紅麴代謝產物 monascin 與 ankaflavin 對於過敏反應中肥大細胞與 T 細胞之影響

Abstract

第一型過敏反應 (type I allergy) 包含氣喘、食物過敏、蕁麻疹等疾病，其盛行率隨著生活方式都市化與現代化而逐漸增加，當人體接觸到過敏原，經由抗原呈現細胞 (antigen presenting cells, APCs) 之辨認、吞噬並呈現給 T 細胞，使得 T 細胞活化及分化，在過敏患者體內 T 細胞趨向分化為 T helper 2 (Th2) 細胞，釋放之 Th2 細胞激素 (cytokines) 包含 interleukin-4 (IL-4)、IL-5、IL-13 等，造成 B 細胞進行型態轉換 (class switching) 而分泌具過敏原專一性之 IgE 抗體，而 IgE 抗體會和肥大細胞表面之 IgE receptor 結合，當過敏病患再次接觸到同一過敏原時，肥大細胞表面的 IgE 會和過敏原形成交互結合 (crosslinking)，導致肥大細胞活化釋放多種發炎介質，使病患發生氣管窄縮、腹瀉、皮膚紅腫等症狀。而先前研究發現，雞卵蛋白 (ovalbuminn) 誘導 BALB/c 鼠呼吸道發炎以模擬氣喘，給予紅麴二次代謝產物monascin 與 ankaflavin 能夠減緩其發炎情形，顯示其具有改善氣喘之潛力，深入研究 monascin 與 ankaflavin 對於參與其中之細胞的影響，發現 monascin 可影響抗原呈現細胞中的樹突細胞之成熟，而可能影響其呈現抗原給 T 細胞，monascin 也可抑制 T 細胞分泌 Th2 細胞激素，但 ankaflavin 對於 T 細胞之影響以及 monascin 和 ankaflavin 對下游之肥大細胞之影響還未被研究，因此期望透過研究這兩種細胞以了解 monascin 及 ankaflavin 對於改善氣喘之可能途徑。實驗結果發現，ankaflavin 能夠抑制經 phorbol 12-myristate 13-acetate (PMA) 及 ionomycin 誘導之 EL4 T 細胞分泌 Th1 與 Th2 細胞激素而具有減緩發炎之潛力，試驗發現 40 μM monascin 與 ankaflavin 對肥大細胞株 RBL-2H3 細胞可以顯著地抑制 PMA 及 ionomycin 誘導之肥大細胞分泌發炎介質組織胺 (histamine) 和腫瘤壞死因子 (tumor necrosis factorα, TNF-α)，因此進一步往上游探討可能的改善機制，結果顯示 monascin 與 ankaflavin 對於細胞內鈣離子與活性氧分子 (reactive oxygen species, ROS) 之產生無影響，但monascin 及 ankaflavin 可藉由抑制 protein kinase C (PKC) 與 mitogen-activated protein kinase (MAPK) family 中之 ERK、p38 及 JNK 蛋白之磷酸化，進而影響肥大細胞釋放 TNF-α 與組織胺等發炎介質。由以上結果得知，紅麴代謝產物 monascin 和 ankaflavin 具減緩 T 細胞分泌 Th1、Th2 細胞激素與調節肥大細胞釋放發炎介質之效果，藉由改善細胞分泌發炎介質而改善病人過敏與發炎之情形，對於第一型過敏反應具改善功效潛力。

Type I allergy includes asthma, food allergy, urticaria and so on. Its prevalence increases with industrialized and modernized lifestyles. The mechanism of type I allergy began with antigen presenting cells (APCs) contacting with allergens, and then APCs recognize, engulfe, and present antigen to naive T cells, which make T cells activated and differentiated. In allergic patients, T cells tended to differentiate into T helper 2 (Th2) cells. Th2 released cytokines included interleukin-4 (IL-4), IL-5, and IL-13 that turn B cells into IgE-releasing plasma cells. The produced IgE bound to IgE receptors on mast cells. As a result, when next time allergy patients contact the same allergen, allergen would cross-link with IgE which is previously bound on mast cells, followed with the activation and inflammatory mediator-secretion of mast cells. These turned out making patients bronchial constriction, diarrhea, and wheal-and-flare reaction. In previous study, treating with Monascus secondary metabolites monascin and ankaflavin to ovalbumin-induced BALB/c mice could ameliorate allergic airway inflammation. It is showed that monascin and ankaflavin may have the anti-asthma potential. In order to know the improving mechanism of monascin and ankaflavin related to which immune cells, our lab investigated the influence of monascin on the APCs dendritic cells and EL4 T cells. We discovered that monascin inhibited the maturation of dendritic cells and reduced the Th2 cytokines release of EL4 T cell. However, the effects of ankaflavin on EL4 T cell and both monascin and ankaflavin on mast cells haven’t been investigated. Therefore, we hope to understand the impact of monascin and ankaflavin on these two cells. According to the results, we found that ankaflavin could reduce the production of PMA/ionomycin-induced Th1 and Th2 cytokine, and may possess anti-inflammatory potential. In RBL-2H3 mast cells, both monasin and ankaflavin (40 μM) decrease PMA/ionomycin-induced mast cell tumor necrosis factor α (TNF-α) release and degranulation. As we further examined the possible mechanism, the results showed that monascin and ankaflavin had no influence over intracellular calcium and reactive oxygen species (ROS) level. Nonetheless, monascin and ankaflvin could improve mast cell activation by controlling the activation and phosphorylation of protein kinase C (PKC) and mitogen-activated protein kinase (MAPK) family ERK, JNK, and p38. To sum up, monascin and ankaflavin had regulating effects on the generation of T cell cytokines and mast cell inflammatory mediators. Both may own the potential to improve type I allergy.