Urinary bladder tissue engineering using natural scaffolds in a porcine model: role of Toll-like receptors and impact of biomimetic molecules.
BACKGROUND
Natural scaffolds have been shown to induce T helper 2 (TH2)-specific immune responses in host tissues; however, the precise mechanisms that underlie this immune response are unknown. Using a porcine animal model, we evaluated the role of Toll-like receptors (TLRs) and matrix remodelling in the implantation of bladder acellular matrix (ACM) grafts and ACMs fortified with biomimetic materials.


METHODS
Bladders were decellularized with detergent and treated in 3 different ways prior to implantation: ACM alone, hyaluronic acid (HA)-ACM and HA-vascular endothelial growth factor (VEGF)-ACM. Animals were sacrificed at 4 or 10 weeks post-implantation and total gene expressions for TH2 (IL-4), TH1 (IFN-γ), TLR2, TLR4, and TGF-β1 were analyzed using real-time RT-PCR. Using histology (H&E and Masson's trichrome) and immunohistochemistry (uroplakin, α-smooth muscle actin, CD31 and factor VIII) the regenerative capacity was correlated with the gene expression of different proteins.


RESULTS
IL-4, TLR2, and TLR4 gene expression were markedly decreased at 4 and 10 weeks in both the HA-ACM group and the HA-VEGF-ACM group compared to ACM alone. IFN-γ expression was negligible in all groups and time periods. TGF-β1 expression was highest in the HA- and VEGF-treated grafts. Recellularization was inversely proportional to TLR and TH2 expression but proportional to TGF-β1.


CONCLUSIONS
ACM alone grafts demonstrated stronger TLR4 expression which may promote a distinct TH2 immune response and a reduced regenerative capacity in grafts. Treatment of grafts with HA and VEGF may help regulate host immune responses by reducing TLR4 and IL-4 and increasing TGF-β1.
Evren
Sevan
S
Developmental and Stem Cell Biology Program, The Hospital for Sick Children, University of Toronto, Toronto, Ontario, Canada.
Loai
Yasir
Y
Antoon
Roula
R
Islam
Syed
S
Yeger
Herman
H
Moore
Katherine
K
Wong
Karrie
K
Gorczynski
Reginald
R
Farhat
Walid A
WA
D000818:Animals; D040761:Biomimetic Materials; D016207:Cytokines / Q000096:biosynthesis / Q000378:metabolism; D005109:Extracellular Matrix; D005786:Gene Expression Regulation / Q000187:drug effects; D048091:Guided Tissue Regeneration / Q000379:methods*; D006820:Hyaluronic Acid / Q000494:pharmacology; D007371:Interferon-gamma / Q000378:metabolism; D015847:Interleukin-4 / Q000378:metabolism; D008264:Macrophages; D018919:Neovascularization, Physiologic; D019408:Platelet Endothelial Cell Adhesion Molecule-1 / Q000378:metabolism; D013552:Swine; D018418:Th2 Cells / Q000276:immunology; D023822:Tissue Engineering; D054457:Tissue Scaffolds; D051195:Toll-Like Receptor 2 / Q000378:metabolism; D051197:Toll-Like Receptor 4 / Q000378:metabolism; D051193:Toll-Like Receptors / Q000378:metabolism*; D053773:Transforming Growth Factor beta1 / Q000378:metabolism; D015854:Up-Regulation; D001743:Urinary Bladder / Q000098:blood supply / Q000276:immunology; D042461:Vascular Endothelial Growth Factor A / Q000494:pharmacology
D016207:Cytokines; D019408:Platelet Endothelial Cell Adhesion Molecule-1; D051195:Toll-Like Receptor 2; D051197:Toll-Like Receptor 4; D051193:Toll-Like Receptors; D053773:Transforming Growth Factor beta1; D042461:Vascular Endothelial Growth Factor A; D015847:Interleukin-4; D007371:Interferon-gamma; D006820:Hyaluronic Acid