Absence of Functional Autoantibodies Targeting Angiotensin II Type 1 Receptor (AT1R) and Endothelin-1 Type A Receptor (ETAR) in Circulation and Purified IgG from Patients with Systemic Sclerosis

Systemic sclerosis (SSc) is a rare but severe autoimmune disease characterized by immune dysregulation, fibrosis, and vasculopathy. While previous studies have highlighted the presence of functional autoantibodies targeting the angiotensin II type 1 receptor (AT1R) and endothelin-1 type A receptor (ETAR), leading to autoantibody-mediated receptor stimulation and subsequent activation of endothelial cells (ECs), a comprehensive understanding of the direct interaction between these autoantibodies and their receptors is currently lacking. Moreover, existing data confirming the presence of these autoantibodies in SSc often rely on similar methodologies and assays. Our aim was to replicate previous findings and to investigate the functional effects of SSc patient-derived IgG (SSc IgG) on AT1R- and ETAR signaling, the downstream EC response, as well as presence of AT1R-binding autoantibodies in circulation.OBJECTIVESystemic sclerosis (SSc) is a rare but severe autoimmune disease characterized by immune dysregulation, fibrosis, and vasculopathy. While previous studies have highlighted the presence of functional autoantibodies targeting the angiotensin II type 1 receptor (AT1R) and endothelin-1 type A receptor (ETAR), leading to autoantibody-mediated receptor stimulation and subsequent activation of endothelial cells (ECs), a comprehensive understanding of the direct interaction between these autoantibodies and their receptors is currently lacking. Moreover, existing data confirming the presence of these autoantibodies in SSc often rely on similar methodologies and assays. Our aim was to replicate previous findings and to investigate the functional effects of SSc patient-derived IgG (SSc IgG) on AT1R- and ETAR signaling, the downstream EC response, as well as presence of AT1R-binding autoantibodies in circulation.Quantitative PCR (qPCR) and cytokine ELISA, alongside a real-time cell analyzer, were utilized to assess receptor-specific functional characteristics of purified IgG from SSc patients (n=18). Additionally, a novel protein capture assay using solubilized epitope-tagged AT1R was developed to detect AT1R-binding autoantibodies in plasma samples from SSc patients (n=28) and healthy donors (n=14).METHODSQuantitative PCR (qPCR) and cytokine ELISA, alongside a real-time cell analyzer, were utilized to assess receptor-specific functional characteristics of purified IgG from SSc patients (n=18). Additionally, a novel protein capture assay using solubilized epitope