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: Proteoglycans
http://kentsimmons.uwinnipeg.ca/cm1504
and polysaccharides
 Collagen
 Collagen : most abundant protein found in the
human body. About 1/3rd 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 α-polypeptide chains arranged in
right-handed supercoil
 Glycine, proline, hydroxyproline
 1.5 nm diameter
 At least 28 different collagens found
 The three α-chains could be same
(collagen II) or different (collagen I)
Collagen molecule
 Classification of collagen
1. Fibril-forming collagens
 No interruptions in triple helix
 Regular arrangement results in characteristic “D” period of 67 nm
 Diameter : 50-500 nm
 Example : Types I, II, III, V, XI
 Classification of collagen
2. Network-forming collagens
 Forms network in basement (Collagen IV) and Descemet’s
membrane (Collagen VIII)
 Molecular filtration
 Example : Types IV, VIII, X
 Classification of collagen
3. Fibril-associated collagens with interrupted triple helices (FACITs)

 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
 Classification of collagen
4. Anchoring collagens

 Provides functional integrity by connecting epithelium to

stroma
 Example : Type VII
 Classification of collagen
5. Beaded-filament-forming collagens
 Form structural links with cells
 Example : Type VI
 Collagen VI crosslink into tetramers that assemble into long
molecular chains (microfibrils) and have beaded repeat of 105 nm
 Type I Fibril assembly
Fibril assembly is determined by chain recognition sequence in C-propeptide

Fish scale

Bone osteon

Tendon

Chain recognition sequence

Skin
 Diseases associated with
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)

 Role of MMP in metastasis

Metastasis

Metastasis
 Spread of cancer from a primary tumor to distant sites of the body
 A defining feature of cancer
 Role of MMP in metastasis
 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)
 Role of MMP in metastasis
 Major role of MMPs was to facilitate the breakdown of physical barriers,
thus promoting invasion, intravasation, extravasation and migration
 MMPs targeted for antimetastasis therapies
 Role of MMP in metastasis
 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
 Modification of collagen for
therapeutics
Uses Second-harmonic generation (SHG) for imaging only collagen fibers

Red SHG: Blue

Conditions : Wavelength=800 nm Wavelength=400 nm

Non-centrosymmetric (collagen,
microtubules)
Lasers (high intensity) SAMPLE

Collagen stained red and

Advantages : imaged by fluorescence
Collagen imaged by SHG
microscopy
microscopy
No staining
3D imaging
No photobleaching
 Modification of collagen for
therapeutics

 SHG intensity collected from live imaging of collagen fibers

provides an good estimate of diffusion coefficient in tumors
 Modification of collagen for
therapeutics
0th day 3rd day 6th day 9th day 12th day

 Chronic relaxin treatment degrades tumor matrix

and improve macromolecular diffusion in tumors

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