LPS诱导马冈鹅法氏囊损伤相关基因的筛选

doi:10.16431/j.cnki.1671-7236.2023.07.005

摘要/Abstract

摘要： 【目的】研究脂多糖（LPS）对马冈鹅法氏囊组织损伤和氧化应激的影响，并探究其调控机制。【方法】选择100只1日龄马冈鹅（公母各半），随机分为对照组和LPS组，每组5个重复，每个重复10只。LPS组雏鹅分别于24、26和28日龄时腹腔注射2 mg/kg LPS，对照组注射等量生理盐水。于28日龄注射1 h后，无菌采集法氏囊进行组织学观察、氧化应激指标检测，并构建法氏囊转录组文库。通过差异表达基因鉴定、GO功能和KEGG通路富集及蛋白互作网络分析筛选出与LPS诱导法氏囊损伤有关的候选基因；随机挑选10个差异表达基因，利用实时荧光定量PCR验证其表达情况。【结果】马冈鹅法氏囊组织学观察结果表明，LPS组法氏囊结构完整性明显降低，囊泡内细胞排列紊乱、空泡数量增加。与对照组相比，LPS组法氏囊谷胱甘肽过氧化物酶（GSH-Px）活性显著降低（P<0.05）；LPS组共获得507个差异表达基因，其中277个基因上调，230个基因下调。GO功能和KEGG通路富集分析显示，差异表达基因主要富集在与炎症和免疫反应相关的条目，涉及Cytokine-cytokine受体相互作用、Notch和Hedgehog等信号通路。结合蛋白互作网络分析，共筛选到TNF超家族成员10（TNFSF10）、CREB结合蛋白（CREBBP）、中心粒旁物质1（PCM1）、蛋白酪氨酸磷酸酶非受体6型（PTPN6）等27个差异表达基因。实时荧光定量PCR结果显示，转录组测序结果准确可靠。【结论】本研究探究了LPS诱导马冈鹅法氏囊损伤前后法氏囊组织中的差异表达基因，筛选到了影响法氏囊组织损伤的Cytokine-cytokine受体相互作用、Notch和Hedgehog等信号通路以及TNFSF10、CREBBP、PTPN6等相关基因，为了解LPS诱导鹅法氏囊损伤的分子机制提供理论参考。

关键词: 脂多糖; 马冈鹅; 法氏囊损伤; 转录组测序

Abstract: 【Objective】 The purpose of this experiment was to study the effects of lipopolysaccharide (LPS) on tissue injury and oxidative stress of bursae of Fabricius of Magang geese and to explore its regulatory mechanism.【Method】 One hundred 1-day-old Magang geese (half female and half male) were randomly divided into control and LPS group with 5 replicates in each group and 10 geese in each replicate.The goslings in LPS group were injected intraperitoneally with 2 mg/kg LPS at 24, 26 and 28 days, respectively, while the control group was injected with the same amount of saline.After 1 hour of injection at 28 days, the bursa of Fabricius was collected aseptically for histological observation and oxidative stress index detection and transcriptome library construction.Candidate genes related to LPS-induced bursa of Fabricius injury were screened by differentially expressed gene identification, GO function and KEGG pathway enrichment, and protein interaction network analysis.Ten differentially expressed genes were randomly selected and the expressions were verified by Real-time quantitative PCR.【Result】 Histological observation of bursae of Fabricius of Magang goose showed that the structural integrity of bursae of Fabricius was significantly decreased, the cell arrangement in vesicles was disordered and the number of vacuoles increased in LPS group.Compared with the control group, the activity of glutathione peroxidase (GSH-Px) of bursae of Fabricius was significantly decreased in LPS group (P <0.05).A total of 507 differentially expressed genes were obtained in LPS group, of which 277 genes were up-regulated and 230 genes were down-regulated.GO function and KEGG pathway enrichment analysis showed that differentially expressed genes were mainly concentrated in items related to inflammation and immune response, and related to Cytokine-cytokine receptor interaction pathway and Notch and Hedgehog signaling pathways. A total of 27 differentially expressed genes, including TNF superfamily member 10 (TNFSF10), CREB-binding protein (CREBBP), paracentric substance 1 (PCM1), protein tyrosine phosphatase non-receptor type 6 (PTPN6), were screened by binding protein cooperative network analysis.The results of Real-time quantitative PCR showed that the transcriptome sequencing results were accurate and reliable.【Conclusion】 This study explored the differential expression genes in bursa of Fabricius in Magang goose before and after LPS-induced, and identified the Cytokine-cytokine receptor interaction, Notch and Hedgehog signaling pathways and related genes such as TNFSF10, CREBBP and PTPN6 affecting bursa of Fabricius injury, which provide theoretical reference for understanding the molecular mechanism of LPS-induced bursae of Fabricius injury in geese.

Key words: lipopolysaccharide; Magang goose; bursa of Fabricius injury; transcriptome sequencing

中图分类号:

S835

余梓榆, 张冰琪, 蔡远荣, 洪龙胜, 李婉雁, 曹楠, 黄运茂, 许丹宁, 田允波, 陈文斌, 李冰心. LPS诱导马冈鹅法氏囊损伤相关基因的筛选[J]. 中国畜牧兽医, 2023, 50(7): 2639-2650.

YU Ziyu, ZHANG Bingqi, CAI Yuanrong, HONG Longsheng, LI Wanyan, CAO Nan, HUANG Yunmao, XU Danning, TIAN Yunbo, CHEN Wenbin, LI Bingxin. Screening of Genes Related to Bursa of Fabricius Injury Induced by LPS in Magang Geese[J]. China Animal Husbandry and Veterinary Medicine, 2023, 50(7): 2639-2650.

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