樟芝免疫調節蛋白ACA1對小鼠腹腔巨噬細胞的活化作用

Mouse Peritoneal Macrophage Activation Induced by ACA1, an Immunomodulatory Protein Isolated from Antrodia cinnamomea

黃婉婷(Wan-Ting Huang)

指導教授 ： 許輔

國立臺灣大學/生物資源暨農學院/園藝研究所/碩士(2007年)

https://doi.org/10.6342/NTU.2007.02426

摘要

樟芝 (Antrodia cinnamomea) 為台灣真菌特有種，為具有保健療效潛力之藥用真菌。本研究室之前已自樟芝菌絲體發酵液純化出免疫調節蛋白 ACA1 並選殖出其核酸序列，且於大腸桿菌中進行異體表現，以大量生產重組 ACA1 蛋白。本研究即以重組 ACA1 之融合蛋白 (frACA1) 為試驗樣品，希望進一步瞭解樟芝免疫調節蛋白對於活體巨噬細胞的活化作用。利用 15 μg/mL frACA1 處理小鼠腹腔巨噬細胞 (peritoneal macrophage，pMΦ)，可增進 pMΦ

關鍵字

樟芝 ； 免疫調節蛋白 ； 巨噬細胞 ； ACA1

並列摘要

Antrodia cinnamomea is not only an endemic fungus but also a potential medicinal mushroom in Taiwan. Our laboratory had previously isolated an protein, named ACA1 and demonstrated its immunomodulatory ability to activate murine macrophages, from the mycelia of A. cinnamomea. The gene encoding ACA1 had also been cloned and expressed in E.coli for the production of recombinant ACA1 protein. In this study, the macrophage-stimulating activity of recombinant ACA1 fusion protein (frACA1) was thoroughly studied. The results showed that treatment with 15 μg/mL frACA1 increased the phagocytic ability and CD86/B7-2 expression of mouse peritoneal macrophages (pMΦ). In addition, real-time PCR results demonstrated that frACA1 could up-regulate the mRNA expression of cytokines, including IL-1β, TNF-α, IL-6, and iNOS, in a time dependent manner, which suggested that frACA1 could activate pMΦ without hyperinflammation. Moreover, frACA1 promoted the mRNA expression of IL-12p35 and M1 type chemokines genes, including CCL3, CCL4, CCL5 CXCL10, in murine pMΦ. These results revealed that frACA1 was capable to promote Th1 immune response by inducing M1 polarization of pMΦ. Furthermore, we investigated whether frACA1-induced pMΦ activation was mediated with TLR2- or TLR4-signaling pathway or not. The results showed that frACA1 could induce TNF-α secretion by the pMΦ of both TLR2 and TLR4 deficiency mouse. It was also observed that frACA1 showed insignificant effect on increasing the mRNA expression of TLR2, TLR4, MyD88, and NF-κB gene in BALB/c mouse pMΦ (p>0.05), suggesting that TLR2- and TLR4-signaling pathway were not involved in frACA1-stimulating pMΦ activation and the possibility of LPS-contamination in frACA1 was also excluded. However, flowcytometry indicated that there was no interaction between frACA1 and cell surface of pMΦ. Therefore, further investigation could be carried out to discover the activation pathway of frACA1 on murine pMΦ.

並列關鍵字

Antrodia cinnamomea ； Immunomodulatory Protein ； macrophage ； ACA1

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