Femtosecond laser cutting of multiple ultrathin corneal stromal lamellae for endothelial graft bioengineering

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Abstract

Purpose

The bioengineering of endothelial grafts (EG) is developing in several countries as an answer to the shortage of donor corneas worldwide. They are composed of a thin transparent biocompatible carrier evenly covered with tightly packed fully functional endothelial cells. Multiple options exist for the carriers. In our laboratory, we are focused on the optimization of human donor tissue because the well-organised cornea donation in France is not the most limiting point. Aim: to present the preparation of ultrathin stromal lamellae from a single human cornea

Methods

Human organ-cultured corneas unsuitable for graft were used either with our without deswelling in Dextran containing medium. They were mounted on a artificial anterior chamber and cut with a femtosecond laser (femto LDV Z6, Ziemer) with aplanation. Six horizontal parallel cuts were realized on a 9 mm diameter area without vertical section. Cornea was then trephine at 8.00mm on a punch trephine. Thickness map was obtained before and after the cut with an AS-OCT equipped with a custom adapter for ex vivo corneas. Lamellae were separated under an operating video-microscope and again observed with OCT. Quality of surface were analysed with scanning electron microscopy and multiphotonic confocal microscopy with SHG for collagen fiber imaging

Results

Seven regular lamellae of approximately 100μm were obtained on a single cornea and easily detached. Characterisation of surface quality is ongoing

Conclusion

Femtosecond laser cutting of multiple lamellae on a single human cornea could allow obtaining a sufficient number of thin transparent and fully biocompatible carriers for EG bioengineering

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