藏猪促炎因子(IL-1β、IL-6、TNF-α)多态性及其表达与免疫性状的关联分析  

Polymorphism of Pro-Inflammatory Factors(IL-1β,IL-6,TNF-α)in Tibetan Pigs and Its Association Analysis with Immune Traits

作　　者：孙雯莉 王浩奇 泽里磋 高雨樊 张非凡 张健 段梦琪 商鹏 强巴央宗 SUN Wenli;WANG Haoqi;ZE Licuo;GAO Yufan;ZHANG Feifan;ZHANG Jian;DUAN Mengqi;SHANG Peng;QIANG Bayangzong(College of Animal Science,Xizang Agricultural and Animal Husbandry College,Linzhi 860000,China;Key Laboratory of Tibetan Pig Genetic Improvement and Reproduction,Linzhi 860000,China;Tibet Collaborative Innovation Center for the Research and Development of Unique Agricultural and Animal Resources,Linzhi 860000,China)

机构地区：[1]西藏农牧学院动物科学学院,林芝860000 [2]藏猪遗传改良与繁殖工程重点实验室,林芝860000 [3]西藏特色农牧资源研发省部共建协同创新中心,林芝860000

出　　处：《畜牧兽医学报》2024年第5期1958-1969,共12页ACTA VETERINARIA ET ZOOTECHNICA SINICA

基　　金：国家重点研发项目(2022YFD1600903);国家自然基金地区基金项目(32160773);西藏自治区科技计划项目(XZ202202JD0002N);西藏自治区重大科技专项项目(XZ202101ZD0005N)。

摘　　要：本研究以约克夏猪为对照,旨在分析藏猪和约克夏猪的血液生理指标和免疫相关器官组织表达谱方面的差异,结合藏猪外周血淋巴细胞在LPS刺激下,促炎因子IL-1β、IL-6和TNF-α的表达情况,探究IL-1β、IL-6及TNF-α对藏猪免疫性状的影响。试验选择180日龄的藏猪和约克夏猪各40头按品种分为两个试验组,采集新鲜血液用于检测血液生理指标;采集猪外周淋巴血用于培养外周血淋巴细胞。随机选取藏猪和约克夏猪各8头按品种分为两个试验组,前腔静脉采集新鲜血液、屠宰后采集黄豆样大小的肝脏、脾脏、下颌淋巴结组织Trizol法提取RNA。每个个体样品设置3个重复,借助RT-qPCR和一代测序技术检测IL-1β、IL-6、TNF-α基因表达情况和基因多态性。设置4组浓度的LPS(0、1、10、100μg·mL^(-1)),每组设置3个重复去刺激外周血淋巴细胞,于0、24、36、48、72 h收集细胞,用RT-qPCR检测促炎因子mRNA的表达情况。结果表明:1)藏猪的WBC、RBC、HGB、HCT和PLT水平极显著高于约克夏猪(P<0.01),同时,平均MCV和PCT也在藏猪中显著高于约克夏猪(P<0.05)。2)RT-qPCR结果表明,在藏猪的血液、脾脏和肝脏组织中,IL-1β、IL-6和TNF-α基因的mRNA表达量也均显著高于约克夏猪(P<0.01);在藏猪的下颌淋巴结中,IL-1β和TNF-α基因的表达量显著高于约克夏猪(P<0.05),IL-6基因的mRNA表达量极显著高于约克夏猪(P<0.01)。3)SNP位点筛选结果发现,在IL-1β基因3′侧翼区存在5个突变位点,其中G^(690)T、C^(1383)G、C^(1480)T、A^(1497)G位点在藏猪和约克夏猪群体间存在极显著差异(P<0.01),C^(1454)T位点在藏猪和约克夏猪群体存在显著差异(P<0.05);IL-6基因3′侧翼区存在1个有义突变位点C^(265)T,两品种间呈显著差异(P<0.05),在5′侧翼区存在1个有义突变位点A-72G,两品种间呈极显著差异(P<0.01);TNF-α基因在5′侧翼区存在1个有义突变位点,但两品种间差异不显著(P>0.0This study,using Yorkshire pigs as controls,aimed to explore the differences in blood physiological indicators and the expression profiles of immune-related organ tissues between Tibetan pigs and Yorkshire pigs.Additionally,the expression levels of pro-inflammatory cytokines IL-1β,IL-6,and TNF-αin Tibetan pig peripheral blood lymphocytes stimulated with LPS was investigated,to understand their impact on the immune characteristics of Tibetan pigs.Forty Tibetan pigs and forty Yorkshire pigs aged 180 days were divided into two experimental groups based on breed.Fresh blood samples were collected for detecting blood physiological indicators,and peripheral blood lymphocytes were cultured for further analysis.Moreover,8 Tibetan pigs and 8 Yorkshire pigs from each breed were randomly selected and divided into two experimental groups.Fresh blood was collected from the anterior vena cava,and liver,spleen,and submandibular lymph node tissues were collected post-slaughter for RNA extraction using the Trizol method.Each individual sample was subjected to 3 replicates for RT-qPCR and next-generation sequencing analysis to assess the expression levels and genetic polymorphisms of IL-1β,IL-6,and TNF-αgenes.Peripheral blood lymphocytes were stimulated with 4 concentrations of LPS(0,1,10,100μg·mL^(-1)),with 3 replicates per group.Cells were collected at 0,24,36,48,and 72 h,and RT-qPCR was performed to assess the expression of pro-inflammatory cytokine mRNA.Results indicate that:1)Tibetan pigs exhibited significantly higher levels of WBC,RBC,HGB,HCT,and PLT compared to Yorkshire pigs(P<0.01).Additionally,the average MCV and PCT were significantly higher in Tibetan pigs than those in Yorkshire pigs(P<0.05).2)RT-qPCR results revealed significantly higher mRNA expression levels of IL-1β,IL-6,and TNF-αgenes in the blood,spleen,and liver tissues of Tibetan pigs compared to Yorkshire pigs(P<0.01).In the submandibular lymph nodes of Tibetan pigs,the expression levels of IL-1βand TNF-αgenes were significantly higher(P<0.05),whil

关 键 词：藏猪 LPS 促炎因子 血液生理指标 SNP 

分 类 号：S828.2[农业科学—畜牧学]