AIM2 accelerates the atherosclerotic plaque progressions in ApoE−/− mice

https://doi.org/10.1016/j.bbrc.2018.03.005Get rights and content

Highlights

  • Our study demonstrated that AIM2 plays an important role in atherosclerosis.

  • We found that AIM2 could mediate GSDMD-N and pyroptosis in VSMCs for the first time.

  • We first found that AIM2 could affect ICAM-1 and macrophage recruitment in atherosclerosis.

  • We first found that ox-LDL can promote AIM2 expression through NF-κB signaling.

Abstract

Plaque formation is initiated and triggered by cell death in the vascular wall, which gradually leads to the progression of atherosclerosis. Pyroptosis is a newly discovered form of programmed cell death. Absent in melanoma 2 (AIM2), a member of the HIN-200 protein family, plays an important role in activating inflammasomes. However, the role and mechanism of AIM2 in atherosclerotic plaque progression has not been thoroughly elucidated to date. The effect of pyroptosis and the mechanism for this effect were investigated in apolipoprotein E-deficient (ApoE−/−) mice. AIM2 overexpression and inhibition were studied in ApoE−/− mice that were fed a high-fat diet. The specific role of AIM2 in vascular smooth muscle cells (VSMCs) was explored in vitro. The results showed that high fat diet increases the expression of AIM2, ICMA-1, GSDMD-N, which could be mediated by AIM2 expression. The plaque lesion area is lager with AIM2 overexpression. Moreover, TUNEL-positive cells were increased when AIM2 was overexpressed. With increased AIM2, macrophages were enhanced. In vitro studies showed that AIM2 and GSDMD-N expression correlated with ox-LDL levels in a concentration dependent manner. AIM2 expression is associated with NF-κB signaling activity and can be inhibited by NF-κB inhibitor. AIM2 mediated GSDMD activity through ASC, caspase1 pathway. EthD-III and TUNEL staining showed that AIM2 mediates pyroptosis in VSMCs. Our study suggests that AIM2 is not only a regular of inflammasome but also an active participant in atherosclerosis.

Introduction

Atherosclerosis (AS) is a complex pathological condition in vascular disease [1,2], leading to many chronic and acute ailments, such as unstable angina pectoris, hypertension, myocardial infarction and sudden coronary death. The progression of AS is characterized by the accumulation of lipids in vessel walls, increased inflammation, changes in artery structure and cell death [1,[3], [4], [5]]. Hyperlipidemia is a major risk factor for atherosclerosis with low-density lipoprotein (LDL) levels playing a pivotal role [6].

Apoptosis and pyroptosis are two forms of programmed cell death. Although a variety of studies have indicated that apoptosis is the primary form of programmed cell death, cell death in atherosclerosis is more complex. Pyroptosis is a novel form of programmed cell death and is uniquely dependent on GSDMD [7]. While caspase-1 is not involved in apoptosis, since caspase-1-deficient cells still respond to apoptotic signals [8], it can promote the activation and release of GSDMD-N, a cytokine that plays an important role in pyroptosis [9]. Pyroptosis progression is characterized by the loss of cell membrane potential and DNA fragmentation, the latter of which can be detected by positive terminal transferase-mediated deoxyuridine triphosphate-biotin nick end labeling (TUNEL) staining. Pyroptosis was initially discovered in immunological cells with bacterial or viral infections, but a recent study showed that ox-LDL can promote pyroptosis [10]. Previous studies of pyroptosis focused on endothelial cells and macrophages [6,11]; however, the role of ox-LDL in promoting pyroptosis in VSMCs has not been thoroughly elucidated to date.

Absent in melanoma 2 (AIM2) is a cytosolic DNA sensor, the activation of which leads to inflammasome formation in macrophages [[12], [13], [14]]. AIM2 belongs to the HIN200 family of hematopoietic interferon (IFN)-inducible genes and plays an important role in host defense against DNA viruses and certain cytosolic bacteria [15,16]. Previous studies focused on the role of AIM2 in inflammation involving the innate immune system [[17], [18], [19], [20]]. Alternatively, other studies found that AIM2 promotes pyroptosis in tumor cells [21,22]. AIM2 has also been shown to play a role in the progression of abdominal aortic aneurysm [23]. Despite intense interest in this DNA sensor, the role of AIM2 in atherosclerosis has not been thoroughly elucidated to date.

We hypothesized that AIM2 promotes pyroptosis and ICAM-1 expression in VSMCs and that this effect could affect the progression of atherosclerosis. In this study, we investigated the mechanism and effect of pyroptosis and ICAM-1 in VSMCs.

Section snippets

Reagents

Mice were purchased from HFK Bioscience Company (Beijing, China), and ox-LDL was purchased from Yiyuan Biotechnology (Guangzhou, China). Rabbit anti-mouse primary antibodies against AIM2, caspase-1, GSDMD-N, and α-SMA were obtained from Abcam (Shanghai, China). Rabbit anti-mouse primaries antibodies against ASC and AIM2 were purchased from Cell Signaling Technology (Shanghai, China). A mouse anti-GAPDH antibody was purchased from Bioss (Beijing, China). Lentivirus AIM2 and AIM2-shRNA were

High-fat diet increases LDL levels in ApoE−/−mice

LDL levels in mice fed a high-fat diet were increased compared to mice fed a chow diet (Fig. 1A). AIM2 overexpression or inhibition had no effect on the LDL level.

ICAM-1 are dependent on AIM2 expression in ApoE−/− mice

RT-PCR showed that AIM2 expression was increased in lvAIM2 and decreased in shRNA group mice. AIM2 expression was also increased in mice that were fed a high-fat diet compared to the control group (Fig. 1B). Next, the role of AIM2 in regulating inflammasome activation in response to a high-fat diet was examined. ICAM-1 increased in

Discussion

The progression of atherosclerosis contributes to a variety of chronic diseases, including hypertension, coronary heart disease and sudden cardiac death [1,25]. Recent studies have shown that a high-fat diet promotes the progression of atherosclerosis due to the accumulation of lipids in the vessel wall, which promotes inflammation, changes in artery structure and cell death [1,[3], [4], [5]]. Cell death in the vascular wall, particularly in VSMCs, can initiate plaque formation and aggravate

Conflicts of interest

All authors declare no financial or commercial conflicts of interest.

Funding

This project was supported by the National Natural Science Foundation of China (No. 81670325 and No. 81370325).

Acknowledgments

The authors thank Yun Zhang for his help in editing the manuscript.

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