Возможность применения криопротекторов для хранения роговичных тканеинженерных конструкций

Abstract

Purpose. Conduct a comparative analysis of corneal tissue–engineered constructs that have undergone cryopreservation of three different cryoprotectants. Materials and methods. After refractive surgery Relex SMILE, corneal tissue was obtained which is called a lenticule. Corneal tissue engineering constructs (RTK) have created using decellularization methods that relate to the technologies of tissue and regenerative medicine. For decellularization of the lenticules, a protocol with a 1.5 M solution of Sodium Chloride with DNase 5 U / ml and RNase 5 U / ml was used. After decellularization, RTK was cryopreserved using: 1) 10% DMSO and 90% “corneal storage solution” (Borzenok–Moroz medium), 2) «Cryoderm» cell cryopreservation medium, 3) 100% glycerin. Transparency was assessed using spectrophotometry. The thickness of collagen fibers was assessed by scanning electron microscopy (SEM). Results. When assessing the transparency of the control and experimental groups, a statistically significant decrease in transparency was revealed in the groups using cryoprotectants – Glycerin and Cryoderm (p<0.001), the groups using DMSO did not differ from the control (p=0.99). Also, statistically significant differences were found between the experimental groups «Cryoderm»–DMSO (p=0.02), «Cryoderm»–Glycerin (p<0.001) and Glycerin–DMSO (p<0.001). When evaluating the SEM data, it was found that the thickness of collagen fibers did not differ in the DMSO–Control and Cryoderm–Control groups (p>0.05). The Glycerol–Control group was statistically different (p<0.001). Conclusions. In our work, we have shown the use of DMSO for storing RTK causes thickening of collagen fibers after decellularization of similar to native lenticules. This protocol can be useful for long–term storage and creation of the RTK cryobank. Key words: storage, lenticule, cryoprotectants, cryopreservation, cornea.