Identifies CCN5 (matricellular protein) as an inhibitor of vascular restenosis (in-stent re-narrowing) through a mechanism involving thymosin β4 and CD9. Single-cell RNA sequencing of injured arteries showed CCN5 expression in endothelial cells correlated inversely with neointima formation. CCN5 inhibited VSMC proliferation and promoted endothelial repair via upregulation of Tβ4 and CD9 signaling. In porcine stent models, CCN5 delivery reduced restenosis. Positions Tβ4 upregulation as a downstream effector of CCN5's vascular protective mechanism.
Abstract
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.