Document Type
Dissertation - Open Access
Award Date
2025
Degree Name
Doctor of Philosophy (PhD)
Department / School
Chemistry and Biochemistry
First Advisor
Adam D. Hoppe
Abstract
CD47-SIRPα signaling is a well-known immune checkpoint that suppresses macrophage-mediated antibody-dependent cellular phagocytosis (ADCP). In this study, we explored how CD47 influences reactive oxygen species (ROS) production driven by Fcγ receptor (FcγR) signaling. Our results show that macrophages produce significantly more ROS when encountering IgG coated target cells lacking CD47.Polarization with IFNγ further amplified this effect. ThusCD47-SIRPα interactions limit phagocytosis and suppress oxidative ROS production by macrophages. We found that CD47 blockade and IFNγ stimulation enhanced ADCP through distinct mechanisms. CD47 blockade removed the inhibitory "don't eat me" signal supporting ADCP and ADROS. Similarly, IFNγ enhances ADCP by upregulating FcγRI (CD64) and FcγRIV. Despite significant increases in ROS following IFNγ treatment and CD47 blockade, we found that ROS production does not play a major role in ADCP efficiency of B cells and B lymphoma cells. This was further supported by in vivo experiments demonstrating that NADPH oxidase-derived ROS is not required for macrophage-mediated B cell depletion, indicating that ADCP can occur independently of oxidative burst activity. To better understand how IFNγ-mediated ADCP is regulated, we performed a CRISPR-Cas9 knockout screen in macrophages engulfing opsonized target cells. This screen confirmed that FcγRI (Fcgr1) contributes to ADCP in IFNγ polarized macrophages, whereas FcγRIV (Fcgr4) was not among the top regulators. Further mechanistic studies demonstrated that interactions between CD47 on target cells and SIRPα on macrophages are the primary regulators of NADPH oxidase activity, whereas cis interactions between CD47 and SIRPα both on the macrophage surface had no effect. Additionally, genetic ablation of SHP1, a phosphatase that associates with SIRPα, did not inhibit NADPH oxidase function, but rather it was essential for ROS production revealing a complex role in CD47-SIRPa signaling. Overall, our study provides new insights into how CD47-SIRPα signaling, Fc receptor upregulation, and ROS production shape macrophage effector functions. These findings have important implications for macrophage-targeted therapies in cancer, inflammatory diseases, and immune regulation.
Publisher
South Dakota State University
Recommended Citation
Akter, Seuly, "Target Cell-Associated Cd47 Suppresses NADPH Oxidase Activity in Macrophages During Antibody Dependent Cellular Phagocytosis" (2025). Electronic Theses and Dissertations. 1502.
https://openprairie.sdstate.edu/etd2/1502
Rights Statement
