IL-36R and IL-17A/F: Key pro-inflammatory signaling axes in autoimmune skin and barrier diseases

• Gene Information: The B-hIL36R/hIL17A/hIL17F mouse model features humanized genes for human IL36R (IL1RL2, chromosome 2q12.1), IL17A (chromosome 6p12.2), and IL17F (chromosome 6p12.2).
• Protein Expression: IL-36R is predominantly expressed on epithelial cells (especially keratinocytes) and barrier mucosal cells, as well as DCs and macrophages. IL-17A and IL-17F are key effector cytokines secreted mainly by activated Th17 cells and ILC3s, acting directly on keratinocytes, fibroblasts, and endothelial cells.
• Signaling Pathway: Binding of IL-36 agonists (α, β, γ) to IL-36R recruits IL-1RAcP to activate the MyD88-NF-κB/MAPK pathway. Simultaneously, IL-17A and IL-17F form homodimers or heterodimers (IL-17A/F) that signal through the IL-17RA/RC complex to activate NF-κB, MAPK, and C/EBP pathways. Together, these pathways act synergistically to strongly induce neutrophilic chemokines (CXCL1, CXCL8), antimicrobial peptides (S100A7/8/9, DEFB4), and pro-inflammatory cascades.
• Therapeutic Inhibition: Dual or combination targeting of the IL-36R and IL-17A/F axes represents a highly promising strategy for severe, refractory inflammatory skin and barrier disorders, including generalized pustular psoriasis (GPP), plaque psoriasis, hidradenitis suppurativa (HS), and severe asthma.

IL17A

• Exons 1-3 of the mouse Il17a gene that encode the extracellular domain were replaced by human IL17A exons 1-3 in B-hIL17A/hIL17F mice.
• The endogenous mouse promoter, 5' UTR, and 3' UTR regions are retained, allowing human IL17A expression to be driven by the native mouse Il17a promoter, while endogenous mouse Il17a transcription and translation are abolished.

IL17F

• Exons 2-3 of the mouse Il17f gene that encode the full-length protein were replaced by human IL17F exons 2-3 in B-hIL17A/hIL17F mice.
• The endogenous mouse promoter, 5' UTR, and 3' UTR regions are retained, allowing human IL17F expression to be driven by the native mouse Il17f promoter, while endogenous mouse Il17f transcription and translation are abolished.

IL36R

• A chimeric CDS that encodes the human IL36R signal peptide, extracellular domain, and transmembrane domain, along with the mouse IL36R cytoplasmic domain, was inserted immediately after the mouse Il36r ATG to replace exon 2 of the mouse Il36r gene.
• Chimeric IL36R protein expression is driven by the endogenous mouse Il36r promoter, while mouse Il36r gene transcription and translation are disrupted.

Note: This model was generated by cross-breeding B-hIL36R mice with B-hIL17A/hIL17F mice.

Strain specific analysis of IL1RL2 mRNA expression in C57BL/6 mice and homozygous B-hIL36R/hIL17A/hIL17F mice by RT-PCR.
Skin RNA was isolated from C57BL/6 mice (+/+) and homozygous B-hIL36R/hIL17A/hIL17F mice (H/H, H/H, H/H), then cDNA libraries were synthesized by reverse transcription, followed by PCR with mouse or human IL36R primers. Mouse Il36r mRNA was only detectable in wild-type mice. Human IL36R mRNA was exclusively detectable in homozygous B-hIL36R mice but not in wild-type mice.

Strain-specific IL17A expression analysis in wild-type C57BL/6 mice and homozygous B-hIL36R/hIL17A/hIL17F mice by ELISA. Serum was collected from wild-type C57BL/6 mice and homozygous B-hIL36R/hIL17A/hIL17F mice (H/H, H/H, H/H) (male, 9-week-old, n=3) stimulated with anti-mCD3ε and anti-mCD28 antibodies in vivo, and analyzed by ELISA with species-specific IL17A ELISA kits (Mouse IL-17A ELISA Kit, Proteintech, KE10020; LEGEND MAX Human IL-17A ELISA Kit, Biolegend, 433917). Mouse IL17A was detectable in wild-type mice. Human IL17A was detectable in homozygous B-hIL36R/hIL17A/hIL17F mice. Values are expressed as mean ± SEM. ND: not detectable.

Strain-specific IL17F expression analysis in wild-type C57BL/6 mice and homozygous B-hIL36R/hIL17A/hIL17F mice by ELISA. Th17 cell culture supernatants were collected from wild-type C57BL/6 mice (+/+) and homozygous B-hIL36R/hIL17A/hIL17F mice (H/H) (male, 9-week-old, n=3). Expression levels of mouse and human IL17F were analyzed by ELISA (anti-mouse IL17F ELISA kit: R&D Systems, DY2057; anti-human IL17F ELISA kit: Biolegend, 435707). Mouse IL17F was exclusively detectable in wild-type C57BL/6 mice. Human IL17F was exclusively detectable in homozygous B-hIL36R/hIL17A/hIL17F mice. Values are expressed as mean ± SEM. ND: not detectable.