Neutrophil dysregulation is implicated in the pathogenesis of systemic lupus erythematosus (SLE). SLE is characterized by elevated levels of a pathogenic neutrophil subset known as low-density granulocytes (LDGs). The origin and phenotypic, functional, and pathogenic heterogeneity of LDGs remain to be systematically determined. Transcriptomics and epigenetic assessment of lupus LDGs, autologous normal-density neutrophils, and healthy control neutrophils was performed by bulk and single-cell RNA sequencing and assay for transposase-accessible chromatin sequencing. Functional readouts were compared among neutrophil subsets. SLE LDGs display significant transcriptional and epigenetic heterogeneity and comprise 2 subpopulations of intermediate-mature and immature neutrophils, with different degrees of chromatin accessibility and differences in transcription factor motif analysis. Differences in neutrophil extracellular trap (NET) formation, oxidized mitochondrial DNA release, chemotaxis, phagocytosis, degranulation, ability to harm the endothelium, and responses to type I interferon (IFN) stimulation are evident among LDG subsets. Compared with other immune cell subsets, LDGs display the highest expression of IFN-inducible genes. Distinct LDG subsets correlate with specific clinical features of lupus and with the presence and severity of coronary artery disease. Phenotypic, functional, and pathogenic neutrophil heterogeneity are prevalent in SLE and may promote immune dysregulation and prominent vascular damage characteristic of this disease.

Type III IFN lambdas (IFN-λ) have recently been described as important mediators of immune responses at barrier surfaces. However, their role in autoimmune diseases such as systemic lupus erythematosus (SLE), a condition characterized by aberrant type I IFN signaling, has not been determined. Here, we identify a nonredundant role for IFN-λ in immune dysregulation and tissue inflammation in a model of TLR7-induced lupus. IFN-λ protein is increased in murine lupus and IFN-λ receptor (Ifnlr1) deficiency significantly reduces immune cell activation and associated organ damage in the skin and kidneys without effects on autoantibody production. Single-cell RNA sequencing in mouse spleen and human peripheral blood revealed that only mouse neutrophils and human B cells are directly responsive to this cytokine. Rather, IFN-λ activates keratinocytes and mesangial cells to produce chemokines that induce immune cell recruitment and promote tissue inflammation. These data provide insights into the immunobiology of SLE and identify type III IFNs as important factors for tissue-specific pathology in this disease.

Differences between female and male immunity may contribute to variations in response to infections and predisposition to autoimmunity. We previously reported that neutrophils from reproductive-age males are more immature and less activated than their female counterparts. To further characterize the mechanisms that drive differential neutrophil phenotypes, we performed RNA sequencing on circulating neutrophils from healthy adult females and males. Female neutrophils displayed significant up-regulation of type I IFN (IFN)-stimulated genes (ISGs). Single-cell RNA-sequencing analysis indicated that these differences are neutrophil specific, driven by a distinct neutrophil subset and related to maturation status. Neutrophil hyperresponsiveness to type I IFNs promoted enhanced responses to Toll-like receptor agonists. Neutrophils from young adult males had significantly increased mitochondrial metabolism compared to those from females and this was modulated by estradiol. Assessment of ISGs and neutrophil maturation genes in Klinefelter syndrome (47, XXY) males and in prepubescent children supported that differences in neutrophil phenotype between adult male and female neutrophils are hormonally driven and not explained by X chromosome gene dosage. Our results indicate that there are distinct sex differences in neutrophil biology related to responses to type I IFNs, immunometabolism, and maturation status that may have prominent functional and pathogenic implications.