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 STEM CELLS AND MYOCARDIAL REGENERATION BY Hollie Groves Shaun Alexander RESEARCH PAPER BASED ON PATHOLOGY LECTURES AT MEDLINK 2014 Grade awarded: Pass with merit 1 ABSTRACT Every six minutes someone dies of a heart attack in the UK. One in three people who have a heart attack die before reaching hospital [1]. A heart attack is primarily caused by the death of cardiac tissue-­‐ would it be possible to regenerate the heart in the near future? We believe so! Since the discovery of stem cells in the early 1900s, scientists have proven that stem cells have the remarkable potential to develop into many different cell types in the body during early life and growth and hold endless possibilities and future cures in medicine. Our research paper outlines the prospective use of stem cells, collected from embryonic fluid, to regenerate myocardial tissue after a myocardial infarction. This theory has previously been tested on lab mice and proved a huge success. However, there are many ethical issues surrounding this development, including arguments that encompass the problems with animal testing and using foetal stem cells, which are also highlighted in our research paper. INTRODUCTION TO MYOCARDIAL INFARCTIONS What is a Myocardial Infarction? A heart attack is medically known as a myocardial infarction. It’s a serious medical emergency triggered by the death of myocytes (cardiac cells). The death of myocytes is caused by five main stages. Firstly, cholesterol and other insoluble lipids collect on the inside of a coronary artery. This deposit is called an atheroma, and it narrows the lumen of the artery, restricting blood flow – atherosclerosis. Secondly, the atheroma can collect minerals and become hardened to form a rough plaque. The plaque then weakens the wall of the artery, so the pressure of blood causes a local swelling called an aneurism. If the wall is particularly weak the aneurism may burst causing blood loss and probable death. The plaque can also encourage the formation of a blood clot called a thrombus. Alternatively, a mobile clot from elsewhere in the blood stream (an embolism) can become lodged in the atheroma. The clot grows until it completely blocks the artery, forming a coronary thrombosis. Finally, the thrombosis prevents oxygen reaching the cardiac cells “downstream” of the blockage, so they cannot respire and consequently die. The severity of the heart attack depends on how far along the coronary artery the thrombosis is. If only a small part of one ventricle is killed then the patient will recover, but a thrombosis early in the coronary artery will always be fatal [2]. Figure 1 shows the effect of a myocardial infarction on the myocardial tissue-­‐ you can see the area of death of the myocytes (notice the colour of the myocardial tissue is a deeper red/brown indicating it’s deoxygenated and deprived of nutrients) and the aneurism(s) of the left ventricle wall. Area of death of the myocytes. Aneurism(s) of the left ventricle wall. [Figure 1] How are Myocardial Infarctions currently treated? [3] The treatment options for a myocardial infarction depend on the type of myocardial infarction. An ST segment elevation myocardial infarction (STEMI) is the most serious form of heart attack and requires emergency assessment and treatment. It is important that treatment is rapid to minimise damage to the cardiac tissue. A STEMI shows up on an electrocardiogram (ECG) as a wave that is markedly raised above 2 the baseline (refer to Figure 2). [Figure 2] The treatment used will depend on when the symptoms started and how soon the patient can access treatment. There are three treatment pathways: 1. If symptoms started within the past 12 hours– they will usually be offered primary percutaneous coronary intervention (PCI). 2. If symptoms started within the past 12 hours but they cannot access PCI quickly–
medication will be offered to break down blood clots. 3. If symptoms started more than 12 hours ago– they may be offered a different procedure, especially if symptoms have improved. The best course of treatment will be decided after an angiogram and may include medication, PCI or bypass surgery. Current treatment is effective but there are many implications. Surgical treatment, i.e. PCI or bypass surgery, is very complex and requires specialist staff and equipment; not all hospitals have the facilities needed to perform the surgery. Also, a coronary angioplasty may not be technically possible sometimes if the anatomy of the arteries is different from normal. This may be the case if there are too many narrow sections in the arteries or if there are lots of branches coming off the arteries that are also blocked. In addition, using medications to treat a heart attack is quick and effective but it is a short-­‐term treatment. “What if there’s an alternative treatment which is at present, out of reach? A treatment that is effective in the long-­‐term, easily accessible and self-­‐administered? A treatment that opens the doors to advanced discoveries?” Well, we believe there is and it comes in the form of fourteen micrometre-­‐sized cells-­‐ stem cells! The use of stem cells, collected from embryonic fluid, to regenerate myocardial tissue after a myocardial infarction has recently been discovered. It is in the early stages of testing and could take up to ten years to be classed as a licensed medication by The World Health Organisation (WHO). INTRODUCTION TO STEM CELL RESEARCH [4] Why is Stem Cell Research essential? Stem cell research is important in generating new cells and tissues for repair. A few years ago, scientists began to research the use of stem cells to treat Alzheimer’s, Parkinson’s, Diabetes, Spinal Cord Injury, Stroke and Arthritis. More recently, stem cell research has expanded in the hope that a treatment for myocardial infarctions is originated. Stem cell research is also needed to help us gain understanding of cell development. Furthermore, scientists will be able to test new drugs on generated tissue which will erase the issues that arise with animal testing. “Stem cell research holds enormous promise for easing human suffering, and federal support is critical to its success” ~ Tom Harkin [5] 3 “Stem cell research can revolutionize medicine, more than anything since antibiotics”~ Ron Reagan [5] Stem cell research first began in the early 1900s and since then scientists have discovered their endless potential. Below is a summary of the latest stem cell research: 1908-­‐ The term stem cell is first coined. 1960s-­‐ Evidence of neurogenesis. 2001-­‐ First early human embryos cloned. 2003-­‐ New source of adult stem cells discovered. 1963-­‐ Presence of self-­‐renewing cells in mice 2005-­‐ Paralysed rats are able to walk again discovered. after stem cell therapy. 1978 -­‐ Haematopoietic stem cells discovered in 2006-­‐ First ever artificial liver cells created. human cord blood. 1981-­‐ Mouse ESCs derived from inner cell 2007-­‐ Skin cells in mice are reprogrammed to mass. an embryonic-­‐like state. 1992-­‐ Neural stem cells cultured in vitro. 2008-­‐ First human ESCs created without destruction of the embryo. 1997-­‐ First direct evidence of cancer stem cells. 2009-­‐ New way to make embryonic-­‐like stem cells from adult cells is discovered. 1998-­‐ First human stem cell line derived. 2010-­‐ First human trials of ESCs. 1998-­‐ Germ cells extracted from foetal tissue. 2011-­‐ First stem cells of endangered species produced. 2000s-­‐ Several reports of adult stem cell plasticity. Cells Prokaryotic cells make up single celled organisms whereas eukaryotic cells make up more complicated life forms. A Stem Cell is a eukaryote. Figure 3 is a diagram of a simple eukaryotic cell (animal cell): Cell Membrane: regulates movement of substances in and out of the cell. Cytoplasm: a gel-­‐like substance where most of the chemical reactions happen. Nucleus: houses the DNA: the genetic information, the blueprint for the organism. Mitochondria: sites of aerobic respiration. [Figure 3] Ribosomes: sites of protein manufacture. Deoxyribonucleic Acid (DNA) DNA is the complex chemical that carries genetic information. DNA is contained in chromosomes (Figure 4), which are found in the nucleus of most cells. The gene is the unit of inheritance and different forms of the same gene are called alleles [6]. Human DNA is arranged into forty-­‐six chromosomes in twenty-­‐three pairs. At the ends of chromosomes are telomeres. These shorten whenever a cell divides and this process is called senescence (cellular ageing). Immortal cells like cancer or stem cells do not age. 4 [Figure 4] A group of three base pairs forms a codon and each codon codes for one of twenty amino acids. Proteins are formed from strings of amino acids (polypeptides). There are four bases-­‐ Adenine (A), Thymine (T), Cytosine (C) and Guanine (G)-­‐ bases always pair the same way (A-­‐T, T-­‐A, C-­‐G and G-­‐C). A useful section of DNA is referred to as an exon whereas a junk DNA section, an intron. Protein Production (see Figure 5) Protein is an important component of every cell in the body. The body uses protein to build and repair tissue and to produce enzymes, hormones, and other body chemicals. Protein is an important building block of bones, muscles, cartilage, skin and blood. 1) There are two strands of DNA in the nucleus. 2) Spindle fibres separate the strands and a complementary mRNA strand is transcribed. 3) The mRNA leaves the nucleus (by export proteins) and goes to the ribosomes. 4) At the ribosomes, tRNA molecules attached to amino acids pair up with matching stretches of mRNA. 5) The amino acids are ordered into chains which are proteins. These are called polypeptide chains. Epigenetic (relating to or arising from non-­‐genetic influences on gene expression [7]) change to the DNA changes the level of gene expression. [Figure 5] What are Stem Cells? [8] [Figure 6] 5 Stem cells (Figure 6) are functionally defined as having the capacity to self-­‐renew and the ability to generate differentiated cells. More explicitly, stem cells can generate daughter cells identical to their mother (self-­‐renewal), as well as produce progeny and more restricted potential (differentiated cells). Such a simple and broad definition may be satisfactory for embryonic or foetal stem cells that do not persist for the lifetime of an organism but it breaks down when trying to describe other types of stem cells (e.g. adult stem cells). Another functional parameter that should be included in a definition of stem cells is potency, or its potential to produce differentiated progeny. Stem cells are capable of giving rise indefinitely to more identical cells from which certain other kinds arise from differentiation. Figure 7 shows an example of stem cell division; a single cell divides into two identical daughter cells and those identical cells both divide into two undifferentiated cells (replication). However, some stem cells differentiate into many other cells (differentiation). By contrast, Figure 8 represents the hierarchy of stem cells. Totipotent cells can form all the cell types in a body, plus the extraembryonic or placental cells. [Figure 7] [Figure 8] Embryonic cells within the first couple of cell divisions after fertilisation are the only cells that are totipotent. Pluripotent cells can give rise to all of the cell types that make up the body; embryonic stem cells are considered pluripotent. Multipotent cells can develop into more than one cell type, but are more limited than pluripotent cells; adult stem cells and cord blood stem cells are considered multipotent [9]. Embryonic Stem Cells versus Adult Stem Cells [4] [Figure 9] [Figure 10] There are two types of stem cells that scientists use in their research-­‐ embryonic stem cells and somatic stem cells (stem cells collected from the human body). Embryonic stem cells are pluripotent and are harvested from the early embryo whereas somatic stem cells can be multi-­‐ or pluripotent and are harvested from postnatal tissue. Unlike somatic stem cells, embryonic stem cells are easier to cultivate but are ethically more problematic. Embryonic stem cells (Figure 9) are sourced from the embryo after embryonic development; firstly, the haploid gametes are brought together to form a diploid cell with a full set of chromosomes. The fertilised 6 egg is now called a zygote. Secondly, the egg divides several times into a group of totipotent blastomeres and continues to divide until there’s a solid lump called a morula. After this point, cells begin to specialise. A blastocyst (Figure 11) is an example of a specialised cell-­‐ the inner cell mass of pluripotent cells form the foetal tissue and the outer layer of multipotent cells (called the trophectoderm) forms the extraembryonic structures, such as the placenta. [Figure 11] Somatic (post-­‐natal) stem cells (Figure 10) are found in the brain, the blood, the cornea, the retina, the heart, in fat, skin, dental pulp, bone marrow, blood vessels, skeletal muscle and the intestines. Cord blood is another important source. STEM CELLS AND MYOCARDIAL REGENERATION [10] In contrast to human skeletal muscle, which regenerates injured tissue through the activation of inactive myogenic multipotent adult stem cell populations, cardiac muscle does not retain a large enough cell population to promote muscle cell repair. Fibrotic scar formation causes a loss of muscle. As a result, cardiomyocytes enlarge leading to a heart failure, an increasingly prevalent disease in the World today. In recent years, advances in stem cell research have moved the field of regenerative medicine closer towards using stem cells to regenerate the heart. However, there have been many drawbacks, including the incomplete differentiation of stem cells, the lack of organ-­‐specific stem cell resources in adult/aged organs and graft rejection. Contracting cardiomyocytes have to divide in order for heart growth. The heart is considered fully developed after cell enlargement. The rate of cell division is reduced to counteract the effects of a myocardial infarction. The reversal of cardiac damage requires a new generation of contractile myocytes and a capillary network able to support its greater demands. Currently, there is a lack of new myocytes but scientists are working on collecting some from the replication of pre-­‐existing myocytes. Myocardial regeneration is always happening but at a very slow rate, therefore the research could be opening doors to a rarely visited natural process. Could the use of stem cell technology further this process? [Figure 12] This image of the fluorescence microscope depicts a section of the heart tissue of a mouse. The green colouring of the cells in the middle shows that the cell originated from a so-­‐called Sca1 stem cell. The lack of information about the origin of cardiac cells has encouraged animal experiments in which stem cells were collected from bone marrow and analysed for their regenerative properties suitable for restoring cardiac function. 7 The outcome of human trials cannot be assessed by the success of animal experimentation; the anatomy and functioning of a mouse/rat heart in comparison to a human heart is quite different. For example, a rodent’s heart beats 300 to 600 times per minute whereas human cardiomyocytes beat 60 to 100 times per minute. In our opinion, a rodent is therefore an unrealistic model. On the other hand, animal tests have demonstrated an improvement in stem cell technology because a significant reduction in graft rejection has been proven and recorded. In general, all risks have been reduced over previous years. Human cardiomyocytes (originating from embryonic stem cells) have shown, through analysis, that they have the ability to contract and share the same specific fibre and protein structure as the adult stem cells. Embryonic stem cells are pluripotent and differentiate into cardiomyocytes. The limited restorative capacity of the adult mammalian heart has been attributed to the loss of cardiomyocyte adaptation after birth. This is why stem cells are collected from cord blood. Human clinical trials started in 2001 to treat cardiac ischemia (the decreased blood flow and oxygen to the heart muscle) in patients with a lack of stem cells. It did work-­‐ as in it accelerated the recovery process/period-­‐ but the effects were not long-­‐term. As evidence, the initial trials showed an improvement in the global percentage of blood leaving the left ventricle as it contracted from 6% to 9%. These results were confirmed in a larger, double-­‐blind, randomised trial called REPAIR-­‐AMI. In this study the global and regional ejection fraction in the bone marrow-­‐treated group improved modestly but significantly, by 2.9% compared to the placebo-­‐treated group. Despite that, long-­‐term studies showed that the beneficial effect of injected bone marrow cells observed after 6 months were no longer present at 18 months. The regenerative potential of the human heart is a rapidly evolving concept. In the near future, cardiac repair is likely to be made hugely important in the field of myocardial surgery. Multipotent stem cells differentiate into vascular and muscular tissue with the capability of restoring the three major cell types of the myocardium-­‐ myocytes, smooth muscle and endothelial vascular cells [11]. The success of trials has improved yet the biggest problem is still that there is an insufficient number of stem cells originating from within the organism. For stem cells to be able to regenerate the heart, we need to replicate more multipotent stem cells. IS THE RESEARCH INTO ‘STEM CELLS AND MYOCARDIAL REGENERATION’ ETHICAL? Using different sources of human embryonic stem cells for research raises different ethical problems. Experimenting on embryos created for in vitro fertilization but left unused, or embryos created especially for research raises ethical questions. In the first case – whether using “spare” human embryos for research means a lack of respect for the beginning of human life, and in the second – whether creation of embryos for research is morally worse than experimentation on already created, but unused human embryos. The possibility of therapeutic cloning also raises the question as to whether it is ethical to create human embryos for therapeutic purposes. When balancing the possible benefit of embryonic stem cell research inventing new therapies, and the ethical problems raised by this research, a question is posed as to whether there are any equally effective alternatives to research on viable human embryos that could avoid or at least decrease these problems [12]. Firstly, religious groups claim that because embryonic stem cell research requires the subsequent destruction of the embryos used, it is a form of abortion. They maintain that creating embryos for the sole purpose of commercial use, followed by their disposal is morally unacceptable, and strongly oppose the area of study while adult and cord stem cells are readily available instead. Also, with any developing form of medical research, the biggest practical risk is the unknown. Because stem cells injected into a patient are permanent, long term side effects may not be fully understood for years. A French study ten years ago found that recipients of genetically altered bone marrow transplants developed leukaemia years after their allegedly successful transplants had cured their severe combined 8 immunodeficiency, one of many examples demonstrating the possibility of long term unforeseen consequences of treatments like these that do not fade the way most medicines do with time. Moving on, significant embryonic stem cell research carried out on rats by the California-­‐based research company “Geron” found a successful cure for paralysis, with the appearance of small cysts in the area near the site where the embryonic cells were injected – while research claims the cysts are harmless and water filled, the implications of a similar occurrence in humans over a long period of time remains questionable. Furthermore, a number of studies have found the injection of embryonic stem cells to result in minor miscalculations resulting in the growth of strange objects such as teeth, bones and hair in areas where they were not intended, and often resembling tumours. With embryonic cells, these growths continue for the remainder of the patient’s life. Approximately 20% of rats injected with embryonic stem cells later die from some form of cancerous tumour. CONCLUSION Indubitably, stem cells epitomise the advancement of our medical and scientific knowledge effectively through the numerous ways that it can help to treat a patient. Four decades after Neil Armstrong was able to step foot on the moon it is remarkable to realise that the new frontier is to come from under the microscope as opposed to beyond the clouds. According to the Heart Research Institute, every six minutes someone dies of a heart attack in the UK, henceforth despite the ethical issues raised about stem cell research, the number of modern day medical issues can be dealt with more effectively. To conclude, more research has to be carried out in order to expand our knowledge of prospective stem cell utilisation. Reinstating a point made beforehand, there is a lack of organ-­‐specific stem cell resources (specifically in adult/aged organs) and therefore to enable stem cell research to continue to thrive, more people need to donate their stem cells. Subsequently more cures will be found and, as a result, more lives will be saved. It will take many years until new developments and discoveries make a significant difference but every stem cell will be worth it! More information about donating cord blood to aid stem cell research is found on The NHS Cord Blood Bank (NHSBT) website: http://www.nhsbt.nhs.uk/cordblood/faq/ [13]. [Figure 13] More information about donating stem cells or bone marrow can be found on the Anthony Nolan website: http://www.anthonynolan.org/8-­‐ways-­‐you-­‐
could-­‐save-­‐life/donate-­‐your-­‐stem-­‐cells [14]. [Figure 14] 9 REFERENCES • Background reading-­‐ Goldstein, L.S.B., Schneider, M. ‘What are Stem Cells? (Part of the Stem Cells For Dummies Cheat Sheet)’. http://www.dummies.com/how-­‐to/content/what-­‐are-­‐stem-­‐cells.html 1) “Every six minutes someone dies of a heart attack in the UK. One in three people who have a heart attack die before reaching hospital”, facts by HRI (UK) (Heart Research Institute UK). http://www.hriuk.org/about-­‐heart-­‐disease/heart-­‐facts/ 2) AQA AS Biology Unit 1, Reference to Pages 58 & 59 (The five main stages of a Myocardial Infarction). https://chemstuff.files.wordpress.com/2014/12/unit_1_notes.pdf 3) Information on treating a heart attack, NHS Choices. http://www.nhs.uk/Conditions/Heart-­‐attack/Pages/Treatment.aspx 4) Introduction to Stem Cells, Medlink. http://medlink-­‐uk.net/wp-­‐content/uploads/2015/01/stem-­‐cells-­‐intro.pdf 5) Quotes about Stem Cell research, Google images. https://www.google.co.uk/search?q=medical+quotes+about+myocardial+infarction&biw=1366&
bih=643&source=lnms&tbm=isch&sa=X&ei=Hnv8VIK0NsK0acCbgJAI&ved=0CAYQ_AUoAQ#tbm=
isch&q=medical+quotes+about+stem+cells 6) DNA, BBC GCSE Bitesize. http://www.bbc.co.uk/schools/gcsebitesize/science/edexcel_pre_2011/genes/dnarev1.shtml 7) Definition of Epigenetic, Google. https://www.google.co.uk/?gws_rd=ssl#q=epigenetic+definition 8) Lanza, R. (2013) Essentials of Stem Cell Biology Third Edition, Academic Press. Reference to Chapter 2 (‘Stemness’: Definitions, Criteria, and Standards), Melton, D., Harvard University. https://books.google.co.uk/books?id=aR6cAAAAQBAJ&printsec=frontcover&dq=what+are+stem
+cells&hl=en&sa=X&ei=8lvLVILAPMKs7AaxtYCwCw&sqi=2&ved=0CEAQ6AEwAg#v=onepage&q=
what%20are%20stem%20cells&f=false 9) Definition of ‘totipotent’ and ‘pluripotent’. http://stemcell.ny.gov/faqs/what-­‐difference-­‐between-­‐totipotent-­‐pluripotent-­‐and-­‐multipotent 10)
Lanza, R. (2009) Essentials of Stem Cell Biology, Academic Press. Reference to Chapter 31, Pages 259-­‐263, Santini, M. Poudel, B. Rosenthal, N. https://books.google.co.uk/books?id=auyfanDzHlIC&pg=PA542&dq=stem+cells+myocardial&hl=
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%20v=onepage&q=stem%20cells%20myocardial&f=false#v=onepage&q&f=false 11)
What type of cells do CSCs differentiate into? http://www.ncbi.nlm.nih.gov/pubmed/16050255 12)
Hug, K. Department of Social Medicine, Faculty of Public Health, Kaunas University of Medicine, Lithuania (2005) ‘Sources of human embryos for stem cell research: ethical problems and their possible solutions’. www.eurostemcell.org 13)
The NHS Cord Blood Bank website (NHSBT): http://www.nhsbt.nhs.uk/cordblood/faq/ 14)
Anthony Nolan website: http://www.anthonynolan.org/8-­‐ways-­‐you-­‐could-­‐save-­‐life/donate-­‐your-­‐stem-­‐cells 10 FIGURES 1) Drawing of the heart showing anterior left ventricle wall infarction. http://upload.wikimedia.org/wikipedia/commons/9/92/Heart_ant_wall_infarction.jpg 2) An ECG showing a STEMI. http://www.rbain.org.uk/Normal12leadecg.jpg 3) Diagram of a simple Eukaryotic cell (animal cell). http://cache4.asset-­‐cache.net/gc/165814604-­‐diagram-­‐of-­‐plant-­‐and-­‐animal-­‐cells-­‐
gettyimages.jpg?v=1&c=IWSAsset&k=2&d=ojj5rY53fzfyxpZgjtUE%2BYVIr7D7%2FqwFCf6osUJT
%2F88%3D 4) Chromosomes under a microscope. https://www.ocf.berkeley.edu/~edy/genome/chromosomes.jpg 5) Protein production. http://dharmacon.gelifesciences.com/uploadedImages/Applications/Gene_Expression/gene-­‐
expression-­‐pathway-­‐lg.png 6) Stem cells under a microscope. http://www.cfne.unimelb.edu.au/research-­‐laboratories/stem-­‐cells/images/stem-­‐cells01.jpg 7) Stem Cell Division Example. http://blog.canacad.ac.jp/wpmu/15matser/files/2013/08/What-­‐Is-­‐Download1.jpg 8) Hierarchy of Stem Cells. http://www.chxa.com/img/stem-­‐cells-­‐hierarchy.jpg 9) Embryonic Stem Cells under a microscope. http://www.laboratoryequipment.com/sites/laboratoryequipment.com/files/legacyimages/Reso
urces/Featured_Editorial/2011/06/062911_stem.jpg 10)
Somatic Stem Cells (from Human Bone-­‐Marrow) under a microscope. http://epigenome.eu/media/images/large/40.jpg 11)
A Blastocyst under a microscope. http://www.advancedfertility.com/images/hatched-­‐blastocyst.jpg 12)
A section of the heart tissue of a mouse under a microscope. http://www.mpg.de/7634230/heart-­‐stem-­‐cells 13)
NHSBT Campaign Poster. http://www.nhsbt.nhs.uk/cordblood/images/content/life_save.gif 14)
Anthony Nolan Campaign Poster. http://www.anthonynolan.org/8-­‐ways-­‐you-­‐could-­‐save-­‐life/donate-­‐your-­‐stem-­‐cells 11