Interleukin-17 pathway activation in Equus caballus supporting limb laminitis

Abstract Supporting Limb Laminitis (SLL) is a painful and crippling secondary complication of orthopedic injuries and infections in horses, often resulting in euthanasia. Due to altered weight bearing, SLL causes structural alternations and inflammation of the interdigitating layers of specialized epidermal and dermal tissues, the lamellae, which suspend the equine distal phalanx from the hoof capsule. Activation of the interleukin-17 (IL-17)-dependent inflammatory pathway is an epidermal stress response that contributes to physiologic cutaneous wound healing as well as pathological skin conditions. To test the hypothesis that IL-17 pathway activation is involved in equine epidermal lamellae in SLL, we analyzed the expression of the IL-17 receptor subunit A and 11 genes upregulated by IL-17 in lamellar tissue isolated from Thoroughbreds euthanized due to naturally occurring SLL and in age and breed matched non-laminitic controls. The IL-17 Receptor A subunit was expressed in both non-laminitic and laminitic tissues. In severe acute SLL (n=7) compared to non-laminitic controls (n=8), quantitative PCR demonstrated ∼20-100 fold upregulation of ß defensin 4 ( E. caballus gene DEFB4B ) and S100A9 genes. DEFB4B was also upregulated in developmental (n=8), moderate acute (n=7), and severe chronic (n=5) samples. By RT-PCR, S100A8, MMP9 , and PTSG2 (COX2) expression was upregulated in most or all severe acute SLL samples, whereas several other genes, CCL2, CxCL8, TNFα, IL6 and MMP1 were detected in some, but not all, severe acute samples. PTGS2, CCL2, TNFα and IL6 were also expressed in some, but not all, developmental and moderate acute disease stages. Moreover, expression of DEFB4 by in situ hybridization and calprotectin (S100A9/S100A8) protein by immunofluorescence was detected in keratinocytes, primarily in suprabasal cell layers, from SLL samples. These data support the hypothesis that the IL-17 inflammatory pathway is active in equine SLL, and that similarities exist between equine and human epidermal tissue responses to stresses and/or damage.