Guangdong Provincial Water Environment and Aquatic Products Security Engineering Technology Research Center, Guangzhou Key Laboratory of Aquatic Animal Diseases and Waterfowl Breeding, College of Animal Sciences and Technology, Zhongkai University of Agriculture and Engineering, Guangzhou, Guangdong Province, 510222, China.
Guangdong Provincial Water Environment and Aquatic Products Security Engineering Technology Research Center, Guangzhou Key Laboratory of Aquatic Animal Diseases and Waterfowl Breeding, College of Animal Sciences and Technology, Zhongkai University of Agriculture and Engineering, Guangzhou, Guangdong Province, 510222, China; School of Biological Sciences, Lake Superior State University, Sault Ste. Marie, MI, 49783, USA.
Fish Shellfish Immunol. 2021 May;112:159-167. doi: 10.1016/j.fsi.2020.09.036. Epub 2020 Oct 2.
The red blood cells (RBCs) of fish make up around 95% of the total peripheral blood cells, and the long-held paradigm is that RBCs are mainly responsible for transporting oxygen. Previous studies have showed that the RBCs can be involved in the immune response against bacterial infection; however, this mechanism remains enigmatic. Here, we explored the structure of grass carp RBCs (GcRBCs). The results showed that the GcRBCs released a pseudopodia-like structure when grown in a 24-well plate, and the transmission electron microscopy (TEM) result showed that GcRBCs contained some organelle-like structures. To further verify the organelle-like structures might be the mitochondria and lysosome which similar to other immune cells, a fluorescent labeling assay was used to verify it. To decipher the antibacterial immunity of GcRBCs, transcriptomic profiling of grass carp RBCs after the incubation with E. coli was analyzed. The results showed that there were 4099 differently expressed genes (DEGs) of GcRBCs upon E. coli incubation, including 2041 up-regulated and 2058 down-regulated genes. In addition, to validate our transcriptomic data, we checked the expression of several cytokines, such as CCL4, CCL20, IL4, IL12 and IFN-α, and the results showed that all the selected gens were significantly up-regulated after E. coli incubation. Furthermore, E. coli incubation induced hemoglobin oxidation and increased the heme in GcRBCs, which further activated the expression of heme oxygenase 1 (HO-1), autophagy related genes 5 (ATG5), and ferritin. In contrast, E. coli incubation inhibited the expression of Ferroportin-1 (FPN1), which increased intracellular iron levels, induced Fenton reaction to release reactive oxygen species (ROS), and activated the ferroptosis signaling pathway in GcRBCs. Herein, we demonstrate that E. coli can induce teleost RBCs cell death through an iron-mediated ferroptosis pathway, which sheds new light on the interaction between bacteria and teleost RBCs.
鱼类的红细胞（RBC）约占外周血总细胞的 95%，长期以来的观点认为 RBC 主要负责运输氧气。先前的研究表明，RBC 可以参与针对细菌感染的免疫反应；然而，这种机制仍然很神秘。在这里，我们研究了草鱼 RBC（GcRBC）的结构。结果表明，GcRBC 在 24 孔板中生长时会释放出伪足样结构，透射电子显微镜（TEM）结果表明 GcRBC 含有一些类似细胞器的结构。为了进一步证实这些类似细胞器的结构可能是线粒体和溶酶体，与其他免疫细胞类似，我们使用荧光标记实验进行了验证。为了解析 GcRBC 的抗菌免疫机制，我们分析了草鱼 RBC 与大肠杆菌孵育后的转录组谱。结果表明，在大肠杆菌孵育后，有 4099 个 GcRBC 的差异表达基因（DEGs），包括 2041 个上调基因和 2058 个下调基因。此外，为了验证我们的转录组数据，我们检查了几种细胞因子的表达，如 CCL4、CCL20、IL4、IL12 和 IFN-α，结果表明，所有选定的基因在大肠杆菌孵育后均显著上调。此外，大肠杆菌孵育诱导血红蛋白氧化并增加 GcRBC 中的血红素，从而进一步激活血红素加氧酶 1（HO-1）、自噬相关基因 5（ATG5）和铁蛋白的表达。相比之下，大肠杆菌孵育抑制了 Ferroportin-1（FPN1）的表达，这增加了细胞内铁水平，诱导芬顿反应释放活性氧（ROS），并激活了 GcRBC 中的铁死亡信号通路。在此，我们证明大肠杆菌可以通过铁介导的铁死亡途径诱导硬骨鱼类 RBC 细胞死亡，这为细菌与硬骨鱼类 RBC 之间的相互作用提供了新的视角。
