基于RNA-Seq技术研究枸杞多糖对环磷酰胺致雏鸡免疫抑制的拮抗机制  

The Mechanism of Lycium barbarum Polysaccharide against Immunosuppression Induced by Cyclophosphamide in Chicks was Studied Based on RNA-Seq Technique

作　　者：王建东 唐玉林 王敏 张宝锁 杨富强 高海慧 于洋 郭延生[2] WANG Jiandong;TANG Yulin;WANG Min;ZHANG Baosuo;YANG Fuqiang;GAO Haihui;YU Yang;GUO Yansheng(Institute of Animal Science,Ningxia Academy of Agriculture and Forestry Sciences,Yinchuan 750002,China;College of Agriculture,Ningxia University,Yinchuan 750021,China;Maternal and Child Health Care Hospital of Ningxia,Yinchuan 750001,China;Ningxia Shunbao Modern Agriculture Co.Ltd.,Qingtongxia 751600,China)

机构地区：[1]宁夏农林科学院动物科学研究所,银川750002 [2]宁夏大学农学院,银川750021 [3]宁夏银川市妇幼保健院,银川750001 [4]宁夏顺宝现代农业股份有限公司,青铜峡751600

出　　处：《畜牧兽医学报》2023年第8期3519-3532,共14页ACTA VETERINARIA ET ZOOTECHNICA SINICA

基　　金：宁夏自然科学重点基金项目(2020AAC02032)。

摘　　要：本试验旨在基于RNA-Seq技术研究枸杞多糖对环磷酰胺致雏鸡免疫抑制的拮抗机制,分析脾中与免疫相关差异表达基因的变化,为枸杞多糖拮抗雏鸡免疫抑制机制提供参考依据。将120只7日龄海兰褐蛋鸡随机分成3个组,分别为空白组(NC)、环磷酰胺组(CY)及枸杞多糖组(CYLbGp),CY和CYLbGp两组连续3 d每日胸肌注射80 mg·kg^(-1)环磷酰胺建立免疫抑制雏鸡模型,NC组肌注等体积生理盐水,CYLbGp组在模型建立后通过饮水的方式每天给予5 mg·kg^(-1)枸杞多糖,连续30 d。给药结束后,每组随机选取6只雏鸡采集脾,采用RNA-Seq技术检测3组在给予枸杞多糖后的差异表达基因。通过KEGG通路富集分析和蛋白质互作(PPI)网络分析,筛选关键差异基因和通路。结果表明,枸杞多糖干预后共有178个差异表达基因显著回调,显著富集在线粒体自噬-动物、趋化因子信号通路、C型凝集素受体信号通路、B细胞受体信号通路、血小板活化、自然杀伤细胞介导的细胞毒性和Th1和Th2细胞分化等免疫相关通路上。PPI分析结果表明,磷酸肌醇3激酶调节亚基(PIK 3CD)、Janus激酶3(JAK 3)、GATA结合蛋白3(GATA 3)、低氧诱导因子(HIF 1A)、NK2同源异型框5(NKX 2-5)和肌醇多磷酸磷酸酶样1(INPPL 1)为其中的关键基因。RT-qPCR结果与转录组学数据变化趋势基本一致,进一步表明转录组数据有较高的可靠性。综上,枸杞多糖通过作用于PIK 3CD、JAK3、GATA3、HIF 1A、和INPPL 1等关键靶点,参与线粒体自噬、趋化因子信号通路、C型凝集素受体信号通路、B细胞受体信号通路、血小板活化、自然杀伤细胞介导的细胞毒性、Th1和Th2细胞分化等信号通路的调控,进而起到缓解雏鸡免疫抑制的作用。(Objective)The aim of this research was to study the antagonistic mechanism of Lycium barbarum polysaccharide(LBP)on cyclophosphamide induced immunosuppression in chicks based on RNA-Seq technology,and to analyze the changes of immune-related differentially expressed genes in spleen,so as to provide reference for the antagonistic mechanism of LBP on immunosuppression of chicks.(Methods)120 One hundred and twenty 7 days old Hy-line brown laying hens were randomly divided into three groups:blank group(NC),cyclophosphamide group(CY)and Lycium barbarum polysaccharide group(CYLbGp).The immunosuppressive chick model was established by injecting 80 mg·kg^(-1) cyclophosphamide everyday into the pectoralis muscle for three consecutive days,and the same volume of normal saline was injected into the to the hens in NC group.After the establishment of the model,the chicks in the CYLbGp group were supplemented with given Lycium barbarum polysaccharide(LBP)5 mg·kg^(-1) by drinking water everyday for 30 consecutive days.At the end of administration,spleens were collected from 6 chicks in each group,and the differentially expressed genes(DEGs)in the three groups were detected by RNA-Seq.KEGG pathway enrichment analysis and protein-protein interaction(PPI)network analysis were used to screen key differentially expressed genes and pathways.The results showed that after LBP intervention a total of 178 genes significant difference correction DEGs significantly reduced,and were significantly enriched in mitochondrial autophagy-animals,chemokine signaling pathways,C-type lectin receptors signaling pathways,B cell receptor signaling pathways,platelet activation,natural killer cell mediated cytotoxicity and Th1 and Th2 cells differentiation and other related immune pathways.It PPI analysis showed that PIK3 CD,JAK3,GATA3,HIF1 A,NKX2-5 and INPPL1 were the key genes by PPI analysis.The results of RT-qPCR were consistent with the trend of transcriptomic data,which further indicated the high reliability of transcriptomic data.It can be conc

关 键 词：RNA-Seq技术 免疫抑制 枸杞多糖 环磷酰胺 脾 

分 类 号：S852.4[农业科学—基础兽医学]