Role of Reactive Oxygen Species in Lymphocyte Development and Function

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This program explores roles of reactive oxygen species (ROS) as specific signaling molecules in B and T lymphocytes through genetic manipulation of NOX/DUOX family NADPH oxidases. These enzymes catalyze NADPH-dependent reduction of molecular oxygen to generate superoxide or hydrogen peroxide. Phagocytes produce large amounts of ROS in response to infectious or inflammatory stimuli through the prototypic NADPH oxidase containing gp91phox (a.k.a., NOX2). Although originally understood as an anti-bacterial mechanism deployed by phagocytes, our research revealed that ROS intentionally generated by several NOX family members play specific signaling roles in B cell receptor (BCR)-stimulated B cells and T cell receptor (TCR)-stimulated T cells. Our studies in lymphocytes are exploring roles of NOX family members in adaptive immune responses to diverse pathogens as well as in autoimmunity or immunodeficiencies. We recently showed that DUOX1- and NOX2-based NADPH oxidases serve distinct signaling functions in splenic B lymphocytes co-stimulated ex vivo with interleukin-4 (IL-4) and anti-IgM. NOX2-deficient B cells showed enhanced apoptosis and diminished IgM, IgG1 and IgG2a production relative to wild type or DUOX1-/- cells. DUOX1-deficient cells exhibited enhanced proliferation, which was correlated with enhanced expression of several BCR signaling intermediates (Akt, BCAP and RGS16). In 2017, we extended the above findings by examining whole animal responses to immunization with T cell dependent and independent antigens (nitrophenyl-keyhole limpet hemocyanin (NP-KLH) and nitrophenyl-lipopolysaccharide (NP-LPS), respectively). No significant differences in serum anti-NP-LPS (IgM, 7 days post-immunization) or anti-NP-KLH (IgG, 14 days post-immunization) were detected when comparing these responses in DUOX1-deficient, NOX2-deficient and wild type mice. However, immuno-histochemical staining of spleens from immunized mice revealed distinct differences in DUOX1 knockout mice, which showed markedly enhanced numbers of follicles with intense reactive germinal center and proliferation marker staining relative to that observed in wild type and NOX2 knockout mouse spleens. These findings are consistent with the enhanced stimulated splenic B cell proliferation observed in vitro with DUOX1 knockout mice. Together, our findings suggest that DUOX1 can suppress B cell proliferation, but may have little influence on systemic immunoglobulin production or class switching. Given the unique induction of DUOX1 by IL-4 in B lymphocytes, future work will examine tissue-specific adaptive immune functions of DUOX1 in response to infection.