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Scaffolds for Bone-Tissue

Engineering: Preparation,
Characterization, Modeling

Department of Chemical Engineering and Materials


Science
Amrita School of Engineering
Coimbatore – 641 105
May 2009
Final Review
By
Divya Haridas (CB105PE012)
Karthikeyan G (CB105PE023)
Krishna Priya C (CB105PE025)
Premika G (CB105PE028)

Guide
Dr. Murali Rangarajan

Co-Guide
Dr. Nikhil K Kothurkar
Polymeric Scaffolds

Three dimensional scaffolds play important roles


as extracellular matrices onto which cells can
attach, grow, and form new tissues and degrade
either during or after healing
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

Experiment: Synthesis and Characterization of PLLA and


PU Scaffolds
 Synthesis: Solid/Liquid Phase Separation/Freeze

Drying
 Characterization: Density and Porosity Measurements,

Surface and Pore Morphology using Optical


Microscope

Modeling of Cell Adhesion on Scaffolds


 One-dimensional Peeling Model


 Linear Model: Linear force- bond displacement profile
 Nonlinear Model: Ligand-receptor force interactions
Characterization of Scaffolds
Characterization of Scaffolds
Porous PLLA and PU scaffolds at 100X magnification
characterized by optical microscopy

P P
LLA U
Characterization of Scaffolds
Porous PLLA and PU scaffolds at 200X magnification
characterized by optical microscopy

P P
LLA U
Inferences
n Both PLLA and PU scaffolds are highly porous

n PLLA scaffolds are lighter compared to PU scaffolds

n High density of PU scaffolds are suitable for implantable


urinary tracts and bladders

n 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.

n Modifying the scaffold by adding HAp will improve growth of


bone tissue
Modeling of cell adhesion on
scaffolds

n Importance of modeling cell adhesion


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

n During cell adhesion there is specific binding between the


cell surface receptors and the counter adhesion molecules
Forces involved in cell adhesion
n Forces that contribute to biological interactions
are
n Non-specific forces/interactions
n Electrostatic forces
n van der Waals forces
n Steric interactions
n Specific forces/interactions

n Adhesion involves two phases


n Attachment phase
n Adhesion phase
Leckband D., 2000. Measuring
the forces that control protein
interactions, Annual Review
Biophysics Biomolecular
Structure 29, 1-26
One-Dimensional Peeling model
n Peeling is opposite of adhesion. At equilibrium,
the structure of the cell surface is the same since
the process is reversible
n Assumptions:
n Cell membrane – rigid
and elastic
n Scaffold – flat, rigid,
nonporous surface

Dong Kong, Baohua Ji, Lanhong Dai, 2008. Nonlinear mechanical model
of cell adhesion, Journal of Theoretical Biology 250, 75-84
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
Evan A. Evans, 1985. Detailed
function of the mechanism of membrane- memb
displacement of adhesion and separation,
bond length (ζ) from Biophysical Journal, 48, 175-183
Governing Equations

Adhesive Free
Zone Zone
Free
Zone

s = arc length f = adhesive bond force of a single


bond
k= local curvature n = bond density
Solution for free zone

n Local contact angle - θ


n External force - Tex
n 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
n More number of ligand- n Less number of ligand-
receptor bonds receptor bonds
n Long adhesive zone, short n Short adhesive zone, long
free zone free zone
n Microscopic contact angle n Microscopic contact angle
is large is small
n More adhesion energy is n Less adhesion energy is
required required
Results Obtained –Strong/Weak
adhesion
Results expected for shape of the
cell
Strong adhesion Weak adhesion
n The effective width δ of the n The effective width δ of the
boundary layer for which boundary layer for which
bonds are stretched at the bonds are stretched at the
edge of the contact zone is edge of the contact zone is
small large

Relationship between δ
and θα ratio of adhesive
energy(γ) to bending
modulus (B)
Results obtained for shape of the
cell
Inferences

Weak adhesion Strong adhesion


n The result n We cannot have –
obtained by ve values for ζ
linear model is n This implies that
acceptable and the cell
works well membrane has
penetrated the
rigid
impenetrable
scaffold
Nonlinear Peeling model

n Nonlinear model for osteoblasts adhesion is


carried out by considering the integrin-fibronectin
force profile

n 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

Li et al., 2003. Force Measurements of the α5β1 Integrin–


Fibronectin Interaction,
Biophysical Journal 84(2), 1252-1262
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 bridge when
the membranes are bought together from a large
separation distance to planar, equilibrium contact is given
by,

n Total adhesion energy Γ = n W

n 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 The shape of the membrane for an osteoblasts
adhesion can be determined by solving the
nonlinear peeling model
n Nonlinear model can be further extended to
curved scaffolds
n The properties of the PLLA and PU scaffold can
be further enhanced by modifying the them
with suitable extracellular matrix proteins like
collagen
n In-vitro cell culture can be done on the
scaffolds to study biocompatibility
Acknowledgement
n Prof. R. R. Subba Rao
n Dr. Murali Rangarajan
n Dr. Nikhil K Kothurkar
n Mr. K. Jayanarayanan
n Ms. Meera B. Sasikumar
n Mr. M. Kannan
n Dr. G. Prema (Dept. of Mathematics)
n Mr. Sanjivi Arul (Dept. of Mechanical
Engineering)
n Mr. R. Padmanaban (Dept. of Mechanical
Engineering)