基于转录组学分析驴油生酮饮...T26-+结肠癌肿瘤的变化_张华宸.pdf

现代食品科技 Modern Food Science and Technology 2023,Vol.39,No.6 18 基于转录组学分析驴油生酮饮食下 CT26+结肠癌肿瘤的变化 张华宸1，张宁2，邢敬亚3，谢兰1，祁兴震1，路侹1，程杰4，李兰杰1*，刘桂芹1*（1.聊城大学农学与农业工程学院，山东省黑毛驴高效繁育与生态饲养工程技术研究中心，山东省驴产业科技协同创新中心，山东聊城 252059）（2.聊城大学生物制药研究院，山东聊城 252059）（3.内蒙古农业大学动物科学学院，内蒙古自治区马属动物遗传育种与繁殖学重点实验室，内蒙古农业大学马属动物研究中心，内蒙古呼和浩特 010018）（4.东阿阿胶股份有限公司，山东聊城 252200)摘要：该研究旨在通过转录组测序技术筛选出驴油生酮饮食（Donkey Oil Ketogeni Diet，DOKD）影响CT26+结肠癌肿瘤生长的相关基因，挖掘对DOKD 响应的基因及其代谢通路，为解析驴油生酮饮食抑制肿瘤生长的机制提供依据。该研究选取一致性较好的CT26+结肠癌BALB/c 模型小鼠，随机分为2 组，每组3 个重复，分别饲喂驴油生酮饮食（DOKD）与正常饮食（ND）。饲喂10 d后处死摘取肿瘤组织，测量肿瘤的质量及体积，并利用 RNA-Seq 技术和生物信息分析方法进行转录组测序及结果分析。DOKD 组与 ND 组小鼠体质量无显著差异，肿瘤明显小于ND 组。与ND 组相比，DOKD 组共18 个基因差异表达，其中12 个DEGs 上调，6 个DEGs 下调。这些DEGs 注释到170 条GO 条目，其中生物过程（BP）类别124 条，细胞组成（CC）类别2 条，分子功能（MF）类别44 条；涉及18条KEGG通路，主要富集在IL-17 信号通路、趋化因子信号通路、细胞因子受体相互作用信号通路、肿瘤坏死因子信号通路等重要通路。此外，该研究通过Western blot 进一步研究了TLR4/NF-B 信号通路在在肿瘤中的作用。综上，DOKD 对CT26+结肠癌肿瘤有明显抑制作用，该作用可能与IL-17 信号通路、趋化因子信号通路、细胞因子受体相互作用信号通路、TNF 信号通路等通路密切相关。关键词：驴油生酮饮食；肿瘤；转录组学；CT26+结肠癌 文章编号：1673-9078(2023)06-18-26 DOI:10.13982/j.mfst.1673-9078.2023.6.0827 Transcriptomic Analysis of Changes in CT26+Colon Cancer Tumors under Donkey Oil Ketogenic Diet ZHANG Huachen1,ZHANG Ning2,XING Jingya3,XIE Lan1,QI Xingzhen1,LU Ting1,CHENG Jie4,LI Lanjie1*,LIU Guiqin1*(1.College of Agriculture,Liaocheng University,Shandong Engineering Technology Research Center of Efficient Breeding and Ecological Rearing of black Donkey,Shandong Province Donkey Industry Science and Technology Collaborative Innovation Center,Liaocheng 252059,China)(2.Institute of Biopharmaceutical Research Liaocheng University,Liaocheng 252059,China)(3.Equine Research Center,Inner Mongolia Key Laboratory of Equine Genetics,Breeding and Reproduction,College of Animal Science,Inner Mongolia Agricultural University,Hohhot 010018,China)(4.Dong-E-E-Jiao Co.Ltd.,Liaocheng 252200,China）引文格式：张华宸,张宁,邢敬亚,等.基于转录组学分析驴油生酮饮食下CT26+结肠癌肿瘤的变化J.现代食品科技,2023,39(6):18-26.ZHANG Huachen,ZHANG Ning,XING Jingya,et al.Transcriptomic analysis of changes in CT26+colon cancer tumors under donkey oil ketogenic diet J.Modern Food Science and Technology,2023,39(6):18-26.收稿日期：2022-06-29 基金项目：山东省现代农业产业技术体系驴产业创新团队项目资助（SDAIT-27）；山东省乡村振兴科技创新提振行动计划项目（2021TZXD012）；聊城大学博士科研启动项目（318052120）；聊城大学畜牧学学科开放课题（319312101-03）作者简介：张华宸（1998-），女，硕士研究生，研究方向：动物学，E-mail： 通讯作者：李兰杰（1988-），男，博士，讲师，研究方向：动物科学，E-mail：；共同通讯作者：刘桂芹（1975-），女，博士，教授，研究方向：动物科学，E-mail： 现代食品科技 Modern Food Science and Technology 2023,Vol.39,No.6 19 Abstract:Screening of the genes related to the donkey oil ketogenic diet(DOKD)that affects the growth of CT26+colon cancer tumors using transcriptome sequencing technology was investigated in this study.Mining the genes and metabolic pathways that respond to DOKD provides a basis for analyzing the mechanism of DOKD in inhibiting tumor growth.BALB/c model mice with CT26+colon cancer with high consistency were selected and randomly divided into two groups with three replicates,and they were fed either a DOKD or normal diet(ND).The animals were sacrificed after 10 days of treatment.Subsequently,the tumors were excised,and tumor weights and volumes were recorded for analysis.RNA-Seq technology and bioinformatic analysis were used for transcriptome sequencing and results analysis.No significant difference was found in the body weight between the DOKD and ND groups,and the tumors were significantly smaller in the DOKD than those in the ND group.Compared with the ND group,a total of 18 genes were differentially expressed in the DOKD group,of which 12 differentially expressed genes(DEGs)were upregulated,and 6 DEGs were down regulated.These DEGs were annotated into a total of 170 GO items,including 124 in the biological process category,44 in the molecular function category,and 2 in the cellular composition category.A total of 18 KEGG pathways were involved,mainly enriched in the interleukin(IL)-17 signaling,chemokine signaling,cytokine receptor interaction signaling,tumor necrosis factor signaling,and other important pathways.In addition,this study further examined the role of the TLR4/NF-B signaling pathway in tumors using western blotting.In summary,DOKD has a significant inhibitory effect on CT26+colon cancer tumors,and this effect may be closely related to the IL-17,chemokine,cytokine receptor interaction,and tumor necrosis factor signaling pathways,among others.Key words:donkey oil ketogenic diet;tumor;transcriptomics;CT26+colon cancer 生酮饮食（Ketogenic Diet，KD）由高比例脂肪、低比例糖类及适量蛋白质等营养元素构成1，常见 KD配方中脂肪通常是糖类及蛋白质二者能量总和的 34倍2。KD 最早被应用于癫痫3的治疗，随着对 KD 代谢机制的深入研究，其应用逐步扩展到糖尿病4、心血管疾病5、肥胖6等多种慢性代谢疾病7的治疗。此外，大量基础研究已证实了 KD 的抗肿瘤疗效中，如神经胶质瘤8、结肠癌9、前列腺癌10、子宫内膜癌11等。KD 利用肿瘤细胞线粒体内膜缺失、无法利用酮体的特征12，通过限制葡萄糖的供应、哺乳动物雷帕霉素靶蛋白（Mammalian Target of Rapamycin，mTOR）信号通路、调节炎症及免疫反应等机制抑制肿瘤的生长。Marsh 等13在小鼠恶性星型细胞瘤模型中采用糖酵解抑制剂和 KD 联合治疗，发现 KD 可以通过降低葡萄糖转运蛋白-1 的表达减少肿瘤对葡萄糖的摄取，从而达到抑制肿瘤生长的效果。mTOR 是一种丝氨酸/苏氨酸蛋白激酶，可通过抑制炎症、血管生成与细胞上皮间-充质转化（EMT）等多条通路抑制肿瘤生 长14，目前已经成为肿瘤治疗的标志性靶点15。Zou等16研究发现，KD 与雷帕霉素药物联合疗法可通过抑制 mTOR 信号通路有效地抑制乳腺癌小鼠肿瘤的生长，延长小鼠的生存期。大量研究表明，炎症反应对肿瘤的作用是把双刃剑。一方面，炎症因子白细胞介素-8（Interleukin-8，IL-8