摘要
【目的】基于转录组测序探究高精料饲粮中添加苦豆子总碱对绵羊肝损伤的缓解机制及其对糖代谢关键通路的调控作用。【方法】选用18只3月龄杜蒙公羊随机分为G1组(精粗比50∶50)、G2组(精粗比70∶30)和S3组(精粗比70∶30,添加121 mg/kg苦豆子总碱),每组6只羊。预饲期15 d,试验期60 d,试验结束后每组随机选取3只羊屠宰,采集肝脏组织进行转录组测序,筛选差异表达基因(DEGs),并进行GO功能和KEGG通路富集分析;分析糖代谢基因表达量倍数变化,评估关键酶基因的表达差异,并从中选取4个基因通过实时荧光定量PCR检测进行验证。【结果】G2 vs G1组中共筛选到411个DEGs(上调233,下调178),S3 vs G2组中共筛选到3 664个DEGs(上调1 799,下调1 865)。GO功能富集分析显示,G2 vs G1组的DEGs主要富集于细胞外基质和炎症免疫反应等条目;而S3 vs G2组的DEGs则主要富集于免疫系统调控及生物过程正调控相关条目。KEGG通路富集分析表明,G2 vs G1组中未显著富集到糖代谢相关通路;而S3 vs G2组的DEGs显著富集到6条与糖代谢相关通路,包括糖酵解/糖异生、丙酮酸代谢、果糖和甘露糖代谢、抗坏血酸和醛酸代谢、戊糖和葡萄糖醛酸的相互转化和肌醇磷酸盐代谢;其中糖异生通路中PCK1、PCK2、PC、G6PC1和FBP1基因表达量显著上调(P<0.05);糖酵解通路中HK1、HK2、HK3和PFKP基因表达量显著下调,PKLR基因表达量显著上调(P<0.05);柠檬酸循环通路中MDH2基因表达量显著上调(P<0.05)。PCK1、G6PC1、IDH3B、SDHC基因的实时荧光定量PCR验证结果与转录组测序结果一致。【结论】苦豆子总碱通过富集免疫系统调控及生物过程正调控相关功能,可减轻高精料诱导的绵羊肝脏炎症与纤维化;通过提高糖异生关键酶基因的表达,抑制糖酵解上游酶基因的表达,同时促进其下游酶基因的表达,并促进肝脏柠檬酸循环,进而改善高精料导致的糖代谢紊乱,维持机体糖代谢稳态。
【Objective】To investigate the mechanism by which liver injury in sheep was alleviated and the key pathways of glucose metabolism were regulated through the addition of total alkaloids from Sophora alopecuroides to high-concentrate diets based on transcriptome sequencing.【Method】Eighteen 3-month-old Dumont male sheep were randomly divided into three groups:G1(concentrate to roughage ratio was 50∶50),G2(concentrate to roughage ratio was 70∶30),and S3(concentrate to roughage ratio was 70∶30,added with 121 mg/kg total alkaloids of Sophora alopecuroides).After a 15-day pre-feeding period and a 60-day experimental period,three sheep were randomly selected from each group for slaughter.Liver tissue was collected for transcriptomic sequencing,with differentially expressed genes(DEGs)identified and subjected to GO functional and KEGG pathway enrichment analyses.The fold changes in gene expression levels for glucose metabolism were further analyzed to evaluate expression differences in key enzyme genes,with four genes selected for validation via Real-time quantitative PCR.【Result】In the G2 vs G1 group,411 DEGs were identified(233 up-regulated,178 down-regulated).In the S3 vs G2 group,3664 DEGs were identified(1799 up-regulated,1865 down-regulated).GO functional enrichment analysis revealed that DEGs in the G2 vs G1 group were primarily enriched in entries such as extracellular matrix and inflammatory and immune responses.Whereas the DEGs in the S3 vs G2 group were mainly enriched in entries related to immune system regulation and positive regulation of biological processes.KEGG pathway enrichment analysis indicated that no pathways related to glucose metabolism were significant enriched in the G2 vs G1 group.Whereas the S3 vs G2 group exhibited significant enrichment of DEGs across six glucose metabolism-related pathways,including glycolysis/gluconeogenesis,pyruvate metabolism,fructose and mannose metabolism,ascorbate and aldarate metabolism,pentose and glucuronate interconversions,and inositol phosphate metabolism.In the glycolysis pathway,the expression levels of HK1,HK2,HK3,and PFKP genes were significantly downregulated,while the expression level of PKLR gene was significantly upregulated(P<0.05).In the citrate cycle pathway,the expression level of MDH2 gene was significantly increased(P<0.05).The results of Real-time quantitative PCR validation for PCK1,G6PC1,IDH3B,and SDHC genes were consistent with those of transcriptomic sequencing.【Conclusion】The total alkaloids of Sophora alopecuroides could alleviate high-concentrate diet induced inflammation and fibrosis in sheep liver by enriching functions related to immune system regulation and positive regulation of biological processes.The expression of key gluconeogenesis enzyme genes was increased,the expression of upstream glycolysis enzyme genes was inhibited,and the expression of downstream glycolysis enzyme genes was promoted,while the liver citrate cycle was enhanced.Through these effects,the glucose metabolism disorders induced by high-concentrate diet were ameliorated,and systemic glucose metabolic homeostasis was maintained.
作者
孙璐
李述方
王丽娜
赵建鑫
卢赫男
王海荣
SUN Lu;LI Shufang;WANG Lina;ZHAO Jianxin;LU Henan;WANG Hairong(Key Laboratory of Animal Nutrition and Feed Science,Higher Education Institutions of Inner Mongolia Autonomous Region,College of Animal Science,Inner Mongolia Agricultural University,Hohhot 010018,China)
出处
《中国畜牧兽医》
北大核心
2026年第2期671-681,共11页
China Animal Husbandry & Veterinary Medicine
基金
国家自然科学基金项目(32460853)
内蒙古自治区教育厅一流学科科研专项项目资助(YLXKZX-NND-007)。
关键词
苦豆子总碱
高精料饲粮
绵羊
糖代谢
转录组
totalalkaloidsfrom Sophora alopecuroides
high-concentrate diet
sheep
glucose metabolism
transcriptome
