Unit of Tissue Engineering & Chemistry For Engineering A.rainer@unicampus - It

Alberto Rainer, PhD
Unit of Tissue Engineering & Chemistry for Engineering
[email protected]
• transplantation of
tissue engineering
REGENERATIVE constructs
MEDICINE
• scaffold acts as a
supporting framework
for cell delivery
• integration of
chemical and
ADVANCED IN VITRO biochemical cues in a
MODELS 3D system
• recapitulation of a 3D
tissue models
Multidisciplinary unit composed of chemists, engineers and biologists.
Chemical & biological

Synthesis of novel
characterization of
biomaterials
biomaterials
Advanced manufacturing
processes Cell/organ-on-a-chip models
• electrospinning
• additive manufacturing
• Electrospinning is a popular scaffold fabrication technique, giving
direct access to nanomaterials.
• Electrospinning exploits an intense electric field to extract polymeric

nanofibers from a solution, and depositing them as a non-woven
material. Fibers mimic the ECM and find wide application in TE.
However, the process is hampered by a ‘trial and error’ approach.
• Our work was an attempt to rationalize electrospinning process,

describing the influence of process variables on the resulting
microstructures using a Design of Experiments (DOE) method.
Seyedmahmoud R, Rainer A, Mozetic P, et al. A primer of statistical methods for correlating parameters and properties of electrospun
poly(L-lactide) scaffolds for tissue engineering--PART 1: design of experiments. J Biomed Mater Res A. 2015 Jan;103(1):91-102.
Seyedmahmoud R, Mozetic P, Rainer A, et al. A primer of statistical methods for correlating parameters and properties of electrospun
poly(L-lactide) scaffolds for tissue engineering--PART 2: regression. J Biomed Mater Res A. 2015;103(1):103-14.
ideally: “to describe scaffold
morphology as a function of
process parameters…”
Discrete values assigned to X1–5 process parameters.
Linear transformation to obtain “coded” variables from linear ones. Standard Order of Treatments in Coded Variables and Mean Response Y1–4
Representative results for
Y1 (mean diameter):
Seyedmahmoud
strong dependence uponR, Rainer A, Mozetic P, et al. A primer of statistical methods for correlating
parameters
X5 (molecular weight)and properties of electrospun poly(L-lactide) scaffolds for tissue engineering--PART 1:
design of experiments. J Biomed Mater Res A. 2015;103(1):91-102.
Seyedmahmoud R, Mozetic P, Rainer A, et al. A primer of statistical methods for correlating

parameters and properties of electrospun poly(L-lactide) scaffolds for tissue engineering--PART 2:
regression. J Biomed Mater Res A. 2015;103(1):103-14.
• Application of additive manufacturing (also known as rapid prototyping)
techniques to tissue engineering is rapidly revolutionizing tissue
engineering.
• Free form fabrication of multi-material constructs with complex shapes,

that were unachievable with conventional techniques.
• Production of 3D porous substrates (scaffolds) for tissue engineering.
• Example: highly compliant polyurethane scaffolds for myocardial tissue

engineering (MIUR FIRB “Future in Research” 2010).
Chiono V, Mozetic P et al. Polyurethane-based scaffolds for myocardial tissue

engineering. Interface Focus, 2014:4;20130045.
• While 3D printing of thermoplastic biopolymers is an established
technique for the production of e.g. bone grafts, the process has
severe limitations for soft tissues.
• Major issues regard: i) limited choice of materials; ii) low resolution;

iii) smooth surface hampering cell adhesion.
• Combination of additive manufacturing with other techniques to

improve scaffold performances  hybrid additive manufacturing.
Giannitelli S, Mozetic P, Trombetta M, Rainer A, Combined additive manufacturing Centola et al. Biofabrication (2010).
approaches for tissue engineering, Acta Biomaterialia, 2015;24:1-11.
• Another major criticality to the widespread diffusion of CATE is
represented by the processing of hydrogels.
• Current technologies are often hampered by low resolution and

harsh processing conditions.
• Incorporation of living cells in the hydrogel (i.e. bioprinting) still

represents a major challenge.
• As an outcome of FIRB project, we have developed a process

exploiting thermosensitive hydrogels for a cell-compatible
bioprinting.
Italian patent application 102015000020718: Metodo per la preparazione di costrutti cellularizzati
a base di idrogeli termosensibili tramite tecnica di prototipazione rapida (2015).
• Development of composite bioinks containing a thermosensitive polymer (e.g.
a commercial poloxamer) blended to other biopolymers (e.g. alginate).
• The poloxamer confers instant gelation and shape fidelity, while a second
crosslinking step (Ca++-induced alginate gelation) provides long term stability.
• The system has been successfully tested in combination with different cells,
showing excellent biocompatibility.
• The templating poloxamer component is rapidly eluted from the hydrogel,

which maintains its 3D structure.
• The material has also been investigated as a potential drug delivery system.
Giannitelli SM, Mozetic P, Trombetta M, Rainer A, Additive Manufacturing of Pluronic/Alginate

Composite Thermogels for Drug and Cell Delivery, in Additive Manufacturing: Innovations, Advances,
and Applications. Srivatsan TS, Sudarshan TS (eds.), Taylor & Francis, 2015.
Awarded at Advanced Functional Polymers in Medicine, Galway, 2015 (best poster presentation)
• Convergence between microfluidics and additive manufacturing.
• At the heart of the process, there is a microfluidic printing head,

with a multi-inlet configuration and terminated with a coaxial needle.
• Bioink contains alginate as a gelling agent, in blend with photo-

curable biopolymers (methacrylate derivatives) and a photo-
catalyst.
1. Bioink is extruded through the inner needle, while a Ca++ solution
is fed through the outer needle
2. Instant gelation of the bioink, which is extruded as a single fiber
3. Post-process curing by mild UV (365 nm) irradiation.
• The process is cell compatible and multi-material cell-laden

constructs can be obtained.
Microfluidic assisted bioprinting of C2C12 murine myogenic cell line
using PEG-Fibrinogen photo-curable bioink.
RATIONALE: mechanical confinement of cells into highly aligned

hydrogel fibers may be sufficient to drive cell fusion and alignment.
CAST HYDROGEL BIOPRINTING myosin heavy chain
laminin
Transverse sections
CAST HYDROGEL BIOPRINTING myosin heavy chain

laminin
Longitudinal section
myosin heavy chain
BIOPRINTING
Technological advances introduced by improved manufacturing techniques
may play a role in the diffusion of tissue engineering approaches for
regenerative medicine.
Tissue Engineering Unit @ UCBM COLLABORATIONS
Marcella TROMBETTA Univ. Tor Vergata (Sivia Licoccia, Cesare Gargioli, Stefano Cannata)
Francesco BASOLI IFN-CNR (Luca Businaro, Annamaria Gerardino, Francesca R. Bertani,
Marco COSTANTINI Adele De Ninno)
Sara M. GIANNITELLI Politecnico di Torino (Valeria Chiono, Gianluca Ciardelli)
Pamela MOZETIC ENEA Casaccia (Antonio Rinaldi)
Manuele GORI Univ. Bari (Antonio Scilimati, Maria Grazia Perrone)
Franca ABBRUZZESE Sapienza Univ. Roma (Mariella Dentini, Andrea Barbetta)
Claudia CICIONE FNUSA-ICRC, Brno (Giancarlo Forte, Vladimír Vinarský)
Maria Chiara SIMONELLI Warsaw Univ. Technology (Jakub Jaroszewicz)
FAU Erlangen-Nürnberg (Aldo R. Boccaccini)
Units @ UCBM
Translational Oncology FUNDING
Biochemistry & Molecular Biology MIUR PRIN 2010/11; PRIN 2012
Nonlinear Physics & Mathematical Modeling MIUR FIRB Futuro in Ricerca 2010
Gastroenterology Unit RICERCA FINALIZZATA RF 2011/12
Microscopic & Ultrastructural Anatomy AIRC MFAG2015
Medical Laboratory UCBM - bando interno
Cardiac Surgery

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Alberto Rainer, PhD

Unit of Tissue Engineering & Chemistry for Engineering

Chemical & biological

• Electrospinning exploits an intense electric field to extract polymeric

• Our work was an attempt to rationalize electrospinning process,

Seyedmahmoud R, Mozetic P, Rainer A, et al. A primer of statistical methods for correlating

• Free form fabrication of multi-material constructs with complex shapes,

• Production of 3D porous substrates (scaffolds) for tissue engineering.

• Example: highly compliant polyurethane scaffolds for myocardial tissue

Chiono V, Mozetic P et al. Polyurethane-based scaffolds for myocardial tissue

• Major issues regard: i) limited choice of materials; ii) low resolution;

• Combination of additive manufacturing with other techniques to

• Current technologies are often hampered by low resolution and

• Incorporation of living cells in the hydrogel (i.e. bioprinting) still

• As an outcome of FIRB project, we have developed a process

• The templating poloxamer component is rapidly eluted from the hydrogel,

Giannitelli SM, Mozetic P, Trombetta M, Rainer A, Additive Manufacturing of Pluronic/Alginate

• At the heart of the process, there is a microfluidic printing head,

• Bioink contains alginate as a gelling agent, in blend with photo-

• The process is cell compatible and multi-material cell-laden

RATIONALE: mechanical confinement of cells into highly aligned

CAST HYDROGEL BIOPRINTING myosin heavy chain

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