RNA-binding protein RBM47 promotes endothelial-to-mesenchymal transition and neointimal hyperplasia in arteriovenous fistulas.

backgroundArteriovenous fistula (AVF) is the preferred vascular access for hemodialysis in patients with end-stage renal disease. However, AVF stenosis, a severe complication, significantly impairs the patient's quality of life. The mechanisms underlying AVF stenosis are multifactorial, with neointima (NI) hyperplasia within the fistula being a key contributor. While the involvement of RNA-binding proteins in the pathogenesis of various vascular diseases has garnered increasing attention, their role in autogenous AVFs remains poorly understood.

methodsDifferentially expressed genes in a mouse AVF model were identified through public database analysis, and quantitative real-time polymerase chain reaction (qRT-PCR) confirmed RBM47 as the most significantly altered gene. Endothelial-specific RBM47 overexpression was achieved by local delivery of adeno-associated virus (AAV)-RBM47 using F127 hydrogel. Hematoxylin-eosin staining was used to assess neointimal thickness in human and mouse AVF tissues. RBM47 expression and function were evaluated by qRT-PCR, Western blotting, and assays of proliferation and migration. Mechanistic studies, including transcriptome sequencing, Replicate Multivariate Analysis of Transcript Splicing (rMATS) analysis, RNA-Binding Protein Immunoprecipitation (RIP) Assay, molecular docking, immunohistochemistry, and immunofluorescence, were performed to investigate the relationship between RBM47, SEC31A splicing, and downstream signaling.

resultsRBM47 expression was markedly increased in stenotic AVF tissues from both patients and mouse models. Endothelial overexpression of RBM47 significantly aggravated neointimal hyperplasia. In vitro, RBM47 promoted human umbilical vein endothelial cell (HUVEC) proliferation, migration, and EndMT. Mechanistically, RBM47 was predominantly localized in the nucleus, suggesting a role in alternative splicing. Transcriptome sequencing revealed that RBM47 promoted retention of exon 13 in SEC31A, thereby suppressing the generation of the SEC31A-Δ13 splice variant. This splice variant inhibited SMAD3 phosphorylation and its downstream signaling cascade.

conclusionsRBM47 promotes neointimal hyperplasia in autogenous AVF by inducing EndMT. Specifically, RBM47 regulates alternative splicing of SEC31A and reduces the production of the SEC31A-Δ13 splice variant, which inhibits SMAD3 phosphorylation. These findings reveal a novel mechanism by which RNA-binding proteins contribute to AVF stenosis and suggest RBM47 as a potential therapeutic target for preventing AVF failure.