Yuanfeng Zhu,Shijun Fan,Ning Wang,Xiaoli Chen,Yongjun Yang,Yongling Lu,Qian Chen,Jiang Zheng,Xin Liu
Activation of NADPH oxidase 2 (NOX2) triggers reactive oxygen species (ROS) generation, both of which are essential for robust microbial clearance by phagocytes. However, it is unknown whether inhibition of NOX2 activation or ROS generation affects cellular phagocytosis. Here, we found that the classic NOX2 inhibitor diphenyleneiodonium (DPI) induced uptake of E. coli by murine peritoneal macrophages through enhancing phagocytosis, and this effect was temperature-sensitive and attenuated by cytochalasin D as well as chemical inhibition of Syk and PLCγ, two downstream kinases involved in actin polymerization during phagocytosis. DPI also decreased the production of TNF-α and IL-6 resulting from E. coli stimulation. The DPI-induced enhancement of phagocytosis was independent of NOX2 inhibition or ROS generation but depended on increased intracellular calcium and activation of the p38 MAPK signaling pathway. Furthermore, DPI enhanced bacterial elimination and ameliorated inflammation in E. coli -infected mice, leading to improved survival. Our results demonstrate that DPI facilitates ROS-independent bacterial phagocytosis by macrophages through activation of calcium and p38 MAPK signaling pathways.
The incomplete degradation of tumour cells by macrophages (Mφ) is a contributing factor to tumour progression and metastasis, and the degradation function of Mφ is mediated through phagosomes and lysosomes. In our preliminary experiments, we found that overactivation of NADPH oxidase 2 (NOX2) reduced the ability of Mφ to degrade engulfed tumour cells. Above this, we screened out liquiritin from Glycyrrhiza uralensis Fisch , which can significantly inhibit NOX2 activity and inhibit tumours, to elucidate that suppressing NOX2 can enhance the ability of Mφ to degrade tumour cells. We found that the tumour environment could activate the NOX2 activity in Mφ phagosomes, causing Mφ to produce excessive reactive oxygen species (ROS), thus prohibiting the formation of phagolysosomes before degradation. Conversely, inhibiting NOX2 in Mφ by liquiritin can reduce ROS and promote phagosome-lysosome fusion, therefore improving the enzymatic degradation of tumour cells after phagocytosis, and subsequently promote T cell activity by presenting antigens. We further confirmed that liquiritin down-regulated the expression of the NOX2 specific membrane component protein gp91 phox, blocking its binding to the NOX2 cytoplasmic component proteins p67 phox and p47 phox, thereby inhibiting the activity of NOX2. This study elucidates the specific mechanism by which Mφ cannot degrade tumour cells after phagocytosis, and indicates that liquiritin can promote the ability of Mφ to degrade tumour cells by suppressing NOX2.
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