Klebsiella pneumoniae induced ferroptosis by inhibition of the Nrf2/xCT/GPX4 pathway in bovine mammary epithelial cells

Klebsiella pneumoniae (K. pneumoniae), a prominent causative agent of mastitis in dairy cattle, remains enigmatic in its pathogenic mechanisms. This study aimed to reveal the effects of K. pneumoniae on mammary glands via the induction of ferroptosis, as well as the protective role of Ferrostatin-1 (Fer-1) against this pathogen-mediated damage in bovine mammary epithelial cells (BMECs). Holstein cows were used to establish an intramammary infection model of K. pneumoniae. In vitro, primary BMECs were treated with 10 μM Fer-1 and K. pneumoniae alone or in combination. The results showed that mammary glands infected with K. pneumoniae exhibited increases in interleukin (IL) -1B, IL-6, IL-8, and tumor necrosis factor-α (TNFA), along with significantly increased iron, 4-hydroxynonenal, and reactive oxygen species (ROS) levels. Conversely, nuclear factor erythroid 2-related factor 2 (Nrf2), cystine/glutamate antiporter (xCT), glutathione peroxidase 4 (GPX4), and glutathione levels were downregulated following K. pneumoniae infection. Following the invasion of K. pneumoniae, intracellular iron and lipid ROS accumulated in the BMECs, which also impeded the activation of Nrf2/xCT/GPX4 signal transduction. Additionally, Fer-1 facilitated the nuclear migration of Nrf2, leading to the upregulation of the Nrf2/xCT/GPX4 pathway and downregulation IL-1B, IL-6, IL-8, and TNFA. In conclusion, the findings revealed the ability of Fer-1 to alleviate K. pneumoniae-induced inflammatory factor activation and ferroptosis in BMECs via upregulation of the Nrf2/xCT/GPX4 pathway, indicating that inhibition of ferroptosis holds promise as a feasible therapeutic agent for the control of mastitis.