Extra Cellular Matrix Proteins and Proteinases

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EXTRACELLULAR MATRIX PROTEINS AND

PROTEINASES

By,

Raghu Ambekar

Photonics Research of Bio/nano Environments

Department of Electrical & Computer Engineering

University of Illinois Urbana - Champaign

BioE 506



Outline

Extracellular matrix proteins

Collagen

Classification

Fibril assembly and collagen diseases

Extracellular matrix proteinases

Role of MMP in metastasis

Modification of tumor collagen for therapeutics



Extracellular matrix (ECM)

Surrounds cell

Provides mechanical support

Controls the flow of nutrients

and signals to the cells

Consists of

Fibrous: collagen,

elastin, fibronectin, laminin

Non-fibrous: Proteoglycansand polysaccharides

http://kentsimmons.uwinnipeg.ca/cm1504



Collagen

Collagen : most abundant protein found in the human

body. About 1/3 rd of the total proteins.

Found abundantly in tendon, cartilage, bone and skin

Functions: cell migration

cell adhesion

molecular filtration tissue repair



Structure of collagen

It has a triple-helix structure containing

three -polypeptide chains arranged in

right-handed supercoil

Glycine, proline, hydroxyproline

1.5 nm diameter

At least 28 different collagens found

The three -chains could be same

(collagen II) or different (collagen I)Collagen molecule



Classification of collagen

No interruptions in triple helix

Regular arrangement results in characteristic ³D´ period of 67 nm

Diameter : 50-500 nm Example : Types I, II, III, V, XI

1. Fibril-forming collagens




Forms network in basement (Collagen IV) and Descemet¶s

membrane (Collagen VIII)

Molecular filtration

Example : Types IV, VIII, X

2. Network-forming collagens




Short collagens with interruptions

Linked to collagen II and carries a GAG chain

Found at the surface of fibril-forming collagens

Example : Types IX, XII, XIV

3. Fibril-associated collagens with interrupted triple helices (FACITs)




Provides functional integrity by connecting epithelium to

stroma

Example : Type VII

4. Anchoring collagens




Form structural links with cells

Example : Type VI

Collagen VI crosslink into tetramers that assemble into longmolecular chains (microfibrils) and have beaded repeat of 105 nm

5. Beaded-filament-forming collagens



Type I Fibril assembly

Chain recognition sequence

Fibril assembly is determined by chain recognition sequence in C-propeptide

Fish scale

Bone osteon

Tendon

Skin



seases assoc ate w t

collagen

Diseases caused by mutations

Subtypes of osteogenesis imperfecta (collagen I)

Ehlers-Danlos syndrome (collagen I and V)

Alport syndrome (collagen IV)

Certain arterial aneurysms (collagen III)

Ullrich muscular dystrophy (collagen VI)

Certain chondrodysplasias (collagen IX and XI)

Kniest dysplasia (collagen II)




Metastasis

Metastasis

Spread of cancer from a primary tumor to distant sites of the body

A defining feature of cancer




Understanding the molecular mechanisms of metastasis is crucial for the

design of therapeutics

Extracellular matrix metalloproteinases (MMP) associated with metastasis

MMPs are capable of digesting ECM and basement membrane under

physiologic conditions

Collagenases degrade fibrillar collagen

Stromelysins degrade proteoglycans and glycoproteins

Gelatinases degrade nonfibrillar and denatured collagens

At tumor sites, experiments have found

Increased number of MMPs

Increased levels of MMPs

Reduced levels of TIMPs (Tissue inhibitors of metalloproteinases)




Major role of MMPs was to facilitate the breakdown of physical barriers,

thus promoting invasion, intravasation, extravasation and migration

MMPs targeted for antimetastasis therapies




Clinical trials of inhibiting MMPs to cure cancer have failed

Metastasis is a complicated process

MMPs contribute to every stage in tumor progression at both

primary and metastatic sites

Specific MMPs play a role in each stage of metastasis

MMP 13, 14 ± invasion

MMP 9± angiogenesis

Understand the role of the MMPs in each cancer setting



Modification of collagen for therapeutics

Structure and content of collagen governs the delivery of

therapeutic molecules in tumors

Penetration of therapeutic molecules improved by

developing agents that modify ECM and increase diffusion

Detect tumor collagen noninvasively to quantify collagen

content and estimate drug delivery characteristics




Uses Second-harmonic generation (SHG) for imaging only collagen fibers

Conditions :

Non-centrosymmetric (collagen,

microtubules)

Lasers (high intensity)

Advantages :

No staining

3D imaging

No photobleaching

SAMPLE

Red

Wavelength=800 nm

SHG: Blue

Wavelength=400 nm

Collagen stained red and

imaged by fluorescence

microscopy

Collagen imaged by SHG

microscopy




SHG intensity collected from live imaging of collagen fibers provides an good estimate of diffusion coefficient in tumors




Chronic relaxin treatment degrades tumor matrix

and improve macromolecular diffusion in tumors

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Extra Cellular Matrix Proteins and Proteinases