Contribution of macrophages and naive T cells during infection with E. coli in presence of exogenous cytokines on the autophagy-apoptosis pathway.

Escherichia coli (E. coli), a gram-negative bacterium typically found in the gastrointestinal tract, encompasses pathogenic strains that can lead to serious infections such as foodborne illnesses, sepsis, and urinary tract infections. The interaction between E. coli and immune cells, especially macrophages and T lymphocytes, is crucial for bacterial survival and evasion of the immune response. In this research, we utilized an in vitro co-culture model involving primary murine splenic macrophages and T cells to explore the contribution of different cytokines on autophagy, apoptosis, and phagocytic activity during E. coli infection. The cytokine administered included IFN-γ, IL-12, IL-4, IL-10, and their combinations. Flow cytometric analysis indicated a consistent increase in the proportion of macrophages compared to lymphocytes across all cytokine treatment conditions. Regarding the macrophage signalling pathway, IFN-γ+IL-12 fostered a proinflammatory environment characterized by a cytokine amplification loop and heightened ROS levels, which enhanced apoptosis and disrupted autophagy, as demonstrated by increased cleaved caspase-3 and decreased LC3-II and Beclin1 expression. Conversely, IL-4+ IL-10 facilitated an anti-inflammatory phenotype, leading to suppressed IL-12 production, lowered ROS levels, enhanced STAT3 and Nrf2 signalling, and a restoration of autophagic flux. Importantly, co-stimulation with IFN-γ and IL-4 resulted in conflicting signalling, where IL-4 alone was inadequate to mitigate the autophagy impairment caused by IFN-γ. These results underscore how varying cytokine environments influence macrophage functionality during E. coli infection, particularly by modulating the interplay between apoptosis and autophagy, which may have significant implications for the regulation of immune responses and bacterial persistence.