銀耳免疫調節蛋白 TFP 活化小鼠骨髓衍生性樹突細胞及作為腫瘤疫苗佐劑之探討

銀耳 (Tremella fuciformis) 為亞洲傳統藥用菇類，具有多種藥理療效。文獻指出純化自銀耳之免疫調節蛋白 TFP 能透過與細胞表面 TLR4 受器結合，活化小鼠腹腔巨噬細胞產生細胞激素、提升吞噬活性及提升細胞表面分子表現量。本研究之目的為探討 TFP 是否能誘導 Th1 反應及增強免疫治療時疫苗效果之能力。首先發現 TFP 可誘導小鼠脾細胞分裂及產生 IFN-γ ，但無法直接刺激 CD90+ T 細胞增生和 IFN-γ 產生，顯示 TFP 無法直接活化 T 細胞，故推測 TFP 需透過活化抗原呈獻細胞如樹突細胞 (Dendritic cells, DCs) 使 T 細胞分化。故接著探討 TFP 對 DCs 之影響，由體外實驗得知，TFP 可增加小鼠 DCs 之 CD40、CD80 及 MHC class II 表現及發炎前驅物質 TNF-α, IL-1β, IL-6 及IL-10 分泌量，但此效果會因 TLR4 缺陷而降低，證實 TFP 具有活化樹突細胞之作用且可被 TLR4辨識。再者，預先以 TFP 及 OVA 活化過之 DCs，在 DCs 調節 T 細胞免疫反應試驗中，可較對照組增加更多 DO11.10 CD4+ T 細胞增生及 IFN-γ 分泌量，顯示 TFP 可提升 DCs 抗原呈獻能力，並將免疫反應導向 Th1。另一方面，體內試驗發現以 TFP 及 OVA 刺激之 DCs 免疫 E.G7 胸腺癌小鼠，可顯著抑制腫瘤生長並延長存活率。此 DCs 可降低罹癌小鼠血清 IL-6 含量，提升其體內 CD8+CD44high T 細胞量及抗原特異性 IFN-γ 分泌量。以上結果證實銀耳免疫調節蛋白 TFP 可透過活化 DCs 之成熟促使 Th1 型細胞分化，TFP 並可作為佐劑增強 DCs 疫苗之功效。

關鍵字

銀耳；免疫調節蛋白；小鼠骨髓衍生性樹突細胞；腫瘤疫苗；佐劑

並列摘要

Tremella fuciformis is one of the fungus that used in traditional medicine in Asian and possesses a variety of pharmacological effects. Tremella fuciformis protein (TFP), an immunomudulatory protein purified from T. fuciformis, has been revealed that can induce cell cytokine production, phagocytic activity and surface marker expression in macrophages via the Toll-like receptor 4 (TLR4). In this study, the immune-activating ability of TFP on lynphocytes and promoting the efficacy of immunotherapeutic vaccines were investigated. We first found that TFP stimulated cell proliferation and IFN-γ production in splenocytes but not in CD90+ T cells. This indicating that antigen-presenting cells (APCs) such as dendritic cells (DCs) could involve in T cell activation. We then examined the TFP-induced activation on murine bone marrow derived DCs and demonstrated that TFP up-regulate the expression of CD40, CD80 and MHC class II and the production of proinflammatory cytokines such us IL-6, TNF-α, IL-1β and IL-10. On the other hand, cytokine production induced by TFP was reduced in TLR4-/- DCs, indicating TFP is recognized mainly by TLR4 to stimulate DCs maturation. Furthermore, TFP-treated DCs activated DO11.10 CD4+ T cells to secrete IFN-γ and further induced T cell proliferation in mixed lymphocyte reaction (MLR), revealing TFP-treated DCs provoke Th1 immune response in vitro. Immunization of mice with TFP-stimulated and ovalbumin (OVA)-pulsed DCs induced an effective regression of tumor growth and long-term survival in a murine E.G7 thymoma model. Injecting TFP-treated OVA-pulsed DCs in tumor-bearing mice reduced serum Il-6 levels. In addition, the number of CD8+CD44hi T cells in splenocytes had increased and the antigen-specific response in vitro had enhanced. These findings provide evidences that TFP induces Th1 response through DC maturation, and highlight TFP as an effective adjuvant for enhancing the efficacy of DC-based antitumor immunotherapy.

並列關鍵字

Tremella fuciformis ； immunomodulatory protein ； bone marrow-derived dendritic cells ； therapeutic cancer vaccine ； adjuvant

參考文獻

Akira, S., Takeda, K., and Kaisho, T. (2001). Toll-like receptors: critical proteins linking innate and acquired immunity. Nature immunology, 2 (8), 675-680.

Alderton, G.K., and Bordon, Y. (2012). Tumour immunotherapy—leukocytes take up the fight. Nature Reviews Immunology, 12 (4), 237-237.

Banchereau, J., and Steinman, R.M. (1998). Dendritic cells and the control of immunity. Nature, 392 (6673), 245-252.

Belz, G.T., and Nutt, S.L. (2012). Transcriptional programming of the dendritic cell network. Nature Reviews Immunology, 12 (2), 101-113.

Berger, F.G. (2004). The interleukin-6 gene: a susceptibility factor that may contribute to racial and ethnic disparities in breast cancer mortality. Breast cancer research and treatment, 88 (3), 281-285.

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銀耳免疫調節蛋白 TFP 活化小鼠骨髓衍生性樹突細胞及作為腫瘤疫苗佐劑之探討

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