(1) Biomaterial for corneal endothelial monolayer transplantation and graft integration.
(2) Advanced hydrogel engineering for 3D culture and morphogenesis.
(3) Bio-fabrication of hepatobiliary organoids.
(4) Spheroid engineering.
Previous Research Projects
Synthetic hydrogel for spatio-temporal control of liver cells differentiation
Liver cholangiocytes form an intricate network of bile ducts to enable proper liver function; yet, recapitulating human stem cell differentiation to cholangiocytes in vitro requires Notch signaling and soluble ligands do not activate the Notch pathway. To overcome these limitations, jagged1 is immobilized on a chemically defined hyaluronan to specifically differentiate human embryonic stem cell-derived hepatoblasts to cholangiocytes. Hepatoblasts cultured on the jagged1-hydrogels upregulate Notch target genes and express key cholangiocyte markers including cystic fibrosis transmembrane conductance regulator. Moreover, cholangiocytes adopt morphological changes that resemble liver biliary structures. To emulate natural biliary system development, a new strategy is developed to achieve spatiotemporal control over the Jagged1–Notch2 interaction: jagged1 is first caged with a photocleavable streptavidin and then it is uncaged photochemically to restore the biological function of Jagged1, which is confirmed with Notch2 activation in a fluorescent reporter cell line. Moreover, the differentiation of human embryonic stem cell-derived hepatoblasts to cholangiocytes is temporally controlled with photochemical uncaging of this streptavidin-Jagged1-immobilized hyaluronan hydrogel. This strategy defines a framework to control protein signaling in time and space and specifically for Notch signaling for ultimate use in regenerative medicine strategies of the liver.
Previous Research Projects
Synthetic hydrogel for spatio-temporal control of liver cells differentiation
Liver cholangiocytes form an intricate network of bile ducts to enable proper liver function; yet, recapitulating human stem cell differentiation to cholangiocytes in vitro requires Notch signaling and soluble ligands do not activate the Notch pathway. To overcome these limitations, jagged1 is immobilized on a chemically defined hyaluronan to specifically differentiate human embryonic stem cell-derived hepatoblasts to cholangiocytes. Hepatoblasts cultured on the jagged1-hydrogels upregulate Notch target genes and express key cholangiocyte markers including cystic fibrosis transmembrane conductance regulator. Moreover, cholangiocytes adopt morphological changes that resemble liver biliary structures. To emulate natural biliary system development, a new strategy is developed to achieve spatiotemporal control over the Jagged1–Notch2 interaction: jagged1 is first caged with a photocleavable streptavidin and then it is uncaged photochemically to restore the biological function of Jagged1, which is confirmed with Notch2 activation in a fluorescent reporter cell line. Moreover, the differentiation of human embryonic stem cell-derived hepatoblasts to cholangiocytes is temporally controlled with photochemical uncaging of this streptavidin-Jagged1-immobilized hyaluronan hydrogel. This strategy defines a framework to control protein signaling in time and space and specifically for Notch signaling for ultimate use in regenerative medicine strategies of the liver. Bioactive, hybrid crosslinked hydrogel for human corneal tissue growth ad transplantation
The transplantation of tissue-engineered primary HCEC monolayer is a potential option to treat corneal endothelium associated blindness. We designed a robust hydrogel thin film that (i) allows primary human corneal endothelial cell growth and monolayer formation (ii) is mechanically robust to survive rupture during transplantation (iii) and enzymatically cleavable to allow gradual dissolution following transplantation. We discovered that activation of the supramolecular chain associations in form of hydrogen bonding in gelatin methacrylate (GelMA) at 4℃ followed by light triggered crosslinking could lead to remarkable increase in the strength of GelMA hydrogel. The presence of low temperature-induced physical network of gelatin triple helices coupled with microstructural changes in the hydrogel improved the subsequent UV covalent crosslinking reactions, which imparted excellent strength to hydrogels. The primary human corneal endothelial cell monolayers grown on 1 µm pillars of square-array GelMA films demonstrated improved ZO1 and Na+/K+-ATPase expression, higher cell density and homogeneity of cell size, which are indications of functionally-superior monolayers. GelMA films were transplanted in rabbit animal models to demonstrate that they were non-cytotoxic and they could gradually dissolve over the period of 12 weeks.
High resolution hydrogel topographies
Micro- and nano-topography on cell culture dishes surface also enhanced proliferation and functional markers of donor-derived primary human corneal endothelial cells.
Engineering a “Disease in a Dish” model to evaluate corneal cell therapy for Fuchs Dystrophy
The transplantation of tissue-engineered primary HCEC monolayer is a potential option to treat corneal endothelium associated blindness. We designed a robust hydrogel thin film that (i) allows primary human corneal endothelial cell growth and monolayer formation (ii) is mechanically robust to survive rupture during transplantation (iii) and enzymatically cleavable to allow gradual dissolution following transplantation. We discovered that activation of the supramolecular chain associations in form of hydrogen bonding in gelatin methacrylate (GelMA) at 4℃ followed by light triggered crosslinking could lead to remarkable increase in the strength of GelMA hydrogel. The presence of low temperature-induced physical network of gelatin triple helices coupled with microstructural changes in the hydrogel improved the subsequent UV covalent crosslinking reactions, which imparted excellent strength to hydrogels. The primary human corneal endothelial cell monolayers grown on 1 µm pillars of square-array GelMA films demonstrated improved ZO1 and Na+/K+-ATPase expression, higher cell density and homogeneity of cell size, which are indications of functionally-superior monolayers. GelMA films were transplanted in rabbit animal models to demonstrate that they were non-cytotoxic and they could gradually dissolve over the period of 12 weeks.
High resolution hydrogel topographies
Micro- and nano-topography on cell culture dishes surface also enhanced proliferation and functional markers of donor-derived primary human corneal endothelial cells.
Engineering a “Disease in a Dish” model to evaluate corneal cell therapy for Fuchs Dystrophy
Fuchs dystrophy – a degenerative disease of corneal endothelial cells -affects millions of people worldwide. Cell therapy is one potential treatment options for Fuchs. We developed a new method to recreate Fuchs environment in a dish by controlled melting of the micropatterned polystyrene surfaces at the glass transition temperature of the polymer. We used this approach to recreate the early stage and late-stage Fuchs disease environment in a dish and injected cells on them to study the regeneration of corneal endothelial monolayer. The primary human corneal endothelial cells were unable to form a monolayer in cell injection therapy approach on densely-packed synthetic guttata, which mimicked the late stage Fuchs. However, the cells could form a monolayer and express ZO1 on dome-shaped sparsely-spaced synthetic guttata micro-structures of a lower height, mimicking the early stage Fuchs. This research suggested that the cell therapy approach is more likely to succeed in early stage Fuchs patients, as compared to late-stage patients. These findings have been reproduced by other groups by using actual Fuchs affected corneas. My research was the first instance of applying micro-fabrication and polymer surface design to recreate Fuchs disease environment that could accurately predict cell therapy outcomes.