Background Systemic sclerosis (SSc) is an autoimmune connective tissue disease characterised by fibrosis, vascular dysfunction and abnormal activation of immune cells. We and others have shown that monocytes along with fibroblasts play an important role in the production of pro-fibrotic factors such as IL-6 and TIMP-1 (tissue-inhibitor of metalloproteinase-1). Importantly, the balance between TIMP-1 and MMPs is altered in SSc leading to abnormal ECM formation. However, the exact factors which drive pro-fibrotic TIMP-1 in monocytes have not been fully established. Previous studies showed that AP-1 transcription factors including Fra-2 and JunD are involved in TIMP-1 gene expression in hepatic stellate cells contributing to liver fibrosis. Also Fra-2 transgenic mice develop a proliferative vasculopathy of the lung and skin fibrosis resembling similar disease manifestations seen in SSc patients.

Objectives The aim of this study is to determine the role of Fra-2 in TIMP-1 production. We also plan to determine if TIMP-1 is a direct target gene of Fra-2 upon TLR8 stimulation in SSc monocytes. Additionally, we want to investigate if altered Fra-2 signalling results in enhanced TIMP-1 production thus contributing to ECM deposition and fibrosis development observed in systemic sclerosis.

Methods 20 patients with SSc, one IRAK4 deficient patient and 20 HC (healthy control) were included in this study. Peripheral blood monocytes were further separated by CD14+ microbeads. Expression of TIMP-1 and Fra-2 by monocytes was determined by ELISA, qRT-PCR in response toTLR8 agonists (ssRNA).

Results The results from our group demonstrated that circulating monocytes from SSc patients contribute to the imbalance between TIMP-1 and MMPs and to an increased pro-fibrotic IL-6 production upon TLR8 agonist stimulation (ssRNA). Interestingly, monocytes from an IRAK-4 (a key kinase in TLR-signalling) deficient patient did not produce IL-6 and TIMP-1. In addition, the results showed that the basal gene expression level of Fra-2 in SSc monocytes is higher than in healthy or IRAK-4 deficient monocytes. Interestingly, TLR8 activation induced even stronger Fra-2 gene expression and correlated with increased expression of TIMP-1 in SSc monocytes, suggesting that the TLR signalling pathway may be involved in Fra-2 mediated TIMP-1 induction and therefore promotes fibrosis. Also, we plan to pre-treat monocytes with the chemical AP-1 inhibitor (T-5224) and assess both gene and protein level of TIMP-1 upon TLR8 activation.

Conclusions This study helps to understand the molecular mechanisms of Fra-2 mediated fibrogenesis in SSc.

Disclosure of Interest None Declared