Fig. 4

Effects of T cell EVs on apoptosis, oxidative stress, and inflammation in A549 cells were attenuated by PKC/NOX4 inhibition. A549 cells were stimulated with EVs isolated from CD4⁺ T cells isolated from healthy subjects treated with 10 μg/mL LPS (LTE) in the absence or presence of LXS-196, GLX351322, or NOX4 siRNA (taking siRNA to nonspecific sequence as control). A, B Flow cytometric analysis with Annexin V-FITC/PI double staining indicated that the elevation of cell apoptosis by LTE treatment was restored by LXS-196, GLX351322, or NOX4 RNAi. Representative plot images from flow cytometry (A) and statistical analysis (B) are shown. C, D DCFH-DA staining with flow cytometric analysis indicated that the elevation of ROS by LTE treatment was restored by LXS-196, GLX351322, or NOX4 RNAi. Representative images from flow cytometry (C) and statistical analysis (D) are shown. E The elevation of MDA content by LTE treatment was restored by LXS-196, GLX351322, or NOX4 RNAi. The reduction of SOD (F) and GPX activities (G) by LTE treatment was restored by LXS-196, GLX351322, or NOX4 RNAi. H The elevation of DAG content by LTE treatment was not affected by LXS-196, GLX351322, or NOX4 RNAi. I The elevation of PKC activity by LTE treatment was restored by LXS-196, but was not affected by GLX351322 or NOX4 RNAi. The increase of (J) mRNA expression and (K) secretion of TNF-α, IL-1β, and IL-6 by LTE treatment was restored by LXS-196, GLX351322, or NOX4 RNAi. Data presented as mean ± SD. ***P < 0.001 versus control. ^###P < 0.001 versus LTE + vehicle. ^ΔΔΔP < 0.001 versus LTE + siNC