Professor, Department of Pharmacology, Addiction Science and Toxicology, College of Medicine, The University of Tennessee Health Science Center, Memphis, TN, United States
Alex M. Dopico
Van Vleet Chair of Excellence, Department of Pharmacology, Addiction Science and Toxicology, College of Medicine, The University of Tennessee Health Science Center, Memphis, TN, United States
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Contents
Contributors xiii
Preface xvii
Afterword xix
SECTION 1
Cholesterol chemistry and cell function
1. Cholesterol chemistry and laboratory synthesis
Hélio M.T. Albuquerque, Clementina M.M. Santos, and Artur M.S. Silva
Introduction 4
Cholesterol structural characterization 6
Cholesterol laboratory synthesis 9
Concluding remarks 21
References 22
2. Molecular evolution of cholesterol and other higher sterols in relation to membrane structure
Ole G. Mouritsen
Introduction: The overlooked lipids 25
Key features of the cholesterol molecule 27
Evolution and streamlining of a molecule 27
Phase equilibria in lipid membranes 29
Cholesterol and lipid membrane phase equilibria: The liquid-ordered phase 30
Cholesterol, transverse membrane order, permeability, and mechanics 32
Cholesterol and lateral membrane organization 33
Other higher sterols: Universality in sterol function 35
A case study: Cholesterol, lateral membrane structure, and the functioning of Na+/K+-ATPase 36
Cholesterol and membrane evolution 36
References 38
3. Role of cholesterol in maintaining the physical properties of the plasma membrane
Witold K. Subczynski, Marta Pasenkiewicz-Gierula, Justyna Widomska, and Natalia Stein
Introduction 42
Phase diagram for cholesterol/phospholipid mixtures 43
Basic membrane physical properties and how cholesterol regulates them 45
Lateral organization of membranes: Effect of cholesterol 57
New information from the saturation recovery electron paramagnetic resonance stretched exponential function approach 59
Concluding remarks 61
Summary 64
References 64
4. The fundamental interaction of cholesterol with lipid membranes: The umbrella model
Juyang Huang
Introduction 73
Maximum solubility of cholesterol in phosphatidylcholine (PC) and phosphatidylethanolamine (PE) bilayers 74
The umbrella model 77
Monte Carlo simulation of chemical potential of cholesterol using multibody interactions 81
A family of small headgroup molecules: Sterols, ceramides, and diacylglycerols 85
Molecular dynamics (MD) simulation of the umbrella effect 87
Measurement of the chemical potential of cholesterol in PC bilayers 90
Concluding remarks 95
References 96
5. Model peptides and cholesterol
István P. Sugár and Parkson Lee-Gau Chong
On the energy and entropy parameters of the free energy of the membrane 107
References 109
6. Cholesterol and ceramide: An unlikely pair
Aritz B. García-Arribas, Alicia Alonso, and Félix M. Goñi
Introduction: Membrane heterogeneity 111
Sphingolipids and cholesterol: Biological impact 112
Sphingolipids and cholesterol: Membrane biophysics 114
Cholesterol and ceramide in fluid membranes 117
Conclusions and future perspectives 123
References 123
7. Cholesterol-recognizing amino acid consensus motifs in transmembrane proteins: Comparative analysis of in silico studies and structural data
Fodil Azzaz, Henri Chahinian, Nouara Yahi, Coralie Di Scala, Carlos J. Baier, Francisco J. Barrantes, and Jacques Fantini
CARC and CRAC algorithms: Basic principles at work 132
Cholesterol-binding motifs in 3D: What structural studies reveal 134
Moving to the 3rd dimension 139
Conclusion 140
References 141
8. Effects of cholesterol on the GPCR AT1 receptor and its interplay with AT1 antagonists
Sofia Kiriakidi, Zoe Cournia, and Thomas Mavromoustakos
Introduction 148
Experimental results and their discussion 156
Cholesterol influence on the physicochemical properties of the cell membrane in the presence of AT1R and sartans 160
Molecular dynamics (MD) simulations on AT1R embedded in a DPPC:Cholesterol (60:40mol%) bilayer reveal a putative binding site for AT1R blockers on the receptor 161
Conclusions 162
References 163
9. Principles of cholesterol regulation of ion channels
Qiu-Xing Jiang and Irena Levitan
Introduction 169
Lipids as structural components and functional regulators of membrane proteins 173
A thermodynamic consideration of lipid-binding sites on ion channels and their generic effects on the gating of the channels 175
Cholesterol-binding sites on ion channels 177
Functional impact of cholesterol on the activities of ion channels 186
Technical limitations and potential solutions for further developments 195
Conclusions 197
References 198
10. Fluorescent probes for microscopy visualization of cholesterol topography and dynamics in membranes
Francisco J. Barrantes
Introduction 206
The elusive ideal probe 208
Cholesterol probes 208
Indirect cholesterol probes 209
Direct imaging of intrinsic fluorescent cholesterol analogs 211
Concluding remarks 219
References 220
11. Cholesterol transport in blood, lipoproteins, and cholesterol metabolism
Mark T. Mc Auley and Amy E. Morgan
Introduction 228
Lipoproteins an overview 230
Low-density lipoprotein cholesterol 235
High-density lipoprotein cholesterol 236
How does aging impact cholesterol metabolism? 238
Obesity and aging: Two sides of the same coin? 240
Using mathematical modeling to explore cholesterol metabolism 242
Discussion of future perspectives 245
References 247
12. Common laboratory research methods for detection and quantification of cholesterol
Anna N. Bukiya, Hanxuan Li, Steven Mysiewicz, and Wei Li
Introduction to cholesterol detection and quantification 259
The early era of cholesterol detection and quantification: Colorimetric approaches 262
MS-based approaches 264
Imaging-based approaches 270
Enzymatic methods 276
Summary and concluding remarks 279
References 280
13. Approaches for modifying cellular cholesterol levels and their application to mechanistic studies: Examples from the ion channel field
Avia Rosenhouse-Dantsker, Alexandria Slayden, and Anna N. Bukiya
Introduction 290
In vitro methods for modifying cholesterol levels in cell membranes 291
In vivo modification of cholesterol levels 303
Applications to mechanistic studies on the effect of cholesterol on ion channel targets 312
Outlook 319
References 320
SECTION 2
Cholesterol homeostasis and its disruption
14. Diet-induced hypercholesterolemia in small laboratory animal models
Tina Herfel
Introduction 343
Mouse models 344
Rat models 348
Hamsters 349
Guinea pigs 352
Rabbits 354
Additional diet considerations 356
Control diets 358
References 359
15. Nutrition and cholesterol metabolism
Ghada A. Soliman
Introduction 372
Cholesterol structure 373
Nutritional recommendations for maintaining healthy blood cholesterol levels 374
Dietary cholesterol intake 375
Saturated fat intake 377
Cholesterol functions 381
Digestion and absorption of cholesterol 381
Transport of cholesterol 382
Endogenous cholesterol synthesis 384
Blood cholesterol and atherosclerosis 385
Guinea pigs as a preclinical model 386
Advances in cholesterol research 387
Explanatory and predictive models approach 393
Conclusions 394
References 396
16. Cholesterol and early development
Isabella Ellinger and Waranya Chatuphonprasert
Introduction 404
The fetus and cholesterol 406
Steroid hormones in reproduction and early life 414
Extraembryonic structures, secondary yolk sac and placenta, and materno-fetal cholesterol transport 422
Maternal cholesterol levels in healthy pregnancies, maternal hyper- and hypocholesterolemia, and consequences for the fetus 426
Effect of selected medicines related to cholesterol metabolism on the fetal development 430
Summary 436
References 437
17. Clinical and biochemical diagnostic methods: What do blood lipid levels tell us?
Marshall B. Elam
Introduction 452
Use of conventional laboratory methods to measure lipoproteins by determining their cholesterol content 454
Beyond standard lipid profiles: Determination of atherogenic lipoproteins by advanced lipoprotein testing 454
Fasting versus nonfasting lipid measurements 458
Lipid and lipoprotein testing in laboratory animals 460
What do lipid/lipoprotein levels tell us? 460
References 462
18. Familial hypobetalipoproteinemia and abetalipoproteinemia
Diabetes mellitus, metabolic syndrome and HIV 1004
Conclusion 1012
References 1013
Index 1021
Contributors
Luis G. Aguayo Department of Physiology, Universidad de Concepción, Barrio Universitario, Concepción, Chile
Marium Ahmed Department of Rehabilitation, Saint Agnes Hospital, Baltimore, MD, United States
Mohamed H. Ahmed Department of Medicine and HIV Metabolic Clinic, Milton Keynes University Hospital NHS Foundation Trust, Milton Keynes, United Kingdom
Musaab Ahmed College of Medicine; Center of Medical and Bio-allied Health Sciences Research, Ajman University, Ajman, United Arab Emirates
Hélio M.T. Albuquerque LAQV-REQUIMTE, Department of Chemistry, University of Aveiro, Aveiro, Portugal
Alicia Alonso Instituto Biofisika (UPV/EHU, CSIC); Departamento de Bioquímica y Biología Molecular (UPV/EHU), Leioa, Spain
Rana Ashkar Department of Physics and Center for Soft Matter and Biological Physics, Virginia Tech, Blacksburg, VA, United States
Fodil Azzaz INSERM UMR_S 1072; AixMarseille Université, Marseille, France
Carlos J. Baier Laboratory of Toxicology, Instituto de Ciencias Biológicas y Biomédicas del Sur (INBIOSUR), Universidad Nacional del Sur-CONICET, Bahía Blanca, Argentina
Francisco J. Barrantes Laboratory of Molecular Neurobiology, Biomedical Research Institute (BIOMED), Catholic University of Argentina (UCA), National Scientific and Technical Research Council (CONICET), Buenos Aires, Argentina
Małgorzata Bednarska-Makaruk Department of Genetics, Institute of Psychiatry and Neurology, Warsaw, Poland
Andrew S. Bell Section on Clinical Genomics and Experimental Therapeutics, National Institute on Alcohol Abuse and Alcoholism, National Institutes of Health, Bethesda, MD, United States
Asier Benito-Vicente Department of Molecular Biophysics, Biofisika Institute (University of Basque Country and Consejo Superior de Investigaciones Científicas (UPV/EHU, CSIC)); Department of Biochemistry and Molecular Biology, University of the Basque Country (UPV/EHU), Leioa, Spain
Michael F. Brown Department of Chemistry and Biochemistry; Department of Physics, University of Arizona, Tucson, AZ, United States
Anna N. Bukiya Department of Pharmacology, Addiction Science, and Toxicology, College of Medicine, The University of Tennessee Health Science Center, Memphis, TN, United States
Carlos F. Burgos Department of Physiology, Universidad de Concepción, Barrio Universitario, Concepción, Chile
Henri Chahinian INSERM UMR_S 1072; Aix-Marseille Université, Marseille, France
Waranya Chatuphonprasert Faculty of Medicine, Mahasarakham University, Maha Sarakham, Thailand
Parkson Lee-Gau Chong Department of Medical Genetics and Molecular Biochemistry, The Lewis Katz School of Medicine at Temple University, Philadelphia, PA, United States
Arrigo F.G. Cicero Medicine and Surgery Sciences Department, Alma Mater Studiorum University of Bologna; S. Orsola-Malpighi University Hospital, Bologna, Italy
Ivan R. Cincione Department of Clinical and Experimental Medicine, University of Foggia; A.O.U. Ospedali Riuniti, Foggia, Italy
Contributors
Tatiana M. Clemente Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, NE, United States
George A. Cook College of Medicine, University of Tennessee Health Science Center, Memphis, TN, United States
Zoe Cournia Biomedical Research Foundation, Academy of Athens, Athens, Greece
Coralie Di Scala Neuroscience Center-HiLIFE, University of Helsinki, Helsinki, Finland
Milka Doktorova Department of Molecular Physiology and Biological Physics, University of Virginia School of Medicine, Charlottesville, VA, United States
Fathima T. Doole Department of Chemistry and Biochemistry, University of Arizona, Tucson, AZ, United States
Alex M. Dopico Department of Pharmacology, Addiction Science, and Toxicology, College of Medicine, The University of Tennessee Health Science Center, Memphis, TN, United States
Alexandros A. Drosos Rheumatology Clinic, Department of Internal Medicine, Medical School, University of Ioannina, Ioannina, Greece
Hong Du Department of Pathology and Laboratory Medicine; Simon Comprehensive Cancer Center, Indiana University School of Medicine, Indianapolis, IN, United States
Marshall B. Elam Department of Pharmacology and Medicine, Cardiovascular Diseases, University of Tennessee HSC, Memphis, TN, United States
Isabella Ellinger Institute for Pathophysiology and Allergy Research, Center for Pathophysiology, Infectiology and Immunology, Medical University of Vienna, Vienna, Austria
Jacques Fantini INSERM UMR_S 1072; Aix-Marseille Université, Marseille, France
Filipe Ferrari Postgraduate Program in Cardiology and Cardiovascular Sciences, School of Medicine, Federal University of Rio Grande do Sul; Hospital de Clínicas de Porto Alegre, Porto Alegre, Brazil
Unai Galicia-Garcia Department of Molecular Biophysics, Biofisika Institute (University of Basque Country and Consejo Superior de Investigaciones Científicas (UPV/EHU, CSIC)); Department of Molecular Biophysics, Fundación Biofísica Bizkaia, Leioa, Spain
Aritz B. García-Arribas Instituto Biofisika (UPV/EHU, CSIC); Departamento de Bioquímica y Biología Molecular (UPV/EHU), Leioa, Spain
Stacey D. Gilk Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, NE, United States
Félix M. Goñi Instituto Biofisika (UPV/EHU, CSIC); Departamento de Bioquímica y Biología Molecular (UPV/EHU), Leioa, Spain
Juliana Gonzalez-Sanmiguel Department of Physiology, Universidad de Concepción, Barrio Universitario, Concepción, Chile
Gregory A. Grabowski Department of Pediatrics; Department of Molecular Genetics, Biochemistry and Microbiology, University of Cincinnati College of Medicine; The Division of Human Genetics, Cincinnati Children’s Hospital Research Foundation, Cincinnati, OH, United States
Sudipta Gupta Department of Physics and Center for Soft Matter and Biological Physics, Virginia Tech, Blacksburg, VA, United States
Tina Herfel Technical Services, Envigo, Teklad Diets, Madison, WI, United States
DanRong Hu State Key Laboratory of Biotherapy and Cancer Center, West China Hospital, Sichuan University, and Collaborative Innovation Center, Chengdu, Sichuan, PR China
Juyang Huang Department of Physics and Astronomy, Texas Tech University, Lubbock, TX, United States
Shifa Jebari-Benslaiman Department of Molecular Biophysics, Biofisika Institute (University of Basque Country and Consejo Superior de Investigaciones Científicas (UPV/EHU, CSIC)); Department of Biochemistry and Molecular Biology, University of the Basque Country (UPV/EHU), Leioa, Spain
Qiu-Xing Jiang Laboratory of Molecular Physiology and Biophysics, and the Cryo-EM Center, Hauptman-Woodward Medical Research Institute; Departments of Materials Design and Innovation, and Physiology and Biophysics, University of Buffalo, Buffalo, NY; Department of Medicinal Chemistry, College of Pharmacy, University of Florida, Gainesville, FL, United States
Samantha Karr Midwestern University, College of Pharmacy—Glendale, Glendale, AZ; Medical Science Liaison, Sanofi, Bridgewater, NJ, United States
George Khelashvili Department of Physiology and Biophysics; Institute of Computational Biomedicine, Weill Cornell Medical College, New York, NY, United States
Sofia Kiriakidi Department of Chemistry, National and Kapodistrian University of Athens; Biomedical Research Foundation, Academy of Athens, Athens, Greece
Tamas Kovacs Department of Biophysics and Cell Biology, Faculty of Medicine, University of Debrecen, Debrecen, Hungary
Teshani Kumarage Department of Physics and Center for Soft Matter and Biological Physics, Virginia Tech, Blacksburg, VA, United States
Claude K. Lardinois University of Nevada School of Medicine, Reno, NV, United States
Asier Larrea-Sebal Department of Molecular Biophysics, Biofisika Institute (University of Basque Country and Consejo Superior de Investigaciones Científicas (UPV/EHU, CSIC)); Department of Molecular Biophysics, Fundación Biofísica Bizkaia, Leioa, Spain
Irena Levitan Department of Medicine, Pharmacology and Bioengineering, University of Illinois College of Medicine, Chicago, IL, United States
Hanxuan Li Department of Pharmaceutical Science, College of Pharmacy, The University of Tennessee Health Science Center, Memphis, TN, United States
Wei Li Department of Pharmaceutical Science, College of Pharmacy, The University of Tennessee Health Science Center, Memphis, TN, United States
Falk W. Lohoff Section on Clinical Genomics and Experimental Therapeutics, National Institute on Alcohol Abuse and Alcoholism, National Institutes of Health, Bethesda, MD, United States
Agnieszka Ługowska Department of Genetics, Institute of Psychiatry and Neurology, Warsaw, Poland
Cesar Martin Department of Molecular Biophysics, Biofisika Institute (University of Basque Country and Consejo Superior de Investigaciones Científicas (UPV/EHU, CSIC)); Department of Biochemistry and Molecular Biology, University of the Basque Country (UPV/EHU), Leioa, Spain
Vítor M. Martins Hospital de Clínicas de Porto Alegre, Porto Alegre, Brazil
Thomas Mavromoustakos Department of Chemistry, National and Kapodistrian University of Athens, Athens, Greece
Mark T. Mc Auley University of Chester, Chester, United Kingdom
Dushyant Mital Department of HIV and Blood Borne Viruses, Milton Keynes University Hospital NHS Foundation Trust, Milton Keynes, United Kingdom
Bob M. Moore, II College of Pharmacy, University of Tennessee Health Science Center, Memphis, TN, United States
Amy E. Morgan University of Chester, Chester, United Kingdom
Ole G. Mouritsen Department of Food Science, University of Copenhagen, Frederiksberg, Denmark
Steven Mysiewicz Department of Pharmacology, Addiction Science, and Toxicology, College of Medicine, The University of Tennessee Health Science Center, Memphis, TN, United States
Kelsey C. North Department of Pharmacology, Addiction Science, and Toxicology, College of Medicine, The University of Tennessee Health Science Center, Memphis, TN, United States
Emma M. O’Connell Section on Clinical Genomics and Experimental Therapeutics, National Institute on Alcohol Abuse and Alcoholism, National Institutes of Health, Bethesda, MD, United States
Contributors
Marta Pasenkiewicz-Gierula Department of Computational Biophysics and Bioinformatics, Jagiellonian University, Krakow, Poland
ZhiYong Qian State Key Laboratory of Biotherapy and Cancer Center, West China Hospital, Sichuan University, and Collaborative Innovation Center, Chengdu, Sichuan, PR China
Jorge P. Roa Department of Physiology, Universidad de Concepción, Barrio Universitario, Concepción, Chile
Avia Rosenhouse-Dantsker Department of Chemistry, University of Illinois at Chicago, Chicago, IL, United States
Clementina M.M. Santos LAQV-REQUIMTE, Department of Chemistry, University of Aveiro, Aveiro; Centro de Investigação de Montanha (CIMO), Instituto Politécnico de Bragança, Bragança, Portugal
Artur M.S. Silva LAQV-REQUIMTE, Department of Chemistry, University of Aveiro, Aveiro, Portugal
Alexandria Slayden Department of Pharmacology, Addiction Science, and Toxicology, College of Medicine, The University of Tennessee Health Science Center, Memphis, TN, United States
Ghada A. Soliman Department of Environmental, Occupational, and Geospatial Health Sciences, City University of New York (CUNY) Graduate School of Public Health and Health Policy, New York, NY, United States
Natalia Stein Department of Biophysics, Medical College on Wisconsin, Milwaukee, WI, United States
Ricardo Stein Postgraduate Program in Cardiology and Cardiovascular Sciences, School of Medicine, Federal University of Rio Grande do Sul, Porto Alegre, Brazil
Witold K. Subczynski Department of Biophysics, Medical College on Wisconsin, Milwaukee, WI, United States
István P. Sugár Department of Neurology, Icahn School of Medicine at Mount Sinai, New York, NY, United States
Lajos Szente CycloLab Cyclodextrin R&D Laboratory Ltd., Budapest, Hungary
Kepa B. Uribe Center for Cooperative Research in Biomaterials (CIC biomaGUNE), Basque Research and Technology Alliance (BRTA), Donostia San Sebastián, Spain
Zoltan Varga Department of Biophysics and Cell Biology, Faculty of Medicine, University of Debrecen, Debrecen, Hungary
Aliki I. Venetsanopoulou Rheumatology Clinic, Department of Internal Medicine, Medical School, University of Ioannina, Ioannina, Greece
Paraskevi V. Voulgari Rheumatology Clinic, Department of Internal Medicine, Medical School, University of Ioannina, Ioannina, Greece
Francine K. Welty Division of Cardiology, Beth Israel Deaconess Medical Center, Boston, MA, United States
Justyna Widomska Department of Biophysics, Medical University of Lublin, Lublin, Poland
Nouara Yahi INSERM UMR_S 1072; AixMarseille Université, Marseille, France
Zdenek Zadak University Hospital Hradec Kralove and Charles University in Prague, Faculty of Medicine in Hradec Kralove, Hradec Kralove, Czech Republic
Florina Zakany Department of Biophysics and Cell Biology, Faculty of Medicine, University of Debrecen, Debrecen, Hungary
Preface
This project started on an otherwise uneventful day when an Elsevier invitation to edit a book on “cholesterol” landed in the pile of our daily emails. We have edited several books throughout our scientific careers, and working on yet another book in the midst of grant renewal, paper resubmission, fellow mentoring, and other regular loads in the daily life of a scientist sounded quite challenging. Our initial hesitancy to pitch in drastically changed when the first details of the book came to light. Upon discussions with Elsevier, it became apparent that there was not an updated volume that could be called “The Book of Cholesterol” expanding from very basic scientific questions to clinical problems associated with cholesterol dyshomeostasis in humans. Thus, while cholesterol is generously covered in many sources from popular literature to the most highly scholastic research articles, there is no single source of knowledge that is available to the wide audience. This gap constituted the “call of the wild” for us to start and organize this book; we expect that this single volume will connect chemical and biophysical knowledge on cholesterol structure and the function of this vital lipid as a regulator of protein-driven signaling with its relevance to tissue and system physiology, pathology, and medicine. We also expect that the writing style, while driven by experts in the field, would be appealing to a wide readership that starts with curious undergraduate students and culminates with seasoned professionals who work on cholesterol whether at the lab bench or by the hospital bed.
The initial draft of the book chapters’ layout underwent a rigorous review by
Elsevier-recruited reviewers. While they expressed high enthusiasm for the book, they also pointed at several weak spots in our initial vision of the list of topics to include. Following the reviewers’ feedback, the book layout has been significantly expanded. From a collection of chapters, it grew to its current structure comprising three sections. The first introduces readers to “Cholesterol Chemistry and Cell Function.” The section covers fundamental knowledge about cholesterol endogenous synthesis and laboratory bench approaches that led to the current understanding of cholesterol’s role in maintaining biophysical properties of cellular membranes. The second section deals with “Cholesterol Homeostasis and Its Disruption” and provides a detailed account of currently known diseases and conditions associated with disruption of cholesterol biosynthesis, availability, trafficking and metabolism. Finally, the third section provides “Pharmacological Considerations and Perspectives” on currently available therapeutic options for treating cholesterol-related disorders. Our integrative effort is summarized in the book’s title Cholesterol: From Chemistry and Biophysics to the Clinic. As the book covers a huge landscape across many disciplines, some omissions are unavoidable. Conversely, as each chapter is supposed to “stand on its own,” so are the overlaps. It should be noted, however, that repeated topics (e.g., structure of cholesterol, pharmacological actions of statins) are presented from a different standpoint and with a drastically different perspective between chapters.
We are very grateful to all authors who made their contribution to this book for their enthusiasm, spark, and dedicated work on respective chapters. Some of them even helped us by recommending or directly inviting their colleagues to pitch in. We would also like to thank the book proposal reviewers who remained anonymous yet must be entitled to share this book’s success if any. Last but not least, we would like to express our endless gratitude to the Elsevier support team, in particular Barbara Makinster, for their round-theclock work and engagement on this project. Before the first page is flipped, we apologize
to the many brilliant scientists and clinicians that have worked in the cholesterol field who did not author a chapter. We hope that most are cited throughout the book. It is their passion for the cholesterol field that built a decades-long foundation for this book. As for the reader, we hope that you enjoy the journey through this book on cholesterol as much as we did during our writing and editing.
Anna N. Bukiya, Editor
Alex M. Dopico Editor Memphis, TN, United States
Afterword
The main objective of this book has been to present a comprehensive state-of-the-art evaluation of a variety of topics dealing with cholesterol, from its synthetic pathway to therapeutic approaches aimed at fighting disease and conditions associated with cholesterol dyshomeostasis in humans. In doing so, we tried to set the tone to a level that, while delivered by experts on highly specific topics, could be accessible for cross-reading by both specialists and the general readership. Now 36 chapters in the making, we can only hope that we have achieved this goal. One thing to be sure of, however, is that as many hypotheses have been tested in the content of these pages, many more will be raised. Even questions that appeared to be settled keep delivering fertile ground for discovery. Thus, while the biosynthetic paths going from a simple linear structure to the complex heterocyclic molecule that makes cholesterol have been largely resolved, their study is still of great interest for the purpose of creating new drug delivery systems, or selective and sensitive cholesterol-sensing probes. Likewise, cholesterol’s well-understood hydrophobic character is a stepping stone when including sterols as novel carriers to improve drug bioavailability. Collectively, examination of cholesterol biosynthetic pathways, structure, and/or its receptor targets and downstream signaling keeps delivering an increasing set of novel agents aimed at controlling cholesterol levels in the body, whether by inhibiting specific biosynthetic steps, sterol-sequestering, or other methods. Detailed knowledge of cholesterol biosynthetic pathways and homeostasis in the
body has helped researchers and clinicians alike to understand the differences between animal models and humans and the limitation of findings in the former to directly translate onto human diagnosis and therapeutics. Finally, the multiple enzymatic steps involved in cholesterol biosynthesis led to the discovery of a wide variety of genetic disorders with poor prognosis. While genetic therapy has a major role in dealing with these conditions, understanding the underlying common pathophysiological process will eventually help in delivering therapeutic approaches when gene therapy is not amenable or recommended.
Cholesterol’s unique role in animal evolution and cell function, as well as its central place in common human pathologies, stems from the particular chemical features of this sterol, which allows it to organize cell membranes, determine lipid phase equilibria, form lipid domains and myelin, act as a precursor of steroid hormones and vitamin D, regulate protein receptor function and trafficking, and affect both pathophysiology and the biological responses to drugs used to treat human disease, in some cases by directly controlling drug passage through membranes and living cells. So far, the study of cholesterol interactions with signaling molecules has been limited by resolution limitations (e.g., cholesterol concentrations that exert regulatory control of proteins are high enough to challenge conventional binding methods, absence of highresolution protein structures), so a multitude of approaches including magnetic resonance methods, spectroscopy, fluorescence probes,
computational modeling, and systematic mutations in receptor targets have been used to tackle many key interactions between cholesterol and other lipids or proteins; in many cases, each of these methods has been found insufficient yet necessary to answer a given question. As methodologies of higher analytical resolution become available and allow us to explore lipid and protein function in more detail, newer parameters and variables will become more accurate readers of the presence of small amounts of cholesterol in any given biological entity. Unavoidably, newer models of cholesterol interaction with other biological players will be advanced. Should other sterols, such as lanosterol and sitosterol, interact with biological entities in ways different from cholesterol, newer studies might also inform us on whether and how incorporation of such sterols into animal organisms through diet affect normal physiology.
A thorough understanding of cholesterolrelated conditions constitutes an urgent public-health matter that affects all mankind: We are witness to an increasing gap between the haves and haves-not, and while many transmitted diseases have been eradicated or drastically reduced, noncommunicable diseases linked to cholesterol dyshomeostasis are all but growing; they include nutritional deficits leading to demyelinization and poor brain development, obesity and type II diabetes, cardiovascular disease and stroke, colon cancer, and some forms of dementia. Understanding the role of cholesterol in human disease
and eventual therapeutic approaches become particularly challenging when cholesterol dyshomeostasis coexists with another major biological process of its own, such as human body colonization by pathogens (e.g., HIV or SARS-CoV-2 viruses), systemic inflammation (e.g., patients with rheumatoid arthritis at risk for cardiovascular disease), some forms of dementia (Alzheimer’s disease), or substance use disorders. Not only therapeutics become multifactorial but human conditions may also lead to potentiation of cholesterol action and other noxa (e.g., cholesterol levels and alcohol presence during intra utero development).
Since the diseases and conditions stemming from cholesterol dyshomeostasis may present, at least during their early stages, without symptoms, genetic mapping and biochemical profiling (e.g., blood lipid panels that enable accurate detection and quantification of blood cholesterol level and various lipoprotein fractions and subfractions) become an increasing necessity for early intervention and to implement dietary and lifestyle recommendations that adjust to each particular individual. This exemplifies the increasing practice of personalized medicine.
In closing, we also hope that the pages herein conceived will help to further trigger intellectual spark and human health concern around this always intriguing molecule termed cholesterol.
Anna N. Bukiya, Editor
Alex M. Dopico, Editor
SECTION 1
Cholesterol chemistry and cell function
Cholesterol chemistry and laboratory synthesis
Hélio M.T. Albuquerquea, Clementina M.M. Santosa,b, and Artur M.S. Silvaa
aLAQV-REQUIMTE, Department of Chemistry, University of Aveiro, Aveiro, Portugal bCentro de Investigação de Montanha (CIMO), Instituto Politécnico de Bragança, Bragança, Portugal
Abbreviations
Ac acetyl
ABSA acetamidobenzenesulfonyl azide
BBN 9-borabicyclo[3.3.1]nonane
Bn benzyl
Bu butyl
COSY correlation spectroscopy
DEPT distortionless enhancement by polarization transfer
DMSO dimethylsulfoxide
DMF N,N-dimethylformamide
DMAP 4-dimethylaminopyridine
Et ethyl
HMBC Heteronuclear Multiple Bond Correlation
HSQC Heteronuclear Single Quantum Coherence
HMPA hexamethylphosphoramide
IUPAC International Union of Pure and Applied Chemistry
The name cholesterol derives from the Ancient Greek chole- (bile) and stereos (solid), followed by the chemical suffix of the functional group alcohol (-ol). Known also by the name cholesterin, cholesteryl alcohol, cholest-5-en-3β-ol, among others, this interesting natural molecule is a type of modified sterol belonging to the heterogeneous group of organic compounds known as lipids. With a bulky, rigid, and asymmetric structure, the cholesterol skeleton possesses four fused rings aligned from A to D, corresponding to three six-membered and one five-membered. As a whole, the four rings comprise the 1,2-cyclopentane perhydrophenanthrene system (Fig. 1A) (Nes, 2011). The rings are trans-connected and create an almost planar structure (Fig. 1C). The C-18 and C-19 methyl substituents are linked at C-10 and C-13, in relative cis configuration. Due to this structural prolife, the flat face of cholesterol is called the smooth α-face, and all substituents located on this face (in trans-conformation relative to C-19) are called α, while the substituents located on the rough β-face (presence of the two methyl substituents) are called β (in cis-conformation relative to C-19). The cholesterol moiety bears an additional polar 3β-hydroxy group and a C5C6 double bond in B-ring (Róg, Pasenkiewicz-Gierula, Vattulainen, & Karttunen, 2009).
From a chemical point of view, the cholesterol molecule comprises four essential domains (Fig. 1B). The 3-hydroxy group of domain I constitutes not only an important active site for hydrogen bond interactions with several biological molecules but also a versatile functional group for derivatization. In domain II, the absence of methyl groups at C-4 and C-14 influences the planarity of the molecule, and the C5 C6 double bond is an attractive carbon center to several addition reactions. The natural ( R )-configuration at C-20 observed in domain III determines the “right-handed” conformation of the side chain, while in domain IV, the conformation and length of the side chain are of high importance to intermolecular contacts (Cerqueira et al., 2016). The recommended name by the International Union of Pure and Applied Chemistry (IUPAC) for natural cholesterol is (3 S ,8 S ,9 S ,10 R ,13 R ,14 S ,17 R )10,13-dimethyl-17-[(R)-6-methylheptan-2-yl]-2,3,4,7,8,9,11,12,14,15,16,17-dodecahydro-1 Hcyclopenta[ a ]phenanthren-3-ol. In its pure state, it is a white and crystalline powder
FIG. 1 (A) Cholesterol tetracyclic nucleus with numbering of carbon atoms and rings labelling; (B) cholesterol four structural domains; (C) cholesterol crystal structure obtained from https://www.ccdc.cam.ac.uk/structures/ search?id=doi:10.5517/cc66d1t&sid=DataCite
that is odorless and tasteless, with a melting point of 148–149°C (“[cholesterol],”, 2016; Barton, 1976).
Historically, the first identification of cholesterol is attributed to the French chemist François Poulletier de la Salle, who collected it as a crystalline component from human gallstones, in 1769. In 1815, the chemist Michel Eugène Chevreul isolated a crystalline compound of bile stones and named it cholesterine, which was renamed to cholesterol after knowing that the substance was a secondary alcohol. The correct chemical formula of C27H45O was only proposed in 1888 by F. Reinitze, and the first steric representations of the molecule were published by Heinrich Wieland and Adolf Windaus, their efforts leading the two scientists to win the Nobel Prize in Chemistry in 1927 and 1928, respectively (Nes, 2011). The steroid nucleus proposed by Wieland in his Nobel lecture presented some limitations. In 1932, however, his research group corrected it to the skeleton known nowadays (Vaupel, 2007). The research in steroids by Konrad Bloch and Feodor Lynen granted them the Nobel Prize in Physiology or Medicine in 1964, for their discoveries concerning the mechanism and regulation of cholesterol and fatty acid metabolism. Later in 1985, Michael S. Brown and Joseph L. Goldstein were also awarded with the Nobel Prize in Physiology or Medicine for their findings relating to the regulation mechanisms of cholesterol metabolism (“Feodor Lynen—Biographical, 2021,” “Joseph L. Goldstein—Biographical, 2021,” “Konrad Bloch—Biographical, 2021,” “Michael S. Brown—Biographical, 2021”).
Cholesterol is synthesized by all animal cells and is an essential structural component of animal cell membranes, where it contributes to the order of phospholipid chains and overall membrane (dis)order, integrity and heterogeneity. It is also used as a precursor for the biosynthesis of steroid hormones, bile acids and vitamin D (Cerqueira et al., 2016; Ercole, Whittaker, Quinn, & Davis, 2015; Róg et al., 2009). Although cholesterol has eight stereocenters (Fig. 1B) that could rise to 256 stereoisomers, only the natural enantiomer with the (3R,20R)-configurations, is used as a membrane constituent (Xu et al., 2005).
As an amphiphilic molecule, having a hydrophobic hydrocarbon body and a hydrophilic hydroxy headgroup, cholesterol occupies a position at polar-nonpolar interfaces, as observed in cell membranes. The crystal structure of one form of cholesterol monohydrate published by Craven (1976) is based on a local pseudosymmetry arrangement of eight independent molecules in the triclinic cells, similar to the structure reported by Shieh, Hoard, and Nordman (1977) for anhydrous cholesterol crystals at room temperature (25°C). This molecular packing in the crystal structures is in some way in line to the tendency toward double layer structures with an end-for-end arrangement of nearly parallel molecules (Bernal, Crowfoot, & Fankuchen, 1940). On the other hand, cholesterol crystals at 37°C have a remarkably large unit cell containing 16 independent cholesterol molecules, and the transition preserves a closely obeyed pseudosymmetry present in the structure (Hsu & Nordman, 1983). Garti et al. studied phase transitions in cholesterol crystallized from various solvents, characterizing the effect of several solvents (e.g., carbon tetrachloride, acetonitrile, methanol, ethanol) and conditions of crystallization (Garti, Karpuj, & Sarig, 1980). Using differential thermal analysis, infrared spectroscopy and polarization microscopy, Barton had found that the phase transitions of cholesterol and other sterols subjected to heating and cooling in a range of 20°C to + 150°C were dependent on the state of hydration and on the structure of the aliphatic side chain (Barton, 1976).
Below, we will review major milestones in characterization of cholesterol structure, cholesterol laboratory synthesis, and synthetic routes for production of enantiomeric cholesterol.
