POS0482 LONG NON-CODING RNA H19X IS A MEDIATOR OF ENDOTHELIAL CELL ACTIVATION IN SYSTEMIC SCLEROSIS

Background In one of our previous studies, we demonstrated that long non-coding RNA (lncRNA) H19X plays a crucial role in the development TGFβ driven fibrosis systemic sclerosis (SSc) and other fibrotic diseases 1 . Objectives To define functional relevance endothelial cell (EC) activation as decisive process SSc vasculopathy. Methods Correlation expression microvascular gene signature was computed on bulk RNA-Seq data derived from skin biopsies patients enrolled multicentre Prospective Registry Early Systemic Sclerosis cohort (PRESS, n=48 vs. n=33 healthy controls, HCs). Single sequencing (scRNA-seq) were collected 27 diffuse cutaneous (dcSSc) 10 HC biopsies. cells barcoded encapsulated droplets using 10X Genomics system. After cDNA synthesis, libraries prepared sequenced Illumina NovaSeq-500 platform. Seurat package R (v.3.0) used to perform analysis. EC identified by enrichment markers CLDN5 , VWF PECAM1 One thousand five hundred eighty-three 3398 patients, respectively. Cells analysed for markers. Additionally, differential pathway analysis between expressing negative carried out. The function investigated human dermal (HDMEC) silencing, locked nucleic acid antisense oligonucleotides (LNA GapmeRs). Gene measured qPCR. Protein levels adhesion molecules Western Blot. Endothelial evaluated co-culture HDMEC fluorescently labelled peripheral blood mononuclear (PBMCs). Results found significantly upregulated PRESS (p<0.0001). positively correlated with all subjects (SSc HC, R=0.43, p<0.0001), confirming is expressed this type. determine if might be an important factor dysfunction scRNAseq performed. This revealed significant upregulation compared (p=0.0095). several subclusters including arterial ( SEMA3G HEY1 ), capillary CA4 RGCC venous ACKR1 VCAM1 ) lymphatic PROX1 LYVE1 ). displayed highest injured EC. Co-expression scRNA-seq higher ICAM JAM3 KEGG differentially genes highly associated ‘Cell molecule’ (p=2.209e-7). silencing lead downregulation mRNA encoding (n=7, p<0.05) E-selectin p<0.01) at 48h after transfection. VCAM1, but not E-Selectin, also reduced protein level Blot (n=3). confirmed PBMCs silenced where able demonstrate decrease leucocyte-to-endothelial (n=5, p<0.05). Conclusion Our results show lncRNA could contribute vasculopathy, acting regulator References [1]Pachera E, et al. Long noncoding key mediator TGF-β-driven fibrosis. J Clin Invest. 2020 Sep 1;130(9):4888-4905. Disclosure Interests Francesca Tirelli: None declared, Elena Pachera: Robert Lafyatis Consultant of: Pfizer, Bristol Myers Squibb, Boehringer-Ingleheim, Formation, Sanofi, Boehringer-Mannheim, Merck Genentech/Roche, Grant/research support from: Corbus, Moderna, Regeneron, Pfizer Kiniksa, Menqi Huang: Shervin Assassi: Eva Camarillo: Francesco Zulian: Gabriela Kania: Oliver Distler Speakers bureau: Bayer, Boehringer Ingelheim, Janssen, Medscape, Abbvie, Acceleron, Alcimed, Amgen, AnaMar, Arxx, AstraZeneca, Baecon, Blade, CSL Behring, 4P Science, Galapagos, Glenmark, Horizon, Inventiva, Kymera, Lupin, Miltenyi Biotec, Mitsubishi Tanabe, MSD, Novartis, Prometheus, Roivant, Sanofi Topadur, Ingelheim.