HMGB1/RAGE轴在结直肠癌肥大细胞浸润中的作用机制研究

Mechanism of HMGB1/RAGE axis in mast cell infiltration of colorectal cancer

目的:探究高迁移率族蛋白B1(HMGB1)/晚期糖基化终产物受体(RAGE)轴在结直肠癌肥大细胞浸润中的作用机制。方法:将雄性BALB/c裸鼠分为正常对照组、结直肠癌模型组、NC shRNA组、HMGB1 shRNA组、PDTC组(NF-κB抑制剂),除正常对照组外,均建立结直肠癌模型。甲苯胺蓝(TB)染色观察结直肠癌组织肥大细胞浸润情况,免疫组化检测结直肠癌组织肥大细胞活性标志物类胰蛋白酶(Tryptase)表达,Western blot检测结直肠癌组织HMGB1、RAGE及其下游NF-κB p65蛋白表达,ELISA检测结直肠癌组织炎症因子TNF-α和IL-1β表达。结果:正常对照组无肿瘤生长,模型组和NC shRNA组裸鼠肿瘤体积增长基本同步,HMGB1 shRNA组与PDTC组裸鼠肿瘤体积较模型组增长缓慢。模型组和NC shRNA组肥大细胞数、Tryptase水平及HMGB1、RAGE、NF-κB p65蛋白表达、TNF-α和IL-1β水平差异均无统计学意义(P>0.05),显微镜下可观察到大量紫色脱颗粒肥大细胞和Tryptase阳性染色颗粒;HMGB1 shRNA组与PDTC组相较于模型组肥大细胞数、Tryptase水平及HMGB1、RAGE、NF-κB p65蛋白表达、TNF-α和IL-1β水平均显著下降(P<0.05),显微镜下可见少量紫色脱颗粒肥大细胞和Tryptase阳性染色颗粒。结论:HMGB1与RAGE结合可激活NF-κB信号通路,促进结直肠癌肥大细胞浸润。

Objective:To investigate mechanism of high mobility group box 1(HMGB1)/receptor of advanced glycation end products(RAGE)axis in mast cell infiltration of colorectal cancer.Methods:Male BALB/c nude mice were divided into normal control group,colorectal cancer model group,NC shRNA group,HMGB1 shRNA group,PDTC group(NF-κB inhibitor). Colorectal cancer models were established except for normal control group. Infiltration of mast cells in colorectal cancer was observed by toluidine blue(TB)staining,expression of mast cell activity marker Tryptase in colorectal cancer tissues was detected by immunohistochemistry,protein expressions of HMGB1,RAGE and their downstream NF-κB p65 were detected by Western blot,expressions of inflammatory cytokines TNF-α and IL-1β in colorectal cancer tissues were detected by ELISA.Results:There was no tumor growth in normal control group,and tumor volume growth in model group and NC shRNA group was basically synchronous,compared with model group,growth of tumor volume in HMGB1 shRNA group and PDTC group was slower. There were no significant differences in mast cell number,Tryptase level,HMGB1,RAGE,NF-κB p65 protein expressions,TNF-α and IL-1β levels between model group and NC shRNA group(P>0.05),a large number of purple degranulated mast cells and Tryptase positive granules were observed under microscope;compared with model group,number of mast cells,level of Tryptase,protein expressions of HMGB1,RAGE and NF-κB p65,levels of TNF-α and IL-1β in HMGB1 shRNA group and PDTC group were significantly decreased(P<0.05),and a few purple degranulated mast cells and Tryptase positive staining granules were observed under microscope.Conclusion:HMGB1 combined with RAGE can activate NF-κB signaling pathway and promote mast cell infiltration in colorectal cancer.