Background:A fundamental role of mitochondria in systemic lupus erythematosus (SLE) was recently demonstrated (1). In brief, mitochondrial ROS participate in the formation of neutrophil extracellular traps (NETs) (2), while extrusion of cell-free mitochondria and highly oxidized interferogenic mtDNA causes disease in an animal model of SLE (3-5).Objectives:The diagnostic and prognostic value of cell-free DNA in SLE is still unknown. The aim of the present study was therefore to examine the clinical utility of cell-free DNA quantification as a non-invasive biomarker in SLE.Methods:Total DNA was isolated from platelet-free plasma samples of healthy individuals (HC) and consecutive SLE patients. Plasma and clinical data were collected at baseline and follow-up. Copy numbers were quantified by qPCR for mitochondrial (mt) DNA (ATP-6 gene) and nuclear (n) DNA (GAPDH gene).Results:Fifty-six HC (median age 48.3 ± 13.5, 64% female) and 103 SLE patients (median age 46.8 ± 15.8, 99% female, mean SLEDAI: 3 ± 4) were available for analysis.mtDNA levels were significantly elevated in SLE plasma (1.3x108 copies/ml plasma, 95% CI: 7.3x107 to 1.7 x108)), compared to HC plasma (8.6x106 copies/ml plasma, 95% CI: 6.9x106 to 1.0x107, p<0.0001). nDNA levels in contrast did not differ between SLE (8.3x106 copies/ml plasma, 95%CI: 5.9x106 to 1.4 x107) and HC (1.0x107 copies/ml plasma, 95%CI: 2.0x106 to 1.5 x107, p=0.61). Receiver operating characteristic curve analysis showed that a cut-off value of 1.9x107 mtDNA copy numbers differentiated between SLE and HC with 87.4% sensitivity, 94.6% specificity and an AUC of 0.95 (Figure 1a).mtDNA levels correlated with the SLE Disease Activity Index 2000 (SLEDAI-2K) (r=0.29, p=0.0026), less so also nDNA copy numbers (r=0.24, p=0.014). There was no association of mtDNA elevation with any particular type of SLE organ involvement and no correlation between mtDNA copy numbers in SLE plasma and dsDNA antibody levels.Follow-up data were available for 32 SLE patients (median follow-up 4.0 months, IQR: 4.0). delta mtDNA-levels robustly correlated with changes in SLEDAI-2K (r=0.51, p=0.0012, Figure 1b).Conclusion:The quantification of cell free mtDNA, but not nDNA copy numbers allows a sensitive and specific distinction between healthy individuals and patients with SLE. mtDNA levels correlate cross sectionally with disease activity in SLE patients and within individual SLE patients longitudinally with the SLEDAI. Plasma mtDNA quantification may therefore aid in the diagnosis of SLE and in monitoring SLE activity.References:[1]Riley JS, Tait SW. Mitochondrial DNA in inflammation and immunity. EMBO Rep. 2020;21(4):e49799.[2]West AP, Shadel GS. Mitochondrial DNA in innate immune responses and inflammatory pathology. Nat Rev Immunol. 2017;17(6):363-75.[3]Crow MK, Olferiev M, Kirou KA. Type I Interferons in Autoimmune Disease. Annu Rev Pathol. 2019;14:369-93.[4]Kim J, Gupta R, Blanco LP, Yang S, Shteinfer-Kuzmine A, Wang K, et al. VDAC oligomers form mitochondrial pores to release mtDNA fragments and promote lupus-like disease. Science. 2019;366(6472):1531-6.[5]Lood C, Blanco LP, Purmalek MM, Carmona-Rivera C, De Ravin SS, Smith CK, et al. Neutrophil extracellular traps enriched in oxidized mitochondrial DNA are interferogenic and contribute to lupus-like disease. Nat Med. 2016;22(2):146-53.Declaration of conflict of interest:UW is coinventor of patents owned by Freiburg University; NV is coinventor of patents owned by Freiburg University.Figure 1.(a) Receiver operating characteristic curve for mtDNA plasma concentrations to discriminate between HC and SLE patients. AUC: area under the curve. (b) Changes in plasma mtDNA levels in SLE patients correlate with the evolution of disease activity at follow-up.Disclosure of Interests:None declared
Neutrophil Extracellular Traps-DNase Balance and Autoimmunity
