Abstrakt
Purpose: The main goal of our study is to prove the possibility of using donor corneal tissue for more than one patient. We want to build an approach allowing to create more corneal lenticules after the preparation of a corneal lamellae for transplantation.
Methods: We provided morphological (histology, scanning electron microscope) and microbiological analysis in order to compare different methods of corneal lenticule and corneal stromal lamellae preparation and preservation. We also tested the surgical handling of the tissue to secure a safe manipulation of the tissue for clinical use.
We compared two methods of corneal lenticule preparation: microkeratome dissection and femtosecond laser. As preservation methods we tested hypothermia, cryopreservation in -80 degrees Celsius in DMSO (dimethyl sulfoxide) and storage in room temperature with glycerol.
Some intrastromal lenticules and lamellae in each group were previously irradiated by gamma irradiation of 25 kGy (KiloGray). Results: The corneal stromal lamellae prepared by microkeratome had smoother surface of the incision than after femtosecond laser.
Femtosecond laser preparation caused more irregularities on the surface and we detected more conglomerates of the fibrils, while lamellae made by microkeratome had more sparse network. Using femtosecond laser we were able to make more than five lenticules from one donor cornea.
Gamma irradiation led to damage of collagen fibrils in corneal stroma and a loss of their regular arrangement. Corneal tissue stored in glycerol showed collagen fibril aggregates and empty spaces between fibrils caused by dehydration.
Cryopreserved tissue without previous gamma irradiation showed most regular structure of the fibrils comparable to storage in hypothermia. Conclusion: Our results suggest that formation of a corneal lenticule lamellae by microkeratome results in smoother corneal lenticules, while being much cheaper than formation by femtosecond laser.
Gamma irradiation 25 kGy caused damage of the collagen fibres as well as their network arrangement which correlated with a loss of transparency and stiffer structure. These changes impair possible surgical utilisation of gamma irradiated corneas.
Storage in glycerol in room temperature and cryopreservation had similar outcomes and we believe that both methods are appropriate and safe for further clinical use.