马春松, 程珂, 郭勋, 王春芳. miR-194a介导饲料维生素D3调节黄颡鱼JAK-STAT免疫通路的研究[J]. 水生生物学报, 2023, 47(1): 11-24. DOI: 10.7541/2022.2021.0291
引用本文: 马春松, 程珂, 郭勋, 王春芳. miR-194a介导饲料维生素D3调节黄颡鱼JAK-STAT免疫通路的研究[J]. 水生生物学报, 2023, 47(1): 11-24. DOI: 10.7541/2022.2021.0291
MA Chun-Song, CHENG Ke, GUO Xun, WANG Chun-Fang. miR-194a MEDIATES DIETARY VITAMIN D3 IN THE JAK-STAT IMMUNE PATHWAY OF YELLOW CATFISH (PELTEOBAGRUS FULVIDRACO)[J]. ACTA HYDROBIOLOGICA SINICA, 2023, 47(1): 11-24. DOI: 10.7541/2022.2021.0291
Citation: MA Chun-Song, CHENG Ke, GUO Xun, WANG Chun-Fang. miR-194a MEDIATES DIETARY VITAMIN D3 IN THE JAK-STAT IMMUNE PATHWAY OF YELLOW CATFISH (PELTEOBAGRUS FULVIDRACO)[J]. ACTA HYDROBIOLOGICA SINICA, 2023, 47(1): 11-24. DOI: 10.7541/2022.2021.0291

miR-194a介导饲料维生素D3调节黄颡鱼JAK-STAT免疫通路的研究

miR-194a MEDIATES DIETARY VITAMIN D3 IN THE JAK-STAT IMMUNE PATHWAY OF YELLOW CATFISH (PELTEOBAGRUS FULVIDRACO)

  • 摘要: 为了研究miRNA如何介导饲料维生素D3在黄颡鱼(Pelteobagrus fulvidraco)JAK-STAT通路中调节免疫应答的机制, 选择黄颡鱼幼鱼为主要研究对象, 设计了3组维生素D3浓度分别为1120、3950和16600 IU/kg的饲料, 开展了为期12周养殖实验, 养殖实验结束后进行鮰爱德华氏菌(Edwardsiella ictaluri)攻毒实验。对攻毒后的头肾和脾脏组织进行Illumina高通量测序, 筛选差异表达的miRNAs, 并对差异表达miRNAs进行靶基因的生物信息学预测和富集分析, 发现miR-194a的靶基因富集在JAK-STAT信号通路中; 进一步通过双荧光素酶报告基因实验验证了miR-194a对靶基因jak2atyk2具有靶向性调控的关系, 能够发挥对jak2atyk2的抑制作用; 通过体外巨噬细胞实验, 在细胞中同样检测到miR-194a可以响应培养基中不同浓度的活性维生素D3, 并对靶基因jak2atyk2同样发挥抑制作用; 通过进一步对靶基因在JAK-STAT通路中的下游基因表达检测, 发现随着JAK-STAT信号通路的激活, 抗炎症反应(AIR)因子及被其诱导激活的靶基因的表达量随着培养基中维生素D3浓度升高呈现上升趋势(P<0.05), 炎症通路NF-κB通路被抑制(P<0.05), 促炎因子IL-1β、IL-6TNF-α等表达量显著下降(P<0.05), 并且调节了巨噬细胞由M1型极化向M2型极化的转变(P<0.05)。研究结果发现miR-194a能在体内体外实验中响应维生素D3的变化, 维生素D3通过促进miR-194a的表达, 负调控靶基因jak2atyk2, 并对JAK-STAT通路中的下游基因发挥间接作用, 阐释了miR-194a介导饲料维生素D3通过JAK-STAT信号通路调控黄颡鱼免疫应答的机制。

     

    Abstract: In order to investigate the mechanism about the regulation of immune response by miRNAs mediating feed vitamin D3 in the JAK-STAT pathway of yellow catfish (Pelteobagrus fulvidraco), three diets with vitamin D3 content at 1120, 3950 and 16600 IU/kg were designed for juvenile Pelteobagrus fulvidraco. The growth trial was carried out over a 12-week period, following by an Edwardsiella ictaluri challenge experiment. Illumina high-throughput sequencing of head kidney and spleen after bacteria challenge identified differentially expressed miRNAs. The target genes of miR-194a was enriched in the JAK-STAT pathway. The relationship between miR-194a and the possible target genes jak2a and tyk2 had been verified by a dual luciferase reporter gene assay, which indicated the inhibitory role of miR-194a on jak2a and tyk2. In vitro, miR-194a was also found to respond with different concentrations of vitamin D3, inhibiting the expression of the target genes of jak2a and tyk2. With the activation of the JAK-STAT signaling pathway, the expression of anti-inflammatory response (AIR) factors and the target genes were also activated, the NF-κB pathway was inhibited, with significant decrease in expression of pro-inflammatory factors IL-1β, IL-6 and TNF-α and the transformation of macrophages from M1-type to M2-type polarization was accelerated with the increasing of vitamin D3 concentration (P<0.05). In conclusion, the results of this study demonstrated that miR-194a could respond to different concentration of vitamin D3 in vivo and in vitro experiments. Dietary vitamin D3 up-regulated the expression of miR-194a, which negatively regulated the target genes of jak2a and tyk2, and exerted indirect effects towards the downstream genes in the JAK-STAT pathway, illuminating the mechanism of miR-194a-mediated regulation of immune response by dietary vitamin D3 through the JAK-STAT signaling pathway in Pelteobagrus fulvidraco.

     

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