Evidence of epigenetic alterations in thrombosis and coagulation: A systematic review

Highlights

• Thrombosis and anticoagulation related genes have differential methylation in CVDs.

• In vitro models further support these findings.

• Top ranked genes in several studies are the thrombin receptors.

Abstract

Thrombosis in the context of Cardiovascular disease (CVD) affects mainly the blood vessels supplying the heart, brain and peripheries and it is the leading cause of death worldwide. The pathophysiological thrombotic mechanisms are largely unknown. Heritability contributes to a 30% of the incidence of CVD. The remaining variation can be explained by life style factors such as smoking, dietary and exercise habits, environmental exposure to toxins, and drug usage and other comorbidities.

Epigenetic variation can be acquired or inherited and constitutes an interaction between genes and the environment. Epigenetics have been implicated in atherosclerosis, ischemia/reperfusion damage and the cardiovascular response to hypoxia. Epigenetic regulators of gene expression are mainly the methylation of CpG islands, histone post translational modifications (PTMs) and microRNAs (miRNAs). These epigenetic regulators control gene expression either through activation or silencing. Epigenetic control is mostly dynamic and can potentially be manipulated to prevent or reverse the uncontrolled expression of genes, a trait that renders them putative therapeutic targets.

In the current review, we systematically studied and present available data on epigenetic alterations implicated in thrombosis derived from human studies. Evidence of epigenetic alterations is observed in several thrombotic diseases such as Coronary Artery Disease and Cerebrovascular Disease, Preeclampsia and Antiphospholipid Syndrome. Differential CpG methylation and specific histone PTMs that control transcription of prothrombotic and proinflammatory genes have also been associated with predisposing factors of thrombosis and CVD, such us smoking, air pollution, hypertriglyceridemia, occupational exposure to particulate matter and comorbidities including cancer, Chronic Obstructive Pulmonary Disease and Chronic Kidney Disease. These clinical observations are further supported by in vitro experiments and indicate that epigenetic regulation affects the pathophysiology of thrombotic disorders with potential diagnostic or therapeutic utility.

Introduction

Thrombosis mainly occurs in the blood vessels supplying the heart, brain and peripheries, resulting in clinical entities which are collectively referred to as cardiovascular disease (CVD). Atheromatous plaque formation, which precedes the thrombotic event, usually occurs in multiple sites and despite the plurality of clinical manifestations (myocardial infarction (MI), stroke, thrombosis and cardiac arrhythmia), these diseases share a common pathophysiological mechanism. Cardiovascular diseases are the world's bigger killers, with 85% of these deaths attributed to heart attack and stroke according to the World Health Organization statistics [1]. Stroke is a clinical syndrome characterized by an acute loss of neurological function, with symptoms lasting longer than 24 h, whereas transient ischemic attack (TIA) lasts less than 24 h without permanent neurological deficit [2]. Ischemic stroke and TIA are usually secondary to thrombosis or embolism of the arteries supplying the brain. Intensive research efforts have revealed several genetic components and environmental predisposing factors, such as smoking, that contribute to the thrombotic process, however key aspects of the thrombotic mechanism, including its deregulation in a pathophysiological context, remain to date largely unknown. Heritability estimates suggest that inherited genetic factors account for approximately 30% of the variation in the incidence of most cardiovascular diseases [3]. The remaining variation is believed to be explained by life style factors such as dietary and exercise habits, environmental exposure to toxins, and drug usage.

Epigenetic variation can be acquired or inherited and constitutes an interaction between genes and the environment. Epigenetic alterations were first described approximately 80 years ago, but mechanistic studies only recently have shed light on the field. The role of epigenetics in determining a range of processes that are believed to be critical in the development and outcome of CVD and thrombosis is being increasingly elaborated [4]. Epigenetic factors have been implicated in influencing atherosclerosis, angiogenesis, ischemia/reperfusion damage, cardiovascular response to hypoxia and fluid shear stress. The reversibility of epigenetic alterations renders them valuable therapeutic targets in the era of precision medicine.

Epigenetic regulators of gene expression are mainly the following three: (a) methylation of CpG islands, (b) histone post translational modifications (PTMs) and (c) microRNAs (miRNAs). These epigenetic regulators control gene expression either through activation or silencing. Epigenetic control is mostly dynamic and can potentially be manipulated to prevent or reverse the uncontrolled expression of genes [5].

The most studied aspect of epigenetics is methylation of DNA at CpG islands. Gene promoter methylation is generally considered to lead to gene silencing. DNA methylation is the process where a methyl group is added to the fifth position of the six-atom ring of cytosine (5-methylcytosine). A group of enzymes known as DNA methyltransferases (DNMTs) catalyze the addition of the methyl group to the cytosine a process which is dynamic [6]. Methyl groups on DNA can be removed either by methyltransferases or by DNA excision and repair [7,8]. DNMTs are subdivided into two groups either for maintenance of DNA methylation (DNMT1) or de novo methylation (DNMT 3a and 3b). DNA methylation results in more compact DNA structure which prevents the interaction of transcription factors with their binding sites. Moreover, methyl binding proteins (MBPs) that bind methylated DNA and subsequently recruit histone deacetylases (HDACs) can result in gene silencing [9].

The effect of histone modifications on gene transcription is more complex due to the different types of PTMs. The principal PTMs are acetylation, methylation and phosphorylation [10]. The effect that these modifications have on gene regulation depends on which residue has been modified and to what extent. Histone modifications provide binding sites for several proteins, affecting chromatin configuration gene expression. This cumulative effect is referred as the ‘histone code’ [11].

The most extensively studied histone PTM is acetylation of lysine residues. Acetylation alters the charge of the histone from positive to neutral. This process weakens the interaction between histones causing the chromatin to become more accessible. The acetylation is regulated by histone acetyltransferases (HATs) and deacetylation is regulated by HDACs [12,13]. Histone methylation takes place on lysine or arginine residues without affecting the electrical charge of the histones. Either one, two or three methyl-groups can be added on the Lysine residues whereas either one or two on the arginine residues [14], a process linked to both positive and negative gene regulation [11]. Histone phosphorylation changes the charge of the histone proteins from positive to negative. This mark is mostly associated with transcriptional repression [15]. Finally, several other histone PTMs have been reported to a much lesser extent. For example, histones can be ubiquitinated, sumoylated, ADP-ribosylated and glycosylated on lysine residues, citrullinated on arginine residues [16] and proline residues can undergo isomerisation, butyrylation, formylation, 2-hydroxyisobutyrylation malonylation, glutathionylation, succinylation and glutarylation [17].

In the current review, we systematically studied and present available data derived from human studies on epigenetic alterations in thrombosis and coagulation. The main goal of the manuscript is to summarize current knowledge and reveal potential therapeutic targets in the context of thrombotic disorders.

After systematic literature research we identified research papers regarding three main categories: epigenetic alterations observed in the various forms of CVD, epigenetic alterations induced by predisposing factors of CVD and molecular alterations observed in in vitro models.