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The Extracellular Matrix

Lecture 4

Objectives

! Understanding of the constituents of the ECM and the role it plays in physiological conditions.

! What do we need to know for tissue engineering.

! Understanding the types of extracellular matrices and unique characteristics for each.

! The challenge of mimicking nature.

! Knowledge on how cells interact with the matrix for signaling.

! Can we signal them artificially?

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ECM: Menu Summary

! Glycosaminoglycans (GAGs)

! Packing materials, ground substances

! Cartilages

! Collagen

! Insoluble, high tensile strength! Tendons, ligaments, cartilage, cornea

! Elastin

! Elastic

! vascular wall, skin, lung

! Fibronectin

! Cell adhesion

! Laminin

! Cell adhesion

Extracellular Matrix (ECM)

! Tissues are not made up of solely of cells.

! A substantial volume of their space is

extracellular and filled by an intricate

network of macromolecules called ECM.

! ECM is the non-cellular material present

between cells throughout the body.

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Roles of ECM

! Provide structural support.

! Provide substrates for cell adhesion.

! Regulates cellular differentiation and metabolic function.

! The ECM is composed of a variety of polysaccharides and fibrous proteins.

Types of Tissues

! Epithelial tissue: Cells are tightly attached

together to form sheets (epithelia).

! ECM is scanty and forms the basal lamina.

! Connective tissues

! The matrix carries the mechanical stress to which

the tissue is subjected.

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ECM of the Connective tissue

! Variation in the relative amounts of the different types of matrix macromolecules and the way they are organized give an amazing diversity of forms adapting the requirement of the tissue.

! Examples…….

! The ECM is made and oriented by the cells within it. Thus it is local macromolecular synthesis.

! The cells also pattern the matrix.

! ECM is mainly secreted by fibroblasts but other cells in the family can also make it. Can you think these cells?

The two main classes of ECM

! Polysaccharide chains of the class called Glycosaminoglycans (GAGs) often found covalently linked to protein (i.e. proteoglycans).

! Proteins

! Structural (collagen and elastin) fibrous protein

! Adhesive proteins (fibronectin and laminin)

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Glycosaminoglycans

! They are unbranched polysaccharides with

repeating disaccharide units.

! They resist compression and fill space. Why?

ECM: GAGs

! They are called GAGs because one or two sugar residues in the repeating unit are an amino sugar(N-acetylglucosamine, N-acetylgalactosamine) which are sulfated.

! The other sugar is uronic acid (Glucuronic acid, iduronic acid).

! The consequences of sulfate and carboxylatecombinations is a high overall negative charge.

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!Given the diversity……what else can you think?

ECM: GAGs Summary

! Glycosaminoglycans (GAGs)

! Negatively charged

! Extended conformation

! Attract cations

! Water is sucked to the matrix

! Difficult to compress

! Packing materials, ground substances

! Cartilages, cornea

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ECM: Summary

! Glycosaminoglycans (GAGs)

! Packing materials, ground substances

! Cartilages, cornea

! Collagen

! Insoluble, high tensile strength

! Tendons, ligaments, cartilage, cornea

! Elastin

! Insoluble elastic protein

! vascular wall, skin, lung

! Fibronectin and Laminin

! Cell adhesion

ECM: Fibrous Protein

! Collagen, Elastin, Fibronectin and Laminin.

! Collagen is a structural connective tissue accounting 30% of total body protein.

! (Gly-X-Y) is the primary amino acid sequence, whereby X is oftenproline. It also has two rare amino acids.

! 1000 amino acid residue per chain

! Many types of collagen super-family exist. Types I-III are fiber forming while IV is membrane type.

! Synthesized by fibroblasts, osteoblasts and condrocytes.

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Primary amino acid sequence of collagen

-Gly-Pro-Met-Gly-Pro-Ser-Gly-Pro-Arg-

-Gly-Leu-Hyp-Gly-Pro-Hyp-Gly-Ala-Hyp-

-Gly-Pro-Gln-Gly-Phe-Gln-Gly-Pro-Hyp-

-Gly-Glu-Hyp-Gly-Glu-Hyp-Gly-Ala-Ser-

-Gly-Pro-Met-Gly-Pro-Arg-Gly-Pro-Hyp-

-Gly-Pro-Hyp-Gly-Lys-Asn-Gly-Asp-

Transcription Translation

Post-transnational modification

Procollagen assembly (triple helix)

Hydroxylation

Glycosylation

Enzymatic action

Fibril formationCollagen fiber

Collagen BiosynthesisCollagen Biosynthesis

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Crosslinking methods of collagen

!Extracellular crosslinking

!aldol condensation

!hydroxypyridinium

!Physical crosslinking

!ultraviolet (UV)

! dehydrothermal (interchain)

!Chemical crosslinking

!aldehyde (FA, GA)

!epoxy

!isocyanate

!carbodiimide/NHS

! azide

!others least important.

Collagen as scaffold for tissue engineering

What we wanted to do….

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Extracellular crosslinking

Deamination

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! In contrast to GAGs that resist compression, collagen

fibrils resist tensile forces.

! Cells pattern ECM specially collagen.The cells can

crawl over it and tug on it helping to compact into sheets

and draw it out into cables.

! When fibroblasts are cultured on collagen gel, cells tug on

it eventually forming either a fiber or shrinking it from its

original size (mechanical role).

! Note the challenge we face in synthetic gels.

Properties of Collagen Fiber

Elastin! Hydrophobic protein of 750 amino acid residue.

! Contain proline and glycine like collagen but the protein is not glycosylated and has only very small hydroxylation. It is secreted in the ribosome and crosslinked at the cell surface.

! Gives tissues (bladder, skin, lung blood vessels) their elasticity as they need to be strong to function in a dynamic environment. It gives the required resilience so that they can recoil after the stretch force is released.

! Elastin fibers can stretch 5 times more than rubber bands at thesame cross-sectional area.

! Large inelastic collagen fibrils are interwoven to limit the stretching and prevent the tissue from tearing.

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Courtesy of M. L. Raghavan, University of Iowa

Courtesy of M. L. Raghavan, University of Iowa

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Courtesy of M. L. Raghavan, University of Iowa

Courtesy of M. L. Raghavan, University of Iowa

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ECM: Summary! Glycosaminoglycans (GAGs)

! Packing materials, ground substances

! Cartilages, cornea

! Collagen

! Insoluble, high tensile strength

! Tendons, ligaments, cartilage, cornea

! Elastin

! Insoluble elastic protein

! vascular wall, skin, lung

! Fibronectin and Laminin

! Cell adhesion