Key Laboratory of Agricultural Animal Genetics, Breeding and Reproduction, Ministry of Education, Huazhong Agricultural University, Wuhan, Hubei Province 430070, China.
Key Laboratory of Agricultural Animal Genetics, Breeding and Reproduction, Ministry of Education, Huazhong Agricultural University, Wuhan, Hubei Province 430070, China.
Res Vet Sci. 2021 Dec;141:164-173. doi: 10.1016/j.rvsc.2021.10.021. Epub 2021 Nov 1.
Mycoplasma gallisepticum (MG) is a major poultry pathogen that can induce Chronic Respiratory Disease (CRD) in chickens, causing serious economic losses in the poultry industry worldwide. Increasing evidence suggests that microRNAs (miRNAs) act as a vital role in resisting microbial pathogenesis and maintaining cellular mechanism. Our previous miRNAs sequencing data showed gga-miR-24-3p expression level was significantly increased in MG-infected chicken lungs. The aim of this study is to reveal the cellular mechanism behind the MG-HS infection. We found that gga-miR-24-3p was significantly upregulated and Ras-related protein-B (RAP1B) was downregulated in chicken fibroblast cells (DF-1) with MG infection. Dual luciferase reporting assay and rescue assay confirmed that RAP1B was the target gene of gga-miR-24-3p. Meanwhile, overexpressed gga-miR-24-3p increased the levels of tumor necrosis factor alpha (TNF-α) and interleukin-1β (IL-1β), and significantly inhibited cell proliferation as well as promoted MG-infected DF-1 cell apoptosis, whereas inhibition of gga-miR-24-3p had the opposite effect. More importantly, the results of overexpression and knockdown of target gene RAP1B demonstrated that the presence of RAP1B promoted cell proliferation and it saved the reduced or increased cell proliferation caused by overexpression or inhibition of gga-miR-24-3p. Furthermore, the overexpression of gga-miR-24-3p could significantly inhibit the expression of MG-HS adhesion protein. Taken together, these findings demonstrate that DF-1 cells can resist MG-HS infection through gga-miR-24-3p/RAP1B mediated decreased proliferation and increased apoptosis, which provides a new mechanism of resistance to MG infection in vitro.
鸡毒支原体（MG）是一种主要的家禽病原体，可引起鸡慢性呼吸道病（CRD），给全球家禽业造成严重的经济损失。越来越多的证据表明，微小 RNA（miRNA）在抵抗微生物发病机制和维持细胞机制方面起着重要作用。我们之前的 miRNA 测序数据显示，MG 感染鸡肺中的gga-miR-24-3p 表达水平显著升高。本研究旨在揭示 MG-HS 感染背后的细胞机制。我们发现，MG 感染鸡成纤维细胞（DF-1）中gga-miR-24-3p 显著上调，Ras 相关蛋白-B（RAP1B）下调。双荧光素酶报告实验和挽救实验证实，RAP1B 是 gga-miR-24-3p 的靶基因。同时，过表达 gga-miR-24-3p 增加了肿瘤坏死因子-α（TNF-α）和白细胞介素-1β（IL-1β）的水平，并显著抑制细胞增殖，促进 MG 感染的 DF-1 细胞凋亡，而抑制 gga-miR-24-3p 则产生相反的效果。更重要的是，过表达和敲低靶基因 RAP1B 的结果表明，RAP1B 的存在促进了细胞增殖，并挽救了 gga-miR-24-3p 过表达或抑制引起的细胞增殖减少或增加。此外，gga-miR-24-3p 的过表达可显著抑制 MG-HS 黏附蛋白的表达。综上所述，这些发现表明，DF-1 细胞可以通过 gga-miR-24-3p/RAP1B 介导的降低增殖和增加凋亡来抵抗 MG-HS 感染，为体外 MG 感染的抗性提供了新的机制。
