Biochemistry of Endothelial Cells Doc. Dr. Mine KUCUR

ENDOTHELIUM Provides a cellular lining to all blood vessels in the circulatory system and forms a structural barrier between the vascular space and the tissue. In adults, the endothelium weights approximately 1kg, comprises 1.6x10 13 cells and has a surface area between 1-7 m 2. Each EC comes into contact with numerous smooth cells.

LUMEN Tunica adventitia Tunica media Tunica intima

Note the individual Endothelial Cells

Vasoconstriction and dilatation NormalVasoconstrictionVasodilatation

VasoconstrictionVasodilatation Vasoconstriction and dilatation ↓ Resistance to flow↑ Resistance to flow

Endothelial Apoptosis Normal Apoptosed

The Endothelium As an Endocrine Organ

The inner lining of our bloods vessels is the Endothelium It plays a central role in regulating the vasomotor tone (vasoregulation) Local homeostasis & control of the coagulation process (provision of anti-thrombotic surface and selective permeability to haematopoietic cells and nutrients) Endothelial cells have ‘Sensors’ and release ‘Mediators’ ‘Mediators’ are the functional molecules on the cell surface The Vascular Endothelium

Endothelium Endothelium is metabolically very active. Endothelium produces a large variety of substances which regulate vascular tone, structure & integrity of blood vessels and also affect the growth and metabolism of surrounding tissue. Total mass of endothelium in a healthy adult man of 70 kg. weight is about 1800 gms-2000gms & considered as the largest endocrine organ (paracrine) of the body.

Endothelium The endothelium produces physiologically active biochemical substances which affect the blood vessels and other tissues & play significant role in health and disease. The substances that cause predominantly vasodilatation: Nitric oxide-Endothelial derived relaxing factor (EDRF) Prostacycline (PGI2) Endothelial derived hyper-polarizing factor (EDHF).

Endothelium The substances that cause predominantly vasoconstriction: Endothelin-1 Angiotensin-II (A-II) Thromboxane-A2 (TXA2) Free radicals Normally, there is delicate balance between vasodilatory and vasoconstricting substances and help to regulate vascular tone of blood vessel.

Vascular Endothelial Mediators Include the following Nitric oxide (NO) Cycloxygenase (CxO) Endothelin-1 (ET-1) Endothelium Depolarisation Factor (EDF) And many others - thus It is the largest endocrine gland

NO is generated in ECs by the oxidation of L-arginine to L-citrulline by a family of enzumes, NO synthases (NOS) Half-life of NO, is affected by its chemical reaction and inactivation by superoxide anion NO is the most abundant free-radical in the body It is the only biological molecule in high concentrations to out-compete superoxide dismutase for superoxide NO has an anti-thrombogenic & anti-atherogenic role Nitric Oxide (NO)

Endothelial NO has the following actions Smooth muscle relaxation and vasodilatation- maintenance of basal vasomotor tone Essential for regulation of blood pressure Reduces proliferation of vascular smooth muscle Protects blood vessel intima from injurious consequences of platelet aggregation-inhibition of thrombosis by inhibiting platelet adhesion, activation and agonist-induced secretion. Protective actions of NO

(L-NMMA) = N(G)-mono-methyl-L-arginine

NO deficiency in the vessel wall promotes Inflammation Oxidation of lipoproteins Smooth muscle proliferation Accumulation of lipid rich material Platelet activation and thrombus formation Finally results in atherosclerosis. ED and NO ↓

Endothelins A family of 21 amino acid peptides. Three members: ET-1,ET-2 and ET-3. Regulate vasomotor tone, cellular proliferation and hormone production. ECs produce only ET-1, also syntesized by vascular smooth cells. Production is induced by hypoxia, ischemia. ET-1 binds to specific receptors on smooth cells: vasoconstriction.

Endothelins  Disease Elevated plasma concentrations of ETs are found in congestive cardiac failure. Plasma ET-1 levels are normal in essential hypertension. Endothelins have been implicated in vascular diseases of the kidney and cyclosporin induced nephrotoxicity. Plasma ET levels are elevated after ischaemic cerebral infarction.

Eicosanoid. Synthesis is induced by disturbances in endothelial function or vascular haemodynamics. Released from ECs and acts in a paracrine manner. Binds to a specific receptor on platelets and vascular smooth muscle cells to limit vasoconstriction and influence platelet deposition. Prostacyclin(PGI 2 )

Phospholipid, remains bound to the EC surface. Most important effect is recruiting leucocytes to the EC surface, and its effects on vascular tone are indirect and exerted through the generation of other eicosanoids and leukotrienes. Platelet Activating Factor (PAF)

Leucocyte transmigration to extravascular sites of inflammation. Initial step is the arrest of leucocytes and random contact with the ECs. Mediated by the selectins. Increasing adhesion occurs with activation of the leucocyte integrins. Leucocytes flatten and migrate along the endothelium: diapedesis. ENDOTHELIUM IN INFLAMMATION

Activated endothelium bind leucocytes. Is activated in response to cytokines including IL-1, TNF-  and lipopolysaccharide which expresses adhesion molecules and bind leucocytes. ENDOTHELIUM IN INFLAMMATION

Three members: E-Selectin, L-Selectin and P- Selectin. Characterized by a C-terminal lectin like domain that binds complex carbohydrates. Involved in leucocyte recruitment to sites of inflammation. Selectins

Integrins relevant to leucocyte recruitment are  1 integrins and the  2 integrins.  1 integrins mediate binding to the ECM (fibronectin and laminin). Binds cell surface VCAM-1.  2 integrins are present only on leucocytes and their activity depends on conformational changes that occuron leucocyte activation. Integrin-immunglobulin superfamily interaction is essential for extravasation to occur. Integrins

ECs maintain anticogulant activity: Prevent activation of thrombin- if activated, stimulates coagulation by causing platelet activation and the activation of several coagulation factors. Express heparan suphate- stimulates antithrombin III. Express tissue factor pathway inhibitor (TFPI)- prevents thrombin formation. COAGULATION

Express thrombomodulin. Thrombomodulin-thrombin interaction activates protein C- strong anticoagulant activity. Synthesize protein S, a cofactor for activated protein C. THE BALANCE IS TOWARDS ANTICOAGULANT FACTORS IN HEALTHY ENDOTHELIUM !!! COAGULATION

The Endothelium in Health and Disease

The Universal Damage Coronary Heart Disease Endothelial Dysfunction  NO ↑ Inflammation ↑ Thrombosis Genes Coronary Risk Factors The Essential Components

endothelial NOS (eNOS) is induced by receptor dependent agonists such as thrombin, bradykinin and substance P. NO causes vascular smooth muscle relaxation by binding to guanyl cyclase. plays a critical role in the inhibition of thrombosis by inhibiting platelet adhesion, activation and agonist- induced secretion. NO promotes platelet desegregation.

Regulatory Functions of the Endothelium NormalDysfunction Vasodilation Vasoconstriction NO, PGI2, EDHF, BK, C-NP ROS, ET-1, TxA2, A-II, PGH2 Thrombolysis Thrombosis Platelet Disaggregation NO, PGI2 Adhesion Molecules CAMs, P,E Selectins Antiproliferation NO, PGI2, TGF- , Hep Growth Factors ET-1, A-II, PDGF, ILGF, ILs Lipolysis Inflammation ROS, NF-  B PAI-1, TF-α, Tx-A2tPA, Protein C, TF-I, vWF LPL Vogel R

Clinical Sequelae

Oxidative stress and Endothelial dysfunction Oxidative Stress leads to ED Endothelial dysfunction is mainly due to reduced bioavailability and bioactivity of Nitric Oxide (NO) It is also a physiological process Takes place gradually by age and menopause.

Oxidant stress and Endothelial dysfunction are major factors for atherosclerosis – the common pathway – for most of the cardiovascular risk factors including Hypertension, DM, Dyslipidemia and Smoking. Both endothelial dysfunction and oxidant stress result in clinical conditions - Heart failure, IHD and MI The Effects of ED

Advances in the understanding of endothelial function have been the basis for many therapeutic strategies. Expanding the understanding of endothelial function will lead to targeted therapies to a myriad of diseases such as cancer, cardiovascular disease and inflammatory conditions. CONCLUSIONS