Background/Purpose: Patients with giant-cell arteritis (GCA) are in an unmet need for reduction of GC exposure because of the high incidence of GC-related side effects in the elderly. Blocking C5a receptor 1 (C5aR1) with avacopan has allowed extreme minimization of GC exposure in ANCA-associated vasculitis. The complement system has not been investigated in GCA.Aims: 1) to perform a transcriptomic analysis of cultured temporal artery biopsies from patients with GCA and controls 2) to perform a proteomic analysis of the supernatants of cultured arteries 3) to explore the expression in tissue of the C5aR1 4) to interrogate the transcriptomic data searching for transcripts regulated by C5aR1 signaling and the impact of blocking IL-6 receptor with tocilizumab (TCZ) on their expression.

Methods: Nanostring nCounter Human Inflammation Panel was applied to RNA extracted from ex-vivo cultured arteries (from 11 patients with GCA and 3 controls) with or without exposure to TCZ. Protein array (Ray-Biotech, 640 proteins) was applied to the corresponding supernatants. RT-PCR of C5aR1 was performed in frozen arteries from 10 patients with GCA and 10 controls and in cultured myofibroblasts alone or exposed to PBMCs. C5aR1 protein expression was explored by immunofluorescence. nCounter barcode counts were processed with nSolver 4.0 software. Normalized data were analyzed using R-Studio 4.0.3

Results: Transcriptomic analysis revealed up-regulation of complement pathways in arterial samples from patients with GCA (fig 1A). C5a was present in the supernatants of cultured GCA arteries and virtually absent in control arteries (fig1B). C5aR1 was overexpressed in temporal arteries from patients with GCA compared to controls (fig1C) and was expressed by inflammatory cells, endothelial cells and scattered vascular smooth muscle cells (fig1D).Primary myofibroblasts derived from temporal arteries expressed C5aR1 when co-cultured with PBMCs. We interrogated the nanostring results in search for transcripts regulated by AP-1 (in the signalling cascade driven by C5aR1) but not regulated by transcription factors in the classical IL-6,IFNγ or GM-CSF cytokine signaling, based on ENCODE transcription factor targets dataset. 13 genes explored in the Human Inflammation panel had potential binding sites for AP-1 complex but not for STAT3, STAT1, STAT5 or SPI1). Of those, CYSLTR, MBL2, PTGER3, TLR2 and TNF transcripts were increased in samples from patients with GCA compared to controls and, in our experimental conditions, were not down-regulated by TCZ (fig1E)

Conclusion: Complement pathways are activated in GCA and C5a is present in the supernatant of cultured GCA arteries, indicating local processing of complement molecules. C5aR1 is expressed in GCA-involved arteries by inflammatory and stromal cells, suggesting a role in vascular remodeling. Transcripts regulated by C5aR1 (but not regulated by other cytokines present in GCA) were overexpressed in GCA and are not modulated by TCZ. Our findings suggest that blocking C5aR1 may contribute to abrogate inflammatory pathways in GCA. Exploring the transcriptomic and functional impact of C5aR1 inhibitor avacopan on cultured temporal arteries is in progress.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/does-complement-product-c5a-and-its-receptor-c5a-receptor-1-play-a-role-in-giant-cell-arteritis/