An experimental study on the construction of tissue engineering corneal stromal stent with Smile derived corneal stromal lens

Abstract

AIM: To investigate the feasibility of constructing corneal stromal scaffolds and the optimal preservation conditions of corneal stromal lenses obtained from the small incision lenticule extraction(Smile)surgery.METHODS: Constructing a bilayer lens by adhering together two corneal stromal lenses with human fibrin sealant(FS). Human corneal fibroblasts were isolated and cultured from Smile derived corneal stromal lenses in vitro, and the toxicity of FS on human corneal fibroblasts was detected by MTT method. The bilayer lenses were then placed in anhydrous glycerin, sodium hyaluronate eye drops, a simulated wet room environment and fetal bovine serum groups respectively, and stored at 4℃ for 14d. The transparency, hardness and stability of the scaffolds were then compared. Afterwards, the bilayer lens scaffolds were stored in anhydrous glycerin at room temperature, 4℃ and -20℃. After 14d of preservation, the diverse effects of temperature on the transparency and hardness of the scaffolds were compared.RESULTS: MTT results showed that the cells of the experimental group and the control group had similar proliferation trend within 0-72h. The cytotoxicity rating of the experimental group was 0 at 36-48h and 1 at 24h and 60-72h. The relative survival rate of the cells within 0-72h was over 90%. FS-bonded bilayer lens scaffold had a smooth surface, close bonding, good transparency and suitable hardness. After 14d of storage at 4℃, none of the nine bilayer lens scaffolds in the anhydrous glycerol group showed signs of cracking cracking after rehydration, and their transparency was good. In the sodium hyaluronate group, three of the nine scaffolds cracked and the remaining six were still intact. In the simulated wet room environment group, none of the 9 scaffolds cracked, but there were different degrees of shrinkage, their surface was rough and transparency was lower. In the fetal bovine serum group, all the 9 stents were cracked, and the single corneal stromal lens was soft and edema was serious. Out of the 15 bilayer lens scaffolds preserved in anhydrous glycerol at room temperature, 2 remained colourless and transparent, 5 slightly yellowed but still remained transparent, 8 yellowed substantially with a significant reduction in transparency. Out of the 15 bilayer lens scaffolds preserved in anhydrous glycerol at 4℃, 5 remained colourless and transparent, and 10 slightly yellowed while remaining transparent. Of the 15 bilayer lens scaffolds preserved in anhydrous glycerol at -20℃, none of the scaffolds yellowed, therefore, remaining colourless and transparent.CONCLUSION: FS is a safe and non-toxic bio-gel. It can be used to glue Smile-derived corneal stromal lenses to construct corneal stromal scaffolds with good stability, high transparency and suitable hardness. Anhydrous glycerol at -20℃ is the best preservation condition for corneal stromal lens scaffolds.

Keywords

• tissue-engineered cornea
• corneal stromal lens
• fibrin adhesive
• cytotoxicity
• preservation