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目的体外研究猪苓多糖(polyporus umbellatus polysacharide,PUP)对结直肠癌Colon26细胞肿瘤免疫抑制的影响。方法获取经PUP作用后的Colon26再培养上清,未经作用的同步培养上清作对照,检测PUP对Colon26肿瘤细胞所致NK杀伤和诱导转化(MTT法测定),以及IL-2Rα、CD3ε+ζ+和CD3ε-ζ+表达(流式细胞计数分析)5项免疫功能抑制的影响,定量ELISA法测定上清中TGF-β1、VEGF、IL-4、IL-6和IL-10五种免疫抑制分子含量,多元相关分析PUP下调Colon26免疫抑制与分泌免疫抑制分子的关系。结果对照上清中5种免疫抑制分子均可测到,TGF-β1含量最高,对5项免疫功能均有显著抑制。PUP作用后的第一次再培养上清,TGF-β1含量及对5项免疫功能的抑制均明显降低;与其相比,第二次再培养上清的5种免疫抑制分子含量及对NK杀伤、IL-2Rα和CD3ε-ζ+表达抑制均明显提高。TGF-β1与抑制诱导转化、NK杀伤及CD3ε+ζ+表达正相关。结论通过下调肿瘤细胞分泌免疫抑制分子而削弱肿瘤免疫抑制,可能是PUP抗瘤效应机制之一。
Objective To study the effect of polyporus umbellatus polysacharide (PUP) on the immunosuppression of colorectal cancer Colon26 tumor in vitro. Methods Recombinant Colon 26 resuspended supernatants were treated with PUP. The non-invasive synchronized culture supernatants were used as control to detect the NK killing and induced transformation of Colony tumor cells induced by PUP (MTT assay), and IL-2Rα, CD3ε+ The effects of ζ+ and CD3ε-ζ+ expression (flow cytometric analysis) on the inhibition of five immune functions were measured. Quantitative ELISA was used to determine five immune responses to TGF-β1, VEGF, IL-4, IL-6 and IL-10 in the supernatant. Inhibiting molecular content, multivariate correlation analysis of PUP down-regulates the relationship between Colon26 immunosuppression and the secretion of immunosuppressive molecules. RESULTS: The five immunosuppressive molecules in the control supernatant were all detectable, and the TGF-β1 content was highest, and all of the five immune functions were significantly inhibited. In the first re-cultured supernatant after PUP treatment, the content of TGF-β1 and the inhibition of five immune functions were significantly reduced; compared with the results of the 5th immunosuppressive molecule content and the NK killing of the second re-cultured supernatant. The inhibition of IL-2Rα and CD3ε-ζ+ expression was significantly increased. TGF-β1 is positively related to inhibition-induced transformation, NK killing and CD3ε+ζ+ expression. Conclusion The weakening of tumor immunosuppression by down-regulating the immunosuppressive molecules secreted by tumor cells may be one of the mechanisms of PUP anti-tumor effect.