Endothelial Cells and Angiogenesis - PM132S 2024-25 PDF

Endothelial cells and angiogenesis presented by Dr Ilyas Khan Associate Professor in Regenerative Medicine Swansea University Medical School Anatomy Endothelial cells Vascuologenesis Blood vessels Angiogenesis Pathogenesis ...

Endothelial cells and angiogenesis presented by Dr Ilyas Khan Associate Professor in Regenerative Medicine Swansea University Medical School Anatomy Endothelial cells Vascuologenesis Blood vessels Angiogenesis Pathogenesis  What we will be learning today Vasculogenesis - is the process of de novo blood vessel formation by Anatomy endothelial cells. Endothelial cells - the cells that line the inner surface of blood vessels. Angiogenesis - is the growth of new blood vessels from the existing vasculature. Pathological conditions - tumour angiogenesis and heart disease.  The requirement for a vascular system In the earliest stages of embryonic development tissues are maintained Anatomy by diffusion of gases, nutrients and metabolites. To generate larger and more complex structures an organised means of transferring gases and nutrients is required. The cardiovascular network is the first organ system to develop in the embryo.  How the circulatory system is made Anatomy (angioblasts)  The circulatory system the pulmonary circulation - results in deoxygenated Anatomy blood leaving the heart travelling to the lungs, becoming oxygenated then travelling back to the heart. the systemic circulation is where oxygenated blood leaves the heart and travels through arteries around the body, and as it becomes deoxygenated makes its way back to the heart through veins.  The function of blood vessels carry blood. carry oxygenated blood from the lungs around the body (O2 to tissues). Anatomy remove CO2 from tissues to the lungs. carry nutrients around the body (glucose). remove metabolites from body via kidneys (such as urea). act as a transport network for hormones and the immune system. exchange of materials between the circulatory system and tissues takes place only across the capillary endothelium.  Endothelial cells Endothelial cells are: derived from angioblast stem cells. Anatomy are simple, squamous epithelia. (squamous means scale-like). form the lining of all blood vessels and lymphatic vessels, i.e. they form a barrier between the blood and vessel wall. simple epithelium allows small metabolites to pass through by diffusion and filtration, and it secretes lubricating substances.  Endothelial cell function Endothelial cells have unique functions that include: Anatomy acting as a semi-permeable barrier between plasma and the tissue, fluid filtration, such as in the glomerulus of the kidney, blood vessel tone - cells produce NO gas which cause the smooth muscle surrounding to dilate. haemostasis - the mechanism that leads to cessation of bleeding from a blood vessel. neutrophil recruitment - by expressing P-selectin which is recognised by leukocytes , and hormone trafficking, - through pores, by paracellular transport or transcytosis. Anatomy Types of blood vessels O2-rich blood O2-poor blood away from towards the the heart heart Anatomy Varicose veins - weakness of veins walls and their valves  Properties of arteries, veins and capillaries Arteries: thick walls, small lumens larger arteries have an internal Anatomy elastic layer the smooth muscle is usually the thickest layer the outer layer is connective tissue which binds and protects the artery. Veins: thinner walls, large lumens Capillaries no elastic layers, but have valves single cell layer - endothelial cells. smooth muscle layer is usually thinner than the outer connective surrounded by a basement layer.. tissue layer. Anatomy Capillaries Fenestrated capillaries are When bone marrow forms new Continuous capillaries are common in the small blood cells, the cells must enter characterised by a complete intestine, which is the the blood supply and can only do endothelial lining with tight primary site of nutrient so through the large openings of a junctions (see above) absorption, as well as in the sinusoid capillary. between endothelial cells. kidneys, which filter the sinus - latin meaning cavity in organ or tissue blood.  Perivascular (mural) cells pericytes and smooth muscle cells constitute mural (relating to the wall of a blood vessel) or perivascular cells. pericytes stabilise the capillary Anatomy structure and induce vascular quiescence (state of inactivity) localising primarily at endothelial- endothelial junctions. Platelet-derived growth factor (PDGF) receptor beta is expressed by mural cells and PDGF is expressed by immature endothelial cells. PDGF acts as a chemokine (a chemical or protein that induces movement (kinos) in nearby cells). Pericytes make angiopoietin-1 (Ang-1) which binds Tie-2 receptors Akt is involved in cellular survival pathways, on endothelial cells causing it to by inhibiting apoptotic (death) processes.  Angiogenesis: formation of new blood vessels angiogenesis is the formation of new blood vessels from the pre-existing vasculature. Physiology there are two principal forms of angiogenesis: sprouting: existing Angpt1 Angpt2 endothelial cells in vessels are activated and form sprouts that subsequently connect (anastamosis) to neighbouring vessels. intussusceptive: splitting of an existing blood vessel in two. Observed in neonatal animals. Physiology The stages of sprouting angiogenesis ischaemia - a restriction in blood supply to tissues, causing a shortage of oxygen that is needed for cellular metabolism (to keep cells and tissues alive).  what stimulates angiogenesis? Hypoxia: lack of oxygen causes cells to produce vascular endothelial growth factor Physiology (VEGF). Cells sense the presence of O2 by the ability of an enzyme called) prolyl hydroxylase-2 (PHD2 to add an O2 molecule to proline amino acids in HIF1α (hypoxia inducible factor alpha.) When there is plenty of O2 HIF1α in the cytoplasm is hydroxylated and then recognised by von Hippel- Landau protein (VHL) which ubiquinates it - this targets HIF1α for degradation. When O2 is scarce then unmodified HIF1α migrates to the nucleus where it promotes the expression of VEGF.  Vascular endothelial growth factor (VEGF) VEGF: 5 isoforms A-E Physiology 3 receptors VEGFR1-3 VEGFA and VEGFR2 are the principal angiogenic ligand and receptor h endothelial migration h endothelial proliferation h protease activity creates fenestrations creates vessel lumens  Step 1: endothelial cell activation the first step in angiogenesis is Physiology endothelial cell activation. this is followed by the production of proteases that degrade the perivascular extracellular matrix Angiopoietin-2 (Ang2) is released This leads to the production (connective tissue). from Weibel-Palade bodies and of proteases which break interfere with Ang1-Tie2 stability down the surrounding VEGF also stimulates signals. Loss of Ang1-Tie2 signalling matrix, allowing endothelial the production of nitric causes pericytes to detach and opens cells to migrate towards the endothelial cells up to new signals from chemokine signal. oxide by cells which the surrounding matrix. causes vasodilation and increased permeability. Physiology Step 2: endothelial cell proliferation and migration Physiology Step 3: tubulogenesis (lumen formation) Electrostatic repulsion is the key to lumen formation. CD34 is a glycosylated protein (has many covalently attached sugar units) and is highly negatively charged. The negative charge helps to push apart the luminal surfaces.  Step 4: vessel fusion Anastomosis is the formation of an interconnected luminal space, Physiology allowing the subsequent Anastomosis can occur between two circulation of blood. sprouts and involve two tip cells. Physiology Step 5: vessel maturation Angpt2  Vascular homeostasis Angiogenic activators: Angiogenic inhibitors: Physiology hypoxia. soluble VEGF receptor-1 (sflt-1) VEGF - especially VEGFA. binds VEGFA - it acts as a decoy. FGF-1 - potent inducer of angiogenesis endostatin - fragment derived from in endothelial cells. type XVIII collagen of the basement membrane. Placental growth factor - PIGF is from the same family as VEGF but expressed angiostatin - fragment of a larger mainly during pregnancy. protein, plasmin, that is activated to degrade fibrin clots. Angiopoetin-2 - Ang2 antagonises Ang1-Tie2 signalling between endothelial cells and promotes loss of pericytes. Pathophysiology Tumour angiogenesis Tumour angiogenesis refers to the ability of a tumor to 1. self sufficiency in growth signals - oncogene activation. stimulate new blood vessel formation. 2. evasion of cell death signals - produce IGF growth factors. Enables tumour expansion, local 3. insensitivity to anti-growth signals - invasion, and dissemination. retinoblastoma Rb mutations. Angiogenesis is one of the six 4. limitless proliferation - gain of telomerase cellular transformations that function. lead to malignant growth of 5. sustained angiogenesis - VEGF, FGF tumours. production. 6. tissue invasion and metastasis - inactivation of E-cadherins. Pathophysiology The transition from the prevascular to the vascular phase is referred to as the “angiogenic switch”. Lack of oxygen and nutrients prevents the growth of tumours in excess of 1–2 mm in diameter Pathophysiology Drugs to treat tumour angiogenesis Monoclonal antibodies (mAbs) are used to target either VEGF receptors (1-3), VEGF isoforms (A or C), Ang2, PDGFβ or its receptor. Bevacizumab, Avastin®, blocks tumour cell- derived VEGF-A, impairing the development of new vessels and leading to tumour starvation and, consequently, growth inhibition. Effective inhibition of tumour angiogenesis might arrest or halt tumour progression but would not eradicate the tumour as a stand- alone therapy. Often used in combination with chemotherapy or radiotherapy but causes significant complications.  Heart disease - atherosclerosis Atherosclerosis is a multistep process ranging from endothelial Pathophysiology dysfunction to plaque development, progression, and rupture, leading to thrombus formation and cardiovascular events. Cholesterol is carried around the body by low density lipoproteins (LDLs). LDLs are usually taken out of the circulation by the liver, if not they can cause injury to endothelial cells which have receptors for LDLs. When endothelial cells engulf LDL, they oxidise it to a product called oxidized LDL. Recent evidence suggests that oxidised LDL contributes to endothelial cell injury, migration of monocytes and lymphocytes into the tunica interna, conversion of monocytes into macrophages,  foam cells and platelets encourage the migration and proliferation of smooth muscle cells to the intima, which in turn ingest lipids, become replaced by collagen. Pathophysiology A protective fibrous cap normally forms between the fatty deposits and the artery lining. ruptures of the fibrous cap exposes thrombogenic material, such as collagen - thrombosis is plaque formation by platelets. plaques can block the blood vessel entirely or move into the bloodstream and block smaller low-density lipoproteins (yellow particles) vessels. invade endothelial, and are oxidised (green particles). blocking the blood vessel leads Oxo-LDL causes circulating monocytes to also to an infarction, and can kill cells invade the sub-endothelial space. They after 5 minutes, if it is a become macrophages and mop up the fat myocardial infarction then the (LDLs), as they do they become enlarged and results can be fatal. are known as foam cells.  Endothelial cells and angiogenesis Endothelial cells (ECs) line all blood Endothelial cells are activated to undergo vessels. They are derived from sprouting by the presence of VEGF (vascular angioblast precursor stem cells. endothelial growth factor) which is released by cells Summary that are in low oxygen (hypoxic) environments. The major vessels are arteries, veins In combination with VEGF, angiopoietins are and capillaries. responsible for assembling and disassembling the endothelial lining of blood vessels Perivascular cells such as pericytes stabilise capillaries, and smooth muscle Pericytes detach from activated ECs which then cells add functionality to arteries and digest the basement membrane. Cells then adopt veins. a tip cell fate and migrate towards the increasing VEGF concentration gradient. Angiogenesis occurs by two EC cells further away express PDGFβ and are processes; sprouting angiogenesis - the stabilised by the attachment of pericytes. formation of new vessels from pre- existing ones, or, intussusception - Eventually, EC tip cells merge and undergo splitting of blood vessel along their anastomosis - forming a connection between length by ingress of matrix. surrounding vessels.