OP0242 Role of MRTF-A Pathway in Scleroderma-Related Fibrosis

Background MRTF-A is a 120kDa transcription factor widely expressed and normally sequestered in the cytosol by binding to G actin. Following actin polymerisation downstream of Rho signalling, or mechanosensing, MRTF-A is released and functions as a signalling molecule partnering serum response factor (SRF) and influencing gene expression via CARG elements in the promoter region of genes. Genes expressing CARG like elements induced by MRTF-A/SRF include CTGF and type I collagen. MRTF-A is also implicated in vascular remodelling and epithelial to mesenchyme transition (EMT). The MRTF-A/SRF axis is highly relevant to fibrosis in systemic sclerosis (SSc). Methods MRTF-A signal transduction was studied in healthy control and SSc fibroblasts. The MRTF-A/SRF small molecule inhibitor CCG1423 was used to block MRTF-A in vitro. SSc fibrosis responses were modelled by collagen gel contraction, CTGF, and type I collagen expression. MRTF-A signalling was assayed by Western blotting of nuclear and cytoplasmic extracts. Wound healing and fibrosis was studied in an MRTF-A knockout mouse (Ollsen lab) and wild type controls. Immunochemistry looking for nuclear localisation of MRTF-A was used to determine presence of active signalling in SSc involved skin biopsy material and healthy control tissue. Results SSc fibroblasts showed enhanced nuclear localisation of MRTF-A at 8 hours following exposure to TGFβ (4ng/ml) not seen in healthy control fibroblasts. Immunochemistry of SSc skin biopsy material revealed enhanced nuclear localisation in dermal fibroblast like cells, keratinocytes within the epidermis, as well as in perivascular cells (pericytes). Following excisional wounding (4mm punch biopsy, basal wound area 12.6 mm2) MRTF-A mice wounds failed to close normally and increased in size during days 1-7, wound area decreasing by day 11. When compared to wild type controls MRTF-A knockout wounds were enlarged at day 7 (wild type area 6mm2, knockout area 12.4 mm2, p