Lecture 3: Degradable Materials with Biological Recognition Last time Theory of hydrolytic polymer erosion Enzymatic degradation of polymers Designing Biodegradable Macromolecules Today Biological recognition in vivo Engineering biological recognition of biomaterials: cell adhesion/migration Readin S.E. Sakiyama-Elbert and J. A Hubbell, ' Functional Biomaterials: Design of Novel biomaterials, Annu. Rev. Mater. Sci. 31, 183-201(2001) J.C. Schense et aL., 'Enzymatic incorporation of bioactive peptides into fibrin matrices enhances neurite extension, Nat. Biotech. 18, 415-419 (2000) Supplementary Reading: The Extracellular Matrix, pp. 1124-1150, Molecular Biology of the Cel, Lodish et al Biological Recognition in vivo Interactions of cells with their environment at the molecular level ECM extracellular matrix Motivation Cell interactions with simple synthetic materials are governed by nonspecific interactions o e.g. surface energies; hydrophobic interactions, charge-charge interactions DRAW OXIDE SURFACE. POL YMER SURFACE but this is not how cells interact with ecm Cells use receptor-receptor/receptor-ligand interactions to guide their functions, including o Adhesion, migration o Differentiation Secretion of molecules Binding of molecules Functions of ECM · Mechanical Support cues for cell survival/function o anchorage-dependent cell growth o differentiation cues organization of tissue o control of tissue morphology, localization of cell types Lecture 3- Biological Recognition 1of15
Lecture 3: Degradable Materials with Biological Recognition Last time: Theory of hydrolytic polymer erosion Enzymatic degradation of polymers Designing Biodegradable Macromolecules Today: Biological recognition in vivo Engineering biological recognition of biomaterials: cell adhesion/migration Reading: S.E. Sakiyama-Elbert and J.A. Hubbell, ‘Functional Biomaterials: Design of Novel biomaterials,’ Annu. Rev. Mater. Sci. 31, 183-201 (2001) J.C. Schense et al., ‘Enzymatic incorporation of bioactive peptides into fibrin matrices enhances neurite extension,’ Nat. Biotech. 18, 415-419 (2000) Supplementary Reading: ‘The Extracellular Matrix,’ pp. 1124-1150, Molecular Biology of the Cell, Lodish et al. Biological Recognition in vivo Interactions of cells with their environment at the molecular level ECM = extracellular matrix Motivation: • Cell interactions with simple synthetic materials are governed by nonspecific interactions: o e.g. surface energies; hydrophobic interactions, charge-charge interactions o DRAW OXIDE SURFACE, POLYMER SURFACE • …but this is not how cells interact with ECM • Cells use receptor-receptor/receptor-ligand interactions to guide their functions, including: o Adhesion, migration o Growth o Differentiation Secretion of molecules Binding of molecules Specialized functions Functions of ECM: • Mechanical Support • cues for cell survival/function o anchorage-dependent cell growth o differentiation cues • organization of tissue o control of tissue morphology, localization of cell types Lecture 3 – Biological Recognition 1 of 15
Structure of native ecm scaffold Prototy pical soft tissue ECM(varies from tissue to tissue) o collagen o other fibers SLIDE) reticular network collagen gel TEM of collagen fibril generalized matrix assembly o fibrils assemble into fibers fibers may be organized or isotropic, and form tight(-10 nm separation between fibers)or open (20-30 um between fibers)meshes o adhesion proteins decorate'fibers o other signals(cytokines, etc )may also be sequestered on fiber surfaces Sequestered Cytokines/ Chemokines/ Collagen fiber hydrogel Adhesion motifs P Friedl et al. Eur J Immuno. 28, 2331(1998) collagen triple helices Lecture 3- Biological Recognition 2of15
Structure of native ECM scaffold Prototypical soft tissue ECM (varies from tissue to tissue): • structural fibers o collagen o other fibers? (SLIDE) collagen gel reticular network of lymph node TEM of collagen fibril • generalized matrix assembly: o fibrils assemble into fibers fibers may be organized or isotropic, and form tight (~10 nm separation between fibers) or open (20-30 µm between fibers) meshes o adhesion proteins ‘decorate’ fibers o other signals (cytokines, etc.) may also be sequestered on fiber surfaces Collagen fiber hydrogel P. Friedl et al.: Eur. J. Immunol. 28, 2331 (1998). Collagen triple helices Collagen fiber Adhesion motifs Sequestered Cytokines/ Chemokines/ Etc. Lecture 3 – Biological Recognition 2 of 15
· Other major matrⅸ k-structure proteins · Key length scales ter of collagen fibrils 50-200nm o diameter of collagen fibers 05-5pm o diameter of collagen triple helices: o diameter ofof collagen chain o length of collagen triple helix 300nm Adhesion proteins Collagen I fibril ligand binding site map H!! lal Binds to surface recep:Drs on 400000 Binds to Heparan sulfate prote Laminin Lodish) o Adhesion proteins designed to bind to structural ECM components, and present binding sites to receptors o Adhesion proteins can present multiple binding sites for different receptors that work Lecture 3- Biological Recognition 3of15
• Other major matrix-structure proteins: o Elastin • Key length scales: o diameter of collagen fibrils: 50-200 nm o diameter of collagen fibers: 0.5-5 µm o diameter of collagen triple helices: o diameter of of collagen chain: o length of collagen triple helix: 300 nm • Adhesion proteins o Complexity of adhesion proteins (SLIDE) Collagen I fibril ligand binding site map laminin structure 1 (from Lodish) o Adhesion proteins designed to bind to structural ECM components, and present binding sites to receptors o Adhesion proteins can present multiple binding sites for different receptors that work in synergy Lecture 3 – Biological Recognition 3 of 15
Interactions of cells with native ecm Signals from the extracellular environment ￥ Extracellular matrix DRAW ON BOARD Cell adhesion: integrin-mediated cell-ECM interactions Cells interact with specific adhesion motifs in adhesion proteins via cell surface receptors; the microsctructure of ECM protein arrangement and its composition can tune cell adhesion o Adhesion in turn regulates growth, differentiation, and migration o Major family of cell-ECM receptors: integrins Composed of noncovalently-associated a and B chains Actin filaments(cytoskeleton integrin (Extracellular space) Adhesion protein ECM fiber o inside-out signaling: biochemical signal triggers affinity change in integrins o focal contacts and signaling integrin clustering drives actin filament assembly and can signal through multiple biochemical pathways, some of which synergize with growth factors to tell the cell where it is Lecture 3- Biological Recognition 4of15
Interactions of cells with native ECM • Signals from the extracellular environment: DRAW ON BOARD: ¥Growth factors ¥Cytokines ¥Extracellular matrix ¥Chemokines Cell adhesion: integrin-mediated cell-ECM interactions • Cells interact with specific adhesion motifs in adhesion proteins via cell surface receptors; the microsctructure of ECM protein arrangement and its composition can tune cell adhesion o Adhesion in turn regulates growth, differentiation, and migration o Major family of cell-ECM receptors: integrins Composed of noncovalently-associated α and β chains Actin filaments (cytoskeleton) α β Ca++ Ca++ Ca++ Ca++ Ca++ Ca++ Ca++ Ca++ Ca++ Ca++ Ca++ Ca++ integrin ECM fiber Adhesion protein Cell membrane (Extracellular space) • Integrins and signaling o ‘inside-out’ signaling: biochemical signal triggers affinity change in integrins o focal contacts and signaling integrin clustering drives actin filament assembly and can signal through multiple biochemical pathways, some of which synergize with growth factors to tell the cell ‘where it is’ Lecture 3 – Biological Recognition 4 of 15
ON BOARD o Length scales in cell adhesion o Size of integrins, focal contacts, relationship to cell size, fiber spacing DRAW ON BOARD focal contacts"(SLIDEl 12-15nm 12-15nm Actin stress fibers LXSN LV3SN Interference reflection microscopy lark spots indicate cell-substrate eparations 50 nm 20 different integrins known, many different pairs possible with different ligand specificities, thus cell-specific adhesion can be modulated by ECM Lecture 3- Biological Recognition 5of15
Ca++ Ca++ Ca++ Ca++ Ca++ Ca ++ Ca+ Ca++ ON BOARD: o Length scales in cell adhesion: o Size of integrins, focal contacts, relationship to cell size, fiber spacing DRAW ON BOARD Integrin structure (Lodish) focal contacts2 (SLIDE) Actin stress fibers Interference reflection microscopy: dark spots indicate cell-substrate separations < 50 nm (stress fibers pics from Maheshwari et al.) • 20 different integrins known, many different pairs possible with different ligand specificities, thus cell-specific adhesion can be modulated by ECM Lecture 3 – Biological Recognition 5 of 15