Human Spinal Enthesis Comparative Biology of IL-17F and IL-17A Reveals Greater T-cell IL-17F Induction and IL-23 Regulation

Nicole McDermott^1*Ala Altaie¹Tom Macleod¹Charlie Bridgewood¹Peter Loughenbury²Robert Dunsmuir²Almas Khan²Vishal Borse²Abhay Rao¹Avneet Manghera³Stevan Shaw³Dennis McGonagle^4*

• ¹University of Leeds Leeds Institute of Rheumatic and Musculoskeletal Medicine, Leeds, United Kingdom
• ²Leeds Teaching Hospitals NHS Trust, Leeds, United Kingdom
• ³UCB Pharma Ltd, Slough, United Kingdom
• ⁴Leeds Biomedical Research Centre, Leeds, United Kingdom

Objective-Although IL-17F is important in psoriasis, the rudimentary biology of IL-17F at the human enthesis remains undefined. We aimed to characterise IL-23-dependent and independent IL-17F production from entheseal innate and adaptive T-cells and determine the impact of IL-17F on entheseal stromal function and mesenchymal stem cell (MSC) osteogenesis. Methods-Anti-CD3 and anti-CD28 activated human spinal entheseal T-cells were immunophenotyped with multi-parameter flow cytometry and 36-marker CyTOF with cytokine profiling, including IL-17A, IL-17F and TNF (n=10). IL-17A, IL-17F and TNF stimulation of entheseal MSC stromal function was evaluated by RNA-seq and measuring CCL20 protein expression following stimulation with TNF in combination with IL-17A or IL-17F. The osteogenic effects of IL-17A and IL-17F on MSC differentiation were assessed. Results-Inducible IL-17A and IL-17F expression was predominantly from CD4 T-cells and γδ T-cells, with higher levels IL-17F at 72hrs. IL-23 significantly increased IL-17F (p=<0.0001) but not IL-17A. Either IL-17A or IL-17F in combination with TNF dramatically upregulated CCL20 protein expression and substantially changed entheseal stromal transcriptome, with differences in gene expression and pathway activation seen between IL-17A or IL-17F stimulation. However, entheseal MSC osteogenesis was not significantly changed. Conclusions-Differential induction of IL-17A and IL-17F from innate and adaptive entheseal T-cells with further significant IL-17F augmentation by IL-23, but not IL-17A. Furthermore, synergistic effects of IL-17A or IL-17F and TNF on stromal function provides the basis for further enthesis IL-17F biology interrogation.