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目的比较研究不同类型抗瘤中药制剂对结直肠癌所致T细胞免疫抑制的影响,寻找能有效逆转结直肠癌T细胞免疫抑制的抗瘤中药制剂组方。方法分别制备三氧化二砷(As2O3)、川芎嗪(LHC)、黄芪(AMB)、氧化苦参碱(MOX)、猪苓多糖(PUPS)、青蒿琥酯(ART)等6种中药制剂作用后的小鼠结直肠癌Colon26细胞再培养上清;MTT法检测中药制剂作用后对Colon26所致T细胞转化抑制的影响,直接免疫荧光FCM法检测对IL-2Rα及CD3ε+ζ+表达抑制的影响;定量ELISA法测定上清中TGF-β1、VEGF、IL-4、IL-6、IL-10和PGE2含量;多元相关分析中药制剂逆转结直肠癌T细胞免疫抑制的相应靶分子。结果 AMB、PUPS及ART对T细胞转化抑制的下调程度较强(下调约50%),维持时间较长;ART对IL-2Rα表达抑制的下调作用最强(下调约50%),维持时间较长;AMB对CD3ε+ζ+表达抑制的下调作用最强,不但可完全消除表达抑制,还可进一步促进表达,维持时间亦较持久。对Colon26肿瘤细胞所致转化抑制及CD3ε+ζ+表达抑制的逆转,LHC的作用靶分子为TGF-β1和IL-10,其他5种中药制剂的靶分子均为TGF-β1。对IL-2Rα表达抑制的逆转,除AMB以外其他5种中药制剂的靶分子均为TGF-β1。结论通过阻碍肿瘤细胞分泌TGF-β1、IL-10等免疫抑制分子,以逆转肿瘤细胞产生的T细胞免疫抑制,应是中药制剂的新型抗瘤机制之一。
Objective To compare the effects of different types of anti-tumor traditional Chinese medicine preparations on T cell immunosuppression induced by colorectal cancer and to find out the anti-tumor traditional Chinese medicine preparations that can effectively reverse the T cell immunosuppression of colorectal cancer. Methods Six kinds of traditional Chinese medicine preparations such as As2O3, LHC, AMB, MOX, PUPS and ART were prepared respectively. Colon26 cells of rat colorectal cancer re-culture supernatant was detected by MTT method Colon26-induced T cell transformation inhibition, direct immunofluorescence FCM assay IL-2Rα and CD3ε + ζ expression inhibition; quantitative The contents of TGF-β1, VEGF, IL-4, IL-6, IL-10 and PGE2 in the supernatant were measured by ELISA, and the correlation between the Chinese herbal preparations and the corresponding target molecules for reversing T cell immunosuppression in colorectal cancer was analyzed by multivariate correlation analysis. Results AMB, PUPS and ART had a strong down-regulation of T cell transformation inhibition (about 50% downregulation) for a long time. ART had the strongest down-regulation of IL-2Rα expression (down to about 50%), maintenance time AMB had the strongest down-regulation of CD3ε + ζ + expression, which could not only completely eliminate the expression inhibition, but also further promote the expression and prolong the maintenance time. The reversal of the inhibition of the transformation of Colon26 tumor cells and the inhibition of the expression of CD3ε + ζ +, the target molecules of LHC for TGF-β1 and IL-10, the other five kinds of Chinese medicine preparation target molecules are TGF-β1. The reversal of inhibition of IL-2Rα expression, except for AMB, was targeted by TGF-β1 among the other five TCM preparations. Conclusion It is one of the new anti-tumor mechanisms of traditional Chinese medicine by hindering tumor cells to secrete immunosuppressive molecules such as TGF-β1 and IL-10 to reverse T cell immunosuppression induced by tumor cells.