Corneal clarity relies on well-functioning and sufficient cell density of endothelial cells to control the hydration of the cornea. Endothelium transplantation using donor tissue is the standard surgical approach for treating endothelium deficiency caused by disease or injuries.
1 More efforts have been made toward a cell therapy approach to circumvent the donor tissue shortage and extend the donor supply for multi-recipients. In cell therapy, endothelial cells are expanded in vitro and transplanted in vivo using a cell carrier or directly injecting cells into the anterior chamber to restore endothelium function.
2
To test whether cell therapy can resume endothelial function by pumping water out of the cornea to restore corneal clarity, it is necessary to establish a wound model for the experiment. Many corneal endothelium-wound models have been proposed, including in vitro, in vivo, and ex vivo. The in vitro endothelium wound models are established by scraping a linear wound into a confluent cell monolayer in a tissue culture plate.
3 This can be suitable for evaluating endothelial cell proliferation and migration and to see if cells could cover the wound area,
4 but it might not be useful for evaluating the endothelial cell pump function. An alternative way to test endothelial pump function in vitro is by measuring the cell potential differences in an Ussing chamber.
5–7 The Ussing chamber technique is used to measure the transepithelial or endothelial electrical resistance related to the integrity of the cellular barriers under investigation.
8 Endothelial pump function can be evaluated in the Ussing chamber by using ouabain.
9 However, this method still cannot directly assess the pump function in relation to reducing corneal thickness.
6 On the other hand, time and special techniques are required to set up a complex Ussing chamber system, whereas the tissue can only survive in the Ussing chamber for hours, which might not be suitable for long-term cell function observation.
10
The in vivo endothelium models use live animals such as rabbits, nonhuman primates, and rats for experiments. The rabbit is the most prevalent animal species used for endothelial wound models. Corneal thickness and clarity changes can be evaluated; however, rabbit corneal endothelial cells possess regenerative and proliferative capacity, which human corneal endothelial cells lack.
11 Therefore using rabbits to establish corneal endothelial models might not be able to recapitulate the biology and immunology of the human cornea. Rats have relatively low costs compared to other animals. However, considering the eye size and the endothelium regeneration ability, it might be difficult to assess the corneal thickness changes in cell therapy studies. Nonhuman primate eyes show similar characteristics to human eyes with limited in vivo endothelium regenerative capacity, whereas using nonhuman primates for research requires special regulation and high expenses.
12 In addition, the tectonic structure of the animal cornea differs from that of the human cornea in terms of corneal curvature, diameter, thickness, and mechanical properties.
13–15 These are also essential parameters that could affect cell attachment in cell therapy transplantation.
The ex vivo endothelium wound model is an alternative to the in vivo animal model, in which native animal or human cornea tissue is either used for testing the feasibility of cell therapy delivery methods
16,17 or establishing an organ culture system for cell attachment and cell morphology observation.
18,19 However, most of the published ex vivo models did not construct the eyeball's anterior segment structure to rebuild the anterior and posterior surfaces of the cornea. Assessing the endothelial cell function is challenging as the whole cornea is submerged in one culture medium.
20,21 Therefore this study aimed to establish a human cornea organ culture model by mounting a cornea in an artificial anterior chamber (to simulate the anterior segment structure) and where the corneal endothelial cell function could be evaluated by assessing the corneal thickness changes.