Scaffolds For Tissue Engineering Applications - Third Review (28 Mar 2009)

Scaffolds for Bone-Tissue Engineering: Preparation, Characterization, Modeling Department of Chemical Engineering and Materials Science Amrita School of Engineering Coimbatore – 641 105 April 2009

Third Review Divya Haridas Karthikeyan G Krishna Priya C Premika G

By (CB105PE012) (CB105PE023) (CB105PE025) (CB105PE028)

Guide Dr. Murali Rangarajan. Ph.D Co-Guide Dr. Nikhil K Kothurkar. Ph.D

Concept of Tissue Engineering n

Life science and Engineering dealing with the development of biological substitutes that restore, maintain and improve tissue functions or a whole organ

Polymeric Scaffolds n

Three dimensional scaffolds play important roles as extracellular matrices onto which cells can attach, grow, and form new tissues

Characterization of Scaffolds

Inferences n

Both PLLA and PU scaffolds are highly porous

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PLLA scaffolds are lighter compared to PU scaffolds

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High density of PU scaffolds are suitable for implantable urinary tracts and bladders

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PLLA scaffolds can serve as a substrate for osteoblasts adhesion, provided these scaffolds are modified with ECM proteins such as collagen which improves the strength and toughness of the scaffolds.

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Modifying the scaffold by adding HAp will improve growth of bone tissue

Characterization of Scaffolds Porous PLLA scaffolds at 100X magnification characterized by optical microscopy

Characterization of Scaffolds Porous PU scaffolds at 100X magnification characterized by optical microscopy

Characterization of Scaffolds Porous PLLA scaffolds at 200X magnification characterized by optical microscopy

Characterization of Scaffolds Porous PU scaffolds at 200X magnification characterized by optical microscopy

Inferences n

The images obtained from optical microscopy show the presence of three dimensional interconnected pores, which is required for the cells to adhere, proliferate and differentiate to form new tissues

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Measuring the pore dimension was not possible due to the unavailability of scale. However, the average pore size is of the order of microns

Modeling of cell adhesion on scaffolds n n n n

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Importance of modeling cell adhesion General introduction to Cell adhesion Forces involved in cell adhesion One-dimensional Peeling Model and its assumptions Linear Peeling model Results Inferences Nonlinear peeling model Results

Importance of modeling Cell adhesion n

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Cell adhesion is one of the most important and preliminary steps for tissue or organ formation Cell adhesion is involved in stimulating signals that regulate cell differentiation, the cell cycle, and cell survival Cell adhesion plays a vital role during the wound healing To have a better understanding of cell interaction with specific protein Once implanted, there is only very limited access to the adhered scaffolds Also any control over the healing response is lost. Surgical intervention to possibly re-engineer any treatment also would prove to be an invasive procedure.

Bone Formation on Scaffolds

None of these steps will proceed if the cells do not adhere well to the scaffold.

Cell adhesion n

Cell adhesion is the contact and firm interaction between cells, between cells and extra cellular matrix (ECM) and between cells and synthetic materials such as scaffolds in tissue engineering

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Adhesion involves two phases § attachment phase § adhesion phase

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During cell adhesion there is specific binding between the cell surface receptors and the counter adhesion molecules i.e. ligands on the cell or substrate.

Forces involved in cell adhesion n

Forces that contribute to biological interactions are n

Non-specific forces/interactions n n n

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Electrostatic forces van der Waals forces Steric interactions

Specific forces/interactions

One-Dimensional Peeling model § Peeling is opposite of adhesion. At equilibrium, the structure of the cell surface is the same since the process is reversible § Assumptions: § Cell membrane – rigid and elastic § Scaffold – flat, rigid, nonporous surface

Linear Peeling model n

Linear, where the cell detachment occurs when the bond force at the leading edge of the cell exceeds the maximum bond force

Force is assumed to be a linear function of the displacement of bond length (ζ) from

Governing Equations

Free Zone

Free Zone

Adhesive Zone

Where s = arc length bond k= local curvature

f = adhesive bond force of a single n = bond density

Solution for free zone

Contact angle θ External force Tex Macroscopic angle θ0

Solution for adhesive zone

Ɵα - ratio of adhesion and bending energies t - ratio of tension and bending s- arc length σn - adhesive stress

Results expected for strong and weak adhesion Strong adhesion

Weak adhesion

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More number of ligandreceptor bonds

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Less number of ligandreceptor bonds

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Long adhesive zone, short free zone

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Short adhesive zone, long free zone

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Microscopic contact angle is large

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Microscopic contact angle is small

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More adhesion energy is required

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Less adhesion energy is required

Microscopic Contact Angle

Results Obtained –Strong/Weak adhesion

Weak Adhesion vs. Bending

Strong Adhesion vs. Bending

Results expected for shape of the cell Strong adhesion n

The effective width δ of the boundary layer for which bonds are stretched at the edge of the contact zone is small Relationship between δ and θα ratio of adhesive energy(γ) to bending modulus (B)

Weak adhesion n

The effective width δ of the boundary layer for which bonds are stretched at the edge of the contact zone is large

Results obtained for shape of the cell

Inferences Weak adhesion n

The result obtained by linear model is acceptable and works well

Strong adhesion n Strong adhesion: n

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We cannot have –ve values for ζ This implies that the cell membrane has penetrated the rigid impenetrable scaffold

•For small displacement, force is linear and linear model works well •But the real adhesion happens only at large forces, where the profile is completely non-linear •Linear model fails here

Nonlinear Peeling model n

Nonlinear model for osteoblasts adhesion is carried out by considering the integrin-fibronectin force profile

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This force-distance relationship will be used along with the peeling model to predict how osteoblasts will adhere to flat, rigid scaffold surfaces or extracellular matrix.

Force displacement profile of integrin and fibronectin

Curve fitting for force displacement profile

The above graph is fitted to Weibull distribution as,

Where x > 0 a = 0.8107 ± 0.192 ; b = 2.147 ± 0.303 The above equation can rewritten in terms of force f, and displacement ζ as

Where a = 0.8107 ± 0.192, b = 2.147 ± 0.303

Solution for adhesive zone – Nonlinear peeling model n

The work done to either break or form cross bridges when the membranes are bought together from a large separation distance to planar, equilibrium contact is given by,

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Total adhesion energy Γ = n W

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Adhesion (normal) stress

Solution for adhesive zone – Nonlinear peeling model n

Governing equations

These equations need to be solved to obtain the contour of the membrane for osteoblast adhesion

Conclusions Three dimensional, interconnected, highly porous PLLA and PU based scaffolds – fabricated Porosity, density and pore morphology of scaffolds – characterized. Membrane contours of cell, based on adhesive force assumptions - studied (Modeling). Solutions for linear peeling model - obtained. Basic equations for non linear model - derived.

Future work n

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The shape of the membrane for an osteoblasts adhesion can be determined by solving the nonlinear peeling model Nonlinear model can be further extended to curved scaffolds The properties of the PLLA and PU scaffold can be further enhanced by modifying the them with suitable extracellular matrix proteins like collagen In-vitro cell culture can be done on the scaffolds to study biocompatibility

Thank you