Differential regulation of 5-lipoxygenase and leukotriene-C4-synthase expression by IFNγ, IL-4 and IL-13 in human monocytes and macrophages from patients with atopic dermatitis.

INTRODUCTION

Skin lesions in atopic dermatitis (AD) are characterized by elevated levels of both Th1 and Th2 cytokines, along with increased concentrations of inflammatory mediators such as cysteinyl leukotrienes (CysLTs). The enzymes 5-lipoxygenase (5-LO/ALOX5) and leukotriene-C4-synthase (LTC4S), which are essential in CysLT biosynthesis, are highly expressed by macrophages. We aimed to investigate the expression of 5-LO/ALOX5 and LTC4S mRNA in lesional AD skin at single cell level. Furthermore, we analyzed the regulatory effects of the Th1 cytokine interferon gamma (IFNγ) and the Th2 cytokines IL-4, IL-13 on 5-LO/ALOX5 and LTC4S expression in monocytes and macrophages derived from AD patients and healthy volunteers.

METHODS

Single-cell RNA sequencing (scRNA-seq) data from lesional AD skin biopsies, as reported in a previously published study, were re-analyzed to evaluate 5-LO/ALOX5 and LTC4S mRNA expression. PBMCs from AD patients, healthy volunteers and anonymous donors were used to isolate monocytes. Macrophages were generated in the presence of GM-CSF or M-CSF for 10 days. Cells were stimulated with IFNγ, IL-4 or IL-13. 5-LO/ALOX5 and LTC4S mRNA expressions were quantified by q-PCR. Intracellular 5-LO/ALOX5 expression was assessed by immunocytochemistry.

RESULTS

Re-analysis of scRNA-seq data revealed high levels of 5-LO/ALOX5 and LTC4S transcripts in monocytes and macrophages. In-vitro, IFNγ induced 5-LO/ALOX5 mRNA expression in blood derived monocytes and macrophages from AD patients, and protein levels in both monocytes and macrophages from anonymous donors, whereas IL-4 and IL-13 suppressed its expression. Vice versa LTC4S mRNA expression was downregulated by IFNγ but upregulated by IL-13. Higher baseline mRNA expressions of 5-LO/ALOX5 and LTC4S were observed in blood derived monocytes from AD patients compared to cells from healthy volunteers .

CONCLUSION

We demonstrate that IFNγ enhances the 5-LO/ALOX5-catalyzed stage of CysLT synthesis, whereas IL-13 promotes the LTC4S dependent pathway in human monocytes or macrophages. These findings suggest a potential role for these cells in driving CysLT production in acute and chronic AD lesions and may give rise for future therapeutic interventions targeting this pathway.