ALTERNATIVE SPLICING IDENTIFICATION IN SYSTEMIC SCLEROSIS AND THEIR ROLES IN SKIN FIBROSIS

Background: Systemic sclerosis (SSc) is a class of heterogeneous autoimmune diseases, and fibrosis is at the centre of the pathogenic triad of SSc. Alternative splicing (AS) is a post-transcriptional regulatory process in which specific exons and/or introns of precursor messenger RNA (pre-mRNA) are selectively included or excluded to generate multiple mature mRNA isoforms. AS encompasses several modes, including splicing exons (SE), alternative 3’ splicing site (A3SS), alternative 5’ splicing site (A5SS), intron retention (IR) and mutually exclusive exons (MXE). AS and IR have recently been recognized as playing important roles in many disorders., but little is known about them in systemic sclerosis and fibrotic disorders. Objectives: To identify AS and IR profiles in SSc, assess their impact on protein and explore their roles in the process of skin fibrosis. Methods: A comprehensive genome-wide analysis of AS and IR was conducted across SSc skin tissue and fibroblasts based on bulk RNA-seq, single-cell RNA-seq, and 4D label-free mass spectrometry. AS events were identified using software rMATS. IR expression was analyzed using iread and validated by real-time PCR (RT‒PCR). Correlations among clinical features, introns, splicing factors (regulators of AS) and proteins were assessed by spearman correlation or the LASSO method. Results: Significant changes were noted in AS profiles within SSc skin tissue, with a particular emphasis on the noteworthy alterations in IR. The levels of AS events and IR demonstrated correlations with the extracellular matrix score, SSc autoantibodies, disease duration, and the clinical local skin score. Beyond skin tissues, distinct IR profiles were discernible among different skin fibroblast subtypes. Upon TGF-β stimulation, fibroblasts experienced substantial changes in IR profiles, influencing genes associated with fibrosis-related pathways, notably collagen fibril organization. An overarching integrated analysis involving introns, exons, and proteome profiles indicated that IR mainly exerted a negative impact on protein expression, with certain changes being intronically regulated. However, IR also manifested other effects on protein expression. Additionally, the regulatory network of splicing factors displayed distinct patterns in both SSc skin tissue and fibroblast subpopulations. Conclusion: Altered AS and IR profiles, along with dysfunctional splicing machinery, were observed in skin fibroblasts of SSc. These findings suggest the potential to directly impact protein expression and may be associated with the fibrosis process. This insight could contribute to a better understanding of the pathogenesis of SSc. REFERENCES: NIL. Acknowledgements: The authors would like to thank the patients and investigators in the multicenter national PRESS registry. Disclosure of Interests: None declared. DOI: 10.1136/annrheumdis-2024-eular.2640 Keywords: '-omics, Skin, Fibroblasts Citation: , volume 83, supplement 1, year 2024, page 1906Session: Systemic sclerosis (Publication Only)

'-omics, Skin, Fibroblasts