Relaxin inhibits fibrogenesis in human lung slices: implications for ipf.

Abstract

Introduction/Aim: Interstitial fibrosis is the primary pathological feature of idiopathic pulmonary fibrosis (IPF). The peptide hormone relaxin inhibits collagen synthesis by human lung fibroblasts and reverses established fibrosis in mouse models of fibrotic lung disease, acting via the RXFP1 receptor (Lam et al., Front Pharmacol, 2018). Human precision-cut lung slices (hPCLS) treated with a 'cocktail' of IPF-relevant mediators to induce fibrosis (Alsafadi et al., AJP Lung, 2017) provide a novel tissue-based model to compare effects of relaxin with current IPF drug pirfenidone. Aim(s): Assess anti-fibrotic efficacy of relaxin in hPCLS and RXFP1 expression in IPF. Method(s): Multiple hPCLS (500 mum thick) were prepared from agarose-inflated non-IPF lung resections or donor lungs not used for transplant (n = 5). PCLS were treated for 120 h with fibrotic cocktail (FC = TGFbeta, LPA, PDGF-AB and TNFalpha) +/- relaxin (100 nM) or pirfenidone (500 muM). Fixed PCLS were stained with Masson's Trichrome for quantitation of collagen. Immunohistochemical staining for RXFP1 in donor and IPF lung sections (n = 10/group) was quantified using a positive pixel count algorithm in Aperio Imagescope. Protein expression of RXFP1 was evaluated using western blotting of donor and IPF fibroblasts (n = 3/group). Result(s): Treatment with FC increased collagen deposition almost 3-fold in hPCLS (%area: untreated 2.8 +/- 0.9; FC 7.0 +/- 1.4; p < 0.05). Both drugs completely prevented the fibrotic response to FC (FC + relaxin 3.4 +/- 0.8, FC + pirfenidone 2.0 +/- 0.4, NS cf untreated). There was no significant difference in RXFP1 expression between donor and IPF lung sections or fibroblasts. RXFP1 was localised to most resident lung cell types especially in the airways, and detected in usual interstitial pneumonia fibrotic lesions in IPF lungs. Conclusion(s): The inhibitory effects of relaxin on fibrogenesis in hPCLS were established, evident at 5000-fold lower concentrations than pirfenidone. Expression of RXFP1 was maintained as a therapeutic target in IPF. Further investigations to assess relaxin-mediated reversal of established fibrosis in hPCLS from IPF lungs are warranted to support clinical translation.