Mechanism of Brucella pathogenesis mediated by outer membrane protein Omp19: Synergistic activation of cytokine storm and dual apoptotic pathways.

Upon host cell invasion, Brucella exhibits dual regulatory effects on apoptosis, demonstrating both pro-apoptotic and anti-apoptotic capabilities. Existing studies have confirmed that Omp31 and Omp25 possess apoptosis-inhibitory properties, while Omp19 can activate the apoptotic pathway through TNFR1 stimulation, thereby inducing apoptosis in astrocytes. Currently, the precise interaction mechanisms between various Brucella virulence factors and host cell apoptosis remain incompletely understood. Therefore, elucidating Brucella's regulatory role in host cell apoptosis represents a crucial aspect for understanding its cellular invasion and intracellular survival strategies. To investigate the regulatory effects of Brucella Omp19 outer membrane protein on host cell biological changes during bacterial invasion. This study first employed MTT assays to determine that the optimal concentration for treating HEK293 cells with purified recombinant Omp19 protein was 10 μg/mL, with a treatment duration of 12 h. Following extracellular stimulation, analysis via RT-qPCR, Western blot, and Annexin V-FITC techniques revealed that Brucella-Omp19 significantly induces apoptosis and inflammation in HEK293 cells. To further examine the intracellular role of Omp19 in apoptosis regulation, we established a Brucella TaqMan qPCR assay to determine its peak expression time. The standard curve confirmed a strong linear correlation between plasmid copy number and Ct values, with high specificity for Omp19 detection. Upon Omp19 overexpression in HEK293 cells, maximal expression was observed at 24 h. Analysis by RT-qPCR, Western blot, and Annexin V-FITC assays revealed that Brucella Omp19 also triggers apoptosis and inflammatory responses within HEK293 cells. In conclusion, Brucella-Omp19 induces apoptosis and inflammation in HEK293 cells through both extracellular and intracellular pathways, highlighting its critical role in host-pathogen interactions.