摘要
目的:探讨亮菌多糖对氧化偶氮甲烷(AOM)/葡聚糖硫酸钠(DSS)诱导的炎症相关性结肠癌小鼠模型及其肠道菌群的影响。方法:将18只6~8周龄小鼠通过随机数字法分为对照组(Control)、模型组(Model),亮菌多糖干预组(ATPS60 mg/kg)。Model和ATPS组一次性腹腔注射AOM 10 mg/kg,对照组注射等量生理盐水,Model和ATPS组以饮用1.8%DSS水溶液5 d,普通水14 d为个一周期,进行三个循环周期。对照组全程饮用普通饮用水。Model组饮用自由水期间ATPS组小鼠每天进行灌胃处理;71天实验结束。收集小鼠结肠并观察其长度和出血情况,记录肿瘤大小和数目,并对结直肠进行HE和Ki-67染色。Elisa检测Control、Model和ATPS组arrb1、IL-10和TNF-α的表达情况。蛋白免疫印迹法(Western blot,WB)检测各组arrb1蛋白表达情况。同时,采集小鼠粪便,进行DNA提取和菌群测序。结果:AOM/DSS处理后小鼠体重明显下降,与Control组相比Model组小鼠结肠长度缩短明显,P=0.002。ATPS干预一定程度上可以抑制小鼠结肠长度的缩短,P=0.046,差别具有统计学意义。病理组织结果显示,对照组小鼠结肠隐窝形态结构正常,排列整齐,均匀分布。模型组小鼠结肠隐窝结构破坏明显,腺体结构扭曲变形,细胞核深染增大,结肠肿瘤形成。ATPS组可见结肠隐窝基本排列整齐,腺体扭曲变形有所改善,未见核大深染的异性细胞,提示ATPS存在抑制炎癌转化的药理作用。同时,Ki-67染色结果显示ATPS能够抑制结肠细胞异常增殖。Elisa和WB结果显示:Model组较Control组相比arrb1和TNF-α水平升高,IL-10水平降低。经过ATPS治疗后arrb1与TNF-α水平明显下降,IL-10水平表达升高。表明ATPS能够通过干预arrb1的表达治疗结肠炎相关性结肠癌。16S rDNA基因测序发现三组微生物区系丰富度和多样性存在显著差异。Control组、Model组和ATPS组共有的物种数量为69个,Model组特有的物种数量为11个,ATPS组特有的物种数量为6个。小鼠在经过AOM/DSS处理后普雷沃菌科_UCG-001菌属较对照组明显增多。ATPS组较Model组拟普雷沃菌属增加明显。结果发现普雷沃菌科_UCG-001是接受亮菌多糖的小鼠中最丰富的属,在小鼠炎癌转化发展过程中明显减少,表明该属的有益作用。结论:亮菌多糖能够通过调节arrb1和恢复AOM/DSS造成的Alpha多样性指数和群落构成变化,治疗慢性非可控性结肠炎,抑制慢性结肠炎炎癌转化。
Objective:To investigate the effects of Armillariella tabescens polysaccharides on the inflammatory associated colon cancer mouse model and intestinal flora induced by azomethane oxide(AOM)/sodium dextran sulfate(DSS).Methods:186~8 week-old mice were randomly divided into Control group,Model group and Armillariella tabescens polysaccharides intervention group(ATPS 60 mg/kg).Model and ATPS groups were injected with AOM 10 mg/kg intraperitoneally at one time,and Control group was injected with the same amount of normal saline.The Model and ATPS groups drank 1.8%DSS aqueous solution for 5 days and ordinary water for 14 days,and carried out three cycles.The Control group drank ordinary drinking water throughout.During the period of Model group drinking free water,ATPS group mice were given intragastric treatment every day.The experiment ended after 71 days.Colon length and bleeding were observed.Tumor size and number were recorded,and colorectal staining was performed with HE and Ki-67.The expression of arrb1,IL-10 and TNF-αin Control,Model and ATPS groups was detected by Elisa.The expression of arrb1 protein in each group was detected by Western blot(WB).At the same time,mouse feces were collected for DNA extraction and microflora sequencing.Results:After AOM/DSS treatment,the body weight of mice decreased significantly,and the colon length of mice in Model group decreased significantly compared with that in Control group,P=0.002.ATPS intervention could inhibit the shortening of colon length to a certain extent,P=0.046,and the difference was statistically significant.The pathological results showed that the colonic crypts of mice in blank group were normal in morphology and structure,arranged neatly and distributed evenly.In the Model group,the colonic crypt structure was destroyed,the glandular structure was distorted and deformed,the nuclear hyperstaining increased,and the colonic tumor formed.In the ATPS group,the colonic crypts were basically arranged,the distortion and deformation of the glands were improved,and no large and deeply stained heterologous cells were found,suggesting that ATPS had pharmacological effects on inhibiting the transformation of inflammatory cancer.Meanwhile,Ki-67 staining showed that ATPS could inhibit the abnormal proliferation of colon cells.Elisa and WB results showed that compared with Control group,the levels of arrb1 and TNF-αin Model group were increased,and IL-10 levels were decreased.After ATPS treatment,the levels of arrb1 and TNF-αdecreased significantly,and the expression of IL-10 increased.These results indicated that ATPS can treat colitis-associated colon cancer by interfering with arrb1 expression.16S rDNA gene sequencing revealed significant differences in microflora richness and diversity among the three groups.The number of species shared by Control group,Model group and ATPS group was 69.The number of species unique to Model group was 11,and the number of species unique to ATPS group was 6.After AOM/DSS treatment,the number of Prevotellaceae_UCG-001 bacteria in mice was significantly higher than that in Control group.The number of Prevotella in ATPS group was significantly higher than that in Model group.The results showed that Prevomyceaceae-UCG-001 was the most abundant genus in mice receiving ATPS,and significantly decreased during the transformation and development of inflammatory cancer in mice,indicating the beneficial effect of this genus.Conclusion:In this study,the Armillariella tabescens polysaccharides can cause changes in Alpha diversity index and community composition by regulating arrb1 and restoring AOM/DSS,treat chronic non-controllable colitis and inhibit the transformation of chronic colitis cancer.
作者
刘利
叶映泉
桂仲旋
张爽爽
张泉
张梅
LIU Li;YE Yingquan;GUI Zhongxuan;ZHNAG Shuangshuang;ZHANG Quan;ZHANG Mei(Department of Oncology Integrated Traditional Chinese and Western Medicine,the First Affiliated Hospital of Anhui Medical University,Anhui Hefei 230000,China;Tumor Center of Integrated Chinese and Western Medicine,Anhui Medical University,Anhui Hefei 230000,China;Graduate School,Anhui University of Traditional Chinese Medicine,Anhui Hefei 230000,China)
出处
《现代肿瘤医学》
CAS
2024年第12期2161-2170,共10页
Journal of Modern Oncology
基金
国家自然科学基金项目(编号:81774051)
2023年安徽省高校协同创新项目(编号:GXXT-2023-078)。
关键词
亮菌多糖
炎症相关性结肠癌
肠道菌群
AOM/DSS
armillariella tabescens polysaccharides
inflammatory associated colon cancer
intestinal flora
AOM/DSS
