Tissue-engineered 3D cornea model to simulate wound healing and scarring of the cornea
Trauma, infection, and wound healing disorders of the cornea can cause scars that significantly impair vision. The development of a standardized in vitro 3D cornea model may allow investigation of new potential agents without animal testing.
Trauma, infection, and corneal wound healing disorders can leave scars in the optical axis that significantly impair vision and - in the worst case - lead to blindness. More effective therapies for the treatment of corneal wounds and especially for the treatment of prolonged wound healing disorders would be a great progress in ophthalmology.
Numerous methods already exist for testing the cornea's response to chemicals, drugs, cosmetics, or other consumer products. The current standard for testing ocular toxicity is the use of various animal models such as the Draize eye test. This is a standardized method in which different concentrations of a substance are applied to the eyes of a rabbits and the reaction of the eyes is evaluated at different time points. Test animals are sacrificed after completion of the test.
The 3D cornea model we developed consists of epithelium and stroma (Figure 1), the two layers in which corneal scarring mainly takes place.
For the stroma, primary human keratocytes are cultured in type 1 collagen. This hydrogel is compressed to a thickness of 500 µm and immortalized or primary human epithelial cells are seeded on top. After one day of growth submerged in medium, the culture is continued as air-lift for 9 days. Meanwhile, a multi-layered, non-keratinizing squamous epithelium has formed, which tightly adheres to the stromal substitute of collagen hydrogel. From day 12 onward, the model can be used for the scenario under investigation, for example, infection with pathogens or simulation of a wound healing process.
The special feature of this model is its high similarity to the natural human cornea: The surface forms a multilayered non-keratinizing squamous epithelium that is histologically very similar to the human cornea and expresses corneal epithelium-specific markers. The cornea models can be kept in culture for another 20 days. In preliminary work we were able to distinguish all UN GHS categories in our 3D cornea model.
Aim of this project is to adapt our existing 3D cornea model to investigate corneal wound healing. In the long run, we aim to develop a standardized test system, but also to gain a better understanding of wound healing and scar formation in general. The test system could allow the investigation of potentially wound healing modulating substances, which so far has only been possible in animal experiments.
Publikation:
Lotz, C., Kiesewetter, L., Schmid, F. F., Hansmann, J., Walles, H. & Groeber-Becker, F. Replacing the Draize eye test: Impedance spectroscopy as a 3R method to discriminate between all GHS categories for eye irritation. Sci Rep 8, 15049, doi:10.1038/s41598-018-33118-2 (2018).
Project management
Dr. Daniel Kampik
PhD in Infectious Diseases and Immunology at the Max von Pettenkofer Institute for Medical Microbiology, LMU Munich. Specialisation in Ophthalmology at the Eye Clinic of Würzburg University Hospital. PhD at the Institute of Ophthalmology, University College London, and Moorfields Eye Hospital, in the field of gene therapy for the cornea and retina. Since 2013, senior registrar at the Eye Clinic of Würzburg University Hospital.
Cooperation
Dr. Ann-Katrin Regensburger
Studied medicine at the University of Lübeck and the Julius Maximilian University of Würzburg. Completed a PhD in the field of autoimmune diseases at the University of Lübeck, within the Research Training Group ‘Modulation of Autoimmunity’. Since 2017, she has been a junior doctor at the Eye Clinic of Würzburg University Hospital.
Dr. Christian Lotz
Studied biomedicine at Julius Maximilian University of Würzburg. Completed a PhD in ocular tissue engineering at the Chair of Tissue Engineering and Regenerative Medicine at Würzburg University Hospital. Since 2018, research associate at Fraunhofer ISC in Würzburg, specialising in areas including ocular tissue engineering and 3D skin models as alternatives to animal testing. Since 2019, Deputy Head of In-vitro Test Systems at the Fraunhofer Translational Centre for Regenerative Therapies (TLZ-RT) in Würzburg.