PI3K/Akt Signaling Pathway Regulates Mitochondrial‑Dependent Apoptosis in Ovine Pulmonary Fibroblasts upon Mycoplasma ovipneumoniae Infection

Mycoplasma ovipneumoniae (Mo) is the major pathogen of ovine mycoplasmal pneumonia, hindering the development of sheep husbandry in China. Its latent and persistent infection characteristics limit the efficacy of conventional drugs and vaccines, making it essential to clarify Mo’s molecular pathogenic mechanism for developing new prevention and control strategies. The correlation between mycoplasma pathogenicity and host cell damage or immune disorders has been confirmed, yet the involved key signaling pathways remain unclear. In this study, in vitro (ovine pulmonary fibroblast OAR-L1 cells) and in vivo (sheep and mouse) Mo infection models were constructed to explore Mo-induced host cell apoptosis and its regulatory mechanism. In vitro results showed that Mo infection triggered time-dependent apoptosis in OAR-L1 cells, upregulated the expression of pro-apoptotic caspase 3, caspase 9, Bak and Bad, and reduced the Bcl-2/Bax ratio. In vivo , Mo infection caused pulmonary pathological lesions and elevated apoptosis-related molecule expression in sheep lungs, with more severe apoptotic changes observed in mice. Transcriptomic KEGG analysis screened the PI3K/Akt signaling pathway as a key differential pathway after Mo infection. Pathway intervention experiments verified that the PI3K inhibitor LY294002 aggravated Mo induced apoptosis, while the Akt agonist SC79 reversed the abnormal expression of mitochondrial apoptotic factors and suppressed cell apoptosis. Mechanistically, Mo infection induced reactive oxygen species (ROS) accumulation and mitochondrial membrane potential decline in OAR-L1 cells. The ROS scavenger NAC restored mitochondrial function, reduced cell apoptosis, and upregulated PI3K phosphorylation, relieving ROS-mediated inhibitory effect on the PI3K/Akt pathway. This study confirms that Mo induces host mitochondrial apoptosis via regulating the ROS mediated PI3K/Akt signaling pathway, which reveals a novel Mo pathogenic mechanism and provides a theoretical basis for targeted drug research and prevention of ovine mycoplasmal pneumonia.