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目的:利用小鼠脾淋巴细胞和结肠癌CT26.WT细胞混合培养体系,研究大黄素对肿瘤细胞核转录因子-κB(NF-κB)途径的影响,探讨其抗肿瘤的可能机制。方法:将密度为1×105/mL的CT26细胞和脾淋巴细胞(SLs)分别按1∶1,1∶2,1∶4比例混合培养24 h后加入大黄素再孵育4 h,大黄素设置5个浓度组:0,10,20,40,80 mmol·L-1。分别检测各组C-C型趋化因子(CCL17)在CT26细胞的表达,C-C型趋化因子受体4(CCR4)在SLs细胞表面的表达比例及CCR4 mRNA表达水平,CT26细胞NF-κB的活化。结果:与单纯CT26组和SLs组比,混合培养各比例组均检测到CCL17(P<0.01);大黄素显著抑制了混合培养组CCL17在CT26细胞的分泌:在1∶1比例组从308 ng·L-1降至124 ng·L-1。单纯SLs组CCR4的表达变化不明显;混合培养组的CD4+CD25+Tregs表达CCR4的细胞比例随着大黄素浓度的增加明显下降(P<0.05),1∶4比例组从44.58%下降到32.86%;相似的CCR4的mRNA表达随大黄素浓度增加出现明显的下调(P<0.05);大黄素显著抑制CT26细胞NF-κB的活化(P<0.05),p65的平均荧光值从358(0 mmol·L-1)降至76(80 mmol·L-1)。结论:大黄素抑制CT26细胞CCL17的分泌,下调CCR4在Treg细胞表面的表达比例和mRNA水平的表达。大黄素抑制结肠癌的机制可能和NF-κB的活化相关。
OBJECTIVE: To study the effect of emodin on the nuclear factor-κB (NF-κB) pathway in murine splenic lymphocytes and colon cancer CT26.WT cells and to explore the possible mechanism of anti-tumor effect. Methods: CT26 cells and splenic lymphocytes (SLs) with a density of 1 × 105 / mL were mixed and cultured at a ratio of 1: 1, 1: 2 and 1: 4 respectively for 24 hours, followed by addition of emodin for another 4 hours. Emodin Five concentration groups: 0,10,20,40,80 mmol·L-1. The expression of C-C chemokines (CCL17) in CT26 cells and the expression of C-type chemokine receptor 4 (CCR4) on the cell surface of SLs and the expression of CCR4 mRNA and the activation of NF-κB in CT26 cells were detected. Results: Compared with CT26 group and SLs group, CCL17 (all P <0.01) was detected in all the mixed culture groups. Emodin significantly inhibited the secretion of CCL17 in CT26 cells in the mixed culture group from 308 ng · L-1 dropped to 124 ng · L-1. The expression of CCR4 in SLs group was not changed obviously. The proportion of cells expressing CCR4 in CD4 + CD25 + Tregs in mixed culture group decreased significantly with the increase of emodin concentration (P <0.05), and decreased from 44.58% to 32.86 in 1: 4 ratio group (P <0.05). Emodin significantly inhibited the activation of NF-κB in CT26 cells (P <0.05). The average fluorescence intensity of p65 increased from 358 (0 mmol / · L-1) to 76 (80 mmol · L -1). CONCLUSION: Emodin inhibits the secretion of CCL17 in CT26 cells and down-regulates the expression of CCR4 on the surface of Treg cells and mRNA expression. The mechanism of emodin inhibiting colon cancer may be related to the activation of NF-κB.