journal of the mechanical behavior of biomedical materials 122 (2021) 104649

Contents lists available at ScienceDirect

Journal of the Mechanical Behavior of Biomedical Materials

journal homepage: www.elsevier.com/locate/jmbbm

Biomechanical and functional comparison of moulded and 3D printed

medical silicones
Alexandra Zühlke a, *, Michael Gasik a, d, Nihal Engin Vrana b, Celine Blandine Muller c,
Julien Barthes c, Yevgen Bilotsky d, Edwin Courtial e, Christophe Marquette e
a
Aalto University Foundation, Espoo, Finland
b
Spartha Medical SAS, Strasbourg, France
c
INSERM, Strasbourg, France
d
Seqvera Ltd. Oy, Helsinki, Finland
e
3dFAB, Lyon, France

A R T I C L E I N F O A B S T R A C T

Keywords: Modern 3D printing of implantable devices provides an important opportunity for the development of person­
Silicones alized implants with good anatomical fit. Nevertheless, 3D printing of silicone has been challenging and the
Mechanical properties recent advances in technology are provided by the systems which can print medical grade silicone via extrusion.
In vitro
However, the potential impacts of the 3D printing process of silicone on its biomechanical properties has not
In vivo
Biocompatibility
been studied in sufficient detail. Therefore, the present study compares 3D printed and moulded silicone
Implants structures for their cytotoxicity, surface roughness, biomechanical properties, and in vivo tissue reaction. The 3D
printing process creates increased nanoscale roughness and noticeably changes microscale topography. Neither
the presence of these features nor the differences in processes were found to result in an increase in cytotoxicity
or tissue reaction for 3D printed structures, exhibiting limited inflammatory reaction and cell viability above the
threshold values. On the contrary, the biomechanical properties have demonstrated significant differences in
static and dynamic conditions, and in thermal expansion. Our results demonstrate that 3D printing can be used
for establishing a better biomechanical microenvironment for the surrounding tissue of the implant particularly
for fragile soft tissue like epithelial mucosa without having any negative effect on the cytotoxicity or in vivo
reaction to silicone. For engineering of the implants, however, one must consider the differences in mechanical
properties to result in correct and personalized geometry and proper physical interaction with tissues.

1. Introduction
There is a growing trend to use 3D printing as the manufacturing
method for medical devices enabling their personalization (Vrana et al.,
2020; Jammalamadaka and Tappa, 2018; Nagarajan et al., 2018;
Kacarevic et al., 2018; DiMarzio et al., 2020). Medical grade silicone is
widely used in health care (Sittel, 2009; Emre et al., 2005) and it can be
processed using 3D printing (Jammalamadaka and Tappa, 2018;
Kacarevic et al., 2018). An example of a medical device requiring a high
personalization is stents used in the upper respiratory tract, whose
function is to keep the respiratory tract open to minimize the harm of
injury caused by trauma or surgery (Sittel, 2009; Debry et al., 2017;
Cooper, 2018). One of the challenges of such stenting is the possibility of
migration, for example, due to coughing or excessive physical loading.
The stent might move from its position or even come out of the patient
entirely. Moreover, the mucosa of the upper respiratory tract is highly
fragile and any excessive pressure applied to the mucosa by stents would
lead to an adverse immune reaction and necrosis. In addition to this,
some patients would have unconventional anatomical features due to
trauma or surgical resection to which normal stents would apply addi­
tional pressure and risk of damage. This problem can be prevented by
creating custom devices with a better fit, which can be achieved with a
3D printing technology. However, beyond the anatomical fit of the
stents, their biomechanical fit should also be considered.
Such 3D printed structures are highly desirable matching the
biomechanical properties of the tissue (Gomes and Reis, 2004) and
closely mimicking in vivo behaviour, assisting the body to rebuild the
surrounding damaged tissue (Chung and Burdick, 2008; Wong, 2007).

* Corresponding author.
E-mail address: alexandra.zuhlke@aalto.fi (A. Zühlke).

https://doi.org/10.1016/j.jmbbm.2021.104649
Received 15 October 2020; Received in revised form 9 June 2021; Accepted 12 June 2021
Available online 16 June 2021
1751-6161/© 2021 Published by Elsevier Ltd.
A. Zühlke et al.
One of the most critical issues to address, however, is the mechanical
properties of 3D printed biomaterials, as additive manufacturing tech­
niques might represent an important potential risk to be considered for
personalized implants (Vrana et al., 2020; Gasik, 2017). Mechanical and
biomechanical stimuli play an important role in organisms’ develop­
ment, homeodynamics and homeostasis (Vrana et al., 2020; van der
Meulen and Huiskes, 2002). On the level of a living cell, mechanical
stimuli are ones of the three fundamental pathways (electrical, (bio)
chemical and mechanical) to communicate with the cell and with its
environment.
One important gap is still a lack of proper understanding of biome­
chanical properties and their meaning in healthcare (Gasik, 2017). All
cells and organisms are mechanosensitive (Ingber, 2006; Orr et al.,
2006), relaying on conversion of mechanical stimuli from their physical
surroundings or from within the organism to electrochemical or
biochemical signals, which then regulate physiological responses (Jaa­
louk and Lammerding, 2009). Mechanotransduction also has a funda­
mental role in regulating physiological phenomena in other specialized
tissues that are not directly involved in sensory functions (Vrana et al.,
2020) and can have a critical influence on cell behaviour, even in tissues
and organs that do not serve an apparent biomechanical role in the body
(Guilak et al., 2014; Kazantseva et al., 2016).
A key issue in the development of engineered implants is the prior­
itization of various biomechanical properties as design parameters, as it
will be difficult (if not impossible) to completely match all the material
properties of native tissues (Guilak et al., 2014). There is also an un­
derstanding that may not be even necessary to match all of the material
properties of native tissues a priori in view of the remodelling potential
of the implanted tissue in vivo – indeed, some degree of ‘mismatch’ is
required to ensure necessary driving forces aiming on the restoring
functionality (Vrana et al., 2020; Gasik et al., 2018). Although me­
chanical strain usually exhibits rather complex patterns in vivo, its im­
pacts and related mechanisms can be investigated with simplified in
vitro models. For safe and widespread application of 3D printed implant
technologies, the underlying mechanical differences stemming from the
production methods should be carefully considered as a part of
personalization protocols.
Due to vast number of biomaterials and their forms, one usually
needs to choose the biomechanical testing methods, and this choice
might not be self-evident. There are constrains in existing standards
(easier to make a test, but more difficult to understand the meaning of
the data for clinical applications) and protocols (ad hoc test devices and
methods which are not easy to replicate in another laboratory) (Vrana
et al., 2020). One may consider elastic modulus, whose definition
originally fits only linear elastic materials and only for very small de­
formations, as has been prescribed by the theory of elasticity for cen­
turies. The guidelines of National Physical Laboratory (UK) list nine
methods of measurement and calculation of elastic modulus (Lord and
Morrell, 2006), all of which naturally lead to different values. Elasticity
theory does not consider the time-dependency of this property, the rate
(speed) of deformation, or its hysteresis due to cyclic load. Almost all
biomaterials and tissues are clearly not elastic ones, it is a significant
simplification trying to artificially reduce experimental data to some
fixed numbers (Vrana et al., 2020). Silicone, a non-linear elastomer,
would thus expect to have different behaviours when it is moulded or
printed based on extrusion; however, to our best knowledge this dif­
ference has not been studied in the literature up to now.
In the present work silicone polymers, intended for otorhinolaryn­
gology applications, namely for respiratory tract stenting, have been
characterised for their biomechanical, surface (micro and nanoscale)
and protein adsorption properties together with their cytotoxicity.
Whereas there are about 100 models of stents known, the problem of
tissue granulation, implant stability and operability has not been solved
yet (Trabelsi et al., 2011; Noppen et al., 2005). It was previously
observed that the biomechanical mismatches between the silicone im­
plants and the surrounding mucosa were inducing serious adverse
2
Journal of the Mechanical Behavior of Biomedical Materials 122 (2021) 104649
Fig. 1. View of the 3D printed (left) and moulded (right) silicone samples as 10
x 10 x 2 ​ mm blocks.
effects including granuloma formation, local necrosis and chronic
inflammation induced restenosis (Nesek-Adam et al., 2010; Alshammari
and Monnier, 2012; Ashfaque and Annju, 2018). In order to provide the
necessary information for the 3D printing based silicone implant
personalization in stenting the biomechanical properties of medical
silicones produced by traditional moulding and via 3D printing have
been studied and compared with dynamic mechanical analysis (DMA)
complemented with biomaterials enhanced simulation testing (BEST)
data analysis (Gasik and Bilotsky, 2019). In addition, surface roughness
and cell behaviour in the presence of 3D printed and moulded implants
were quantified. The results were combined to evaluate how production
method affects potential properties of implants for a given application
and to which extent these differences needed to be considered in the
design and optimization of these medical devices.
2. Materials and methods
2.1. Materials tested
The silicone samples (moulded silicone marked as “SilM” and 3D
printed silicone marked as “Sil3D”) were produced of a medical grade
silicone (Elkem Silicones, Norway, Silbione® LSR 4350) by a) moulding
in aluminium moulds with industry standards and b) using a custom-
made extrusion printer (Tobeca, 3dFAB, Lyon, France). The samples
were ~2 ​ mm thick and 1 x 1 ​ cm in area. For the compression test, the
samples were measured as they were, but for the 3-point bending test the
samples were cut in half, resulting in rectangular samples with 2 x 5 x
10 ​ mm dimensions. Pictures and specifications of the samples, either 3D
printed or moulded, used to perform 3-point bending and compression
experiments are given in Fig. 1.
The surface properties of the moulded and 3D-printed samples were
characterised with a Veeco Multimode Atomic Force Microscope (AFM)
in tapping mode for nanoscale surface features and roughness. The Ra
averages are calculated from 25 x 25 ​ μm ROIs using the Nanoscope
software. For microscale surface features and roughness, a Keyence
VHC-5000 digital microscope was used to have 3D reconstruction of
surface features via z-stacks (up to 700 ​ μm) using a 60x objective.
2.2. Mechanical testing
Before performing the mechanical tests, the samples were cleaned as
follows with 30 ​ min in ultrasound bath in ethanol (70%) and then by
rinsing with deionized water and air dried. All tests were done at
37 ​ ± ​ 1 ◦ C in dry static air. The biomechanical analysis was carried out
using dynamic mechanical analysis (DMA) with apparatus “Artemis”
DMA 242 (models C and E; Netzsch Gerätebau GmbH, Germany) under
pseudo-static (creep) and frequency scans (0.01–20 ​ Hz) at 20 ​ μm dy­
namic amplitude (resolution ±0.5 ​ nm). In the creep test, the material is
subjected to a constant static force over a given time and the resulting
deformation is measured. Dynamic tests foresee stimulation of the
sample by an oscillatory stress and the resulting deformation is
measured together with its phase shift (also commonly expressed as loss
tangent). In the frequency scan the amplitude of the oscillation is kept
A. Zühlke et al.
Fig. 2. An example of a frequency scan test principle, where the amplitude is constant but the frequency (F1 ​ < ​ F2 ​ < ​ F3) varies.
constant but the frequency is varied (Fig. 2). For all combinations,
changes in length, stiffness, loss tangent, dynamic and static stress/
strain ratios, viscosity, creep compliance, aggregate modulus, etc. were
analysed.
Tests were performed for compression mode and for 3-point bending
at 10 ​ mm span. All sample holders are automatically calibrated on
empty system stiffness, static, dynamic and rotation tuning, from which
data are subtracted from the measured signal. In addition, in compres­
sion mode a thermo-mechanical analysis (TMA) mode was applied to
analyse thermal expansion of silicones up to 45 ◦ C. The deformation
values recorded by DMA (in μm) were converted into true logarithmic
strain. True strain is the only measure having solid thermodynamic
grounds among used strain forms (Norris, 2008; Lubarda and Chen,
2008; Xiao, 1995), although others might be used especially when the
deformations are small. True strain in this case is calculated for
quasi-static loading (creep) conditions as
⃒ ( )⃒
⃒ ΔL ⃒⃒
εstat = ⃒⃒In 1 + , (1)
H0 ⃒
and for dynamic loading conditions as (Gasik and Bilotsky, 2019):
( )
1 2adyn
εdyn = In 1 + , (2)
2 H0 + ΔL − adyn
where H0 - initial dimension (height) of the specimen before the test, ΔL -
instant value of change of the static height, adyn - amplitude of the dy­
namic displacement. Equation (2) takes into account the change in the
specimen static dimensions during the test, incorporating loading his­
tory, and accounts for non-symmetry of the harmonic stimulus signal vs.
origin due to this.
2.3. Mechanical data analysis
With the conventional biomechanical readouts obtained directly
from the DMA, advanced data analysis was carried out with BEST
(Biomaterials Enhanced Simulation Testing). Briefly, BEST comprises
steps of 1) defining the proper physiologically relevant conditions for
your clinical application, 2) subjecting the specimen to coherent
biomechanical stimuli, inducing conditions closer to hostile environ­
ment, 3) assessing changes in properties of the specimen in time, phase
and stimulus domains, 4) post-processing experimental readouts with
idempotent analysis (Gasik and Bilotsky, 2019; Maslov, 1970), incor­
porating specimen history and 5) extracting true invariant values, suit­
able for material behaviour prediction in a form of model-free
constitutive equation for given biomaterial.
The differences between BEST and other methods are in direct
3
Journal of the Mechanical Behavior of Biomedical Materials 122 (2021) 104649
experimental observation of materials properties without assumption of
a material mechanistic model, method linearity, and without applica­
tion of Fourier transform or using complex numbers (Gasik and Bilotsky,
2019). Whereas Fourier transform is a very useful and perfectly working
tool for linear operations (in linear viscoelasticity), it fails for properties
as arbitrary functions of time or frequency (Neumark, 1962a). Gener­
alized Fourier transforms such as the Fourier integral operators and
pseudo-differential operators could be used (Maslov, 1970), however,
the drawback of this approach is in its strict adherence to certain types of
pseudo-differential equations and requirements of a deep knowledge of
such functional techniques (Maslov, 1970; Neumark, 1962a). On the
contrary to these approaches, BEST does not employ differential equa­
tions or series expansions, neither requires material homogeneity (the
properties are evaluated for the whole specimen, i.e. closer to the con­
ditions which might be faced upon implantation). The BEST method
does not stipulate that the material has to be compliant with some
pre-selected physical model (elastic, viscoelastic, hyperelastic,
neo-Hookean, etc.). Selection of the material model in any combination
is a must for any conventional calculations in viscoelastic analysis or in
numerical computer simulations, but this may lead to wrong predictions
when the model assumptions do not hold. In this work BEST analysis was
used to extract true (invariant) properties of elasticity, viscosity and
their time and frequency dependences (Gasik and Bilotsky, 2019).
2.4. Biocompatibility testing
Cytotoxicity by indirect contact was analysed with an extraction
vehicle (the supplemented DMEM cell culture medium) in the contact
with the material (moulded and 3D printed silicone samples, PET as
positive control, sodium azide NaN3 as negative control) for 24 ​ h in the
incubator (5% CO2, 37 ◦ C). In parallel, the 3T3 cells are cultured in a cell
culture dish until reaching 70–80% confluence in 24 ​ h. On the second
day, the extraction vehicle was transferred to the cell mat and left in
contact for next 24 ​ h before evaluating possible cell toxicity at the
morphological level (microscopy) and at the level of viability (MTT
test).
Cytotoxicity by direct contact was evaluated with the 3T3 cells first
grown in a cell culture dish to reach 70–80% confluence within 24 ​ h. On
the second day, the materials to be tested were placed in contact with
the cell mat and left for 24 ​ h before evaluating possible cell toxicity at
the morphological level (microscopy) or at the level of viability (MTT
test). For this test, the dimensions of the material should not exceed 10%
of the surface of the cell layer. To keep the material in contact with the
cell mat, a cell culture insert was used which was placed on the material.
A. Zühlke et al.
Fig. 3. Thermal expansion of silicones.
2.5. In vivo experiments
For testing the tissue reaction to moulded and 3D printed silicones in
vivo, subcutaneous paravertebral implantation of samples of 1 ​ cm2 with
5 ​ mm thickness was performed in mice for a duration of 2 weeks after
the obtention of the ethical authorisation. Experiments were performed
in UE-1277 Plateforme d’Infectiologie Expérimentale (INRA, Tours,
France). Protocols were submitted to the C2EA–19 ethics committee
with approval number DAP-2017031321109445, in accordance with
the European directive 2010/63/EC for conducting animal experiments.
Anaesthesia of the animals was performed with an intraperitoneal
injection of ketamine (Imalgène 1000®, 75 ​ mg/kg) and xylazine
(Rompun®, 5 ​ mg/kg). Then mice were placed on a heated pad, shaved,
cleaned with betadine and buprenorphine (Buprécare®, 0.05 ​ mg/kg)
was injected in subcutaneous. Then incision was made in paravertebral
and silicones patches were implanted in subcutaneous. To finish the
operation, muscles and incisions were closed with stitches. Buprécare
Fig. 4. Surface characterization of moulded (SilM) and 3D printed (Sil3D) silicone samples using digital microscope.
4
Journal of the Mechanical Behavior of Biomedical Materials 122 (2021) 104649
(0.05 ​ mg/kg) was injected into the animal subcutaneously to reduce
pain at the end of the implantation and every 12 ​ h for 5 days. Eutha­
nasia was performed after 14 days. Totally 8 mice were implanted for
each condition. The samples were explanted after 2 weeks and charac­
terised by histology together with the measurement of cytokine levels
for determination of systemic effects. Only cytokines with read-outs
above threshold were reported (IL-1β, IL-23, IL-27, MCP-1, TNF-α,
IFN-γ). Histology images were scored by an independent histopatholo­
gist without access to the sample information (blind analysis).
2.6. Statistical analysis
Data of biomechanical measurements, after time convolution pro­
cedure shown below, were fitted with exponential function of memory
values as it is theoretically predicted by idempotent analysis (Gasik and
Bilotsky, 2019). The presence of leverage points was detected with hat
matrix diagonal components, which did not fit Stephen’s rule (these
points were removed). Influence (outlier) points analysis was made by
calculating Cook’s distances, and data points exceeding unity value (if
any) were removed. The consistency of regression coefficients was
independently checked by application of Theil-Shen estimator and the
goodness of fit significance by Nelson-modified Anderson-Darling test
(Nelson, 1998). The algorithm for this test has been improved, as the
original source code (Nelson, 1998) does not allow negative normalized
(studentized) values in the data.
Heteroscedasticity of residuals was estimated with RUNS test and the
residuals autocorrelation with Durbin-Watson parameter. Based on the
mathematical analysis, obtained biomechanical invariant values were
considered to be BLUE (best linear unbiased estimators).
The statistical significance for in vitro assessed using Student’s t-test
and in vivo studies of the obtained data was assessed using one-way
ANOVA with Tukey’s multiple comparisons or Friedman’s test with
Dunn’s multiple comparisons (n ​ ≥ ​ 3). The error bars are representative
of the standard deviation (SD). Differences with values of p ​ ≥ ​ 0.05
were considered statistically insignificant.
A. Zühlke et al. Journal of the Mechanical Behavior of Biomedical Materials 122 (2021) 104649

Fig. 5. Surface characterization and roughness determination (Ra) of moulded (SilM) and 3D printed (Sil3D) silicone samples using AFM.

3. Results and discussion

3.1. Thermal expansion of silicones

Thermal expansion of silicone samples was measured under constant

force of 0.3 ​ N and heating rate of 1 ​ K/min. Data of the expansion (in
μm) were normalized to the initial height of the samples and converted
into true logarithmic strain, expressed in Fig. 3 in micro-strains vs. ab­
solute temperature (K).
It was observed that the expansion of 3D printed silicone samples is
not monotonic, showing jumps in expansion at certain temperatures.
This was seen to originate from a non-uniform surface structure of the
3D sample which wavy surface features might collapse or expand upon
loading and heating (for 3D sample, a rapid increase was observed soon
after heating eventually due to shape irregularities, hence it does no
start from zero strain). Nevertheless, polynomial approximation of the
expansion and its derivation has given coefficient of thermal expansion
(CTE) values (μ/K) as CTESilM ​ = ​ 6.22 ​ T - 1661.45; and
CTESil3D ​ = ​ 15.96 ​ T - 4832.45 in the range of 20–45 ◦ C (293–318 ​ K).
For temperature of 37 ◦ C (310 ​ K) as being of a clinical interest, these
CTE values are 266.75 μ/K and 115.15 μ/K for moulded and 3D printed
specimens respectively. Literature data (Migwi et al., 1994) reported
CTE for silicone at 37 ◦ C of 273.5 μ/K - rather close to moulded material
despite values (Migwi et al., 1994) were measured with a different
method. Lower CTE values for 3D printed material are considered
originating from a non-uniformity of the surface and the structure, as
well as air bubbles, residing inside the material during its
manufacturing.
The probe of the DMA machine is highly sensitive (sub-nm resolu­
tion), therefore a good surface quality of the sample is required for
obtaining consistent data in compression mode. There are differences in
surface topology between the 3D printed and moulded samples. The
surface of 3D printed silicone is “wavy”, with some leftover material
appearing as semi-random protrusions shown in Fig. 4. These “tails”
confused the sensor probe apparently reaching the surface, when in fact
it only had a tiny contact with a very little material. This has caused a
major uncertainty when trying to measure Sil3D samples mechanical
properties in compression. The digital microscopy composite images of
the surfaces of moulded and 3D printed surfaces (Fig. 4) clearly show the
presence of the “waves” due to the printing process, which also signif­
icantly increases their roughness (Ra) to 160-120 ​ nm vs. 20–30 ​ nm for
moulded samples (Fig. 5).
Whereas CTE data for 3D printed material are more scattered, it is
evident that material features coming from the 3D printing process lead
to at least twice lower CTE than its moulded (non-porous) analogue. This
Fig. 6. Example of the creep compliance (ratio of strain to stress, 1/kPa) of
SilM specimens vs. experiment time (sec) on log-log scales. Numbers in legend
are applied creep stresses, kPa.
difference might have an implication for the medical device design and
processing, when the exact dimensions at physiological range are
important.
3.2. Creep data
Creep tests were carried out at 1–60 ​ kPa pressure in compression
mode at 37 ◦ C. This pressure range cover most physiological and over-
physiological conditions, from a very weak contact to extreme
compression. If an implant exerts on surrounding soft tissues pressure of
5–10 ​ kPa (37–75 ​ mmHg), this still would allow hemocirculation, but
long-lasting pressures over >20 ​ kPa (>150 ​ mmHg) would ultimately
lead to tissue necrosis and hypoxia following severe complications
leading to the removal of the implant. Higher pressures however might
be faced for short times during the implant placement and its setting, so
it is reasonable to cover the wider pressure range for materials
evaluation.
The creep compliance data, defined as the ratio of true strain to
applied stress are shown in Fig. 6 for applied creep stresses. It is seen that
creep compliance clearly depends not only on experiment time, but also
on the level of applied stress. An interesting experimental observation is
that the creep compliance level is not proportional to the applied stress
(Fig. 6), indicating that materials behaviour is non-linear.

5
A. Zühlke et al.
Fig. 7. Memory values (α) in creep tests for both silicone materials. Vertical
bars are standard error.
For this loading case (creep), the theoretical idempotent solution
(Gasik and Bilotsky, 2019; Gunawardena, 1996) of the behaviour of the
materials predicts the power-law formal dependence of strain vs. time
for every fixed value of applied stress (Gasik et al., 2018, 2021; Magin,
2010; Nelson, 1998):
∫t
εcreep (t) 1 dτ tα
= α = ,
σstar Γ(α) Cα (t − τ)1− Γ(1 + α)E01− α ηα0
0
where σstat is the applied creep (static) stress, Cα is the viscostiffness
(quasi-property in units of kPa⋅sα) (Gasik and Bilotsky, 2019), Γ(⋅) -
gamma-function, α – material memory, E0 – intrinsic stiffness, η0 –
intrinsic viscosity. The last two values are invariant in a sense that they
do not depend on the applied stress or time of experiment (Gasik et al.,
2018, 2021; Gasik and Bilotsky, 2019; Nelson, 1998). Equation (3) can
be always numerically explicitly computed without need of assumptions
of linearity of Cα (α, t) and α(t).
It is noteworthy that the rhs of (3) in practice is calculated with
convolution integration in time steps, and not by log-log fitting of the
data (“power law”; (Bagley and Torvik, 1983; Rodríguez et al., 2018;
Coussot et al., 2009; Magin, 2010)). The values of memory parameter in
equation (3) must be non-negative to ensure causality principle (Hanyga
and Seredynska, 2003; Nelson, 1998; Gasik et al., 2021), and in the
range 0 ​ < ​ α ​ < ​ 1 they present the fading memory (zero means only
short-time memory, i.e. ideally elastic behaviour, whereas unity means
long-time memory i.e. ideally viscous behaviour). For the range
1 ​ < ​ α ​ < ​ 2 the memory values are covering inertial effects and waves
Fig. 8. Invariant stiffness (a) and viscosity (b) in compression for two silicone materials as function of applied stress. Vertical bars are standard errors.
Journal of the Mechanical Behavior of Biomedical Materials 122 (2021) 104649
propagation (elastic, poroelastic or viscoelastic) (DeAlmeida and Fer­
reira, 2012; Hanyga and Seredynska, 2003), however, such waves are
not present in pseudo-static experiments here.
Experimental memory values (time-invariant) are shown in Fig. 7 for
both materials as function of applied stress. It is seen that higher
compressive stresses results in more short-time memory (the materials
adapt to the stimulus faster), but in low stress range moulded silicone
exhibit more long-time memory (revealing its more viscous nature) with
a steeper dependence from the applied stress. Low memory values for
Sil3D might be also due to presence of structural non-uniformity, uneven
surface, and trapped air bubbles.
With these experimental memory values, data of Fig. 6 and Eq. (3)
the values of invariant modulus and viscosity have been obtained, Fig. 8.
It is seen that moulded material is stiffer and more viscous than 3D
printed one. Both silicones in compression also exhibit substantial
stiffening with increased loading.
Combing all the above for the creep test, the constitutive equation for
calculation of the expected behaviour of silicones as function of stress
and time from (3) could be presented as:
( ) ( t )α(σ)
σ
εcreep t, σ = (4)
Γ(1 + α(α))⋅E0 (α) τ(α)
where the parameters for α(σ), E0(σ) and characteristic time τ0(σ) are
shown in Table 1. These functions are obtained from experiment without
considering a material model or its linearity, and they can be directly
used for the prediction of these silicone materials behaviour in creep
conditions.
(3)
3.3. Three-point bending tests
In dynamic bending mode the behaviour of a polymer is different
from static or creep loading. The results taken from the DMA outputs at
scans from 0.1 to 20 ​ Hz under 20 ​ μm deformation amplitude, are
shown in Fig. 9. This deformation amplitude corresponds to the physi­
ological ranges (Vrana et al., 2020; Gasik et al., 2018; van Mow and
Huiskes, 2005), taking into account proper pressure intervals. Ampli­
tudes of complex elastic modulus by DMA seem to be nearly
Table 1
Parameters of the constitutive creep Eq. (4) for silicones. Invariant (steady)
viscosity η0(σ) can be assessed as a product E0(σ)⋅τ0(σ) where stress variable (σ)
is expressed in kPa.
Function SilM Sil3D
Memory value α(σ) 0.2382⋅σ-1.071 0.0179⋅σ-0.318
Stiffness E0(σ), kPa 1391 ​ + ​ 17.12⋅σ 98.21 ​ + ​ 8.845⋅σ
Characteristic time τ0(σ), sec 71.92⋅σ-0.154 104.26⋅σ-0.205
6
A. Zühlke et al. Journal of the Mechanical Behavior of Biomedical Materials 122 (2021) 104649

Fig. 9. Amplitude of the complex elastic modulus (a) and the loss tangent (b) as obtained by DMA, without data post-processing.

viscoelasticity models) usually leads to wrong predictions, artefacts or

Table 2
just improper conclusions (Neumark, 1962b). Authors (Ewoldt et al.,
Dynamic parameters of the constitutive equation (5) for silicone samples
2015) have emphasized that presentation of oscillatory stress or strain
(bending mode, 0.1–20 ​ Hz).
response in real Fourier series is not at all equivalent to complex Fourier
Parameter SilM Sil3D transformation when only the first harmonic real and imaginary moduli
Dynamic invariant 8.03 ​ ± ​ 0.15 3.44 ​ ± ​ 0.04 are calculated (which itself automatically imposes linearity trans­
stiffness E0ω, MPa formation requirements).
Dynamic invariant 0.022 ​ ± ​ 0.004 0.0125 ​ ± ​ 0.0015
One may understand the meaning of the invariants relating the value
viscosity, η0ω,
MPa⋅s of (E0ω)1-α(ω) as elastic contribution to the dynamic stress/strain ampli­
Dynamic memory (0.06265 ​ ± ​ 0.005)⋅ω0.16 ±
(0.1054 ​ ± ​ 0.010)⋅ω0.483 ± tudes ratio, and (η0ωω)α(ω) - as viscous contribution to the whole
value α(ω) 0.01 0.005
stiffness:
σ dyn
= E01−ω α(ω) ⋅(η0ω ⋅ω)α(ω) (6)
independent on frequency, but there are substantial differences in loss εdyn
tangent (Fig. 9, b). At low frequencies Sil3D material does not generate
The smaller the values of η0ω and α(ω) are, the less dynamic stress/
stable DMA readouts, possibly due to specimens’ uneven surface and
strain ratio (dynamic stiffness) would depend of frequency. There
instability in the sample holder.
intrinsic stiffness E0ω and intrinsic viscosity η0ω themselves do not
Application of the lhs integral of (3) to harmonic stress input (Gasik
depend on frequency or time (being invariants), and they are not com­
and Bilotsky, 2019; Bagley and Torvik, 1983; Rodríguez et al., 2018;
plex numbers.
Coussot et al., 2009) results in the non-linear equation for dynamic
It is noteworthy that this analysis does not have a simple decompo­
strain as function of time, frequency, and stress (without postulating any
sition of (6) into “elastic” and “viscous” parts contribution, alike in
models of elastic behaviour of the material):
simple ‘spring-dashpot’ models because of general non-linearity of the
viscoelasticity of the system. It is also notable that these elastic (E0ω)1-
⎛ ⎞ ( )
∫t ( ) σdyn sin ωτ + πα2(ω) α(ω)
εdyn ⎝t, ω, σ dyn ⎠ =
1 σ dyn sin ωτ d τ
= 1− α(ω) (5) and viscous (η0ωω)α(ω) contributions cannot be simply related to
Γ(α) Cω (t − τ)1− α E0ω (η0ω ω)α(ω) storage E’ = Re(E*) and loss E” = Im(E*) moduli usually calculated in
0
linear viscoelasticity obtained with the Fourier transform, since the
where σ dyn is the applied dynamic stress amplitude, ω - circular fre­ presence of a material memory (even when not being explicitly
quency, Cω is the dynamic viscous stiffness (quasi-property in units of frequency-dependent) makes the product (6) being non-linear.
kPa⋅sα) (Gasik and Bilotsky, 2019), α – dynamic material memory The benefits of using data as in Tables 1 and 2 comprise simpler,
parameter, E0ω – intrinsic dynamic stiffness, η0ω – intrinsic dynamic faster, and more reliable biomechanical estimation of personalized
viscosity. This equation comprises time-convolution of the specimen implant quality, minimizing the risks of complications. It was reported
loading history without a need to involve complex algebra (i.e. without that already during prolonged intubation and pressure, the tracheal
using complex numbers), assumptions of a material model (Maxwell, mucosa becomes ischemic which increases the likelihood of tracheal
Burger, standard linear solid, Prony series, etc.) or use of local differ­ wall damage and subsequent stenosis (Nesek-Adam et al., 2010). These
entiation (Gasik and Bilotsky, 2019; Gasik et al., 2021; Zühlke et al., pathological changes may eventually lead to fibrosis of circumferential
2021). In the case the stress input in (5) is not harmonic, the integral part lesions (Alshammari and Monnier, 2012), resulting in progressive
will have different appearance, but the parameters will keep their tracheal stenosis, mucosal ulceration and cartilage destruction, with
meanings. The integration limit in (5) covers separate periods with injury occurring even within minutes after ballooning of the cuff and
constant frequency, where the same harmonic stimulus is applied. subsequent fibrotic changes within the following 3–6 weeks (Ashfaque
The material parameters in (5) have the same meaning as for the and Annju, 2018) – despite all cautions, glottic stenosis continues to
creep tests in (4), but their values are now different, Table 2, and can be occur at a high frequency of 6–21% in intensive care units.
used for prediction of the bending stiffness of these silicones in marked This critical compressive pressure was mentioned to be about
frequency range. These values do not depend on time, frequency, or 30–35 ​ mmHg (4–5 ​ kPa) (Nesek-Adam et al., 2010; Alshammari and
applied force, except for the memory value α(ω), which frequency Monnier, 2012), however, the original study (Knowlson and Basset,
dependence brings major non-linearity in the materials behaviour. If 1970) noted that clinically accepted capillary flow pressure of
α(ω) ​ = ​ const, the material behaviour is linear and can be equally well 32 ​ mmHg needed to be complemented with pressure resistance of the
described in frequency domain with a Fourier transform as in conven­ surrounding tissues, which was estimated to span of 30–60 ​ mmHg extra
tional linear viscoelasticity. For the opposite case, application of com­ values, resulting in total of 60–95 ​ mmHg (8–13 ​ kPa). Taking ≈10 ​ kPa
plex math transformation (such as in traditional rheology or of extra stress as an average value, one can estimate with Tables 1 and 2

7
A. Zühlke et al. Journal of the Mechanical Behavior of Biomedical Materials 122 (2021) 104649

Table 3 for analogous moulded part. Due to this deformation, the exerted stress
Expected static and dynamic (1 ​ Hz) properties for silicone samples under might drop faster and therefore the primary stability of the implant
10 ​ kPa stress. might be compromised even if biomechanical risk of stenosis would be
Parameter estimations SilM Sil3D minimized in this case. Dynamic stiffness at 1 ​ Hz (i.e. closer to physi­
Creep memory value α 0.020 0.009
ological pulse frequency) in these conditions for moulded silicone would
Creep stiffness E0, MPa 1.56 0.186 be about 3.8 times higher than for 3D printed one, meaning that the
Characteristic time τ0, sec 50.45 65.03 deflection of moulded part would be smaller and the exerted pressure to
Strain at creep at 10 ​ kPa after 5 hours 0.007 0.054 soft tissues would be respectively higher if the deformation of the
Dynamic stiffness E0ω, MPa 8.03 3.44
implant is constrained by some means.
Dynamic invariant viscosity, η0ω, MPa⋅s 0.022 0.0125
Dynamic memory value αω 0.084 0.256 Pseudo-static (creep) and dynamic (frequency scan) analyses were
Dynamic stiffness, MPa 6.80 1.79 made with DMA, but the readouts were also additionally processed with
BEST (Vrana et al., 2020; Gasik and Bilotsky, 2019), which does not use
complex math, artificial models neither Fourier transforms. This leads to
data how much the static and dynamic deformation of the silicone stent a minimal set of parameters describing the mechanical process in static
would be feasible for a selected design, size, geometry and the status of and dynamics. As known, there are tangible practical and experimental
the patient (Table 3). costs for adding extra variables and parameters to the constitutive
It is seen that expected static (creep) deformation of 3D printed sil­ equation for describing the underlaying processes (Magin, 2010). When
icones after 5 ​ h post-implantation would be about 8 times higher than the method uses less parameters, and when most of the parameters are

Fig. 10. Indirect cytotoxicity results for moulded (SilM) and 3D printed silicone (Sil3D), with PET as a positive control (~100% viability) and sodium azide imbibed
silicone as a negative control (~10% viability). Corresponding brightfield images of the cells after exposure.

Fig. 11. Direct contact cytotoxicity tests for moulded (SilM) and 3D printed silicone (Sil3D) with PET as positive and sodium azide imbibed silicone as negative
control. Corresponding brightfield microscopy images (10x).

8
A. Zühlke et al. Journal of the Mechanical Behavior of Biomedical Materials 122 (2021) 104649

Fig. 12. Paravertebral subcutaneous implantation of moulded and 3D printed silicones. A) The implantation sites, the histological scoring and scoring criteria. B)
Histological sections after 2 weeks of implantation with haemotoxylin and eosin and Masson Trichrome staining.

invariants, it provides simpler, more concise and more consistent pre­
diction for the biomaterial system without unnecessary complication of
the calculations or ersatz-models. The appearance of Eqs. (3)–(6) for the
case of the testing modalities is close to known fractional viscoelasticity
(Bagley and Torvik, 1983; Rodríguez et al., 2018; Coussot et al., 2009;
Magin, 2010). However, here it was shown that the memory values α are
not constants and therefore traditional methods of fractional calculus
(Bagley and Torvik, 1983; Magin, 2010) or linear viscoelasticity (Neu­
mark, 1962b; Ewoldt et al., 2015) cannot be directly used. Biome­
chanical data obtained with new method (Gasik and Bilotsky, 2019) do
not impose restrictions on the linearity of the system or material, and
can be extended for the cases dominated by inertial or wave-propagation
conditions (1 ​ < ​ α ​ < ​ 2), when linear approach becomes invalid (such
as with negative loss tangent and strain energy density).
Whereas biomechanical properties and the surface of the materials
show substantial differences, cytotoxicity of 3D-printed and moulded
samples does not alert about cell toxicity by direct contact since the cell
viability is respectively 80% and 95% compared to the TCPS control. In
addition, there is no morphological change in the cells which is another
criterion for confirming that all materials are not cytotoxic (Fig. 10). In
indirect contact, moulded silicone shows no cytotoxicity since full
viability (100%) is maintained compared to the control, without cells

Fig. 13. In vivo systemic cytokine levels in mice at D1, D7 and D14 of implantation of moulded and 3D printed silicone samples compared to baseline read-outs.

9
A. Zühlke et al.
morphological change, with no clear effect by cell density populations
which one could potentially able to determine (Volfson et al., 2008).
There was a slightly higher viability results compared to 3D printed
samples in this configuration but the viability with 3D printed samples
was still above 70% (the cytotoxicity threshold), Fig. 11.
After 2 weeks of implantation, the histology shows good healing
around both moulded and 3D printed silicone. The histological scoring
resulted on a zero score for the 3D printed silicone (good healing, within
the scale of 0–3; where 3 corresponds to chronic inflammation), Fig. 12.
The moulded samples, on the other hand had a score of 0.4; but still
within good healing index. The quantification of systemic cytokine
levels showed a slight increase in the pro-inflammatory cytokines after
implantation at similar levels for both silicones. In most cases the dif­
ferences were insignificant; except on D1 moulded silicone samples
shown a higher TNF-α, MCP-1 and IL-27 levels - all pro-inflammatory
markers (Fig. 13). However, over the course of 2 weeks (D14), the
levels were stable and cannot be considered to induce a systemic high
level of inflammation where the read-outs <100 ​ pg/ml range for any of
the cytokines.
4. Conclusions
In this work biomechanical properties of moulded and 3D printed
silicones in statics (creep) and dynamics have been measured and
quantified for the ranges of boundary and loading conditions which are
physiologically relevant for implants. It was observed that 3D material
has very distinct differences in structure and surface quality which are
immediately reflected in mechanical properties. On the contrary, cyto­
toxicity tests carried out have shown that processing method (moulding
and 3D printing) does not have an effect on the cytotoxicity of silicones,
and the differences in topology do not effectively impact on the cell
behaviour. Thus, biomechanical properties might be the decisive factor
for preference of certain implant types, designs and geometry, depend­
ing on their intended purpose and patient-specific requirements.
Moulded silicone has nearly twice higher thermal expansion coeffi­
cient vs. 3D printed so for a precise fitting this is needed to be considered
when designing a personalized stent. These differences might also have
implications for implants with a complex shape or fine geometric fea­
tures, as uneven thermal expansion might lead to peaks of stress con­
centration which could become potential weak spots for failures.
Obtained experimental biomechanical data of moulded and 3D
printed silicones have demonstrated substantial differences which
needed to be taken into account for design and manufacturing of precise
medical implants. Model-free data for invariant stiffness, viscosity and
material memory can be used for numerical modelling and optimization
of these silicone devices as they enable much better prediction for set
stress and frequency values.
Authors contributions
A.Z. has made the experimental DMA testing, analysed DMA read­
outs and made manuscript figures as well as revise the final version. M.
G. has composed the manuscript, arranged the text, and checked the
numerical outcomes. E.C., C.M. optimized 3D printing process, prepared
and produced 3D-printed silicone samples. N.E.V., J.B., C.B.M. carried
out AFM analysis and studied cell reactions on the samples. Y.B. has
processed the readouts with BEST and wrote the method explanation.
Disclosures
Y.B. and M.G. are shareholders of Seqvera Ltd. N.E.V. is a share­
holder of Spartha Medical SAS. These ownerships did not have any
implication on the data or the interpretation of the results.
10
Journal of the Mechanical Behavior of Biomedical Materials 122 (2021) 104649
Author statement
As the corresponding author I confirm on behalf of all the authors,
that we have read and approved the revisions to this manuscript.
Declaration of competing interest
The authors declare that they have no known competing financial
interests or personal relationships that could have appeared to influence
the work reported in this paper.
Acknowledgments
This project has received funding from the European Union’s Hori­
zon 2020 research and innovation programme under grant agreement
No 760921 (“PANBioRA”; A.Z., M.G., N.E.V., J.B.) and from the “FUI-
FASSIL” project (J.B., C.B.M., N.E.V.)
References
Alshammari, J., Monnier, P., 2012. Airway stenting with the LT-Mold™ for severe glotto-
subglottic stenosis or intractable aspiration: experience in 65 cases. Eur. Arch. Oto-
Rhino-Laryngol. 269, 2531–2538.
Ashfaque, A., Annju, T., 2018. Multimodality surgical approach in management of
laryngotracheal stenosis. Case Rep. Otolaryngol., 4583726
Bagley, R.L., Torvik, P.J., 1983. A theoretical basis for the application of fractional
calculus to viscoelasticity. J. Rheol. 27, 201–210.
Chung, C., Burdick, J.A., 2008. Engineering cartilage tissue. Adv. Drug Deliv. Rev. 60,
243–262.
Cooper, J.D., 2018. Use of silicone tubes in the management of complex airway
problems. Thorac. Surg. Clin. 28, 441–447.
Coussot, C., Kalyanam, S., Yapp, R., Insana, M.F., 2009. Fractional derivative models for
ultrasonic characterization of polymer and breast tissue viscoelasticity. IEEE Trans.
Ultrason. Ferroelectrics Freq. Contr. 56, 715–726.
DeAlmeida, M.F., Ferreira, L.C.F., 2012. Self-similarity, symmetries and asymptotic
behavior in Morrey spaces for a fractional wave equation. Differ. Integr. Equ. 25,
957–976.
Debry, C., Vrana, N.E., Dupret-Bories, A., 2017. Implantation of an artificial larynx after
total laryngectomy. N. Engl. J. Med. 376, 97–98.
DiMarzio, N., Eglin, D., Serra, T., Moroni, L., 2020. Bio-fabrication: convergence of 3D
bioprinting and nanobiomaterials in tissue engineering and regenerative medicine.
Front. Bioeng. Biotechnol. 8, 326.
Emre, U., Zerrin, B., Gurkan, K., Ahmet, K., Haluk, O., 2005. Soft tissue response of the
larynx to silicone, Gore-Tex, and irradiated cartilage Implants. Laryngoscope 115,
1009–1014.
Ewoldt, R.H., Johnston, M.T., Caretta, L.M., 2015. Experimental challenges of shear
rheology: how to avoid bad data. In: Spagnolie, S. (Ed.), Complex Fluids in Biological
Systems. Springer.
Gasik, M., 2017. Understanding biomaterial-tissue interface quality: combined in vitro
evaluation. Sci. Techn. Adv. Mater. 18, 550–562.
Gasik, M., Bilotsky, Y., 2019. In vitro method for measurement and model-free evaluation
of time-invariant biomaterials functions. US Patent, 10379106 B2.
Gasik, M., Zühlke, A., Haaparanta, A.M., Muhonen, V., Laine, K., Bilotsky, Y.,
Kellomäki, M., Kiviranta, I., 2018. The importance of controlled mismatch of
biomechanical compliances of implantable scaffolds and native tissue for articular
cartilage regeneration. Front. Bioeng. Biotechnol. 6, 187.
Gasik, M., Ivanov, R., Kazantseva, J., Bilotsky, Y., Hussainova, I., 2021. Biomechanical
features of graphene-augmented inorganic nanofibrous scaffolds and their physical
interaction with viruses. Materials 14, 164, 1.
Gomes, M.E., Reis, R.L., 2004. Biodegradable polymers and composites in biomedical
applications: from catgut to tissue engineering. Int. Mater. Rev. 49, 261–273.
Guilak, F., Butler, D.L., Goldstein, S.A., Baijens, F.P.T., 2014. Biomechanics and
mechanobiology in functional tissue engineering. J. Biomech. 47, 1933–1940.
Gunawardena, J., 1996. An Introduction to Idempotency. HP Laboratories Bristol,
Publication HPL-BRIMS-96-24, p. 50.
Hanyga, A., Seredynska, M., 2003. Power-law attenuation in acoustic and isotropic
anelastic media. Geophys. J. Int. 155, 830–838.
Ingber, D.E., 2006. Cellular mechanotransduction: putting all the pieces together again.
Faseb. J. 20, 811–827.
Jaalouk, D.E., Lammerding, J., 2009. Mechanotransduction gone awry. Nat. Rev. Mol.
Cell Biol. 10, 63–73.
Jammalamadaka, U., Tappa, K., 2018. Recent advances in biomaterials for 3D printing
and tissue engineering. J. Funct. Biomater. 9, 22.
Kacarevic, Ž.P., Rider, P.M., Alkildani, S., Retnasingh, S., Smeets, R., Jung, O.,
Ivaniševic, Z., Barbeck, M., 2018. An introduction to 3D bioprinting: possibilities,
challenges and future aspects. Materials 11, 2199.
Kazantseva, J., Ivanov, R., Gasik, M., Neuman, T., Hussainova, I., 2016. Graphene-
augmented nanofiber scaffolds demonstrate new features in cells behaviour. Sci.
Rep. 6, 30150.
A. Zühlke et al.
Knowlson, G.T.G., Basset, H.F.M., 1970. The pressures exerted on the trachea by
endotracheal inflatable cuffs. Br. J. Anesth. 42, 834–837.
Lord, J.D., Morrell, R., 2006. Elastic Modulus Measurement: Measurement Good Practice
Guide No. 98. NPL Teddington, UK, p. 100.
Lubarda, V.A., Chen, M.C., 2008. On the elastic moduli and compliances of transversely
isotropic and orthotropic materials. J. Mech. Mater. Struct. 3, 155–170.
Magin, R.L., 2010. Fractional calculus models of complex dynamics in biological tissues.
Comput. Math. Appl. 59, 1586–1593.
Maslov, V., 1970. The characteristics of pseudo-differential operators and difference
schemes. Actes Congrès Int. Math. 2, 755–769.
Migwi, C.M., Darby, M.I., Wostenholm, G.H., Yates, B., Duffy, R., Moss, M., 1994.
A method of determining the shear modulus and Poisson’s ratio of polymer
materials. J. Mater. Sci. 29, 3430–3432.
Nagarajan, N., Dupret-Bories, A., Karabulut, E., Zorlutuna, P., Vrana, N.E., 2018.
Enabling personalized implant and controllable biosystem development through 3D
printing. Biotechnol. Adv. 36, 521–533.
Nelson, L.S., 1998. The Anderson-Darling test for normality. J. Qual. Technol. 30,
298–299.
Nesek-Adam, V., Mršić, V., Oberhofer, D., Grizelj-Stojčić, E., Košuta, D., Rašić, Ž., 2010.
Post-intubation long-segment tracheal stenosis of the posterior wall: a case report
and review of the literature. J. Anesth. 24, 621–625.
Neumark, S., 1962a. Concept of Complex Stiffness Applied to Problems of Oscillations with
Viscous and Hysteretic Damping. A.R.C. Reports and Memoranda No. 3269. Ministry of
Aviation, London, UK, p. 36.
Neumark, S., 1962b. Concept of Complex Stiffness Applied to Problems of Oscillations
with Viscous and Hysteretic Damping. Aeronaut. Res. Council Rep. 3269. Ministry of
Aviation, London, UK, p. 36.
11
Journal of the Mechanical Behavior of Biomedical Materials 122 (2021) 104649
Noppen, M., Stratakos, G., D’Haese, J., Meysman, M., Vicken, W., 2005. Removal of
covered self-expandable metallic airway stents in benign disorders. Chest 127,
938–944.
Norris, A., 2008. Eulerian conjugate stress and strain. J. Mech. Mater. Struct. 3, 243–260.
Orr, A.W., Helmke, B.P., Blackman, B.R., Schwartz, M.A., 2006. Mechanisms of
mechanotransduction. Dev. Cell 10, 11–20.
Rodríguez, R.F., Salinas-Rodríguez, E., Fujioka, J., 2018. Fractional time fluctuations in
viscoelasticity: a comparative study of correlations and elastic moduli. Entropy 20,
28.
Sittel, C., 2009. Larynx: implants and stents. Laryngo-Rhino-Otol. 88, S119–S124.
Trabelsi, O., Prez del Palomar, A., Mena Tobar, A., Lopez-Villalobos, J.L., Ginel, A.,
Doblare, M., 2011. FE simulation of human trachea swallowing movement before
and after the implantation of endoprosthesis. Appl. Math. Model. 35, 4902–4912.
van der Meulen, M.C.H., Huiskes, R., 2002. Why mechanobiology? A survey article.
J. Biomech. 35, 401–414.
van Mow, C., Huiskes, R., 2005. Basic Orthopedic Biomechanics and Mechanobiology,
third ed. Lippincott, Williams & Wilkins, USA, p. 720.
Volfson, D., Cookson, S., Hasty, J., Tsimring, T.S., 2008. Biomechanical ordering of dense
cell populations. Proc. Natl. Acad. Sci. Unit. States Am. 105, 15346–15351.
Biomaterials for organ and tissue regeneration: new technologies and future prospects.
In: Vrana, N.E., Knopf-Marques, H., Barthes, J. (Eds.), 2020. Woodhead Publ., UK,
p. 828.
Wong, J.Y., Bronzino, J.D., 2007. Biomaterials. CRC Press/Taylor & Francis, p. 296.
Xiao, H., 1995. Invariant characteristic representations for classical and micropolar
anisotropic elasticity tensors. J. Elasticity 40, 239–265.
Zühlke, A., Gasik, M., Shahramian, K., Närhi, T., Bilotsky, Y., Kangasniemi, I., 2021.
Enhancement of gingival tissue adherence of zirconia implant posts: in vitro study.
Materials 14, 455, 2.