In vitro study on corneal biomechanical changes after corneal stromal lenticule implantation combined with corneal collagen cross-linking

Objective  To investigate the changes in corneal biomechanics of isolated porcine eyes after stromal lenticule addition keratoplasty (SLAK) combined with corneal collagen cross-linking (CXL).

Methods  A total of 139 porcine eyes from 5- to 7-month-old Duroc pigs were collected within 3 hours after death. Twenty-four eligible eyes were selected and divided into a normal control group, SLAK group, CXL group and SLAK-CXL group, and the tensile strength and elastic modulus of each group were detected via uniaxial tensile test. One hundred eligible eyes were assigned to the SLAK group, SLAK combined with riboflavin soaking (RNF) group (SLAK-RNF group), SLAK after cross-linking of both donor and recipient group (CXL-CXL-SLAK group), SLAK combined with simple ultraviolet A (UVA) irradiation group (SLAK-UVA group), and SLAK-CXL group, and the maximum lap shear strength of each group was measured via the lap shear test. The remaining 15 eligible eyes were divided into SLAK group, SLAK-RNF group and SLAK-CXL group, and the ultrastructure of recipient corneal stroma and donor stromal lens in each group was observed by hematoxylin-eosin staining and Masson staining. Slit-lamp microscope and Sirius corneal topographer were used to observe clinical features of the tested eyes in the normal control group, SLAK group, CXL group and SLAK-CXL group. All experiments were performed in compliance with the guidelines specified in the Association for Research in Vision and Ophthalmology Statement for the Use of Animals in Ophthalmic and Vision Research. The experimental protocol was approved by the Biomedical Ethics Committee of Xi’an Jiaotong University Health Science Center (No. XJTUAE2024-2042).

Results  The tensile strength of the normal control group, SLAK group, CXL group and SLAK-CXL group at 50% strain was (0.60±0.15), (0.45±0.19), (1.56±0.27) and (1.22±0.24)MPa, respectively, and the elastic modulus was (2.01±0.69), (1.69±0.52), (4.16±1.28) and (3.82±0.68)MPa, respectively, showing statistically significant differences ( F=29.39, 11.01; both P<0.001). The tensile strength and elastic modulus of the CXL group and SLAK-CXL group were significantly higher than those of the normal control group (all P<0.001). There was a significant overall difference in lap shear strength among the SLAK group, SLAK-RNF group, CXL-CXL-SLAK group, SLAK-UVA group and SLAK-CXL group ( F=18.45, P<0.001). The lap shear strength of the SLAK-RNF group and CXL-CXL-SLAK group was significantly lower than that of the SLAK group, and the lap shear strength of the SLAK-UVA group was significantly higher than that of the SLAK group, and the lap shear strength of the SLAK-CXL group was significantly higher than that of the CXL-CXL-SLAK group (all P<0.05). Hematoxylin-eosin staining and Masson staining results showed that the collagen fibers in each group were arranged orderly. The diameter of collagen fibers and the proportion of collagen fibrosis area in the anterior corneal stroma and stromal lenticule in the SLAK-CXL group were greater than those in the SLAK group, and the differences were statistically significant (both P<0.05). Slit lamp microscope and corneal topography results showed that the implanted lenticule in the SLAK group and SLAK-CXL group was anastomosed with the recipient, with a visible lens boundary, reduced transparency, and obvious edema in the surrounding stroma.

Conclusions  SLAK-CXL can increase corneal thickness and corneal biomechanical strength, which may result from cross-linking increasing the diameter and area of matrix fibers. However, slight corneal edema may occur after surgery, and cross-linking can not achieve immediate bonding between the lenticule and the recipient cornea.