NOX5 and p22phox are 2 novel regulators of human monocytic differentiation into dendritic cells

Abstract

Dendritic cells (DCs) are a heterogeneous population of professional antigen-presenting cells and are key cells of the immune system, acquiring different phenotypes in accordance with their localization during the immune response. A subset of inflammatory DCs is derived from circulating monocytes (Mo) and has a key role in inflammation and infection. The pathways controlling Mo-DC differentiation are not fully understood. Our objective was to investigate the possible role of nicotinamide adenine dinucleotide phosphate reduced form oxidases (NOXs) in Mo-DC differentiation. In this study, we revealed that Mo-DC differentiation was inhibited by NOX inhibitors and reactive oxygen species scavengers. We show that the Mo-DC differentiation was dependent on p22phox, and not on gp91phox/NOX2, as shown by the reduced Mo-DC differentiation observed in chronic granulomatous disease patients lacking p22phox. Moreover, we revealed that NOX5 expression was strongly increased during Mo-DC differentiation, but not during Mo-macrophage differentiation. NOX5 was expressed in circulating myeloid DC, and at a lower level in plasmacytoid DC. Interestingly, NOX5 was localized at the outer membrane of the mitochondria and interacted with p22phox in Mo-DC. Selective inhibitors and small interfering RNAs for NOX5 indicated that NOX5 controlled Mo-DC differentiation by regulating the JAK/STAT/MAPK and NFκB pathways. These data demonstrate that the NOX5-p22phox complex drives Mo-DC differentiation, and thus could be critical for immunity and inflammation.

NOX5: Molecular biology and pathophysiology

“…The early NOX5 studies demonstrated that mRNA of NOX5 is expressed in pachytene spermatocytes of the testis and in B‐ and T‐lymphocyte‐rich areas of the lymph nodes and spleen (Bánfi et al., ; Cheng et al., ). Whether NOX5 is expressed and functionally active in circulating phagocytic cells has been debated, because original studies failed to identify NOX5 in peripheral lymphocytes, whereas recent studies indicate that NOX5 is expressed in human monocytes and macrophages (Manea et al., ; Marzaioli et al., ) and that NOX5 regulates human monocyte differentiation into dendritic cells (Marzaioli et al., ). However, most of those studies were performed in monocyte/macrophage leukaemia cell lines (Marzaioli et al., ), and the physiological role of neutrophil NOX5 is unclear.…”

“…Whether NOX5 is expressed and functionally active in circulating phagocytic cells has been debated, because original studies failed to identify NOX5 in peripheral lymphocytes, whereas recent studies indicate that NOX5 is expressed in human monocytes and macrophages (Manea et al, 2015;Marzaioli et al, 2017) and that NOX5 regulates human monocyte differentiation into dendritic cells (Marzaioli et al, 2017). However, most of those studies were performed in monocyte/macrophage leukaemia cell lines (Marzaioli et al, 2017), and the physiological role of neutrophil NOX5 is unclear. In the developing zebrafish, Nox5 is expressed throughout the CNS and, together with Nox2, it might be important in neurodevelopment and regeneration (Weaver et al, 2018;Weaver, Leung, & Suter, 2016).…”

Guidelines for the Detection of NADPH Oxidases by Immunoblot and RT-qPCR

“…Human embryonic kidney cell line (HEK 293 cells) or any other cell line with very low or no endogenous expression of NOX/DUOX enzymes.24. DMEM medium, high glucose (4.5 g/L), containing 10% FBS, or similar 25…”

Calcium-dependent reactive oxygen formation and blood-brain barrier breakdown by NOX5 limits post-reperfusion outcome in stroke

“…In humans, Nox5 is broadly expressed in blood vessels, primarily smooth muscle cells, microvascular endothelial cells, fibroblasts [6,8,9] but also in testis [8] and spleen, specifically in monocytes, macrophages [7,10], B- and T-cells [11]. To mimic the physiological human expression pattern of NOX5 in mice, we created a mouse line called NOX5 KI (knock-in) bearing the human NOX5 gene in the Hprt locus and under control of the Tie2 promoter (Fig 1A-B).…”