Interleukin-37 suppresses the inflammatory response to protect cardiac function in old endotoxemic mice

Highlights

• IL-37 protects against cardiac dysfunction in old mice during endotoxemia.

• Endotoxemic old IL-37tg mice have reduced myocardial inflammation.

• IL-37 inhibits NF-κB activation in cardiac cells exposed to endotoxin.

• IL-37 blocks endotoxin-induced inflammatory responses in cardiac cells.

Abstract

Myocardial inflammatory responses to endotoxemia are enhanced in old mice, which results in worse cardiac dysfunction. Anti-inflammatory cytokine interleukin (IL)-37 has a broad effect on innate immunoresponses. We hypothesized that IL-37 suppresses myocardial inflammatory responses to protect cardiac function during endotoxemia in old mice. Old (20–24 month) wild-type (WT), and IL-37 transgenic (IL-37tg) mice were treated with lipopolysaccharide (LPS, 0.5 mg/kg, iv) or normal saline (0.1 ml/mouse, iv). Six hours later, left ventricle (LV) function was assessed using a pressure-volume microcatheter. Levels of monocyte chemoattractant protein-1 (MCP-1), tumor necrosis factor-α (TNF-α), interleukin (IL)-1β and IL-6 in plasma and myocardial tissue, as well as mononuclear cell density in the myocardium, were examined. Cardiac microvascular endothelial cells isolated from WT and IL-37tg mice were treated with LPS (0.2 µg/ml) for 0.5–24 h. Nuclear factor-kappa B (NF-κB) p65 phosphorylation was examined by immunoblotting, and MCP-1 levels in cell culture supernatant was determined using enzyme-linked immunosorbent assay. LV dysfunction in old WT endotoxemic mice was accompanied by up-regulated MCP-1, myocardial accumulation of mononuclear cells and production of TNF-α, IL-1β and IL-6. Expression of IL-37 suppressed myocardial inflammatory responses to endotoxemia in old mice, resulting in improved LV function. Treatment of old WT endotoxemic mice with recombinant IL-37 also improved LV function. In vitro experiments revealed that cardiac microvascular endothelial cells from IL-37tg mice had attenuated NF-κB activation and MCP-1 production following LPS stimulation. In conclusion, IL-37 is potent to suppress myocardial inflammation and protects against cardiac dysfunction during endotoxemia in old mice.

Introduction

Major surgery and trauma can cause endotoxemia [1]. Excessive production of pro-inflammatory mediators caused by bacterial lipopolysaccharide (LPS) frequently leads to cardiac dysfunction. We and others have observed that LPS induces cardiac contractile depression through up-regulation of myocardial production of pro-inflammatory cytokines, such as tumor necrosis factor-α (TNF-α), interleukin (IL)-1β, and IL-6 [2], [3], [4]. Importantly, morbidity and mortality due to the systemic inflammatory response syndrome associated with trauma or sepsis are significantly increased in the elderly [5], [6].

Old animals are found to display enhanced systemic and myocardial inflammatory responses to LPS [4], [7]. We previously reported that enhanced monocyte chemoattractant protein-1 (MCP-1) production in old endotoxemic mice has a critical role in mononuclear cell accumulation and cytokine production in the myocardium and contributes to the mechanism of exaggerated cardiac contractile depression. It appears that the MCP-1-mediated myocardial inflammation, i.e. mononuclear cell infiltration and production of cardiodepressant cytokines, plays an important role in cardiac contractile depression in old endotoxemic animals, and suppression of MCP-1 production and/or the MCP-1-mediated myocardial inflammation may have therapeutic potential for preservation of cardiac function in the elderly during endotoxemia.

Nuclear factor-kappa B (NF-κB) is a master transcription factor that mediates cellular inflammatory responses, including the expression of chemokines and cytokines [8]. Toll-like receptor (TLR) 4 on cell surfaces is activated in response to LPS and interacts with intracellular adaptors to activate NF-κB. Activated NF-κB translocates to the nucleus to initiate the transcription of pro-inflammatory genes [9]. Thus, NF-κB is critical in mediating LPS-induced cytokine production and cardiac dysfunction [10], [11], [12].

IL-37 is a member of the IL-1 family of cytokines. It is expressed in humans, and its expression has been identified in most cell types, including monocytes, dendritic cells, endothelial cells and epithelial cells, and acts as a natural regulator of the inflammatory responses [13]. IL-37 has repressive effects on LPS-stimulated cells, such as macrophages and endothelial cells, which implies that it may interfere with the TLR4 signaling pathway. In vitro studies show that expression of IL-37 in macrophages or epithelial cells dampens constitutive or induced production of pro-inflammatory cytokines, such as IL-1α, IL-1β, TNF-α, and macrophage inflammatory protein (MIP)-2 [14]. In vivo studies indicate that expression IL-37 in mice protects against chemical-induced colitis [15], and metabolic syndrome [16]. Further, mice that express IL-37 transgenic gene exhibit reduced lung, kidney and liver injury during endotoxemic shock [14]. However, it remains unclear whether IL-37 suppresses myocardial inflammatory responses in old animals subjected to endotoxemia. In addition, the effect of IL-37 on cardiac dysfunction caused by endotoxemia has not been determined.

In the present study, we tested the hypothesis that IL-37 suppresses the inflammatory responses in old endotoxemic mice to protect cardiac function against endotoxemic depression. We examined: (1) whether old IL-37 transgenic (IL-37tg) mice have better cardiac function during endotoxemia, (2) whether expression of IL-37 results in reduced MCP-1 production in old mice during endotoxemia and attenuated myocardial mononuclear cell accumulation and cytokine production, (3) whether IL-37 suppresses LPS-induced NF-κB activation and (4) whether recombinant IL-37 can protect old mice against endotoxemic cardiac dysfunction.