黄芪多糖抑制结肠癌肿瘤微环境IDO1的表达增加瘤内CD8^(+)T细胞浸润

Astragalus polysaccharide inhibits IDO1 expression in colon tumor microenvironment to increase intratumoral CD8^(+)T cell infiltration

基于结肠癌肿瘤微环境探讨黄芪多糖(Astragalus polysaccharide,APS)及APS联合5-氟尿嘧啶(5-fluorouracil,5-FU)对结肠癌肿瘤微环境中IDO1的调控作用。将60只Balb/c小鼠随机分为空白组、模型组、APS组、APS联合5-FU组、APS联合低剂量5-FU组和5-FU组,除空白组外,利用CT-26小鼠结肠癌细胞建立皮下移植瘤模型。造模成功后,各组给予相应药物连续治疗7 d;治疗期间每天观察检测小鼠一般情况、体质量和肿瘤体积,治疗结束后处死小鼠,收集所需标本并计算小鼠抑瘤率和脾脏指数;Western blot实验检测各组荷瘤小鼠肿瘤组织中IDO1蛋白表达变化;荧光定量PCR实验检测各组荷瘤小鼠肿瘤组织中IDO1 mRNA表达变化;高效液相色谱实验检测各组荷瘤小鼠肿瘤组织内Trp和Kyn含量变化;HE染色实验观察各组荷瘤小鼠肿瘤组织学变化;免疫组化实验检测各组荷瘤小鼠肿瘤组织内CD4和CD8的表达变化。与模型组相比,各治疗组小鼠肿瘤体积显著降低;APS联合5-FU组和5-FU组小鼠从第4~7天治疗结束体质量显著降低;APS联合5-FU组和5-FU组脾脏指数显著下降;APS组、APS联合5-FU组和APS联合低剂量5-FU组中IDO1蛋白及mRNA表达量均显著下降;各治疗组Trp含量显著升高,各治疗组之间Kyn含量呈现降低趋势,但差异均无统计学意义;APS联合5-FU组CD4^(+)T淋巴细胞的浸润显著降低,CD8^(+)T淋巴细胞的浸润显著升高。APS抑制结肠癌肿瘤微环境IDO1的表达增加CD8^(+)T细胞浸润,且联合5-FU治疗具有协同作用。

This study aims to investigate the regulatory effects of Astragalus polysaccharide(APS)and APS combined with 5-fluorouracil(5-FU)on indoleamine-2,3-dioxygenase(IDO1)in the colon tumor microenvironment.Sixty Balb/c mice were randomized into a blank group,a model group,an APS group,an APS+5-FU group,an APS+low-dose 5-FU group,and a 5-FU group.A tumor model was established by subcutaneous transplantation with CT-26 mouse colon cancer cells in other groups except the blank group.After successful modeling,each group was treated with corresponding drugs for 7 days.The general condition,body weight,and tumor volume of the mice were observed and measured daily during the treatment period.The mice were sacrificed at the end of treatment,and the tumor suppression rate and spleen index of the mice were calculated.Western blot and fluorescence quantitative PCR were employed to determine the protein and mRNA levels,respectively,of IDO1 in the tumor tissue of mice.High performance liquid chromatography was employed to measure the levels of tryptophan(Trp)and kynurenine(Kyn)in the tumor tissue of mice.Hematoxylin-eosin(HE)staining was performed to observe the histological changes of the tumor tissue,and immunohistochemistry to detect the changes of CD4 and CD8 expression in the tumor tissue.Compared with that in the model group,the tumor volume of mice in each treatment group significantly reduced.The body weights of mice in APS+5-FU group and 5-FU group significantly reduced from day 4 to day 7 of treatment.In addition,the APS+5-FU group and 5-FU group showed significantly decreased spleen index.The protein and mRNA levels of IDO1 were significantly down-regulated in the APS,APS+5-FU,and APS+low-dose 5-FU groups.The drug interventions significantly increased the Trp content and decreased the Kyn content.The APS+5-FU group showed significantly reduced infiltration of CD4^(+)T lymphocytes and increased infiltration of CD8^(+)T lymphocytes.APS inhibited the expression of IDO1 in the colon tumor microenvironment to increase CD8^(+)T lymphocyte infiltration,and the combination of APS with 5-FU demonstrated better effect.