Epithelial CELL JUNCTIONS /endodontic courses

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CELL JUNCTIONS

INDIAN DENTAL ACADEMYLeader in continuing Dental Education


REFERENCES

Textbook of Basic pathology. Robins 5th edition“Oral cells and tissues" : P.R Garrant ; 1st edition.Antonio Nanci, Tencate’s Oral Histology ,

Development, Structure, Function , 7th editionEssentials of medical physiology – K

SembulingamBasics of histology – Ham’s


LEARNING OBJECTIVES

At the end of seminar, one should know -Classification of intercellular junctionsTight junctionsAdhesive junctionsDesmosomesHemidesmosomesGap junctions


CONTENTS

Classification of intercellular junctionsTight junctionsAdhesive junctionsDesmosomesHemidesmosomesGap junctions


INTERCELLULAR JUNCTIONS

When cells come into contact with one

another , and sometimes with the

extracellular matrix , specialized

junctions may form at specific sites on

the contacting cell membranes.


CLASSIFICATION

CELL JUNCTIONS

TIGHT JUNCTIONS

ADHERENS JUNCTIONS

GAP JUNCTIONS


ADHESIVE JUNCTIONS

CELL TO CELL

ZONULA ADHERENS

DESMOSOMES

CELL TO MATRIX

FOCAL ADHESIONS

HEMIDESMOSOMES



TIGHT JUNCTIONS It is also known as ‘ZONULA

OCCLUDENS’.Zonula means junction that completely

encircles the cellsHere the membranes of the two cells

become opposed and the outer layer of the membrane fuses.

Thus obliterating the space between the cells.

These junctions are seen typically along the apical margins of the cells.


Epithelia are sheets of cells that provide the interface between masses of cells and a cavity or space (a lumen).

The portion of the cell exposed to the lumen is called its apical surface.


The rest of the cell (i.e., its sides and base) make up the basolateral surface.

Tight junctions seal adjacent epithelial cells in a narrow band just beneath their apical surface.


Tightly aligned rows of tight junction proteins serve to stitch the membrane together effectively sealing the association between adjacent cells.

This serves to block the movement of materials through the intercellular space.


On the molecular level , intercellular junctions typically consist of 3 components -

TRANSMEMBRANE ADHESIVE PROTEIN

CYTOPLASMIC ADAPTERPROTEIN

CYTOSKELETAL FILAMENT


TRANSMEMBRANE ADHESIVE PROTEINS

Transmembrane adhesive proteins include –

- OCCLUDIN

- MEMBERS OF THE CLAUDIN FAMILY

- JUNCTIONAL ADHESION MOLECULE


Tight Junction Proteins


Described for the first time in 1993 by Shoichiro Tsukita.

It is the main component of the tight junctions, along with the members of claudin family.


Transmembrane adhesive proteins interact homotypically with the same proteins on the adjacent cell.


CYTOPLASMIC ADAPTER PROTEINS

Associate with the intracellular portions of the transmembrane proteins.

These include – - cell polarity related proteins - vesicular transport related proteins - kinases - transcription factors - tumor suppression proteins


Some of the cytoplasmic proteins of the

tight junctions bind to the actin

filaments.


FUNCTIONS OF TIGHT JUNCTIONS

They hold cells together.


They prevent the passage of molecules and ions through the space between cells. This pathway provides control over what substances are allowed through.


They block the movement of integral membrane proteins (red and green ovals) between the apical and basolateral surfaces of the cell.

Thus the special functions of each surface, for example - receptor-mediated endocytosis at the apical surface - exocytosis at the basolateral surface can be preserved.


ADHESIVE JUNCTIONS


Adherens junctions are protein complexes that occur at cell–cell junctions in epithelial tissues, usually more basal than tight junctions.


CELL ADHESION MOLECULES

Cells adhere to each other and to the extracellular matrix through cell-surface proteins called cell adhesion molecules (CAMs).

Cell Adhesion Molecules (CAMs) are proteins located on the cell surface involved in binding with other cells or with the extracellular matrix(ECM) in the process called cell adhesion.


These proteins are typically transmembrane receptors and are composed of three domains:

An intracellular domain that interacts with the cytoskeleton,

A transmembrane domain, And an extracellular domain that interacts

either with other CAMs of the same kind (homophilic binding) or with other CAMs or the extracellular matrix (heterophilic binding).


CELL ADHESION COMPLEXES

Adhesion receptorsExtra cellular matrix

molecules{ECM}Adhesion plaque proteins


CELL ADHESION RECEPTORS: Transmembrane glycoproteins that mediate binding to extra cellular matrix molecules or to counter receptors on other cells.

EXTRACELLULAR MATRIX PROTEINS: Usually fibrillar in nature and provide a complex structural and functional network that can interact simultaneously with multiple cell surface receptors.


INTRACELLULAR PLAQUE PROTEINS:

Provide structural and functional linkages between adhesion receptors and the actin microfilaments,microtubules and intermediate filaments of the cytoskeleton.


FAMILIES OF CAMsCalcium-independent

IgSF CAMs Integrins Lymphocyte homing receptors

Calcium-dependent Cadherins Selectins


Cell Adhesion Molecules (CAMs)


CYTOPLASMIC ADAPTER PROTEINSThe cytoplasmic adapter proteins are

members of the catenin family.Catenins are proteins found in

complexes with cadherin cell adhesion molecules of animal cells. The first two catenins that were identified became known as alpha-catenin and beta-catenin.


Alpha-catenin can bind to beta-catenin and can also bind actin.

Beta-catenin binds the cytoplasmic domain of some cadherins.

Additional catenins such as gamma-catenin and delta-catenin have been identified.

The name "catenin" was originally selected ('catena' means 'chain' in Latin) because it was suspected that catenins might link cadherins to the cytoskeleton



TYPES OF CATENINS

Alpha - cateninBeta-cateninDelta-cateninGamma-catenin


Catenins interact with the cytoplasmic domain of the transmembrane cadherin molecule , with the cytoskeleton and with a number of other proteins including kinases and tumor suppressor molecules that are associated with other junctions.


ZONULA ADHERENS

Cadherin family member : E Cadherin

Cytoplasmic adapter proteins : α catenin and

β catenin

Cytoskeleton componennt : Actin filaments


The catenins and actin filaments are concentrated on the cytoplasmic side of the cell membrane at the zonula adherens to form a dense web that is continuous with the terminal web of actin at the apical end of the cells.


Other transmembrane adhesive proteins present in adhesive junctions are –

NECTIN : member of imunoglobulin superfamily.

Nectin has an important role during junction formation, establishing the initial adhesion site and recruiting E cadherin and other proteins to the junction.


• Other cytoplasmic proteins associated with the zonula adherens include –

- p120 catenin: a signaling molecule associated with E cadherin that is important in stabilizing the junction. - Afadin: which links nectin to the actin cytoskeleton - Vinculin and α actinin which are actin

binding proteins. - Ponsin : which links afadin and vinculin


DESMOSOMES

Desmosomes are essential adhesion structures in most epithelia that link the intermediate filament network of one cell to its neighbour, thereby forming a strong bond.


Also known as macula adherens junctions or spot desmosomes


STRUCTURE OF DESMOSOMES

Desmosome consists of three component -

The Transmembrane molecules The Plaque molecules or cytoplasmic

adaptor protein Intermediate filament (IF)

Extracellularly, desmosomes are separated by a 25-30 nm space.


• The interaction of the transmembrane proteins with those from the adjacent cell results in a dense line in the middle of the intercellular space at the demosome


In desmosomes – CADHERINS : desmoglein desmocollin CATENINS : desmoplakin plakoglobin plakophillin This forms an electron dense plaque on

the cytoplasmic side of the desmosome.This plaque serves as an attachment site

for the cytoskeleton components,which in the case of the desmosome are intermediate filaments.


FUNCTIONS OF DESMOSOMES

Desmosomes create a transcellular network throughout a tissue that resist forces of mechanical stress. Hence found in stratified squamous epithelia and myocardium subjected to constant stress.

The desmosome- intermediate filament complex impart tensile strength and resilience to the epithelium


ASSOCIATED PATHOLOGY

Desmosomes are adhesive intercellular junctions prominent in the oral epithelium, skin and heart. Loss of desmosome function is associated with severe congenital and acquired disorders characterized by tissue fragility.

Diseases associated are- Pemphigus Drug-induced pemphigus


PEMPHIGUS

Mutations in genes encoding desmosomal components and autoantibodies directed against the desmosomal cadherins lead to severe epidermal blistering disorders termed pemphigus.


Pemphigus includes a group of autoimmune blistering diseases of skin and mucous membrane characterized histologically by intradermal blisters due to damage to the intercellular area leads to separation of the keratinocytes – acantholysis and immunologically by circulating immunoglobin G ( IgG ) antibody directed against the cell surface of keratinocytes.


The three primary subsets of pemphigus :-

Pemphigus vulgaris ( PV ) Pemphigus foliaceus Paraneoplastic pemphigus

Oral epithelial desmosomal components differ somewhat from those of skin.

The cadherin-type adhesion molecules desmoglein 1 (Dsg1) and Dsg3 are both expressed in skin but in oral epithelium only the 130 kDa molecule Dsg3 is preferentially expressed .


Desmosomes and CarcinomaDesmosomes are intercellular junctions that

have been shown to be down-regulated in certain types of carcinomas and that may play a role in invasion and metastasis.

Immunohistochemical staining for the major desmosomal glycoprotein desmoglein (Dsg) and desmoplakin (Dp) is reduced in some cases of squamous cell carcinoma (SCC) of the head and neck, and that reduced staining correlates with lymph node involvement & is associated with invasion.


EPITHELIAL ATTACHMENTS IN CARCINOMA


FOCAL ADHESIONS

Located at the basal poleAttaches the cell to the basement

membraneFound in epithelial tissues


Here , transmembrane component is a member of the integrin family of adhesion molecules.

The cytoplasmic adapter proteins which include the actin binding proteins α actinin, vinculin and talin, link the transmembrane integrins to the actin cytoskeleton.



HEMIDESMOSOMESHemidesmosomes are specialised junctional complexes that contribute to the attachment of epithelial cell to the underlying BM in stratified and other complex epithelia.


Hemidesmosomes (HD) are very small stud- or rivet-like structures on the inner basal surface of keratinocytes in the epidermis of skin.

While desmosomes link two cells together, hemidesmosomes attach one cell to the extracellular matrix.

Rather than using cadherins, hemidesmosomes use integrin cell adhesion proteins.

Hemidesmosomes are asymmetrical and are found in epithelial cells connecting the basal face to other cells.


STRUCTUREUltra structurally, they appear as small

electron-dense domains on the plasma membrane of the ventral surface of basal keratinocytes.

Hemidesmosomes are associated with a sub-basal dense plate in the lamina lucida and are connected via fine thread-like anchoring filaments to the lamina densa.

The hemidesmosomal plaque, anchoring filaments, and anchoring fibrils constitute a functional unit termed the hemidesmosomal adhesion complex.


Three classes of proteins are involved in the molecular organization of this complex:

(a)Cytoplasmic plaque proteins,

(b) Transmembrane proteins, (c) Basement membrane associated

proteins


Transmembrane adhesive proteins are – integrin α 6 β 4 which binds specifically to the basal lamina glycoprotein laminin and collagen XVII ( BP 180)

Cytoplasmic adapter proteins are bullous pemhigoid antigen 230 ( BP 230) and plectin form a dense plaque on the cytoplasmic surface of the hemidesmosome which functions as an attachment for intermediate filaments.



FUNCTIONS

These multiprotein complexes determine cell-stromal coherence (Plaque components BP230 and Plectin connect intermediate filament to plasma membrane).

Help in spatial organization of cells.


Important for tissue architecture (α6β4 integrin & BP180 involved in assembly of hemidesmosomes and mediate cell adhesion).

Play an important role in tissue morphogenesis and wound healing.

Serve as signalling devices (α6β4 integrin).


DISORDERS

AUTOIMMUNE DISEASES

PEMPHIGOID

Term used to indicate a desquamating or blistering disease in which epithelial separation takes place at the level of the basement membrane.


When immune deposits (mainly IgG and C3) were detected in some patients along the epithelial basement membrane zone of stratified squamous epithelium, the term pemphigoid was coined.

Structural weakness may be caused by the binding of autoantibodies to components of anchoring complex or by aberrant expression of the components can lead to submucosal blistering.


MUCOUS MEMBRANE PEMPHIGOID

• The clinical phenotype that is acquired and consists of vesicles, bullae or erosions, or both, and that mainly affects mucosa, often in the mouth, is called mucous membrane pemphigoid (MMP).

• Frequently affects women.

• At least 10 different and distinct components of the basement membrane zone have now been identified as autoantigens .


The pathogenesis of MMP probably

includes autoantibody induced complement-mediated sequestration of leucocytes (mainly neutrophils).

Resultant release of cytokines and leucocyte enzymes.

Detachment of the basal cells from the basement membrane zone.

Possibly some complement-mediated lysis of cells.


MUCOUS MEMRANE PEMPHIGOID- SHOWS CHARACTERISTIC SUBEPITHELIAL CLEFTING.


BULLOUS PEMPHIGOID

Chronic subepithelial blistering disease affecting predominantly older individuals.

Oral lesions occur in 20-30% patients.

Patients with BP have circulating IgG autoantibodies against BP230 and BP180.

Circulating IgE and IgA antibasement membrane antibodies have also been detected.


PATHOGENESIS

There is strong evidence that binding of pemphigoid antibodies to the basement membrane zone may be followed by complement activation which lead to inflammatory injury and blister formation.

Leucocytes bind to the basement membrane subsequently causing tissue damage.


SQUAMOUS CELL CARCINOMA

A loss, or abnormal expression, of integrins(α6β4, α2β1, and α3β1) in basal cells is a characteristic of oral squamous cell carcinomas.

However the de novo expression of αvβ6 in epithelial cells at the leading edge of SCC tumors suggests that adhesive contacts between αvβ6 and tenascin promote tumor invasiveness.


In vitro studies indicate that the addition of αvβ6 to SCC cells decreases anchorage-independent growth while stimulating cell differentiation.

Thus oral squamous carcinoma cells demonstrate reduced expression of α2, α3, α6, and α6β4 integrin subunits, thereby decreasing their adhesion to basal laminae and enhancing their ability to invade connective tissues.


GAP JUNCTIONS


GAP JUNCTIONS

Gap junctions are plaque like regions of the cell membrane.

Cell membranes are 2 to 3 nm apart.Transmembrane proteins : connexin

family.These form aqueous channels between

the cytoplasm of adjacent cells.


Six connexin molecules forms a connexon which has a central channel approx 2 nm in diameter.

The connexons in one cell pair with connexons in the adjacent cell to create a patent channel.

Small molecules such as ions and signaling molecules can move readily from one cell to another



Properties Allows for direct electrical communication between cells, although different connexin subunits can impart different single channel conductances. Allows for chemical communication between cells, through the transmission of small second messengers, such as inositol triphosphate(IP3) and calcium (Ca2+), although different connexin subunits can impart different selectivity for particular small molecules.


Generally allows molecules smaller than 1,000 Daltons to pass through, although different connexin subunits can impart different pore sizes and different charge selectivity. Large biomolecules, for example, nucleic acid and protein, are precluded from cytoplasmic transfer between cells. Ensures that molecules and current passing through the gap junction do not leak into the intercellular space.


APPLIED Heart

Gap junctions are particularly important in cardiac muscle: the signal to contract is passed efficiently through gap junctions, allowing the heart muscle cells to contract.


Mutations of the connexin genes have been identified as the basis for certain types of deafness , congenital cataracts , a demyelinating disease and occulodentaldigital dysplasia, a disease that exhibits craniofacial abnormalities , syndactly , conductive hearing loss and hair and nail abnormalities.


IHC MARKERS FOR CELL JUNCTIONS


MARKER EXPRESSION IN OSCCSyndecan –I Decreased P-cad Positivity is corelated with

better prognosisE-cad Decreased E/P/N-cad E-cad: underexpressed in

80%; P-cad: overexpressed in 53%; N-cad: negative in 63%.

α/β/y -catenin Absent or Reduced expression

CK8 Positive CK1,5/6,8/1810,14,19

The expression of CK 8/18 and CK 19 were overall correlated with a poor prognosis

CD44 Increased intensity was seen in 90% of tumours.

MMP17 Increased expressionMMP9 4-fold increase in expressionLaminin-5 Positive


ENDOTHELIAL JUNCTIONS Junctions in endothelial and epithelial

cells share common features.

In the endothelium, junctional complexes comprise tight junctions, adherens junctions, and gap junctions.

Epithelial cells also form desmosomes, which are absent in the endothelium.


Endothelial cells adhere to one another through junctional structures formed by transmembrane adhesive proteins that are responsible for homophilic cell-to-cell adhesion.

These transmembrane proteins are linked to specific intracellularpartners,which mediate anchorage to the actincytoskeleton and as a consequence,stabilize junctions.


• In endothelial cells, the junctional architecture is less defined and, along the cleft, adhesive junctions are intermingled with tight junctions, unlike the epithelial cells.

• The organization of endothelial junctions varies along the vascular tree in function of organ-specific requirements.


MOLECULAR COMPONENTS OF A ADHERENS JUNCTION

The cadherin-catenin complex

The nectin-afadin complex


CADHERINEndothelial cells express a specific

cadherin called vascular endothelial (VE)-cadherin (77).

This protein cannot be found in any other cell type.

VE- cadherin is present in all endothelial

cells of all types of vessels.

Besides,N-cadherin,V-cadherin and P-cadherin are also present .


CATENINSβ-cateninsPlakoglobin (gamma-catenin)α-cateninp120Vinculinα –actinin


THE NECTIN-AFADIN COMPLEX

The complex consists of at least three components,i.e.

Nectin,

Afadin and

Ponsin



TIGHT JUNCTIONS1. Integral membrane

proteins: Occludins(ZO-1) Claudins Junctional adhesion

molecule-A2. Intracellular

proteins ZO-2 ZO-3
