Mycoplasma ovipneumoniae belongs to Mycoplasma, a genus containing the smallest self-replicating microorganisms, and causes infectious pleuropneumonia in goats and sheep. Nucleotide-binding oligomerization domain-containing protein (NOD2), an intracellular pattern recognition receptor, interacts with muramyl dipeptide (MDP) to recognize bacterial peptidoglycans and is involved in autophagy induction. However, there have been no reports about NOD recognition of mycoplasmas or M. ovipneumoniae-induced autophagy. In this study, we sought to determine the role of NOD2 in M. ovipneumoniae-induced autophagy using Western blotting, immunofluorescence, real-time PCR (RT-PCR), and color-changing unit (CCU) analysis. M. ovipneumoniae infection markedly increased NOD2 but did not increase NOD1 expression in RAW 264.7 cells. Treating RAW 264.7 cells with MDP significantly increased colocalization of M. ovipneumoniae and LC3, whereas treatment with NOD inhibitor, NOD-IN-1, decreased colocalization of M. ovipneumoniae and LC3. Furthermore, suppressing NOD2 expression with small interfering RNA (siRNA)-NOD2 failed to trigger M. ovipneumoniae-induced autophagy by detecting autophagy markers Atg5, beclin1, and LC3-II. In addition, M. ovipneumoniae infection significantly increased the phosphorylated c-Jun NH₂-terminal kinase (p-JNK)/JNK, p-Bcl-2/Bcl-2, beclin1, Atg5, and LC3-II ratios in RAW 264.7 cells. Treatment with JNK inhibitor, SP600126, or siRNA-NOD2 did not increase this reaction. These findings suggested that M. ovipneumoniae infection activated NOD2, and both NOD2 and JNK pathway activation promoted M. ovipneumoniae-induced autophagy. This study provides new insight into the NOD2 reorganization mechanism and the pathogenesis of M. ovipneumoniae infection.

中文翻译：

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NOD2 / c-Jun NH2末端激酶触发卵原支原体诱导的巨噬细胞自噬。

猪肺炎支原体属于支原体，它是一种具有最小自我复制能力的微生物，可在山羊和绵羊中引起传染性胸膜肺炎。包含核苷酸结合的寡聚域的蛋白质（NOD2），一种细胞内模式识别受体，与鼠李二肽（MDP）相互作用以识别细菌肽聚糖，并参与自噬诱导。然而，没有关于NOD识别支原体或猪肺炎支原体诱导的自噬的报道。在这项研究中，我们试图使用蛋白质印迹，免疫荧光，实时荧光定量PCR（RT-PCR）和变色单元（CCU）分析来确定NOD2在卵形支原体诱导的自噬中的作用。猪肺炎支原体感染显着增加了NOD2的表达，但并未增加RAW 264.7细胞中NOD1的表达。用MDP处理RAW 264.7细胞可显着提高卵形支原体和LC3的共定位，而用NOD抑制剂NOD-IN-1处理可降低卵形支原体和LC3的共定位。此外，抑制与小干扰RNA表达NOD2（siRNA）的-NOD2未能触发M.绵羊肺炎通过检测标记的自噬的Atg5，Beclin1基因，和LC3-II诱导的自体吞噬。此外，卵形支原体感染明显增加了磷酸化的c-Jun NH ₂-RAW激酶（p-JNK）/ JNK，p-Bcl-2 / Bcl-2，beclin1，Atg5和LC3-II在RAW 264.7细胞中的比例。用JNK抑制剂，SP600126或siRNA-NOD2处理不会增加该反应。这些发现表明卵形支原体感染激活了NOD2，而NOD2和JNK途径的激活均促进了卵形支原体诱导的自噬。这项研究提供了新的见解，以NOD2重组机制和卵裂分枝杆菌感染的发病机理。