediators of Inflammation, , 2007:45673Hervert-Hernández, D., Goñi, I., Dietary polyphenols and human gut microbiota: A review (2011) Food Reviews International, 27 (2), pp. 154-169Hou, J., Wang, J., Zhang, P., Li, D., Zhang, C., Zhao, H., Fu, J., Liu, J., Baicalin attenuates proinflammatory cytokine production in oxygen-glucose deprived challenged rat microglial cells by inhibiting TLR4 signaling pathway (2012) International Immunopharmacology, 14 (4), pp. 749-757Hougee, S., Sanders, A., Faber, J., Graus, Y.M., van den Berg, W.B., Garssen, J., Smit, H.F., Hoijer, M.A., Decreased pro-inflammatory cytokine production by LPS-stimulated PBMC upon in vitro incubation with the flavonoids apigenin, luteolin or chrysin, due to selective elimination of monocytes/macrophages (2005) Biochemical Pharmacology, 69 (2), pp. 241-248Hu, F., Hepburn, H.R., Li, Y., Chen, M., Radloff, S.E., Daya, S., Effects of ethanol and water extracts of propolis (bee glue) on acute inflammatory animal models (2005) Journal of Ethnopharmacology, 100 (3), pp. 276-283Jakus, P.B., Kalman, N., Antus, C., Radnai, B., Tucsek, Z., Gallyas, F., Sumegi, B., Veres, B., TRAF6 is functional in inhibition of TLR4-mediated NF-κB activation by resveratrol (2013) Journal of Nutritional Biochemistry, 24 (5), pp. 819-823Joseph, S.V., Edirisinghe, I., Burton-Freeman, B.M., Fruit polyphenols: A review of anti-inflammatory effects in humans (2015) Critical Reviews in Food Science and NutritionJung, Y.C., Kim, M.E., Yoon, J.H., Park, P.R., Youn, H.-Y., Lee, H.-W., Lee, J.S., Anti-inflammatory effects of galangin on lipopolysaccharide-activated macrophages via ERK and NF-κB pathway regulation (2014) Immunopharmacology and Immunotoxicology, 36 (6), pp. 426-432Koeberle, A., Werz, O., Multi-target approach for natural products in inflammation (2014) Drug Discovery Today, 19 (12), pp. 1871-1882Lamothe, B., Besse, A., Campos, A.D., Webster, W.K., Wu, H., Darnay, B.G., Site-specific Lys-63-linked tumor necrosis factor receptor-associated factor 6 auto-ubiquitination is a critical determinant of IκB kinase activation (2007) Journal of Biological Chemistry, 282 (6), pp. 4102-4112Machado, J.L., Assunção, A.K., da Silva, M.C., Dos Reis, A.S., Costa, G.C., Arruda Dde, S., Rocha, B.A., do Nascimento, F.R., Brazilian green propolis: Anti-inflammatory property by an immunomodulatory activity (2012) Evidence-based Complementary and Alternative MedicineMartinez, F.O., Sica, A., Mantovani, A., Locati, M., Macrophage activation and polarization (2007) Frontiers in Bioscience, 13, pp. 453-461Muschietti, L., Gorzalczany, S., Ferraro, G., Acevedo, C., Martino, V., Phenolic compounds with anti-inflammatory activity from Eupatorium buniifolium (2001) Planta Medica, 67 (743), pp. 743-744Natarajan, K., Singh, S., Burke, T.R., Grunberger, D., Aggarwal, B.B., Caffeic acid phenethyl ester is a potent and specific inhibitor of activation of nuclear transcription factor NF-kappa B (1996) Proceedings of the National Academy of Sciences, 93 (17), pp. 9090-9095Nicholas, C., Batra, S., Vargo, M.A., Voss, O.H., Gavrilin, M.A., Wewers, M.D., Guttridge, D.C., Doseff, A.I., Apigenin blocks lipopolysaccharide-induced lethality in vivo and proinflammatory cytokines expression by inactivating NF-κB through the suppression of p65 phosphorylation (2007) The Journal of Immunology, 179 (10), pp. 7121-7127Nishihira, J., Sato-Ueshima, M., Kitadate, K., Wakame, K., Fujii, H., Amelioration of abdominal obesity by low-molecular-weight polyphenol (Oligonol) from lychee (2009) Journal of Functional Foods, 1 (4), pp. 341-348Okamoto, Y., Hara, T., Ebato, T., Fukui, T., Masuzawa, T., Brazilian propolis ameliorates trinitrobenzene sulfonic acid-induced colitis in mice by inhibiting Th1 differentiation (2013) International Immunopharmacology, 16 (2), pp. 178-183Orsatti, C.L., Missima, F., Pagliarone, A.C., Sforcin, J.M., Th1/Th2 cytokines' expression and production by propolis-treated mice (2010) Journal of Ethnopharmacology, 129 (3), pp. 314-318Park, Y.K., Paredes-Guzman, J.F., Aguiar, C.L., Alencar, S.M., Fujiwara, F.Y., Chemical constituents in Baccharis dracunculifolia as the main botanical origin of southeastern Brazilian propolis (2004) Journal of Agricultural and Food Chemistry, 52 (5), pp. 1100-1103Paulino, N., Abreu, S.R.L., Uto, Y., Koyama, D., Nagasawa, H., Hori, H., Dirsch, V.M., Bretz, W.A., Anti-inflammatory effects of a bioavailable compound, Artepillin C, in Brazilian propolis (2008) European Journal of Pharmacology, 587 (1), pp. 296-301Petroski, M.D., Zhou, X., Dong, G., Daniel-Issakani, S., Payan, D.G., Huang, J., Substrate modification with lysine 63-linked ubiquitin chains through the UBC13-UEV1A ubiquitin-conjugating enzyme (2007) Journal of Biological Chemistry, 282 (41), pp. 29936-29945Sawaya, A., Cunha, I., Marcucci, M.C., Analytical methods applied to diverse types of Brazilian propolis (2011) Chemistry Central Journal, 5 (1), p. 27Sforcin, J.M., Bankova, V., Propolis: Is there a potential for the development of new drugs? (2011) Journal of Ethnopharmacology, 133 (2), pp. 253-260Shahidi, F., Ambigaipalan, P., Phenolics and polyphenolics in foods, beverages and spices: Antioxidant activity and health effects - A review (2015) Journal of Functional Foods, 2015. , In pressShi, H., Yang, H., Zhang, X., Yu, L., Identification and quantification of phytochemical composition and anti-inflammatory and radical scavenging properties of methanolic extracts of Chinese propolis (2012) Journal of Agricultural and Food Chemistry, 60 (50), pp. 12403-12410Shigeshiro, M., Tanabe, S., Suzuki, T., Dietary polyphenols modulate intestinal barrier defects and inflammation in a murine model of colitis (2013) Journal of Functional Foods, 5 (2), pp. 949-955Szliszka, E., Mertas, A., Czuba, Z.P., Krol, W., Inhibition of inflammatory response by artepillin C in activated RAW264.7 macrophages (2013) Evidence-based Complementary and Alternative Medicine, 2013, p. 735176Toreti, V.C., Sato, H.H., Pastore, G.M., Park, Y.K., Recent progress of propolis for its biological and chemical compositions and its botanical origin (2013) Evidence-based Complementary and Alternative Medicine, 2013, p. 697390Wang, C., Deng, L., Hong, M., Akkaraju, G.R., Inoue, J., Chen, Z.J., TAK1 is a ubiquitin-dependent kinase of MKK and IKK (2001) Nature, 412 (6844), pp. 346-351Wang, K., Ping, S., Huang, S., Hu, L., Xuan, H., Zhang, C., Hu, F., Molecular mechanisms underlying the in vitro anti-inflammatory effects of a flavonoid-rich ethanol extract from Chinese propolis (poplar type) (2013) Evidence-based Complementary and Alternative Medicine, 2013, p. 127672Wang, K., Zhang, J., Ping, S., Ma, Q.X., Chen, X., Xuan, H., Shi, J., Hu, F., Anti-inflammatory effects of ethanol extracts of Chinese propolis and buds from poplar (Populus×canadensis) (2014) Journal of Ethnopharmacology, 155 (1), pp. 300-311Wang, S., Moustaid-Moussa, N., Mo, H., Shastri, A., Su, R., Bapat, P., Kwun, I., Shen, C., Novel insights of dietary polyphenols and obesity (2014) Journal of Nutritional Biochemistry, 25 (1), pp. 1-18Xia, M.Z., Liang, Y.L., Wang, H., Chen, X., Huang, Y.Y., Zhang, Z.H., Chen, Y.H., Xu, D.X., Melatonin modulates TLR4-mediated inflammatory genes through MyD88-and TRIF-dependent signaling pathways in lipopolysaccharide-stimulated RAW264. 7 cells (2012) Journal of Pineal Research, 53 (4), pp. 325-334Xuan, H., Zhao, J., Miao, J., Li, Y., Chu, Y., Hu, F., Effect of Brazilian propolis on human umbilical vein endothelial cell apoptosis (2011) Food and Chemical Toxicology, 49 (1), pp. 78-85Yin, Q., Lin, S.-C., Lamothe, B., Lu, M., Lo, Y.-C., Hura, G., Zheng, L., Myszka, D.G., E2 interaction and dimerization in the crystal structure of TRAF6 (2009) Nature Structural & Molecular Biology, 16 (6), pp. 658-666Zhang, J., Cao, X., Ping, S., Wang, K., Shi, J., Zhang, C., Zheng, H., Hu, F., Comparisons of ethanol extracts of Chinese propolis (poplar type) and poplar gums based on the antioxidant activities and molecular mechanism (2015) Evidence-based Complementary and Alternative Medicine