Cardiovascular Diseases: Genetic Susceptibility, Environmental Factors and their Interaction

By Nikolaos Papageorgiou

Cardiovascular Diseases: Genetic Susceptibility, Environmental Factors and Their Interaction covers the special heritability characteristics and identifying genetic and environmental contributions to cardiovascular health. This important reference provides an overview of the genetic basis of cardiovascular disease and its risk factors.

Included are important topics, ranging from lifestyle choices, risk factors, and exposure, to pollutants and chemicals. Also covered are the influences of Mendelian traits and familial aggregation and the interactions and interrelationships between genetics and environmental factors which, when compared, provide a sound understanding of the interplay between inherited and acquired risk factors.

The book provides a much needed reference for this rapidly growing field of study. By combining the latest research within the structured chapters of this reference, a better understanding of genetic and environmental contribution to cardiovascular disease is found, helping to substantiate further investigations in the field and design prevention and treatment strategies.

• Provides an overview of the genetic basis of cardiovascular disease and its risk factors
• Reviews several large population-based studies which indicate that exposure to several environmental factors may increase CVD morbidity and mortality, exploring the plausibility of this association by data from animal studies
• Reflects on future studies to help understanding the role of genes and environmental factors in the development and progression of cardiovascular disease

• Language: English
• Publisher: Academic Press
• Release date: Aug 5, 2016
• ISBN: 9780128033135

Nikolaos Papageorgiou

Dr. Papageorgiou has completed his basic training in Greece and continued his cardiology training in Dusseldorf, Germany and thereafter in London, UK. He has trained at the London Heart Hospital, University College London and currently at the Barts Heart Centre, St. Bartholomew's Hospital London UK. He has a strong interest in endothelial function and the mechanisms of atherosclerosis. He also has a strong interest in the role of genetics in cardiovascular disease and holds a PhD degree which is related to that. Currently, he focuses on arrhythmias and the therapeutic approaches which aim to them. He is a reviewer of several international journals, member of the EACVI35. Dr. Papageorgiou currently has over 120 publications in peer reviewed journals, all PubMed listed, and has done more than 100 presentations in international conferences where he has been an invited speaker.

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Cardiovascular Diseases

Genetic Susceptibility, Environmental Factors and their Interaction

Editor

Nikolaos Papageorgiou, MD, PhD

Table of Contents

Cover image

Title page

Copyright

Dedication

List of Contributors

Acknowledgments

Chapter 1. Summary

Chapter 2. Atherosclerosis and Coronary Artery Disease: From Basics to Genetics

Introduction

Normal Artery Structures

Pathogenesis of Atherosclerosis

Initiation of Atheroma

The Formation of Atherosclerotic Plaque

Clinical Implications of Atherosclerosis

Genetics of Coronary Artery Disease

Atherosclerosis: Therapeutic Options

Chapter 3. The Role of Genetics in Acute Coronary Syndrome

Introduction

Clopidogrel

Prasugrel

Ticagrelor

Genotyping Assays

Cost-effectiveness of Genotyping

Genetic Polymorphisms Related to Aspirin

Future Directions

Summary: Role of Genotyping in Acute Coronary Syndrome

Chapter 4. Global Epidemiology and Incidence of Cardiovascular Disease

Introduction

Global Burden of Disease: Reporting Metrics

Measuring the Global Burden of Cardiovascular Disease

Changes in Total Disease Burden Versus CVD

Global Cardiovascular Mortality

Global Cardiovascular DALY

Risk Factors for CVD Globally

Epidemiological Transition

Aging Population

Ethnicity

Migration

Urbanization

Specific CVD Conditions

Conclusions

Chapter 5. Lifestyle Choices, Risk Factors, and Cardiovascular Disease

Introduction

Dietary Patterns and CV Risk

Physical Activity/Fitness and CV Risk

Obesity and CV Risk

Smoking and CV Risk

Psychological Factors and CV Risk

Conclusion

Chapter 6. The Contribution of Pollutants and Environmental Chemicals in Cardiovascular Disease

Introduction

Air Pollutants

Metal Toxicity

Conclusions

Chapter 7. Genetic Susceptibility to Cardiovascular Diseases: From Mendelian Disorders to Common Variants

Introduction

Genetic Studies: A Methodologic Approach

Genetic Polymorphisms in Cardiovascular Diseases

Genome-Wide Linkage and Association Studies: Leaps Toward Individualized Cardiovascular Medicine?

Conclusions

Glossary

Chapter 8. Role of Conventional Risk Factors in Genetic Susceptibility to Cardiovascular Diseases

The Relationship of Risk Factors and Genetics to Disease

Methods for Assessing Causality

Randomized Controlled Trials

Observational Studies

Genetic Studies and Mendelian Randomization

Blood Lipids

Inflammation

Anthropometric Traits as Risk Factors

Behavioral and Environmental Risk Factors

Limitations of Causal Inference Using Genetics

Conclusions

Chapter 9. Genetic Susceptibility in Biochemical and Physiological Traits

Introduction

Hypertrophic Cardiomyopathy

Channelopathies

Connective Tissue Disorders

Genetic Disorders Leading to Atherosclerosis

Cardiovascular Pharmacogenetics and Pharmacogenomics

Chapter 10. Interactions–Interrelationships Between Genetics and Environmental Factors in Cardiovascular Disease

Introduction

Models of Interaction

Environmental Factors

Interactions Between Genotype, Diet, and Coronary Heart Disease

Interactions Between Alcohol and Coronary Heart Disease

Interaction Between APOE Genotype, Smoking, and Risk of Coronary Heart Disease

Environmental Pollutants, Epigenetics, and Cardiovascular Disease

Drug–Genome Interactions Relevant to Cardiovascular Disease

Gene–Environmental Interactions and Congenital Heart Disease

Conclusions

Index

Copyright

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Notices

Knowledge and best practice in this field are constantly changing. As new research and experience broaden our understanding, changes in research methods, professional practices, or medical treatment may become necessary.

Practitioners and researchers must always rely on their own experience and knowledge in evaluating and using any information, methods, compounds, or experiments described herein. In using such information or methods they should be mindful of their own safety and the safety of others, including parties for whom they have a professional responsibility.

To the fullest extent of the law, neither the Publisher nor the authors, contributors, or editors, assume any liability for any injury and/or damage to persons or property as a matter of products liability, negligence or otherwise, or from any use or operation of any methods, products, instructions, or ideas contained in the material herein.

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A catalogue record for this book is available from the British Library

ISBN: 978-0-12-803312-8

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Dedication

To my mentor and friend,

Professor Dimitris Tousoulis

List of Contributors

E. Androulakis,     John Radcliffe Hospital, Oxford, United Kingdom

A. Briasoulis,     Wayne State University/Detroit Medical Center, Detroit, MI, United States

J. Chandrasekhar,     Icahn School of Medicine at Mount Sinai, New York, NY, United States

M. Charakida,     University College London, London, United Kingdom

S. Chiesa,     University College London, London, United Kingdom

T. Christophides,     Barts Heart Centre, Barts Health NHS Trust, London, United Kingdom

R.M. Dumitru,     St Peter’s Hospital, Chertsey, Surrey, United Kingdom

E. Farmaki,     1st Cardiology Department, Hippokration Hospital, Athens University Medical School, Athens, Greece

P. Glennon,     University Hospital Coventry, Coventry, United Kingdom

G. Hatzis

1st Cardiology Department, Hippokration Hospital, Athens University Medical School, Athens, Greece

Philipps University Marburg, Germany

S.A. Hayat,     University Hospital Coventry, Coventry, United Kingdom

K. Karatolios,     Philipps University Marburg, Germany

P.B. Lim,     Imperial College NHS Health Care Trust, London, United Kingdom

C.J. McAloon,     University Hospital Coventry, Coventry, United Kingdom

R. Mehran,     Icahn School of Medicine at Mount Sinai, New York, NY, United States

M. Michail,     Barts Heart Centre, Barts Health NHS Foundation Trust, London, United Kingdom

K. Norrington,     Barts Heart Centre, Barts Health NHS Trust, London, United Kingdom

E. Oikonomou,     1st Cardiology Department, Hippokration Hospital, Athens University Medical School, Athens, Greece

F. Osman,     University Hospital Coventry, Coventry, United Kingdom

N. Papageorgiou

Barts Heart Centre, St. Bartholomew’s Hospital, London, United Kingdom

Athens University Medical School,Athens,Greece

G. Siasos,     1st Cardiology Department, Hippokration Hospital, Athens University Medical School, Athens, Greece

D.I. Swerdlow

University College London, London, United Kingdom

Imperial College London, London, United Kingdom

D. Tousoulis,     1st Cardiology Department, Hippokration Hospital, Athens University Medical School, Athens, Greece

E. Zacharia,     1st Cardiology Department, Hippokration Hospital, Athens University Medical School, Athens, Greece

K. Zacharias,     Croydon University Hospital, London, United Kingdom

M. Zaromitidou,     1st Cardiology Department, Hippokration Hospital, Athens University Medical School, Athens, Greece

Acknowledgments

I would like to record my most sincere thanks to all the people from Elsevier who have contributed to the production of this book.

All the chapter contributors deserve special thanks and are listed on the page above.

Chapter 1

Summary

N. Papageorgiou¹,  and D. Tousoulis²     ¹St. Bartholomew’s Hospital, London, United Kingdom     ²Athens University Medical School, Athens, Greece

Abstract

Cardiovascular disease (CVD) is the leading cause of mortality and morbidity in modern societies and encompasses a range of human pathology. It is highly suspected that CVD is strongly related to genetic and environmental factors, and studies have demonstrated that there is an interaction between genetic and environment factors that can lead to cardiovascular disease.

Keywords

Cardiovascular; Environmental; Factors; Genetic; Interaction; Medicine; Susceptibility; Translational

Cardiovascular disease (CVD) is the leading cause of mortality and morbidity in modern societies and encompasses a range of human pathology. It is highly suspected that CVD is strongly related to genetic and environmental factors, while studies have demonstrated that there is an interaction between genetic and environment factors that can lead to CVD.

In this book, we aim to review the current knowledge on this topic:

Zaromitidou et al. discuss thoroughly the role of atherosclerosis as well as the role of genetics in CVD, providing related mechanistic links. Further to this, Chandrasekhar et al. review the current data on the impact of genetic variations on antiplatelet therapy, as well as the potential role of genotyping in prescription of antiplatelet therapies in acute coronary syndromes. In addition, MacAloon et al. examine the current cardiovascular epidemiology and perform specific analyses on fatal and nonfatal CVD burden. They also examine risk factor exposure and prevalence to determine explanations for the current pattern of CVD. Specific CVD categories and conditions are also examined to elucidate a more definitive analysis of global CVD determinants. Moreover, Chiesa et al. review the current evidence surrounding the impact of lifestyle choices on cardiovascular risk factors and disease, and they discuss the beneficial effects of modifying these behaviors with regard to morbidity and mortality. Hatzis et al. summarize the thus-far-acquired knowledge on the main pollutants and present the basic mechanisms that mediate their actions on the vasculature.

Briasoulis et al. acknowledge the contribution of candidate gene studies to complex CVDs such as acute coronary syndromes, and the fact that the clinical impact and the pathophysiological implications of polymorphisms are elusive. On top of these data, Swerdlow et al. discuss the different relationships that a risk factor can have with disease and the role of genetic studies in causal inference, and they evaluate the insights provided by genetics into the contribution of some prominent risk factors to CVD pathogenesis.

Furthermore, Dumitru et al. provide a few examples of genetic susceptibility in biochemical and physiological traits of CVD. They also examine the opportunities provided by pharmacogenomics for future improvements in treatment of CVD. Finally, Norrington et al. introduce key concepts in gene–environment interactions in CVD. The expounding importance of gene–environment interactions in understanding the missing heritability of multifactorial CVDs is highlighted as well as the fundamental concepts of statistical and biological interactions.

The available data are promising, and there are still ongoing studies aiming to evaluate the interactions between genetics and environmental factors in CVD. We hope that the information and data provided will be interesting for the readers of the book and will stimulate further research on the topic.

Chapter 2

Atherosclerosis and Coronary Artery Disease

From Basics to Genetics

M. Zaromitidou, G. Siasos, N. Papageorgiou, E. Oikonomou,  and D. TousoulisAthens University Medical School, Athens, Greece

Abstract

Despite the tremendous progress in primary and secondary prevention, coronary artery disease (CAD) is unfortunately among the leading causes of death globally. Atherosclerosis, the underlying pathology of CAD, is the result of multiple complex mechanisms, many of which still remain unclear. In order to prevent or treat atherosclerotic complications, it is imperative to clarify and comprehend the mechanisms involved in the pathogenesis. The central role of lipoproteins and inflammation in atherosclerosis has been validated in many studies. Genome-wide association studies identified the first genetic loci associated with CAD, confirming the detrimental role of lipoproteins and underscoring the presence of unknown mechanisms. Further deciphering of the molecular and gene mechanisms that lead to atherosclerosis provided novel therapeutic targets such as proprotein convertase subtilisin/kexin type 9 (PCSK9) inhibitor and mipomersen.

Keywords

Atherosclerosis; Cardiovascular disease; Coronary artery disease; Endothelium; Genetics; Inflammation

Introduction

Cardiovascular disease is the leading global cause of death. According to the World Health Organization, 17.3  million deaths resulted from cardiovascular disease worldwide in 2008, and that number is expected to reach 23.3  million by 2030. The underlying pathology of cardiovascular disease is atherosclerosis and, depending on the artery affected, can manifest as CAD or as cerebrovascular or peripheral disease. Coronary artery disease (CAD) is responsible for 7.3  million deaths and is the second cause of death in people under the age of 59. It is also noteworthy that CAD is listed among the first causes for years of life lost due to premature death or disability, and thus has important social and economic impact. Low- and middle-income countries exhibit a continuous increase in the prevalence of CAD in contrast to developed countries. This discrepancy is attributed to the ineffective primary and secondary prevention measures in addition to poor healthcare systems in the low- and middle-income countries [1].

The mechanisms of atherosclerosis have gained increasing attention over recent decades. Many questions still remain concerning the heterogeneity that atherosclerosis displays in time (when?), in the areas affected (where?), in the factors triggering the initiation (how?), and in defining the natural history and evolution of the disease (why?). Atherosclerosis is a chronic systematic disease involving large and medium-sized arteries that initiates early in life [2]. As it progresses, the structure of normal arteries is modified, and atherosclerotic plaques are formed with consequent narrowing or dilation of the vessel. Atherosclerosis is a systematic disease with local manifestations, but the factors determining the preferential involvement of a vascular region over another (ie, coronary or carotid artery) as well as the development of atherosclerotic lesions in certain sites of a specific artery (eg, branches, curvatures, and proximal areas of left anterior descending) are unclear. In addition, regardless of tremendous research efforts, sparse data are available illuminating the factors responsible for the transition of a stable atherosclerotic plaque to a vulnerable plaque that can cause the acute complications of atherosclerosis (eg, myocardial infarction (MI) in CAD). Diversity also exists in the biological procedures and their clinical manifestations following rupture or erosion of unstable plaques (clinically silent MI, unstable angina, and MI with or without ST elevation).

The biggest question to be answered is why the human arteries are vulnerable to atherosclerotic changes. As most communicable diseases are now successfully treated and average life expectancy has increased, cardiovascular disease burden emerges as one of the most important health issues. It was some decades ago that the Framingham Heart Study provided valuable data regarding the primary prevention of atherosclerotic cardiovascular disease and established the cardiovascular risk factor as a new term. The identification of several risk factors associated with modern lifestyle such as hyperlipidemia, smoking, hypertension, obesity, diabetes mellitus, lack of exercise, anxiety, and depression supports the notion that atherosclerosis is a disease of urbanization [3]. However, well-known risk factors such as older age, family history of cardiovascular disease, male sex, and genetic abnormalities (familial hypercholesterolemia) indicate a genetic contribution to atherosclerosis. The report of atherosclerotic findings in mummies from populations of disparate regions with differences in dietary habits and certainly differences in dietary and lifestyle patterns compared to the present time suggests that human vessels are susceptible to atherosclerotic alterations regardless of the current environmental risk factors. Moreover, these observations underline the ongoing need for novel risk factors [4]. Therefore, both genetic and environmental factors are implicated in the pathogenesis of atherosclerosis, whereas their interaction may account for the heterogeneity that atherosclerosis displays.

Normal Artery Structures

The normal artery wall comprises three layers: the tunica intima, the tunica media, and the tunica adventitia.

• The tunica intima consists of the endothelium, connective tissue (collagen, laminin, fibronectin, and other extracellular matrix molecules), and a basal layer of elastic tissue called internal elastic lamina that separates the tunica intima from tunica media. Endothelium is a thin monolayer of cells that serves as the contact surface with blood. Due to its strategic location, the endothelium has emerged as the main regulator of vascular homeostasis with its structural and functional properties altered in response to local and systemic stimuli [5].

• The tunica media is characterized by the presence of concentric layers of vascular smooth muscle cells (VSMCs) and elastin-rich extracellular matrix. It is the final recipient of signals regulating the vascular tone and is separated from adventitia by the external elastic lamina.

• The tunica adventitia is the outer layer of the vascular wall, and it consists of fibroblasts, collagen, mast cells, nerve endings, and vasa vasorum. It was not until recently that important functions of the adventitia were identified. More specifically, adventitia seems to participate in the cell trafficking through the arterial wall and the signaling between vascular endothelial cells, smooth muscle cells, and the local tissue environment. In addition, this layer is involved in the repair mechanism following vessel injury, in the regulation of the dynamic lumen size (via medial smooth muscle tone), and in the inward or outward wall-remodeling response [6].

Pathogenesis of Atherosclerosis

Atherosclerosis is a complex disease with pieces of the pathophysiology puzzle still missing. In order to prevent or treat atherosclerotic complications, it is imperative to clarify and comprehend the mechanisms involved in the pathogenesis. Endothelial cells, VSMCs, and arterial extracellular matrix macromolecules (collagen, proteoglycans, and elastin) are components of a normal artery that play a crucial role in the atherosclerosis process. Lipid accumulation, leukocyte recruitment, and local flow hemodynamic and inflammatory mechanisms are essential elements of the underlying pathology, whereas plaque angiogenesis and mineralization contribute to the evolution of atheroma.

In brief, altered endothelial function leads to transportation and subendothelium accumulation of low-density lipoprotein (LDL) particles. Retention of LDL results in its modification to oxidized LDL (Ox-LDL), triggering an inflammatory cascade and activating endothelial cells to signal the recruitment and migration of monocytes. The coexistence of monocytes and lipids in the subendothelium leads to the phagocytosis of the latter by monocytes and the formation of foam cells that gradually progress into atherosclerotic lesions (Fig. 2.1).

Initiation of Atheroma

The Role of Endothelium

The endothelium plays a crucial part in the initiation and in all stages of atherosclerosis due to its role as the barrier between blood flow and arterial layers. Normal endothelial cells sense the changes in the microenvironment and regulate important functions such as vascular tone, circulating cell adhesion, coagulation, fibrinolysis, vessel wall inflammation, and response to hemodynamic changes. In addition, endothelial cells are arranged in a specific way, forming a tight seal and thus controlling the transportation of all molecules between the lumen and the intima.

In order for atherosclerosis to initiate, LDL needs to cross the endothelium and reside in the subendothelium. Increased endothelium permeability is part of endothelial dysfunction and a precondition for LDL accumulation. Several factors can impair normal endothelial function, including smoking, hyperlipidemia, arterial hypertension, and diabetes mellitus. It is noteworthy that whereas these factors affect the whole artery, atherosclerosis occurs only in certain vessel areas (curved regions, branches, and bifurcation intersections) that are mainly characterized by disturbed flow patterns. On the contrary, atherosclerotic lesions develop rarely in arterial regions with accordingly few branches (internal mammary artery) and laminar blood flow. Predilection sites are complicated with atherosclerosis in the presence of cardiovascular risk factors.

Figure 2.1  Low shear stress (ESS) promotes endothelial dysfunction, transportation, and subendothelium accumulation of low-density-lipoprotein (LDL) particles. Retention of LDL results in its modification to oxidized LDL, which triggers an inflammatory cascade that activates endothelial cells and signals the recruitment and migration of monocytes. After crossing the endothelium, monocytes differentiate into macrophages, internalize modified LDL, and form foam cells. The infiltration of macrophages within the lipid pool and their subsequent apoptosis result in the development of the lipid-rich necrotic core that combined with a collagen fibrous cap constitute the advanced lesion called fibroatheroma.

Mechanosensors positioned on the cell surface identify endothelial shear stress (ESS) stimuli and activate atheroprotective or atheroprone intracellular pathways. Low ESS promotes an atheroprone endothelial cell phenotype (endothelial activation) via up- or down-regulation of gene expression. Regarding the initiation of atherosclerosis, low ESS upregulates the expression of genes encoding for the LDL receptor of the endothelial membrane. In addition, low ESS contributes to the increased endothelial LDL permeability by inducing structural alterations to the cells that result in the transition from fusiform to polygonal shape and widening of the junctions between the cells. The attenuation of mitotic and endothelial apoptotic cell circles, in accordance with the prolonged residence time of LDL near the endothelium due to low ESS, contributes to the augmented membrane permeability [7]. Thus, in the presence of hyperlipidemia, LDL particles cross the endothelium and start to accumulate in the intima tunica.

Endothelial activation promotes early atherosclerosis not only by affecting extracellular LDL accumulation but also by diminishing the atheroprotective properties of the endothelial-derived nitric oxide (NO). Specifically, NO synthesized in the normal endothelial cells modulates vascular tone (favors vasodilation), hampers leukocyte migration to the intima by reducing the expression of adhesion molecules, and decreases VSMC proliferation. Whereas laminar blood flow upregulates the genes implicated in NO production, disturbed flow and low ESS act in an opposite manner.

Moreover, activated endothelial cells promote the atherosclerotic process by increasing the production of reactive oxygen species (ROS) and the secretion of cytokines and chemokines that trigger the migration and activation of monocytes. An additional atherogenic characteristic of endothelial activation is the release of growth factors such as platelet-derived growth factor (PDGF) that induce VSMC proliferation and extracellular matrix synthesizing. The accumulation of VSMCs and extracellular molecules, especially proteoglycans, is known as intimal thickening or intimal hyperplasia [8].

The Role of LDL Particles

The accumulation of cholesterol in the subendothelium is an essential step for the initiation of atherosclerosis. LDL particles transfer cholesterol through blood circulation and into the arterial intima. The endothelial cell membrane is considered impermeable and highly controls molecule trafficking between the blood flow and the vessel wall. In sites of endothelial dysfunction, LDL can penetrate through the cells and accumulate in the intima. The negative charged proteoglycans of extracellular matrix (ECM) bind LDL, resulting in the retention of the lipoproteins – a key step for the initiation of atherosclerosis. The LDL–proteoglycan interaction entraps LDLs in the subendothelium and prolongs their residence time. Thus, LDLs are susceptible to oxidative modification generated by ROS and a number of enzymes such as nitric oxide synthases, NADPH oxidases, lipoxygenases, and myeloperoxidases [9]. In addition, modified LDL aggregates are produced by lipases and proteases such as secretory phospholipase A2 (sPLA2) and sphingomyelinase under proteoglycan regulation. LDL particles may also be chemically modified by nonenzymatic glycation in patients with chronic hyperglycemia, underlining diabetes mellitus as a major risk factor for cardiovascular disease [10].

Ox-LDL displays pro-atherogenic properties that aggravate atherosclerotic pathology. In particular, Ox-LDL induces tissue factor expression and platelet aggregation, limits the beneficial biological actions of NO, and promotes endothelial damage and apoptosis. Ox-LDL also affects the genetic substrate by upregulating genes associated with inflammation. Chemotactic molecules such as monocyte chemotactic protein-1 (MCP1) and PDGF are secreted, inducing the recruitment and proliferation of monocytes and VSMCs. Macrophages express scavenger membrane receptors for Ox-LDL, whereas VSMCs increase collagen ECM concentration. Overall, Ox-LDL further amplifies endothelial activation, inflammation, and thrombogenicity.

The Role of Inflammation

Monocyte-Derived Macrophages

Modified LDL is identified as a pathogen, and thus an inflammatory response is initiated. The first step in the inflammatory process is the recruitment of leukocytes, mostly monocytes. Endothelial cells with normal function are not susceptible to transmigration of monocytes. The activated endothelial cells and modified LDL reverse the endothelium microenvironment in favor of monocyte adherence and migration by triggering the expression of adhesion molecules on endothelial cell membrane and the secretion of chemoattractant cytokines. Molecules known to be involved in monocyte adhesion are the vascular cell adhesion molecule-1 (VCAM1), the intracellular adhesion molecule-1 (ICAM1), and members of the selectin family (P- and E-). In order for adherent monocytes to penetrate the endothelium, the secretion of chemokines must precede. MCP1 and fractalkine are the main chemokines implicated in the transmigration of monocytes through the endothelial layer [11].

Once monocytes overcome the endothelium barrier and reside in the intima, they differentiate into macrophages. Macrophages in the atherosclerotic lesion present marked phenotype heterogeneity that is defined by the environmental stimuli. Their categorization is based on the surface markers they express, the molecules they secrete, and their biological functions. The most abundant phenotype in the subendothelium is the M1 macrophage, which is induced by a variety of stimuli such as proinflammatory cytokines (eg, interferon-γ (IFNγ) and tumor necrosis factor (TNF)), cholesterol crystals and esters, and modified LDL. M1 macrophages play an essential role in the immune response against pathogens during an infection mediating the production of ROS. In atherosclerotic lesions, modified LDL induces chronic activation of M1 macrophages, resulting in tissue damage. Cholesterol crystals and esters promote the M1 phenotype by triggering the caspase-1 activating NLRP3 inflammasome, the Toll-like receptor-4 (TLR4) or nuclear factor–kappa B (NFκB)-mediated pathway. Oxidized LDL leads to M1 differentiation by hampering Kruppel-like factor-2 and stimulating the TLR4 pathway [12]. Macrophages expressing the M1 phenotype are characterized as proinflammatory and secrete a number of cytokines, such as interleukin-6 (IL6), IL1β, and TNF that further aggravate atherosclerosis [13]. The M2 macrophage is a different phenotype that is induced by stimuli such as IL4 and IL10 and displays antiinflammatory properties, including the clearance of apoptotic cells [14]. Interestingly, the phenotype of monocytes depends on environmental stimuli that in turn can be modulated by the activated macrophages. In addition, macrophages can switch between different phenotypes in response to environmental changes, a function known as plasticity [15].

Monocyte-derived macrophages internalize the LDL particles, forming the so-called foam cells. Although macrophages express LDL receptors, foam cell formation is not mediated through them, as they are downregulated early by the accumulated cholesterol levels. Modified LDL uptake is mediated by scavenger receptors, whereas aggregated LDLs and native LDLs are engulfed by phagocytosis and pinocytosis mechanisms, respectively. The scavenger receptors participating in foam cell formation are scavenger receptor-A (SRA), CD36, scavenger receptor class B member 1 (SRB1), and lectin-type receptor-1 (LOX1), with the first two internalizing the majority of modified LDLs [16]. Once lipoproteins enter the macrophage, they are transferred to the late endosome/lysosome, where they are subjected to hydrolysis by lysosomal acid lipase and form free cholesterol. Subsequently, free cholesterol is re-esterified in the endoplasmic reticulum (ER) by cholesterol acyltransferase-1 (ACAT1) to form cholesteryl fatty acid esters, and it is eventually stored in lipid droplets [17]. The re-esterification in the ER is an essential and prophylactic step against the cytotoxicity of free cholesterol [18]. Neutral cholesterol ester hydrolase transforms cholesterol esters into free cholesterol that can efflux macrophages through ABCA1 and ABCG1 transporters, and thus it comprises a crucial step in the initiation of reverse cholesterol transport. The accumulation of cholesteryl esters in the macrophage due to hypercholesterolemia ultimately provides a foamy appearance (foam cells).

LDL and Inflammation

The internalization of