Date: Monday, November 9, 2015
Session Type: ACR Concurrent Abstract Session
Session Time: 2:30PM-4:00PM
Background/Purpose: Dysregulation in the formation of neutrophil extracellular traps (NETs) may contribute to modification and externalization of autoantigens and to organ damage in diverse autoimmune diseases including systemic lupus erythematosus (SLE). In SLE, a distinct subset of proinflammatory low-density granulocytes (LDGs) displays an exuberant capacity to form NETs in the absence of added stimulation. NETosis appears to require generation of reactive oxygen species (ROS). However, whether the process is strictly NADPH-oxidase dependent is unclear. Indeed, the role of other important sources of ROS like the mitochondria, in autoimmune NETosis, remains to be determined. This is particularly relevant as lack of a functional NADPH-oxidase, as seen in chronic granulomatous disease (CGD), is associated with enhanced predisposition to chronic inflammation and autoimmunity including SLE.
Methods: To clarify the contribution of mitochondrial ROS in the context of autoimmunity, mitochondrial activation and ROS were quantified in lupus LDGs by fluorescent microscopy and flow cytometry. Enrichment in oxidized mitochondrial DNA (ox-mitDNA) in isolated LDG NETs versus healthy control NETs was quantified with 8-Oxo-2′-deoxyguanosine antibodies and by calculating 16S/18S ratio by immunoprecipitation and qPCR. Various inhibitors of mitochondrial ROS and other sources of ROS were tested in lupus LDGs. Similar experiments were performed in neutrophils isolated from patients with CGD, to assess the role of NADPH-oxidase independent sources of ROS production. The type I IFN serum signature was quantified by assessing induction of IFN-inducible genes (ISGs) in THP1 cells. Circulating NET products were quantified by ELISA. Finally, the mitochondrial ROS inhibitor mitoTEMPO was administered systemically, continuously to lupus-prone female MRL/lpr mice for 7 weeks and, at euthanasia, lupus clinical phenotype, induction of ISGs, inflammatory cytokines and NETosis were quantified.
Results: Lupus LDGs displayed prominent enhancement of mitochondrial ROS synthesis when compared to healthy controls and lupus normal-density granulocytes. Lupus LDGs spontaneously formed NETs highly enriched in ox-mitDNA that significantly enhanced ISGs in monocytic cell lines. Emphasizing the role of mitochondria-derived ROS in autoimmune NETosis, CGD patients displayed an LDG population with exuberant spontaneous NET formation through enhanced mitochondrial ROS production and also had elevated type I IFN signature. There was a significant correlation between the development of autoimmunity in CGD patients and the serum type I IFN activity. Supporting the in vitro data, both lupus and CGD patients displayed evidence of enhanced in vivo NETosis. Importantly, lupus-prone mice that received mitoTEMPO displayed a significant decrease in proteinuria, autoantibodies, ISGs and proinflammatory cytokines, in association with hampered NETosis.
Conclusion: Mitochondrial ROS synthesis and ox-mitDNA appear to be key drivers of autoimmunity and proinflammatory responses in SLE and CGD and may represent novel therapeutic targets in these diseases.
To cite this abstract in AMA style:Blanco LP, Lood C, Purmalek M, DeRavin SS, Smith CK, Carmona-Rivera C, Malech H, Ledbetter J, Elkon KB, Kaplan MJ. Mitochondrial Reactive Oxygen Species Modulate Autoimmunity in Systemic Lupus Erythematosus [abstract]. Arthritis Rheumatol. 2015; 67 (suppl 10). https://acrabstracts.org/abstract/mitochondrial-reactive-oxygen-species-modulate-autoimmunity-in-systemic-lupus-erythematosus/. Accessed April 13, 2021.
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