College of Animal Science and Technology, Northwest A&F University, Yangling, 712100, Shaanxi, China.
Power China Northwest Engineering Corporation Limited, Xi'an, 710065, Shaanxi, China.
Fish Shellfish Immunol. 2024 Nov;154:109929. doi: 10.1016/j.fsi.2024.109929. Epub 2024 Sep 26.
Galectin-9 (Gal-9) belongs to a family of the glycan-binding proteins (GBPs) and is known to restrict bacterial activity via interacting with pathogen associated molecular pattern (PAMPs). However, the underlying immune mechanism of endogenous Gal-9 on fish against bacterial infection is still unclear. In this study, effects of Gal-9 from Onychostoma macrolepis (OmGal-9) on expression of immune-related genes were measured by HEK293T. The immune response of O. macrolepis with OmGal-9 overexpression to Aeromonas hydrophila (A. hydrophila) infection (1.65 × 10 CFU/mL) was evaluated by tissue bacterial load, fish survival rate and transcriptome analysis. The results showed that OmGal-9 displayed a punctate distribution in the nucleus and cytoplasm of HEK293T cells. Compared to cells transfected with the empty vector (EV group), recombinant plasmid pEGFP-Gal9 treatment (Gal9 group) significantly down-regulated the expression of immune-related genes TNFα, STAT3, MyD88, LCK, and p52 of HEK293T cells stimulated with LPS at 24 h, while up-regulated IκBα and caspase-1 (P < 0.05). The activities of catalase (CAT), superoxide dismutase (SOD), the total antioxidant capacity (T-AOC), alkaline phosphatase (AKP), acid phosphatase (ACP), and lysozyme (LZM) of O. macrolepis were significantly increased on 7 days in Gal9 group compared to EV group (P < 0.05). The bacterial load of liver, spleen, and kidney of O. macrolepis infected with A. hydrophila in Gal9 group at 24 h was significantly lower than that in EV group (P < 0.05), and the survival rate had increased from 15 % to 35 %. A comparative transcriptome analysis between the Gal9 and EV group identified 305 differentially expressed genes (DEGs). The analysis showed that OmGal-9 might play an important regulatory role in glycolysis/gluconeogenesis, fatty acid degradation, and ascorbate and aldarate metabolism. Moreover, the immune-related DEGs were predominantly enriched in eleven pathways, with the most important three of them being linked to innate immunity: NOD-like, C-type lectin and Toll-like receptor signaling pathway. Taking together, OmGal-9 can enhance the resistance of fish to bacterial diseases by improving immune system function and activating immune-related pathways.
半乳糖凝集素-9（Gal-9）属于糖结合蛋白（GBP）家族，已知通过与病原体相关的分子模式（PAMPs）相互作用来限制细菌的活性。然而，鱼类内源性 Gal-9 对细菌感染的潜在免疫机制仍不清楚。在这项研究中，通过 HEK293T 测量了来自大鳍鱯（Onychostoma macrolepis）的 Gal-9（OmGal-9）对免疫相关基因表达的影响。通过组织细菌载量、鱼类存活率和转录组分析评估了过表达 OmGal-9 的大鳍鱯（O. macrolepis）对嗜水气单胞菌（A. hydrophila）感染（1.65×10 CFU/mL）的免疫反应。结果表明，OmGal-9 在 HEK293T 细胞的核和细胞质中呈现点状分布。与转染空载体（EV 组）的细胞相比，重组质粒 pEGFP-Gal9 处理（Gal9 组）在 LPS 刺激 24 小时后显著下调了 HEK293T 细胞中 TNFα、STAT3、MyD88、LCK 和 p52 的免疫相关基因表达，同时上调了 IκBα 和 caspase-1（P<0.05）。与 EV 组相比，Gal9 组在第 7 天大鳍鱯的过氧化氢酶（CAT）、超氧化物歧化酶（SOD）、总抗氧化能力（T-AOC）、碱性磷酸酶（AKP）、酸性磷酸酶（ACP）和溶菌酶（LZM）的活性均显著升高（P<0.05）。Gal9 组感染嗜水气单胞菌的大鳍鱯肝、脾、肾的细菌载量在 24 小时时明显低于 EV 组（P<0.05），存活率从 15%提高到 35%。Gal9 组和 EV 组之间的比较转录组分析确定了 305 个差异表达基因（DEGs）。分析表明，OmGal-9 可能在糖酵解/糖异生、脂肪酸降解以及抗坏血酸和醛酸代谢中发挥重要的调节作用。此外，免疫相关的 DEGs 主要富集在十一条通路中，其中最重要的三个通路与先天免疫有关：NOD 样、C 型凝集素和 Toll 样受体信号通路。总之，OmGal-9 通过提高免疫系统功能和激活免疫相关途径，增强鱼类对细菌性疾病的抵抗力。
