The role of STAT4 transcription factor in purified human Dendritic cells treated with interferon-beta, an autoimmune disease model

Abstract

STAT4 (Signal Transducer Activator of Transcription factor 4) polymorphisms are risk factors contributing to autoimmune disease. While STAT4 is dominant in promoting interferon-γ in T lymphocytes and NK cells, its existence and function in dendritic cells (DCs) are still absent. To characterize the STAT4 in human DCs, we isolated dendritic cells from buffy coat of healthy blood donors (n = 6). The dendritic cells are enriched and specifically sorted into 3 subtypes including plasmacytoid dendritic cells (pDCs, CD123+), CD8+ priming dendritic cells (cDC1, CLEC9A+), and CD4+ priming dendritic cells (cDC2/3, CD1c+) using magnetic separation and flow cytometry. By quantitative PCR and western blotting, the presence of STAT4 gene expression as well as phosphorylated STAT4 was determined in both IFN-β-induced dendritic cell maturation and immature dendritic cells ex vivo. In addition, we study the role of STAT4 by performing lisofylline inhibition and cytokine secretion. Regarding our analysis, the percentages of each dendritic cell subset are 0.07% for pDC, 0.04% for cDC1, and 0.31% for cDC2/3 in healthy blood donors. The IFN-β-enhanced STAT4 expression and phosphorylated STAT4 were prominent in mature dendritic cells, especially in cDC2/3, but not in pDC and cDC1 (p-value < 0.01). As cDC1 and cDC2/3 are responsible for T-cell priming, this indicated that IFN-β-induced STAT4 might be crucial in regulating dendritic cells producing IFN-γ. In addition, STAT4 and pSTAT4 were not inhibited by lisofylline pretreatment. Lastly, we found that the secretion of IFN-α2 was increased in cDC2/3 after IFN-β stimulation. Our study highlights the significance of STAT4 in dendritic cells which might be involved in the IFN-mediated autoimmune diseases.