TRACTION FORCE

MICROSCOPY
2482 words

11 APRIL 2024
UNIVERSITY COLLEGE LONDON
 (1) ABSTRACT
The biomechanical interactions between tumour cells and the extracellular matrix
(ECM) are partly responsible for its role in cancer progression and metastasis. This study
employs the use of Finite Element Analysis (FEA) how the elastic modulus of the collagen
hydrogel, an ECM substitute affects its deformation and strength distribution due to traction
forces induced by tumour cells which causes tumour cell migration and cellular invasion.Ansys
APDL was the computational software used for FEA and it was discovered that with higher
levels of elastic modulus, the collagen hydrogel experiences less deformation and strain
distribution.

(2) LITERATURE REVIEW (INTRODUCTION & BACKGROUND)

The microenvironment surrounding a tumour is a complex ecosystem which consists

of multiple components such as cells, the extracellular matrix (ECM) and soluble that can
influence the growth and progression of cancer and metastasis (the pathogenic spread of
growth) [Hanahan and Weinberg, 2011]. The extracellular matrix (ECM) serves as a housing
unit that regulates the cellular behaviours and structural support [Discher et al, 2009]. It is
composed of proteins like collagens, elastin, and fibronectin. Changes to the ECM stiffness
indicates early stages of cancer in terms of its initiation, progression, and metastasis [Paszek
et al, 2005].

Interaction of tumour cells with the ECM can be through various mechanisms of
adhesion, migration, and contraction. Through these mechanisms, traction forces are
generated deforming the surrounding ECM further propagating tumour invasion into
neighbouring tissues [Friedl and Wolf, 2003] [Martin et al, 2013]. Studying this phenomenon
in a microscopic cellular level has proven an arduous task and in recent development, collagen
hydrogels which is a natural polymer is a valuable substitute to that of biological ECM
[Lambert, Nusgens and Lapiere, 1998] [Antoine, Vlachos and Rylander, 2014]. This polymer
provides detailed in-vitro studies on the interactions and biomechanics between the cell and
ECM. Simulations like finite element analysis (FEA) is the main approach to study and
investigate the mechanical behaviour and properties of these tumour cells within the collagen
hydrogels providing results of cell-induced deformations and substrate (collagen hydrogels)
stiffness evaluation [Holenstein et al, 2019].

The stiffness of the ECM can alter the cellular behaviour and biomechanic locomotion,
contributing to tumour growth, spread and cellular invasion [Lu,Weaver and Werb, 2012]
[Kopanska et al, 2016]. ECM stiffness is directly correlated with its elastic modulus of the
collagen hydrogel as its representative [Levental et al, 2009]. By investigating the relationship
of the effects of collagen hydrogel’s elastic modulus on cell behaviour, cellular biomechanics
can be understood more deeply.

Traction Force Microscopy (TFM) is the primary investigative scientific tool used for
measuring and quantifying the cellular traction forces exerted by the cell on the surrounding
ECM. This method’s strength lies in its capability to provide quantitative stress maps which
displays the stress distribution within a substrate and assess the deformation of the substrate
caused by cellular forces [Zancla et al]. With the integration of TFM with computational
modelling softwares, insights of the physical interactions between tumour cells and collagen
hydrogel can be obtained.

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 Several research has been conducted in previous years regarding the relationship
between substrate stiffness and strength and deformations of cell-induced behaviour.
Holenstein et al, (2019) proposed in his paper a method for stimulating and evaluating 3-
dimensional traction forces exerted by cells within the collagen hydrogels using finite element
analysis (FEA). Additionally, according to the research Kopanska et al, (2016), there are
substantial evidence that tensile forces originating from cancer spheroids contribute to tumour
invasion.

(3) AIMS & OBJECTIVES

This study aims to investigate the effects of the elastic modulus of the collagen
hydrogel on the strength and distribution of cell-induced deformations within the tumour cell
spheroids using FEA via ANSYS. Varying the elastic modulus of the collagen hydrogel may
provide insights into how alterations in ECM stiffness properties can affect tumour cell
behaviour and cellular dynamics.

(4) METHODOLOGY

Model Creation
The size of the individual tumour cells is microscopic in nature in the scale of
nanometre or micrometre. But the rapid proliferation and growth of these tumour cells can lead
to a macroscopic size resulting in the formation of solid masses [Hanahan and Weinberg,
2011] The design of a 3-dimensional model can be simplified as a sphere (tumour cell) inside
a cube (collagen hydrogel). Ansys APDL was used to create the model by writing APDL scripts
onto the command prompt of the user interface.

[Refer to Image 1]

Setting the Element Type

After the model is created, the element type of the model must be defined. The element
type of the model in Ansys will dictate the physical behaviour representation of the model
during Finite Element Analysis (FEA) when meshing and solving. It specifies and formulates
the finite elements used to discretize the geometry influencing the simulation accuracy and
efficiency. Both the sphere and the cube are 3-dimensional volumetric structures, so the
SOLID element type was chosen.

When picking SOLID element, there is sub-element category to choose from like brick
and tetra elements. Choosing the appropriate type of elements for the models is tantamount
to mimicking real biological scenarios. brick elements are also known as hexahedral elements
that have 6 faces,8 nodes and straight edges being more structured with block-like
geometries. Tetra or tetrahedral elements have 4 nodes and 4 triangular faces. They represent
unstructured elements that can contort to irregular shapes and more complex geometries like
the curved surface of a sphere. Hence, the SOLID-Brick element was chosen for the cube and
SOLID-Tetra was chosen for the sphere.

Material Properties

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 The tumour cell has a Young’s modulus of 29.09kPa [Kwon et al, 2020] and a Poisson
Ratio of 0.5 for a simplified assumption [Mokbel et al] while the Young’s modulus of the
collagen hydrogel ranges up from (0.5-30) kPa [Paek et al, 2024] [Kobayashi et al, 2020] with
a Poisson ratio between 0.2 and 0.3 [Castro et al, 2016]. For simplification in the simulation,
the average of 0.25 Poisson ratio and a range from 5kPa with 5kPa intervals are chosen for
the constant and variables respectively.

[Refer to Image 2]

Meshing

Prior to application of forces, boundary conditions and constraints, the models must be
finely meshed for FEA simulation to run. Meshing is important as it involves the discretization
of complex geometry into smaller, finer elements like triangles and tetrahedrons for 2D and
3D respectively. It approximates the behaviour of the structure the model is based on under
applied loads and boundary conditions [Melchiorre and Duncan, 2021].

A well-refined mesh can capture geometric details and the variations of the material
properties more accurately leading to more accurate predictions of stress, strain,
displacement, and deformations. As previously explained in the element type, the meshing of
the model is dictated by the type of element the model is chosen. During meshing, the
geometry of the model can then represent the properties the model is based on.

[Refer to Image 3]

Boundary Conditions
To accurately imitate the cell biomechanics and its effects on the ECM, boundary
conditions must be established on the meshed models in terms of the forces applied, the
direction and magnitude of forces and the constraint to be fixed in place. In an in-vitro
examination of a tumour cell surrounded by collagen hydrogel, it is often placed on a petri dish
to be examined under an electron microscope. The traction force exerted by the tumour cell
thus do not deform the bottom surface of the collagen hydrogel with it being fixed in place by
the petri dish, but any other directions are deformable. In the 3d models, the bottom surface
of the cube can be constrained to mimic the real clinical phenomenon.

The tumour cell exerts a traction force in 3-dimensions, the X-axis, the Y-axis, and the
Z-axis at around 1kPa [Holenstein et al]. To simulate this, 3 outward forces are applied on the
volume nodes near the surface of the sphere of the X, Y and Z plane onto the cube. Once the
boundary conditions are applied on the model, FEA solving can be performed. The process is
repeated and iterated for all values of the collagen hydrogel’s elastic modulus.

[Refer to Image 4]

(5) RESULTS
In the general post-processing sections, the analysis and FEA results can be measured
and observed quantitatively. Observations from Image 5 shows that the cube underwent a

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significant amount of deformation in all directions. Image 5 also displays the displacement
deformation in the X-axis.

Table 1,2 and 3 shows the tabled data of the collagen hydrogel’s elastic modulus and
its relationship with maximum deformation, maximum stress in 3-dimensions and maximum
strain in 3-dimesnions respectively.

(6) DISCUSSION
The elastic modulus (E) is the characterization of a material’s stiffness and its
𝜎
resistance to deformation. From Hooke’s Law (𝐸 = 𝜀 ) where the elastic modulus [E] is
inversely proportional to the strain (𝜀). Therefore, a higher elastic modulus will result in a lower
strain deformation. The result of the simulation clearly dictates the relationship between the
elastic modulus of the collagen hydrogel and its mechanical response when experiencing
applied loads. As the elastic modulus of the collagen hydrogel increases, the cube exhibits a
significant reduction level in deformation, displacement, and mechanical strain.

Deformation is related to the change is size of the material under load while
displacement is the change in position from the original position. The mechanical strain is the
geometric measurement of a material’s deformation denoting the relative displacement
between particles inside the body. Graph 1 and Graph 2 shows that with increasing magnitude
of elastic modulus, the maximum deformation, displacement, and strain of the cube (collagen
hydrogel) reduces very drastically under a constant applied load. This result is consistent with
the initial hypothesis and the expected behaviour of materials with increasing stiffness in which
it can highly resist permanent deformation.

However, in the simulation study performed to investigate the effects of elastic modulus
and deformation of the collagen hydrogel, there was an anomaly where the internal stress
values remain the same even with varying levels of elastic modulus. This discrepancy may
arise due to potential errors in modelling or applying boundary conditions. One factor may be
due to the simplification of the tumour cell and ECM design. The tumour cells are described
to being spheroid in shape but taking into consideration the cell movement (motility) the shape
is constantly shaping itself. By simply shaping and modelling the tumour cell as a sphere
reduces the accurate mimicry of a real tumour cell. Furthermore, the ECM surrounding the
tumour cell has its design being reduced to just a cube with the sphere (tumour cell) embedded
inside it. These simplifications may be easier to design and simulate but it comes with the risk
of producing relatable results imitating real biological occurrence. This highlights the
complexities of accurately modelling the real-life materialistic behaviour of the models in
computational studies.

Even with the variations of elastic modulus of the collagen hydrogel, there will be
changes in the influence the constant load has to the collagen hydrogel’s capability to resist
deformation and distribute stress accordingly. In simple terms, with varying elastic modulus,
the displacements, strains, and stresses distribution will change corresponding to the elastic
modulus variability. The same stress distribution results for the various elastic modulus just do
not correlate with real cellular mechanics and biophysics. The results for maximum
deformations, displacements and strains seem to meet the expectations of the experiments
but only the stresses remain an outlier.

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(7) CONCLUSIONS

In this experimental study, we investigated the influence of elastic modulus of the

collagen hydrogel and its mechanical and physical response under applied traction forces
induced by the tumour cell embedded inside it. The goal of the study was to understand how
changes in the elastic modulus of the collagen hydrogel affects the strength and distribution
of the cell-induced deformations providing valuable insights to the importance in
biomechanical simulations.

Using the Ansys APDL software, the tumour cell and ECM model was created, and the
elastic modulus of ECM was varied while applying constant forces in 3-dimensions. Notable
differences were observed in its deformation, displacement and strain indicating the material’s
sensitive response to external forces. However, unchanging internal stress of the ECM model
posed as a potential anomaly and an outlier in the simulation results. Although some simulated
results deviated slightly from the expected outcome, this anomaly just undermines the
complexities of accurately modelling and representing real materialistic behaviour and the
need for further refinement and improvement on simulation approaches to encapsulate
biological mimicry.

Discussion from the results dives deep into the implications for tumour cell traction
force microscopy, paving the way for the important role of ECM stiffness and its relationship
with cellular behaviour modulation within microenvironments. Further research and higher
computational simulations can bring forth a more advanced material modelling in the capture
of linear and non-linear elasticity with heterogenous properties that aligns with current or future
literature findings of cellular biomechanics and cell-matrix interactions.

In summary, this study managed to contribute to the mechanical behaviour of collagen

hydrogel albeit with some discrepancies which displays the challenges and obstacles of
translating real-life phenomenon into practical computational simulations. By integrating these
modelling anomalies and properties, the understanding of a more profound biomechanical
phenomenon can be achieved opening the gates to more approached to medical research.

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(9) FIGURES & TABLES (Including Captions)

Image 1: Wireframe View of Sphere (Tumour Cell) inside the Cube (Collagen Hydrogel)

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 Image 2: Material Properties Panel of Ansys APDL

Image 3: Meshed Model of Sphere (Left) and Cube (Right)

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 Image 4: Boundary Conditions of Models. Bottom Surface Constraint of Cube (Top Left),
Applied Forces of Sphere in 3-Dimensions (Top Right), and Overall Display of Sphere Force
on Cube (Bottom)

Image 5: Graphical Display of Cube Deformation (Left) and FEA Results (Right)

Table 1: Tabled Data of Elastic Modulus (Pa) and Maximum Deformation (mm)

Elastic
Modulus Max Deformation
(Pa) (mm)
500 31582.1
1000 15791
5000 3158.2
10000 1579.1
15000 1052.7
20000 789.6
25000 631.6

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 30000 526.4

Table 2: Tabled Data of Elastic Modulus and Stress in 3-Dimensions

X-Axis Y-Axis Z-Axis

Min Max Min Max
Elastic Min Stress Max Stress Stress Stress Stress Stress
Modulus (Pa) (MPa) (MPa) (MPa) (MPa) (MPa) (MPa)
500 -1.89 2.58 -5.94 8.14 -1.89 2.58
1000 -1.89 2.58 -5.94 8.14 -1.89 2.58
5000 -1.89 2.58 -5.94 8.14 -1.89 2.58
10000 -1.89 2.58 -5.94 8.14 -1.89 2.58
15000 -1.89 2.58 -5.94 8.14 -1.89 2.58
20000 -1.89 2.58 -5.94 8.14 -1.89 2.58
25000 -1.89 2.58 -5.94 8.14 -1.89 2.58
30000 -1.89 2.58 -5.94 8.14 -1.89 2.58

Table 3: Tabled Data of Elastic Modulus and Strain in 3-Dimensions

X-Axis Y-Axis Z-Axis

Elastic Min Max Min Max
Modulus (Pa) Strain Strain Min Strain Max Strain Strain Strain
500 -2890.4 1844.7 -10579.1 14484.5 -2630.5 1865.2
1000 -1445.2 922.4 -5289.5 7242.2 -1315.3 932.6
5000 -289 184.5 -1057.9 1447.5 -263.1 186.5
10000 -144.5 92.2 -529 724.2 -131.5 93.3
15000 -96.3 61.5 -352.6 482.8 -87.7 62.2
20000 -72.3 46.1 -264.5 362.1 -65.8 46.3
25000 -57.8 36.9 -211.6 289.7 -52.6 37.3
30000 -48.2 30.7 -176.3 241.4 -43.8 31.1

Max Deformation (mm) of Collagen Hydrogel

Max Deformation

35000
Max Deformation *mm)

30000
25000
20000
15000
10000
5000
0
0 5000 10000 15000 20000 25000 30000 35000
Elastic Modulus (Pa)

Figure 1: Maximum Deformation (mm) of Collagen Hydrogel

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 Max Strain in 3-Dimensions
16030
14030
12030
Max Strain

10030
8030 X-Axis
6030 Y-Axis
4030 Z-Axis
2030
30
0 5000 10000 15000 20000 25000 30000 35000
Elastic Modulus (Pa)

Figure 2: Maximum Strain in 3-Dimensions

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