基于TLR4/MAPK/NF-κB信号通路探讨萝卜硫素抗甲状腺乳头状癌的作用

Exploring the anti-papillary thyroid carcinoma effect of sulforaphane based on TLR4/MAPK/NF-κB signaling pathway

目的探讨萝卜硫素对裸鼠人乳头瘤状甲状腺癌移植瘤的作用及机制。方法采用皮下注射TPC-1细胞的方法构建裸鼠人乳头瘤状甲状腺癌移植瘤模型,随后将模型小鼠随机分为模型组、萝卜硫素低剂量组和萝卜硫素高剂量组,每组10只。萝卜硫素低、高剂量组分别灌胃给予5 mg/kg和20 mg/kg萝卜硫素,模型组灌胃给予等量生理盐水,1次/d,连续25 d。每5天测量肿瘤生长情况;取材后称量肿瘤质量,计算肿瘤抑制率;TUNNEL法观察肿瘤组织细胞凋亡情况;ELISA法检测血清炎症因子和氧化应激指标水平;流式细胞仪检测血液CD4^(+)/CD8^(+)淋巴细胞亚群比值;Western blot法检测肿瘤组织TLR4、P38和p-NF-κB蛋白的表达水平。结果与模型组比较,萝卜硫素给药组的肿瘤生长速度均明显被抑制(P<0.01),肿瘤质量明显减轻(P<0.01),肿瘤抑制率明显升高(P<0.01),肿瘤细胞凋亡阳性率明显升高(P<0.01),血清TNF-α、IL-1β和IL-6水平均明显降低(P<0.01),血清GSH-Px和SOD水平均明显升高(P<0.01),而MDA水平明显降低(P<0.01),血液CD4^(+)/CD8^(+)淋巴细胞亚群比值明显升高(P<0.01),肿瘤组织TLR4、P38、p-NF-κB蛋白相对表达量明显降低(P<0.01),且呈剂量依赖性。结论萝卜硫素具有抗甲状腺乳头状癌的作用,其机制主要与调控TLR4/MAPK/NF-κB信号通路、抗炎和氧化应激以及免疫调节有关。

Objective To investigate the effect and mechanism of sulforaphane on papillary thyroid carcinoma xenografts in nude mice.Methods A nude mouse model of human papillary thyroid carcinoma xenograft was constructed by subcutaneous injection of TPC-1 cells.The model mice were randomly divided into the model group,the low-dose group of sulforaphane,and the high-dose group of sulforaphane,with 10 mice in each group.The low-dose group and high-dose group of sulforaphane were orally administered with 5 mg/kg and 20 mg/kg sulforaphane,respectively.The model group was orally administered with an equal amount of saline once a day for 25 consecutive days.Tumor growth was measured every 5 d.The mass of the tumor was weighed and the tumor inhibition rate was calculated.TUNNEL method was used to observe the apoptosis of tumor tissue cells.ELISA is used to detect the levels of serum inflammatory factors and oxidative stress indicators.Flow cytometry was used to detect the ratio of CD4^(+)/CD8^(+)lymphocyte subsets in the blood.Western blot was used to detect the expression of TLR4,P38,and p-NF-κB proteins in tumor tissues.Results Compared with the model group,the tumor growth rate was significantly inhibited,the tumor mass was reduced,the tumor inhibition rate was increased,and the positive rate of tumor cell apoptosis was increased(P<0.01).The levels of serum TNF-α,IL-1βand IL-6 were lowered,the levels of serum GSH-Px and SOD were increased,and serum MDA levels were decreased(P<0.01).The ratio of CD4^(+)/CD8^(+)lymphocyte subsets in blood was enhanced,and the relative expression of TLR4,P38 and p-NFκB protein in tumor tissue was decreasedin a dose-dependent manner.Conclusion Sulforaphane has anti-papillary thyroid carcinoma effects,and its mechanism is mainly related to the regulation of TLR4/MAPK/NFκB pathway,anti-inflammatory and oxidative stress,and immune regulation.