ROLE OF ENDOTHELIAL CELL IN HEALTH & DISEASE

SWEETY KALANTRI

Simple Squamous epithelium (monolayer)Thinnest possible type of membrane

Endothelium is thin layer of cells which lies the interior surface of blood vessels and lymphatic vessels

Cells which form endothelium are called endothelial cells (EC’s)

Mesodermal in origin

It forms an interface between circulating blood/lymph in the lumen and rest of the vessel wall

Most quiescent & genetically stable cells of the body- turnover time usually 100 days

HISTORY Endothelium- 1st described by Virchow in capillaries

as a simple membrane with flattened nuclei

Swiss Anatomist Wilhelm His- introduced the term “endothelium”

Waldayer -suggested the term restricting to those cells that make up the innermost layer of blood vessels and lymph vessels & posterior lining of cornea

1st pro-angiogenic factor (bFGF)- purified in 1984 from tumor cells by Sching & Klagsbrun

The vascular wallThe vascular wall

Endothelial cells

Liver

/Sinusoidal

EMBRYOGENESIS

Earliest blood vessels develop from Blood islands (insulae sanguineae) which appear in mesoderm surrounding the wall of the yolk sac at 3 weeks of development

Centre of blood islands form

Hematopoeitic stem cells ( Precursor of all blood cells)

Peripheral hemangioblasts form

Mesodermal cells

HEMANGIOBLAST

ANGIOBLAST(Precursor of blood vessels

FGF- 2

VEGF

Angioblasts proliferate and eventually induced to form EC’s (by VEGF, secreted by surrounding mesoderm cells)

Once process of vasculogenesis estalishes a primary vascular bed, additional vasculature is added by angiogenesis, the sprouting of new vessels (by VEGF)

Maturation & modeling of the vasculature is regulated by other GF’s ( PDGF, TGF-b )

VEGF- A

VEGF

Signalling protein- Mediates both vasculogenesis & angiogenesis

Stimulates-- EC mitogenesis EC migration Enhances microvascular permeability (aka Vascular permeabilty factor

VEGF

VEGFR’s( tyrosine kinase receptors )- present on cell surface

VEGFR-2/Flk 1/KDR

Mediates lymphangiogenesis

VEGFReceptor

KDR ( kinase insert domain receptor 2)- It is human gene

encoding Flk-1

VEGFR-2

KDR is designated as CD 309

Flk -1 (fetal liver kinase 1)

Expression of flk-1 represents the earliest marker of the developing endothelial lineage during vasculogenesis

SCL (Stem cell leukaemia) transcription factor/ TAL- 1 i.e T-cell acute lymphocytic leukaemia= crucial for the development of blood cells and blood vessels

AC 133 / / Prominin - 1= Useful marker for isolation of hematopoeitic & endothelial progenitor cell

(homeobox gene)- marker for both hemangioblast and angioblast

present only in endothelial precursors

flk-1

SCL

CD 133

Hex

CD133 is absent on mature endothelial cells and monocytic cells

Microvasculature (Capillaries & post capillary venules

EC’s Pericytes• Contractile function

• Multipotential capabilities- differentiate to adipocytes, osteoblasts, phagocytes, SMC’s

• Pericytes & SMC’s recruit to form periendothelial layer- for vessel maturation & stabilisation

• Imp for BBB formation

/ Mural /Rouget cells

STRUCTURE Epithelial lining of the vascular system

Almost always Simple squamous epithelium

EC’s are very flat and elongate, have central nucleus, thickness is maximum at the level of nucleus(2-3 µm), 10-20 µm in diameter

Elsewhere-thinner and laminar; in capillaries as thin as 0.2 µm

Elongated in the direction of the blood flow, especially in arteries

Cytoplasm is relatively simple with few organelles; mostly concentrated in the perinuclear zone

At ultra structural level, they have few characteristic organelle

e.g.1) transcytotic/ pinocytotic vesicle 2) caveole 3) Weibel-Palade body

TRANSCYTOTIC VESICLE :

- in all ECs but particularly present in exchange vessels - shuttle small amount of extracellular fluid or blood plasma across the endothelial cytoplasm

-facilitates bulk exchange of dissolved gases, metabolites and nutrients -E.g. in the lung capillaries where there is very efficient movement of gases (carbon dioxide, oxygen and anaesthetics etc).

CAVEOLE :

- special type of transcytotic vesicle

- typical in vessels of smooth muscle cells

- vesicular invagination of cell surface

- associated with receptors, enzymes and ion channels

WEIBEL-PALADE BODY :

-also known as rod-shaped (micro) tubulated bodies

-characterizes EC’s

- elongated cytoplasmic vesicle(3 x 0.1) µm single membrane; dense interior,

-stores adhesion molecule, P- selectin Von Willebrand factor( vWF )

INTERCELLULAR JUNCTION BETWEEN ADJACENT EC’s

Syndesmos

Formed by Cadherin-trans membrane gp

Cell to cell contact & with cytoskeleton

Cadherin 5/ VE cadherin/ Endothelial specific cadherin

In large arteries & brain vessels

Composed of- Occludin, Claudin 5, JAM’s( Junctional adhesion molecules

Present more at the apical region of the cell

Function-Seals neighboring cells together to prevent leakage of molecules between them

c/o Connexons- mainly of 37, 40, 43 (detected in EC’s)

Function-cell to cell junction allowing passage of small water soluble ions & molecules

ENDOTHELIAL HETEROGEINITY

EC’s exhibit different phenotype- both in structure & function

heterogeneity is linked to both intrinsic, i.e., genetic factor, and extrinsic factors

Structural heterogeneity- obtained following electron microscopy observations where differences in intercellular junctions led to the classification of-

# continuous endothelium # fenestrated endothelium # discontinuous endothelium

Endocrine glands,GI mucosaGlomerulus

LiverSpleenBM

Specialized EC’s of post capillary venules

plump cuboidal morphology, have large no. of lymphocytes within their walls, basal lamina is continuous, 7-30µm

A) High Endothelial Venules (HEV)-

HEVs are found in all secondary lymphoid organs except spleen

HEVs enable lymphocytes to move in and out of the lymph nodes from the circulatory system

HEV-expresses addressins (specific adhesion molecules) eg CD34 Mad CAM1

Attaches to L-selectin on Lymphocytes

BLOOD BRAIN BARRIER:BBB-

It is highly selective permeability barrier that prevent the entry of lipid insoluble substances to enter brain, SC & peripheral nerve

BBB is formed by capillary EC’s which are connected by tight junctions & relative lack of transcytotic vesicle

Tightness of barrier depends upon the close apposition of astrocytes (astrocyte cell projections surround the EC’s of BBB)

Circumventricular organs lack it.

LUNG : respiratory membrane

have a selective phagocytic activity & are able to extract substances from blood

KIDNEY : finely fenestrated : functions as a selective filter

Principle barrier(≈33µm)--is the BM, the fused endothelium & podocyte basal lamina

And allows the passage of water, small molecules & ions; but not larger & those with -ve charge

/PECAM-1Receptor for TGF-b

/CD105

7B4 antigenACE (angiotensin-converting enzyme)- endothelial enzymeBNH9/BNF13CD31(PECAM-1)CD34 (Gly CAM 1)CD54 (ICAM-1) CD106 (VCAM-1)CD62P (p-Selectin GMP140)CD105 (Endoglin)CD146 (P1H12)E-selectinEndothelial cell autoantibodiesEndoglyx-1

FEW MARKERS

Endosialin (tumor endothelial marker 1, TEM-1, FB5) (VEGFR-1) ICAM-2 (intercellular adhesion molecule 2) Thrombomodulin (TM, CD141) VEGF (Vascular endothelial growth factor) vWF (von Willebrand factor)

CLINICAL ASSESSMENT OF ENDOTHELIAL FUNCTION

by both invasively and non- invasively

involves evaluating measure of endothelial cell behavior in vivo viz endothelium dependent vasodilatation

done using either pharmacological or mechanical agonist that stimulates endothelium to release effector molecules that alter underlying SM cell tone.

INVASIVE PROCEDURE :

agonist that stimulate release of endothelial NO is used e.g. acetyl choline & methacholine (short lived rapidly acting )

intracoronary infusion is given

Change in coronary diameter is measured

NON- INVASIVE PROCEDURE :

Assessed in the forearm circulation

Brachial artery blood flow is occluded with a BP cuff

Then cuff is deflated

change in blood flow and diameter is measured ultrasonographically

depends upon - -shear stress-dependent changes in endothelial release of NO following restoration of blood flow & -the effect of transient adenosine released from ischemic tissue.

RESULTS:

Normally the change is approx. 10%

Endothelial dysfunction( ED) is defined by- smaller change & in extreme cases ,a paradoxical vasoconstriction effect is also seen.

Occurs due to direct effect of cholinergic agonist on vascular SMC

ED seen in pat with atherosclerosis & risk factors(HTN, ↑cholesterolemia, DM, smoking etc.)

PERMEABILITY BARRIER & TRANSPORT provide barrier between the blood and rest of the body

tissues

Simple diffusion- O2, CO2

Active transport- Glucose, AA’s, electrolytes

Pinocytosis- small molecules, soluble proteins

Receptor mediated endocytosis (clathrin dependent process)-

GF’s, Antibodies, LDL, Transferrin, MHC complexes

SYNTHESIS OF-

Role in VASOMOTION-

1. Prostacyclin (PGI2)2. Endothelium derived hyperpolarizing factor( EDHF)3. Nitric oxide (NO)/ (EDRF)

Vasodilator factors

1. Endothelin-12. Thromboxane (TXA2)3. Angiotensin II

Vasoconstrictive factors

NOS 3 subtype present in EC’s

Secretes ECM protein-

1. Basal lamina- collagen, laminin, elastin, fibronectin 2. Glycocalyx - Proteoglycans

• Smoothness of endothelial surface- due to glycocalyx• Negative charge on EC’s – due to GAG’s (mainly heparan

SO4)

(EC’s binds to ECM via Integrin) Secretion of growth stimulating factors-PDGF, FGF, GM-

CSF

Secretion of growth inhibiting factors- heparin, TGF- β

Secretion of IL-1, IL-6, IL-8

PECAM 1 (CD31)- found on the surface of endothelial cell intercellular junctions, platelets, monocytes, neutrophils, macrophages, lymphocytes, megakaryocytes

involved in leukocyte transmigration, angiogenesis & integrin activation

VCAM-1 (CD106)- expressed on both large and small blood vessels only after EC’s are

stimulated by cytokines

mediates the adhesion of lymphocytes, monocytes, eosinophil, and basophils to vascular endothelium

Major BM addressin for hematopoeitic progenitor cells expressing VLA-4 / integrin α4β4

ICAM (CD54)-

is expressed by the vascular endothelium, macrophages, and lymphocytes.

is a ligand for LFA-1 (integrin) , a receptor found on leukocytes.

stabilizing cell-cell interactions and facilitating leukocyte endothelial transmigration

HEMOSTATIC FUNCTIONS-

Intact endotheliumHomeostatic Phenotype

Damaged endotheliumDysfunctional phenotype

Anticoagulants- production of Thrombomodulin (CD141)- (co-factor for thrombin)

Anti thrombogenic agents- production of prostacyclin, heparin, t PA, anti thrombin III

Pro thrombogenic agents( released after damage to cells)- # tissue thromboplastin, vWF, PAI

ClottingRole in CLOTTING-

Role of EC in normal hemostasis

INFLAMMATION

Leucocyte normally repelled by endothelium(for free flow of blood )

Inflammatory states - leucocytes are attracted to the endothelium by leucocyte adhesion molecules( expressed on EC’s)- leucocyte Margination

They leucocyte pass by diapedesis

Hallmark of inflammation- Increased vascular permeability → edema

Vascular leakage occurs due to contraction of EC (M.C.)

ENDOTHELIUM AS AN ORGAN

1-2 trillion EC’s, forming an almost 1.5 kgs organ

Uniquely contains Weibel-palade bodies (stores vWF)

Not only a permeability barrier, also multifunctional paracrine & endocrine organ

Involved in- immune response, growth regulation, coagulation production of extracellular matrix components modulator of blood flow & blood vessel tone

ROLE IN DISEASE

Oxidative Stress leads to Endothelial dysfunction (ED)

ED- 1) decreased NO

2) increased Endothelin ( ET-1 binds to Endothelin A and B receptors in pulmonary vascular bed- potent vasoconstrictor)

It is also a physiological process

Takes place gradually by age and menopause.

ENDOTHELIUM IN CVS DISEASE

Disease of large & medium sized muscular arteries Characterized by- 1. ED 2. vascular inflammation 3. Atheroma's/atherosclerotic plaque--build up of cholesterol, lipids, cellular debris, calcium & fibrin within the intima.intima

Angiotensin-converting-enzyme(ACE) is an endothelial enzyme

Converts angiotensinogen I to angiotensinogen II

A II is a potent vasoconstrictor: important in pathogenesis of hypertension

Hypertension

SMOKING

Nicotine- opens up intercellular junction & allow large molecules to pass through the wall

Such toxins can potentiate degenerative changes in the blood vessels & lead to vascular disease

Prolonged( years ) smoking of one pack of cigarettes daily or more –daily, increase death rate from IHD by 200%

Smoking cessation decreases that risk substantially

ENDOTHELIUM & STROKE

Production of EDCF- counteracts the normal dilator effect of NO

Reduced activity of NO synthase

Presence of Hemoglobin in SAH- # inhibition of NO- vasospasm

DIABETES

ENDOTHELIUM & CKD

ENDOTHELIUM & SEPSIS

TTP & HUS are caused by insult that activates platelets & deposited as thrombi in microcirculation

Superimposition of endothelial injury may further promote platelet micro aggregate formation : initiate or exacerbate

Thrombotic microangiopathies

PHYSIOLOGICAL ANGIOGENESIS

TUMOR ANGIOGENESIS

In disease , EC growth supports metabolic requirement of tumor beyond few mm : growth of primary & metastatic tumor

Several steps= -stimulation of EC -degradation of ECM -proliferation of EC & migration into tumor -Formation of new capillary tubes

Tumor vessels are= -tortuous -dilated, uneven diameter -excessive branching &shunting -lack perivascular cells

ANTIANGIOGENIC THERAPY

Acquired drug résistance of tumor – due to high intrinsic mutation rate -- major cause of treatment failure

But ECs are genetically stable ; ECs apoptosis pathway is intact

EC provides nourishment to many tumor cells; tumor growth dependent on angiogenesis

blockade of a single GF (e.g. VEGF) may inhibit tumor induced vascular growth

von Willebrand disease

vWF required for interaction & adhesion of platelets to ECM

Genetic absence of this factor( AD, rare AR)→von Willebrand disease

The WHO classification of vascular tumors A)Benign=1) Hemangiomas-sub cut/deep-capillary-cavernous-arteriovenous-Venous-intramuscular-synovial2) Epitheloid Hemangiomas3) Angiomatosis4) Lymphangioma

TUMOR OF ENDOTHELIAL CELLS

B) Intermediate (locally aggressive)-Kaposiform hemangioendothelioma

C) Intermediate (rarely metastasizing)-retiform hemangioendothelioma-papillary intralymphatic angioendothelioma-composite hemangioendothelioma-kaposi sarcoma

D) Malignant -Epitheloid hemangioendothelioma-Angiosarcoma of soft tissue

EC’s play a vital role in health and integrity of every tissue of the body because

apart from cartilage, every cell lies within a few µm of a capillary

The diffusion limit of oxygen in tissue is only ≈ 100µm,and can not cross blood vessel thicker than that

fine capillaries ( 10-15 µm ) & consist merely of endothelial cells and a very fine basal lamina, thus helps in providing oxygen, nutrients & metabolites.

Summary

adjust their number & arrangement to accommodate local requirement

Thus, they are life-support tissue extending & remodeling the network of blood vessels to enable tissue growth, motion & repair.

Dysfunction of EC has been implicated in virtually every type vascular disease( Atherosclerosis,HTN etc.)

and hence integrity & proper function of EC is

essential for proper organ function and good health.

Pathological basis of disease : Robbins & Cotran,8th e Harrison’s internal medicine: 17th e WHO classification: tumors of soft tissue and bone Hematology : Basic Principles & Practise,5th e( Hoffman) Hemostasis and Thrombosis –Basic Principles & Practice, 5th

e( Colman , Goldhaber ) Internet sites

References

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