Objective In SLE, monocytes are instructed by type I interferon (IFNα) to be activated and produce inflammatory mediators. Metabolome studies in SLE patient sera have revealed distinct perturbations of lipid metabolism.^{1 2} However, the metabolic alterations of SLE monocytes under the effect of IFNα and their contribution to disease pathogenesis remain ill-defined. Our aim is to delineate the metabolic repercussion of monocytes in response to prolonged IFNα signaling in the context of SLE.

Methods RNA was extracted from CD14⁺ monocytes of active SLE patients and healthy donors and the latter were cultured in the presence or absence of recombinant IFNα. RNA sequencing was performed followed by differential gene expression and Gene Set Enrichment Analysis (GSEA). CD14⁺ monocytes were isolated from the abovementioned groups and cholesterol levels and lipid droplet formation were assessed by colorimetric assays and confocal microscopy. Cytokine release and antigen presenting capacity of IFNα-stimulated monocytes, in the presence or absence of metabolic inhibitors, were evaluated by ELISA and flow cytometry, respectively.

Results Transcriptome analysis in IFNα-treated CD14+ monocytes revealed significant enrichment of inflammation and lipid-related genes, which are implicated in cholesterol homeostasis. These molecular aberrations displayed significant overlap with differentially expressed genes (DEGs) from SLE CD14⁺ monocytes with IFNα^high gene signature as compared to their counterparts with IFNα^low gene signature. The direct effects of IFNα on cholesterol synthesis were confirmed by the increased intracellular cholesterol levels and enhanced lipid droplet formation both in IFNα-stimulated monocytes and in SLE monocytes with IFNα^high-signature (figure 1). Notably, the blockade of cholesterol biosynthesis was able to reduce the IFNα-mediated inflammatory phenotype of monocytes as evidenced by the reduced secretion of IL-6 and TNF-α, as well as reduced expression of the co-stimulatory molecules CD86 and CD40 (figure 2).

Conclusions Our data suggest that IFNα-mediated perturbations in lipid-related pathways may contribute to the development of SLE inflammation. These results may offer a mechanistic explanation for the increased disease burden within IFNα^high SLE patients.

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Abstract P19 Figure 1

(A) The levels of SIGLEC-1 were assessed by monitoring its expression using flow cytometry (n=3). (B) A representative confocal image of immunofluorescence microscopy depicting lipid droplets (green: BODIPY 493/503) is shown. The bar plot shows the number of lipid droplets per donor Healthy donors n=8, SLE patients n=10. Results are shown as mean ± SEM; *p < 0.05; **p < 0.01; ***p < 0.001.

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Abstract P19 Figure 2

(A) CD14+ primary monocytes from healthy donors were stimulated with IFNα (104 U/ml) for 18 hours in the presence of Fluvastatin (2μM) or Simvastatin (10μM). Cell culture supernatants were assayed for human IL-6 (n=7) and TNFα (n=3) using an ELISA. (B) CD14+ primary monocytes from healthy donors were stimulated with IFNα (104 U/ml) for 18 hours in the presence of Fluvastatin (2μM) or Simvastatin (10μM). Extracellular levels of CD86 and CD40 were assessed by monitoring CD86 and CD40 expression using flow cytometry (n=6). Results are shown as mean ± SEM; *p < 0.05; **p < 0.01; ***p < 0.001

References

1. Gkirtzimanaki K, et al. ‘IFNα Impairs Autophagic Degradation of mtDNA Promoting Autoreactivity of SLE Monocytes in a STING-Dependent Fashion,’ Cell Rep, Oct. 2018;25(4):921–933.e5, doi: 10.1016/j.celrep.2018.09.001.

2. Stathopoulou C, Nikoleri D, Bertsias G. Immunometabolism: an overview and therapeutic prospects in autoimmune diseases. Immunotherapy, Jun. 2019, 11(9):813–829, doi: 10.2217/imt-2019–0002.

3. Lee M-S, Bensinger SJ. Reprogramming cholesterol metabolism in macrophages and its role in host defense against cholesterol-dependent cytolysins. Cell Mol Immunol, Mar. 2022;19(3):327–336, doi: 10.1038/s41423–021-00827–0.