CCN5 suppresses injury-induced vascular restenosis by inhibiting smooth muscle cell proliferation and facilitating endothelial repair via thymosin β4 and Cd9 pathway

BACKGROUND AND AIMS: Members of the CCN matricellular protein family are crucial in various biological processes. This study aimed to characterize vascular cell-specific effects of CCN5 on neointimal formation and its role in preventing in-stent restenosis (ISR) after percutaneous coronary intervention (PCI). METHODS: Stent-implanted porcine coronary artery RNA-seq and mouse injury-induced femoral artery neointima single-cell RNA sequencing were performed. Plasma CCN5 levels were measured by enzyme-linked immunosorbent assay. Endothelial cell (EC)- and vascular smooth muscle cell (VSMC)-specific CCN5 loss-of-function and gain-of-function mice were generated. Mass spectrometry and co-immunoprecipitation were conducted to identify CCN5 interacting proteins. Additionally, CCN5 recombinant protein (CCN5rp)-coated stents were deployed to evaluate its anti-ISR effects in a porcine model. RESULTS: Plasma CCN5 levels were significantly reduced and correlated closely with the degree of restenosis in ISR patients. CCN5 expression was significantly decreased in VSMCs of stent-implanted porcine coronary segments and injured mouse femoral arteries, especially in synthetic VSMCs. In contrast, elevated CCN5 expression was observed in regenerating ECs of injured vessels. Endothelial cell- and VSMC-specific CCN5 deletion mice exhibited exacerbation of injury-induced neointimal hyperplasia, while CCN5 gain-of-function alleviated neointimal formation. Mechanistic studies identified thymosin β4 (Tβ4) as a CCN5 interacting protein in ECs and EC-CCN5 promoted injury repair through Tβ4 cleavage product Ac-SDKP. Also, CCN5rp promoted EC repair to suppress neointimal hyperplasia via interaction with Cd9 extracellular domain. Moreover, implantation with CCN5rp-coated stent significantly increased stent strut coverage with ECs, which suppressed neointimal formation and ultimately alleviated ISR. CONCLUSIONS: CCN5 exerts a dual protective effect on ISR by inhibiting VSMC proliferation and facilitating EC repair. CCN5rp-coated stent might be promising in the prevention of ISR after PCI.