Background/Purpose: Systemic lupus erythematosus (SLE) is a complex autoimmune disease involving multiple organ systems and periods of waxing and waning disease activity. The lack of well-defined disease biomarkers to predict disease flares hinders therapeutic optimization. Exosomes are secreted intraluminal vesicles involved in cell-cell communication and can be sampled from patients using minimally invasive methods. By identifying the role of exosomes in driving SLE disease activity, early biomarkers of flare and disease specific pathways can be identified.

Methods: Exosomes were isolated from plasma of healthy controls (n=10) and SLE patients with either elevated disease activity (SLEDAI≥6; n=5; SLE-hi) or low disease activity (SLEDAI≤4; n=7; SLE-lo) using Exoquick Isolation Reagent (SBI). Total RNA from plasma exosomes were extracted and purified using SBI’s Seramir Kit and libraries were constructed using a modified Illumina adapter method and sequenced on an Illumina MiSeq v3 instrument. Cleaned sequences were filtered and sequences with lengths ≥18 nt were aligned using Bowtie 2 against the human genome reference sequences. Hairpin and mature miRNA sequence hits were identified using BLAST in MiRBase. BLAST searches for transkingdom RNAs with known associations in SLE were also performed. Differentially expressed genes were determined using DESeq2 R package. Characterization of exosome size and quantity was determined by electron microscopy and EXOCET Assay (SBI), respectively. Further, isolated exosomes from SLE-hi and SLE-lo disease activity patients and healthy controls were co-cultured with healthy control PBMCS for 48 hours and cellular activation and cytokine production of PBMCs was measured by flow cytometry and ELISA. 

Results: SLE-hi patient exosomes had an increased median size (mdn 115.5nm) compared to SLE-lo patients (mdn 103.0nm) (p<0.0001), however, the abundance of exosomes did not differ between SLE disease activity or healthy controls (mdn 1.4×10¹²/ml). Exosome RNA libraries from SLE-hi patients had significantly more raw and clean RNA reads compared to healthy controls (p<0.0082) and SLE-lo patients (p<0.05). Further, SLE patients had 77 differentially expressed transcripts compared to healthy controls and 28 differentially expressed transcripts between high and low SLE disease activity (p<0.05) including Y RNAs and those associated with Epstein-Barr virus, transcriptional and post-transcriptional regulation, cytokinesis and apoptosis. Interestingly, when exosomes from SLE patients were incubated with healthy control PBMCs, SLE exosomes activated more CD4+ T cells (p=0.0196) and monocytes (p=0.0294). Further, PBMCs treated with SLE-hi exosomes produced greater amounts of IL-6 (p<0.05), but significantly reduced amounts of IL-10 (p<0.005) compared to PBMCs treated with exosomes from SLE-lo patients.

Conclusion: Our findings suggest that exosomes from active SLE patients have distinct characteristics and components that contribute to immune cell activation, inflammatory cytokine production, and impaired regulatory cytokine production which may contribute to lupus disease pathogenesis and activity.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/systemic-lupus-erythematosus-exosomes-contain-distinct-rna-transcripts-that-differentiate-disease-activity-and-modulate-cellular-function/