Dehydrothermal crosslinking of the “Corneoplast”: possibilities of controlling physical, structural and biological properties. Experimental research

Abstract

Collagen crosslinking – the formation of cross-links between polypeptide chains – reduces the latter’s ability to hydrate. This property is universal for any variant of crosslinking (chemical, physical, physico-chemical) and for any form of collagen (gelatin, fibrillar collagen). Dehydrothermal crosslinking (DTC) is the formation of cross-links in biomaterials when they are heated under vacuum. Despite the fact that DTC methods are widespread in tissue engineering, its effect on the properties of the corneal stroma has not been practically studied. The use of DTK at temperatures up to 200 °C is described. For corneal stroma, the temperature limits of DTC treatment that allow its transplantation are unknown.Purpose: to evaluate the effect of dehydrothermal (DHT) cross-linking of stromal corneal grafts based on the “Corneoplast” material at temperatures of 60, 100, 140, 180 and 220 °C on their principal applicability in keratoplasty.Materials and methods. A corneoscleral disc was excised from a porcine eye (<12 h post-mortem) with epithelium and Descemet’s membrane removed, dried, and a 10 mm central corneal graft was cut out. Dry grafts (n = 36) were divided into 6 groups of 6 samples: in five groups they were kept under vacuum for 3 days at 60, 100, 140, 180, 220 °C; group 6 – untreated control; the structure was evaluated according to small-angle X-ray scattering and atomic force microscopy; for other studies, the grafts were immersed in a phosphate-salt buffer for 24 hours. Parameters evaluated: gross appearance, image quality from 5 m, light transmission, water content, central thickness, suture retention load. A test for epithelialization was performed ex vivo after 3 days in fresh porcine cornea organ culture.Results. The 220 °C mode caused charring destruction of the material. The grafts after 180 °C became significantly weaker than the 9-0 nylon thread. In the range of 60–140 °C suture retention load significantly lowered from 913 (control) down to 137 g (140 °C). Grafts increased their light transmission from 30 (control) up to 75% (140 °C); the central thickness went from 3000 (control) down to 320 um (140 °C), water content decreased from 94 (control) down to 44% (140 °C). Subjective image quality of the grafts after treatment at 60, 100, and 140 °C increased and grafts allowed distinguishing optotypes 0.3, 1.0, and 1.0 decimal, respectively. In organ culture control grafts and grafts after 60 and 100 °C treatment were covered with epithelial cells. 140 °C samples showed smooth surface with no cells upon.Conclusion. The limits of DTC processing are determined. Changing the temperature regime of DTK makes it possible to control the basic properties of “Corneoplast” in a wide range to achieve the possibility of using it as a keratoplastic material.