Fig 2: Topical treatment by dexamethasone, vitamin D3, and acitretin suppress IL23Ap19 and EBI3 expressions in NHEKs. Normal human epidermal keratinocytes (NHEKs) were stimulated with the triple combination of TNF-α (50 ng/mL), IL-17A (50 ng/mL), and IFN-γ (50 ng/mL), and simultaneously treated with (A,B) dexamethasone (10-8,10-7, or 10-6 M), (C,D) 1,25(OH)2 vitamin D3 (10-8,10-7, or 10-6 M), or (E,F) acitretin (10-8,10-7, or 10-6 M) for 24 h. The relative mRNA expressions of IL23Ap19 and EBI3 to GAPDH were analyzed with quantitative real-time PCR (qPCR). ** p < 0.01, *** p < 0.001. T, TNF- α; 17A, IL-17A; G, IFN-γ; Triple, T+17A+G; DEX, dexamethasone; VitD, 1,25(OH)2 vitamin D3; Aci, acitretin.

Fig 3: Expressions of IL-23Ap19 and EBI3 are upregulated in psoriatic keratinocytes. Expressions of IL-23Ap19 and EBI3 were examined in normal skin and lesional psoriatic skin sections by immunohistochemistry with polyclonal rabbit anti-IL-23p19 antibody (A,B) and monoclonal mouse anti-EBI3 antibody (C,D).

Fig 4: IL-23Ap19- and/or EBI3-including cytokines in epidermal keratinocytes. IL-23Ap19 and EBI3 were detected by qPCR, ELISA, and LC-Ms/Ms. IL-27Ap28 and IL-12Ap35 were undetected by qPCR. IL-23(p19/p40), IL-27(p28/EBI3), IL-35(p35/EBI3), and IL-39(p19/EBI3) were undetected by ELISA or LC-Ms/Ms. This result raises the possibility that IL-23Ap19 and EBI3 produced from epidermal keratinocytes exist as a monomer or homopolymer, such as a homodimer or homotrimer. ?, Not yet detected and possibility.

Fig 5: Proteins detected by immunoprecipitation with tildrakizumab or anti-human EBI3 antibody and LC-Ms/Ms. Culture supernatant of NHEKs stimulated with the triplet mixture of cytokines (50 ng/mL, TNF-α, IL-17A and IFN-γ) for 48 h were immunoprecipitated with (A) humanized anti-human IL-23Ap19 antibody or (B) rabbit anti-human EBI3 antibody and magnetic beads. The detected proteins were analyzed by liquid chromatography–electrospray tandem mass spectrometry (LC-Ms/Ms). The non-specific proteins detected in common with the control were removed.