於前期研究中，液化澱粉芽孢桿菌（Bacillus amyloliqufaciens）amy-1的胞外多醣（exopolysaccharides, EPS）於動物實驗被發現具有抗發炎效果，但機制不明。於是本論文以人類單核球細胞株THP-1為模型，分析EPS對發炎中的巨噬細胞之抗發炎效果與分子機制。方法是以12-O-tetradecanoylphorbol (TPA)誘導THP-1細胞分化為M0期，再以lipopolysaccharides (LPS)誘導細胞分化為發炎狀態的M1期，分析LPS與EPS共處理時發炎的效果與機制。結果，EPS明顯抑制LPS所誘導的iNOS, TNF-α, IL-6等發炎因子的表現。EPS也抑制LPS所誘導的IκB kinase (IKK) / nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB)路徑的活化。同時EPS也抑制mitogen-activated protein kinase (MAPK)中ERK的活化，但較高濃度的EPS卻活化JNK及p38。另外，EPS能活化抗氧化酵素heme oxygenase-1 (HO-1)的表現，並抑制ROS的生成。研究更進一步的發現，EPS抗氧化的效果至少與活化p38有關。結論是，EPS能抑制LPS所引起的發炎反應，機制與IKK/ NF-κB路徑的抑制有關。同時EPS能藉由活化p38來活化保護性的抗氧化機制，以降低發炎對細胞的傷害。

The exopolysaccharides (EPS) of Bacillus amyloliqufaciens amy-1 were found to have anti-inflammatory effects in animal tests, but the underlying mechanism is unknown. Therefore, this study used the human monocyte cell line THP-1 as a model to investigate the anti-inflammatory effect and molecular mechanism of EPS on macrophages. The cells were induced to differentiate into M0 phase by 12-O-tetradecanoylphorbol (TPA). Lipopolysaccharides (LPS) was then used to induce the differentiation of THP-1 cells into the pro-inflammatory M1 phase. This model was used to analyze the effect of EPS. Consequently, EPS significantly inhibited LPS-induced expression of iNOS, TNF-α and IL-6. Meanwhile, EPS inhibited the activation of IκB kinase (IKK) /nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) pathway by LPS. Moreover, EPS also suppressed the activation of mitogen-activated protein kinase (MAPK), ERK, but higher concentrations of EPS activated JNK and p38. In addition, EPS activated the expression of the antioxidant enzyme heme oxygenase-1 (HO-1) and reduced the generation of ROS. Furthermore, it was found that the antioxidation effect of EPS is at least correlated with the activation of p38. In conclusion, EPS inhibits LPS-induced inflammation, and the underlying mechanism is associated with the suppression of the IKK/NF-κB pathway. Meanwhile, EPS activate the protective antioxidant pathway by activating p38 to reduce cellular damages caused by inflammation.

黃聖達，分析益生菌胞外多醣的降血糖及抗發炎效果，國立屏東科技大學生物科技系碩士論文，2015。
Arulselvan, P., Fard, M.T., Tan, W.S., Gothai, S., Fakurazi, S., Norhaizan, M.E., and Kumar, S.S. (2016). Role of antioxidants and natural products in inflammation. Oxidative Medicine and Cellular Longevity 2016.
Ayroldi, E., Cannarile, L., Migliorati, G., Nocentini, G., Delfino, D.V., and Riccardi, C. (2012). Mechanisms of the anti‐inflammatory effects of glucocorticoids: genomic and nongenomic interference with MAPK signaling pathways. The FASEB Journal 26, 4805-4820.
Baharav, E., Mor, F., Halpern, M., and Weinberger, A. (2004). Lactobacillus GG bacteria ameliorate arthritis in Lewis rats. The Journal of Nutrition 134, 1964-1969.
Barendsen, N., Mueller, M., and Chen, B. (1990). Inhibition of TPA-induced monocytic differentiation in THP-1 human monocytic leukemic cells by staurosporine, a potent protein kinase C inhibitor. Leukemia Research 14, 467-474.
Cao, G., Zhan, X., Zhang, L., Zeng, X., Chen, A., and Yang, C. (2018). Modulation of broilers’ caecal microflora and metabolites in response to a potential probiotic Bacillus amyloliquefaciens. Journal of Animal Physiology and Animal Nutrition 102, e909-e917.
Carmona-Aparicio, L., Pérez-Cruz, C., Zavala-Tecuapetla, C., Granados-Rojas, L., Rivera-Espinosa, L., Montesinos-Correa, H., Hernández-Damián, J., Pedraza-Chaverri, J., Sampieri III, A., and Coballase-Urrutia, E. (2015). Overview of Nrf2 as therapeutic target in epilepsy. International Journal of Molecular Sciences 16, 18348-18367.
Chanput, W., Mes, J.J., and Wichers, H.J. (2014). THP-1 cell line: an in vitro cell model for immune modulation approach. International Immunopharmacology 23, 37-45.
Chen, Y.-C., Huang, S.-D., Tu, J.-H., Yu, J.-S., Nurlatifah, A.O., Chiu, W.-C., Su, Y.-H., Chang, H.-L., Putri, D.A., and Cheng, H.-L. (2020). Exopolysaccharides of Bacillus amyloliquefaciens modulate glycemic level in mice and promote glucose uptake of cells through the activation of Akt. International Journal of Biological Macromolecules 146, 202-211.
Chen, Y.-C., Wu, Y.-J., and Hu, C.-Y. (2019). Monosaccharide composition influence and immunomodulatory effects of probiotic exopolysaccharides. International Journal of Biological Macromolecules 133, 575-582.
Chi, X., Yao, W., Xia, H., Jin, Y., Li, X., Cai, J., and Hei, Z. (2015). Elevation of HO-1 expression mitigates intestinal ischemia-reperfusion injury and restores tight junction function in a rat liver transplantation model. Oxidative Medicine and Cellular Longevity 2015.
Choi, Y.H. (2016). The cytoprotective effect of isorhamnetin against oxidative stress is mediated by the upregulation of the Nrf2-dependent HO-1 expression in C2C12 myoblasts through scavenging reactive oxygen species and ERK inactivation. General Physiology and Biophysics 35, 145-154.
Delgado, S., Sánchez, B., Margolles, A., Ruas-Madiedo, P., and Ruiz, L. (2020). Molecules produced by probiotics and intestinal microorganisms with immunomodulatory activity. Nutrients 12, 391.
Dilna, S.V., Surya, H., Aswathy, R.G., Varsha, K.K., Sakthikumar, D.N., Pandey, A., and Nampoothiri, K.M. (2015). Characterization of an exopolysaccharide with potential health-benefit properties from a probiotic Lactobacillus plantarum RJF4. LWT-Food Science and Technology 64, 1179-1186.
Ghosh, S., May, M.J., and Kopp, E.B. (1998). NF-κB and Rel proteins: evolutionarily conserved mediators of immune responses. Annual Review of Immunology 16, 225-260.
Guha, M., and Mackman, N. (2001). LPS induction of gene expression in human monocytes. Cellular Signalling 13, 85-94.
Guo, C., Yang, L., Wan, C.-X., Xia, Y.-Z., Zhang, C., Chen, M.-H., Wang, Z.-D., Li, Z.-R., Li, X.-M., and Geng, Y.-D. (2016). Anti-neuroinflammatory effect of Sophoraflavanone G from Sophora alopecuroides in LPS-activated BV2 microglia by MAPK, JAK/STAT and Nrf2/HO-1 signaling pathways. Phytomedicine 23, 1629-1637.
Hemarajata, P., and Versalovic, J. (2013). Effects of probiotics on gut microbiota: mechanisms of intestinal immunomodulation and neuromodulation. Therapeutic Advances in Gastroenterology 6, 39-51.
Hill, C., Guarner, F., Reid, G., Gibson, G.R., Merenstein, D.J., Pot, B., Morelli, L., Canani, R.B., Flint, H.J., and Salminen, S. (2014). Expert consensus document: The International Scientific Association for Probiotics and Prebiotics consensus statement on the scope and appropriate use of the term probiotic. Nature Reviews Gastroenterology & Hepatology 11, 506.
Hong, H.A., Duc, L.H., and Cutting, S.M. (2005). The use of bacterial spore formers as probiotics. FEMS Microbiology Reviews 29, 813-835.
Islam, V.H., Babu, N.P., Pandikumar, P., and Ignacimuthu, S. (2011). Isolation and characterization of putative probiotic bacterial strain, Bacillus amyloliquefaciens, from North East Himalayan soil based on in vitro and in vivo functional properties. Probiotics and Antimicrobial Proteins 3, 175-185.
Jiang, G., Hu, Y., Liu, L., Cai, J., Peng, C., and Li, Q. (2014). Gastrodin protects against MPP+-induced oxidative stress by up regulates heme oxygenase-1 expression through p38 MAPK/Nrf2 pathway in human dopaminergic cells. Neurochemistry International 75, 79-88.
Jiang, K., Guo, S., Yang, C., Yang, J., Chen, Y., Shaukat, A., Zhao, G., Wu, H., and Deng, G. (2018). Barbaloin protects against lipopolysaccharide (LPS)-induced acute lung injury by inhibiting the ROS-mediated PI3K/AKT/NF-κB pathway. International Immunopharmacology 64, 140-150.
Khochamit, N., Siripornadulsil, S., Sukon, P., and Siripornadulsil, W. (2015). Antibacterial activity and genotypic–phenotypic characteristics of bacteriocin-producing Bacillus subtilis KKU213: Potential as a probiotic strain. Microbiological Research 170, 36-50.
Kuang, J.-h., Huang, Y.-y., Hu, J.-s., Yu, J.-j., Zhou, Q.-y., and Liu, D.-m. (2020). Exopolysaccharides from Bacillus amyloliquefaciens DMBA-K4 ameliorate dextran sodium sulfate-induced colitis via gut microbiota modulation. Journal of Functional Foods 75, 104212.
Li, Y., Zhang, H., Chen, Y., Yang, M., Zhang, L., Lu, Z., Zhou, Y., and Wang, T. (2015). Bacillus amyloliquefaciens supplementation alleviates immunological stress in lipopolysaccharide-challenged broilers at early age. Poultry Science 94, 1504-1511.
Liu, C.F., Tseng, K.C., Chiang, S.S., Lee, B.H., Hsu, W.H., and Pan, T.M. (2011). Immunomodulatory and antioxidant potential of Lactobacillus exopolysaccharides. Journal of the Science of Food and Agriculture 91, 2284-2291.
Miyamoto, S., and Verma, I.M. (1995). Rel/NF-κB/IκB story. In Advances in cancer research (Elsevier), pp. 255-292.
Moi, P., Chan, K., Asunis, I., Cao, A., and Kan, Y.W. (1994). Isolation of NF-E2-related factor 2 (Nrf2), a NF-E2-like basic leucine zipper transcriptional activator that binds to the tandem NF-E2/AP1 repeat of the beta-globin locus control region. Proceedings of the National Academy of Sciences 91, 9926-9930.
Moynagh, P.N. (2005). The NF-κB pathway. Journal of cell science 118, 4589-4592.
Richter, E., Ventz, K., Harms, M., Mostertz, J., and Hochgräfe, F. (2016). Induction of macrophage function in human THP-1 cells is associated with rewiring of MAPK signaling and activation of MAP3K7 (TAK1) protein kinase. Frontiers in Cell and Developmental biology 4, 21.
Shukla, A., Mehta, K., Parmar, J., Pandya, J., and Saraf, M. (2019). Depicting the exemplary knowledge of microbial exopolysaccharides in a nutshell. European Polymer Journal 119, 298-310.
Stefanson, A.L., and Bakovic, M. (2014). Dietary regulation of Keap1/Nrf2/ARE pathway: focus on plant-derived compounds and trace minerals. Nutrients 6, 3777-3801.
Sun, P., Wang, J., and Zhang, H. (2010). Effects of Bacillus subtilis natto on performance and immune function of preweaning calves. Journal of Dairy Science 93, 5851-5855.
Sun, Z., Huang, Z., and Zhang, D.D. (2009). Phosphorylation of Nrf2 at multiple sites by MAP kinases has a limited contribution in modulating the Nrf2-dependent Antioxidant Response. PloS one 4, e6588.
Surh, Y.-J., Chun, K.-S., Cha, H.-H., Han, S.S., Keum, Y.-S., Park, K.-K., and Lee, S.S. (2001). Molecular mechanisms underlying chemopreventive activities of anti-inflammatory phytochemicals: down-regulation of COX-2 and iNOS through suppression of NF-κB activation. Mutation Research/Fundamental and Molecular Mechanisms of Mutagenesis 480, 243-268.
Valentine, R., Dawson, C.W., Hu, C., Shah, K.M., Owen, T.J., Date, K.L., Maia, S.P., Shao, J., Arrand, J.R., and Young, L.S. (2010). Epstein-Barr virus-encoded EBNA1 inhibits the canonical NF-κB pathway in carcinoma cells by inhibiting IKK phosphorylation. Molecular Cancer 9, 1-17.
Vincenzi, A., Goettert, M.I., and de Souza, C.F.V. (2020). An evaluation of the effects of probiotics on tumoral necrosis factor (TNF-α) signaling and gene expression. Cytokine & Growth Factor Reviews.
Wang, N., Liang, H., and Zen, K. (2014). Molecular mechanisms that influence the macrophage M1–M2 polarization balance. Frontiers in immunology 5, 614.
Wardyn, J.D., Ponsford, A.H., and Sanderson, C.M. (2015). Dissecting molecular cross-talk between Nrf2 and NF-κB response pathways. Biochemical society transactions 43, 621-626.
Yang, H., Deng, J., Yuan, Y., Fan, D., Zhang, Y., Zhang, R., and Han, B. (2015). Two novel exopolysaccharides from Bacillus amyloliquefaciens C-1: antioxidation and effect on oxidative stress. Current microbiology 70, 298-306.
Yunna, C., Mengru, H., Lei, W., and Weidong, C. (2020). Macrophage M1/M2 polarization. European Journal of Pharmacology 877, 173090.