Abstract
Purpose :
Hydrogels are promising biomaterials in regenerative medicine. They are made up of cross-linked polymer chain networks that can retain a high percentage of water content. Their structure can incorporate cells due to their biocompatibility and their permeability to oxygen, nutrients and growth factors, among others. Our aim was to synthetize and characterize a hydrogel-based 3D model of conjunctival extracellular matrix (Conj-ECMh) and to determine its capacity to incorporate and maintain stromal and/or epithelial conjunctival cells.
Methods :
The Conj-ECMh was prepared with collagen type I, hyaluronan, fibronectin, and heparan sulphate, adjusting pH and gelation temperature. Degradation of Conj-ECMh was tested using 0.1 mg/ml of collagenase. Light transmission was measured at 300-700 nm wavelengths and elasticity analyzed by spectral-domain optical coherence tomography. After freeze-drying, Conj-ECMh structure and porosity were observed by scanning electron microscopy. Biocompatibility of human conjunctival mesenchymal stromal cells (Conj-MSCs) and immortalized human conjunctival epithelial cells (IM-HConEpiC) with Conj-ECMh was tested. Conj-MSCs isolated from cadaveric donor tissues were incorporated into Conj-ECMh (1,000,000 cells/ml) and IM-HConEpiC (50,000 cells/cm2) were seeded onto the Conj-ECMh. Cell proliferation was measured for 7 days using the Alamar Blue assay (AB). Cell viability in Conj-ECMh was visualized with the live/dead staining (L/D) using confocal microscopy. Data are shown as mean ± standard deviation.
Results :
Conj-ECMhs were translucent and 1,99±0,15 mm thick in average. Degradation test showed that Conj-ECMhs completely degraded after 2h upon exposure to collagenase. The elastic modulus of Conj-ECMh was 32.05±2.1 kPa and pore size was 5.53±2.16 µm. Conj-MSCs successfully populated the Conj-ECMh and their proliferation rate reached a plateau maintained up to 7 days. IM-HConEpiC platted on Conj-ECMh surface reached confluence on day 4 and were maintained for 7 days. L/D staining showed a majority of both Conj-MSC and IM-HConEpiC living cells after 3 days in Conj-ECMh.
Conclusions :
Our results demonstrate the feasibility of a novel strategy for the production of conjunctiva-mimicking hydrogels that can provide an adequate support for stromal and epithelial conjunctival cells cell survival.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.