Department of Paediatrics & Adolescent Medicine, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong.
J Ethnopharmacol. 2011 Nov 18;138(2):463-71. doi: 10.1016/j.jep.2011.09.031. Epub 2011 Sep 22.
Propolis has long been used as a popular folk medicine by various ethnic groups due to its wide spectrum of alleged biological and pharmaceutical properties including anti-microbial, anti-cancer and anti-inflammatory functions. All these can be linked to the modulation of immune function. Therefore, it will be relevant for us to find out whether there is any novel compound that can account for such action and the mechanism involved.
We investigated the immune modulating effect of Brazilian green propolis (PBrazil) and its constituent Artepillin C (Art-C) by using mixed leukocytes reaction.
The cytotoxic effect of Art-C on non-tumorigenic human liver cell line miHA and non-tumorigenic human kidney cell line HK-2 as well as human peripheral blood mononuclear cells (PBMCs) were measured by XTT cell proliferation assay. The effect of PBrazil and Art-C on T cell proliferation and activation were determined by using carboxyfluorescein succinimidyl ester (CFSE) and by CD25 expression, respectively. Cytokines including tumor necrosis factor-α (TNF-α), interferon-gamma (IFN-γ), interleukins such as IL-2, IL-17 were measured by intracellular cytokine staining and IL-10 was measured by ELISA. The effect of PBrazil and Art-C on regulatory T cells (Treg) induction was determined by the Foxp3 expression. The apoptotic effect of these compounds on CFSE labeled alloreactive T cells was measured by using Annexin V.
Using mixed leukocytes reaction we demonstrated for the first time that both Art-C and PBrazil significantly inhibited the alloreactive CD4 T cell proliferation, activation, and suppressed the expressions of IL-2, IFN-γ and IL-17 in these alloreactive CD4 T cells. The inhibitions of Art-C and PBrazil on CD4 T cells were not due to direct cytotoxic effect on PBMC or inducing regulatory T cells differentiation. Both Art-C and PBrazil were found to selectively induce apoptosis in proliferating T cells. The anti-proliferative effect of Art-C and PBrazil were reversible and were also applied to the activated T cells.
In conclusion, our results indicated that Art-C and PBrazil can suppress alloreactive CD4 T cell responses in vitro, suggesting that Art-C could be used as a potential immunosuppressant, either solely or as adjunct agent in treating graft versus host disease.
由于其广泛的据称具有生物和药物特性，包括抗微生物、抗癌和抗炎功能，蜂胶长期以来一直被不同民族用作流行的民间药物。所有这些都可以与免疫功能的调节联系起来。因此，我们有必要找出是否有任何新的化合物可以解释这种作用及其涉及的机制。
我们通过混合白细胞反应研究了巴西绿蜂胶（PBrazil）及其成分 Artepillin C（Art-C）的免疫调节作用。
通过 XTT 细胞增殖测定法测量 Art-C 对非肿瘤源性人肝细胞系 miHA 和非肿瘤源性人肾细胞系 HK-2 以及人外周血单核细胞（PBMC）的细胞毒性作用。通过使用羧基荧光素琥珀酰亚胺酯（CFSE）和 CD25 表达，分别测定 PBrazil 和 Art-C 对 T 细胞增殖和活化的影响。通过细胞内细胞因子染色测定肿瘤坏死因子-α（TNF-α）、干扰素-γ（IFN-γ）、白细胞介素如 IL-2、IL-17 等细胞因子，通过 ELISA 测定 IL-10。通过 Foxp3 表达测定 PBrazil 和 Art-C 对调节性 T 细胞（Treg）诱导的影响。通过使用 Annexin V 测定这些化合物对 CFSE 标记同种反应性 T 细胞的凋亡作用。
通过混合白细胞反应，我们首次证明 Art-C 和 PBrazil 均能显著抑制同种反应性 CD4 T 细胞的增殖、活化，并抑制这些同种反应性 CD4 T 细胞中 IL-2、IFN-γ 和 IL-17 的表达。Art-C 和 PBrazil 对 CD4 T 细胞的抑制作用不是由于对 PBMC 的直接细胞毒性作用或诱导调节性 T 细胞分化所致。Art-C 和 PBrazil 均被发现选择性诱导增殖 T 细胞凋亡。Art-C 和 PBrazil 的抗增殖作用是可逆的，也适用于已激活的 T 细胞。
总之，我们的结果表明，Art-C 和 PBrazil 可在体外抑制同种反应性 CD4 T 细胞反应，提示 Art-C 可单独或作为佐剂用于治疗移植物抗宿主病。
