ASSEMBLY OF PLATELET LYSATE-LOADED NANOPARTICLES AS A NEW
HYDROGEL SYSTEM FOR CARTILAGE TISSUE ENGINEERING
VITOR E. SANTO*, ELENA G. POPA
Supervisors: Rui L. Reis, João F. Mano and Manuela E. Gomes
University of Minho
School of Engineering
<Nome do Centro>
* [email protected]
Results and Discussion
Main goal
Materials & Methods
To develop a new “hydrogel” for application in the cartilage tissue
2. Assembly of PL-loaded Nanoparticles for hASCs entrapment
engineering field through the self-assembly of nanoparticles loaded
PL-adsorbed CH/CS NPs
with growth factors enabled throught the use of platelets lysates (PL)
2. Histological Characterization
hASCs (isolated from human
lipoaspirates)
hASCs+ PL loaded NPs hASCs+ empty NPs
in two aways : (i) source of growth factors and (ii) as connecting
hASCs pellets
H&E
hydrogel matrix.
100µm
Spin down
Introduction
Alcian Blue
The cartilage tissue engineering (TE) strategy described in this work
relies on the combination of natural polymer-based nanoparticles
(NPs), produced from the complexation of chitosan (CH) with
chondroitin sulfate (CS), for the incorporation and sustained release of
bioactive agents, namely platelet lysate (PL). The CH/CS complex
Safranin-O
3. In vitro culture under chondrogenic stimulus
Formulations:
The H&E staining reveals that after 28 days, the stability of the PL-loaded NPs
autologous source of a cocktail of GFs acting on tissue healing and
(i) hASCs pellets; (ii) assembled empty NPs + hASCs; (iii) assembled
hydrogel is higher in comparison with the empty NPs. Moreover, we can
repair. When used at determined concentrations, the PL-releasing
PL-loaded NPs + hASCs.
observe the formation of cartilage ECM comparable with the positive control.
NPs can assemble in a simple and quick mode to form a three-
Characterization:
dimensional (3D) stable hydrogel while in suspension with human
(i) In vitro GF release; (ii)Histology: Hematoxylin & Eosin (H&E), alcian
Adipose derived Stem Cells (hASCs), following a mild centrifugation,
blue and safranin-O; (iii) real time Polymerase Chain Reaction (rt-PCR)
allowing the cells to be entrapped in this enriched 3D environment.
for evaluation of collagen type II and type I gene expression.
mimics the extracellular matrix (ECM) interactions and PL is an
Materials and Methods
Polyelectrolyte
Complexation
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Collagen type I
Centrifugation
and resuspension
PL Adsorption
PDGF-BB
comparison with col I, showing the formation of hyaline articular cartilage ECM.
and
TGF-β1
is
release
at
day
1,
followed by a more controlled
delivery during the remaining
days. After day 7, the GF
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release was undetectable.
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The GF release profile for
The scaffolds loaded with PL showed a stronger upregulation of col II in
burst
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1. In vitro GF release
characterized by a strong
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CS in distilled
water
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CH in acetic
acid
Collagen type II
Results and Discussion
1. Nanoparticles Production and Platelet Lysate Adsorption
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3. Real time PCR – chondrogenic gene expression
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Conclusions
The presence of PLs in the 3D matrix enhances the in vitro chondrogenic
diferentiation of hASCs. PLs have a multifunctional role as a connective/stability
agent for the matrix and as a GFs supplier for chondrogenic differentiation,
creating an effective hydrogel network for cartilage TE..
Acknowledgments
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The authors thank the FCT grants (SFRH/BD/39486/2007 and SFRH/BD/64070/2009), the European
projects (NMP3-CT-2004-500283 and NMP4-SL-2009-229292), IPS and Hospital da Prelada.
Uma Escola a Reinventar o Futuro – Semana da Escola de Engenharia - 24 a 27 de Outubro de 2011