Homocysteine accelerates atherosclerosis by inhibiting scavenger receptor class B member1 via DNMT3b/SP1 pathway

Highlights

• Downregulation of SCARB1 facilitated Hcy-mediated lipid accumulation in foam cells.

• DNA methylation was not responsible for Hcy-mediated downregulation of SCARB1 expression.

• Inhibition of SCARB1 expression mediated by DNMT3b is independent on its DNA methyltransferases activity.

• DNMT3b interferes SP1 binding to SCARB1 promoter leading to SCARB1 downregulation.

Abstract

Homocysteine (Hcy) is an independent risk factor for atherosclerosis, which is characterized by lipid accumulation in the atherosclerotic plaque. Increasing evidence supports that as the main receptor of high-density lipoprotein, scavenger receptor class B member 1 (SCARB1) is protective against atherosclerosis. However, the underlying mechanism regarding it in Hcy-mediated atherosclerosis remains unclear. Here, we found the remarkable inhibition of SCARB1 expression in atherosclerotic plaque and Hcy-treated foam cells, whereas overexpression of SCARB1 can suppress lipid accumulation in foam cells following Hcy treatment. Analysis of SCARB1 promoter showed that no significant change of methylation level was observed both in vivo and in vitro under Hcy treatment. Moreover, it was found that the negative regulation of DNMT3b on SCARB1 was due to the decreased recruitment of SP1 to SCARB1 promoter. Thus, we concluded that inhibition of SCARB1 expression induced by DNMT3b at least partly accelerated Hcy-mediated atherosclerosis through promoting lipid accumulation in foam cells, which was attributed to the decreased binding of SP1 to SCARB1 promoter. In our point, these findings will provide novel insight into an epigenetic mechanism for atherosclerosis.

Introduction

Atherosclerosis, a complex chronic disease characterized by lipid deposition in the arterial wall, is a major contributor to morbidity and mortality worldwide [1]. Emerging evidence demonstrated that homocysteine (Hcy), a key intermediate in the metabolism of sulfur-containing amino acid, has been recognized as an independent risk factor for atherosclerosis [2]. Although it was reported that Hcy-induced oxidative stress, endothelium dysfunction and vascular smooth muscle cells (VSMCs) proliferation also played important roles in the pathogenesis of atherosclerosis [[3], [4], [5]], the underlying mechanisms regarding Hcy-accelerated lipid deposition in atherosclerosis still need to be further elucidated.

Scavenger receptor class B type 1 (SCARBI) is a cell-surface glycoprotein which belongs to the CD36 superfamily together with scavenger receptor CD36 and lysosomal membrane protein 2 [6]. It was evidenced that SCARB1 is widely expressed in mammals including macrophages and endothelial cells. Additionally, SCARB1 was reported to play a vital role in cholesterol homeostasis [7,8]. Ubiquitous deletion of SCARB1 can disturb HDL metabolism which reverses the transportation of cholesterol [6,9]. On the other hand, overexpression of SCARB1 in SCARB1-deficient animals can reduce plasma HDL cholesterol levels [10]. Thus, it is reasonable to suspect that SCARB1 might be involved in atherosclerosis due to its regulating role in the metabolism of LDL and HDL which maintain cellular cholesterol balance.

Gene transcriptional regulation involves epigenetics-transcriptional factor interaction, which plays a pivotal role in gene regulation processes and disease development at the molecular level [11]. DNA methylation is the most highly studied epigenetic marker in the modulation of gene transcription, which was regulated by DNA methyltransferases (DNMTs) and intermediate metabolites of the one-carbon pathway including Hcy. Meantime, several studies have suggested the tight correlation between plasma Hcy and DNA methylation [12]. In our previous study, it was found that Hcy-mediated FABP4 gene demethylation via DNMT1 can accelerate atherosclerosis in ApoE^-/- mice [13]. However, a variety of DNA methylation aberrations at a genome-wide level and particular loci under similar pathological conditions was observed [14]. Besides, aberrant expression of DNMT3b, one kind of functional DNMTs in mammals, can inhibit gene expression and onset the bladder cancer by decreasing global DNA methylation [15]. More importantly, several investigations have documented that epigenetic modification including DNA methylation is initiated by the specific transcription factor that binds to the promoter of the gene [16]. As a member of the family of SP/KLF transcription factors, specificity protein 1 (SP1) can drive various cellular processes, such as development, proliferation, apoptosis, as well as lipid metabolism [17]. Meanwhile, it can trigger gene expression through binding to the GC-rich motifs present in the proximal promoter of a gene [18]. In breast cancer, Sp1-induced DNMT1 activity increase was responsible for p53-induced expression inhibition of p125 and the methylation of POLD1 gene promoter [19], while little is known about the association between SP1 and DNMT3b during the development of in atherosclerosis.

Therefore, this study is aimed to elucidate the role of SCARB1 in Hcy-mediated atherosclerosis. The evidence indicated that the decreased recruitment of SP1 to SCARB1 promoter mediated by DNMT3b was responsible for the downregulation of SCARB1, which finally result in the lipid accumulation in foam cells. These findings provide new insight into Hcy-associated lipid accumulation and a new potential molecular target of SCARB1 for the therapy of Hcy-related cardiovascular diseases.