Rationale: Targeting inflammation has been shown to provide clinical benefit in the field of cardiovascular diseases. Although manipulating regulatory T-cell function is an important goal of immunotherapy, the molecules that mediate their suppressive activity remain largely unknown. IL (interleukin)-35, an immunosuppressive cytokine mainly produced by regulatory T cells, is a novel member of the IL-12 family and is composed of an EBI3 (Epstein-Barr virus–induced gene 3) subunit and a p35 subunit. However, the role of IL-35 in infarct healing remains elusive. Objective: This study aimed to determine whether IL-35 signaling is involved in healing and cardiac remodeling after myocardial infarction (MI) and, if so, to elucidate the underlying molecular mechanisms. Methods and Results: IL-35 subunits (EBI3 and p35), which are mainly expressed in regulatory T cells, were upregulated in mice after MI. After IL-35 inhibition, mice showed impaired infarct healing and aggravated cardiac remodeling, as demonstrated by a significant increase in mortality because of cardiac rupture, decreased wall thickness, and worse cardiac function compared with wild-type MI mice. IL-35 inhibition also led to decreased expression of α-SMA (α-smooth muscle actin) and collagen I/III in the hearts of mice after MI. Pharmacological inhibition of IL-35 suppressed the accumulation of Ly6C low and major histocompatibility complex II low /C-C motif chemokine receptor type 2 − (MHC II low CCR2 − ) macrophages in infarcted hearts. IL-35 activated transcription of CX3CR1 (C-X3-C motif chemokine receptor 1) and TGF (transforming growth factor) β1 in macrophages by inducing GP130 signaling, via IL12Rβ2 and phosphorylation of STAT1 (signal transducer and activator of transcription family) and STAT4 and subsequently promoted Ly6C low macrophage survival and extracellular matrix deposition. Moreover, compared with control MI mice, IL-35–treated MI mice showed increased expression of α-SMA and collagen within scars, correlating with decreased left ventricular rupture rates. Conclusions: IL-35 reduces cardiac rupture, improves wound healing, and attenuates cardiac remodeling after MI by promoting reparative CX3CR1 + Ly6C low macrophage survival.

Background: The development of thoracic aortic dissection (TAD) is closely related to the extracellular matrix (ECM) degradation and the vascular smooth muscle cell (VSMC) transformation from contractile to synthetic type. Legumain (Lgmn) degrades ECM components directly or by activating downstream signals. However, the role of Lgmn in the VSMC differentiation and the occurrence of TAD remains elusive. Methods: Microarray datasets concerning vascular dissection or aneurysm were downloaded from the Gene Expression Omnibus (GEO) database to screen differentially expressed genes (DEGs). Four-week-old male Lgmn knockout mice (Lgmn -/- ), macrophage-specific Lgmn knockout mice (Lgmn F/F ; LysM Cre ) and RR-11a treated C57BL/6 mice were given β-aminopropionitrile monofumarate (BAPN, 1g/kg/day) in drinking water for four weeks for TAD modeling. RNA-Seq analysis was performed to recapitulate transcriptome profile changes. Cell interaction was examined in macrophage and VSMC coculture system. The reciprocity of macrophage derived Lgmn with integrin αvβ3 in VSMCs was tested by co-immunoprecipitation assay and colocalization analyses. Results: Microarray datasets from the GEO database indicated up-regulated Lgmn in aorta from TAD patients and Ang II induced aortic abdominal aneurysm (AAA) mice. Elevated Lgmn was evidenced in the aortas and serums from TAD patients and BAPN-induced TAD mice in this study. BAPN induced TAD progression was significantly ameliorated in Lgmn deficient or inhibited mice. Macrophage specific deletion of Lgmn alleviated BAPN induced ECM degradation. Unbiased profiler PCR-array and GO analysis displayed that Lgmn regulated VSMC phenotype transformation. Macrophage specific deletion of Lgmn ameliorated VSMC phenotypic switch in BAPN treated mice. Macrophage derived Lgmn inhibited VSMC differentiation in vitro as assessed by macrophages and VSMCs coculture system. Mechanically, macrophage derived Lgmn bound to integrin αvβ3 in VSMCs and blocked integrin αvβ3, therefore attenuating Rho GTPase activation, down-regulating VSMCs differentiation markers and eventually exacerbating TAD development. Rho-kinase (ROCK) inhibitor Y-27632 reversed the protective role of Lgmn depletion in vascular dissection. Conclusions: These findings indicate that Lgmn signaling may be a novel target for the prevention and treatment of TAD.