POS0484 LUNG ORGANOIDS: A NOVEL APPROACH TO STUDY THE MOLECULAR PATHOLOGY OF PULMONARY FIBROSIS IN SYSTEMIC SCLEROSIS

Background Pulmonary fibrosis is one of the major manifestations in Systemic Sclerosis (SSc) associated with high mortality. Mesenchymal transformation of the airway epithelial cells has been implicated as one of the causes for developing pulmonary fibrosis. Though several animal models shed light towards some of these aspects, an in vitro airway epithelial model would provide a novel experimental platform for the understanding and molecular and genetic changes that occur in SSc associated pulmonary fibrosis. Objectives To establish a functional model for airway epithelium from patient with diffuse cutaneous SSc (dSSc)and healthy volunteers derived nasal stem cells. Subsequently to induce Epithelial Mesenchymal transformation (EMT). Methods Nasal stem cells harvested from healthy volunteers(HV) and dSSc patients were differentiated into ciliated airway epithelium in an Air -Liquid Interface (ALI) using a transwell system. 4 HV cultures were then stimulated with TGF beta (5ug/ml) for 10 days at a basal stage and when differentiated. Markers of mesenchymal transformation including loss of E cadherin, and gain of N cadherin, fibronectin and vimentin were analysed by flow cytometry and image stream, and mean expression intensities given as (MFI). Secreted Type 1 collagen and fibronectin were measured by ELISA. Results Ciliated epithelial cultures could successfully be established from nasal stem cells (Figure 1). TGF beta induced a phenotypic change in the epithelial cells towards a mesenchymal one in HV cultures. This was observed by significantly increased expression of fibronectin and vimentin and loss of expression of E cadherin on the ciliated cells with 7 days of stimulation with TGF beta at a basal stage (Figure 1b). When cells, stimulated with TGF beta for 7 days, were analysed at Day 35 a similar trend was seen in their Delta MFI (Figure 1c). Stimulating the ALI cultures with TGF beta for 20 days completely repressed epithelial cell growth and disrupted their microstructure. Figure 1. Conclusion This novel ALI differentiated Airway epithelial model serves as a functional organoid to test various pulmonary manifestations of Systemic Sclerosis. The ability to induce Epithelial Mesenchymal Transformation of these cultures provides a proof of concept for TGF beta mediated fibrosis in dSSc. Moreover, this model can be utilized to explore, at the cell and molecular level, the impact of various autoantibodies and therapeutics on epithelial cells. References [1]Mehmet Kesimer,1 Sara Kirkham,2 Raymond J. Pickles,3 Ashley G. Henderson,4 Neil E. Alexis,5 Genevieve DeMaria,1 David Knight,2 David J. Thornton,2 and John K. Sheehan1 Tracheobronchial air-liquid interface cell culture: a model for innate mucosal defense of the upper airways?; Am J Physiol Lung Cell Mol Physiol 296: L92–L100, 2009 Disclosure of Interests None declared.