Open Access

ARTICLE

Three-month effects of corneal cross-linking on corneal fibroblasts

XINYAN CHEN^1,2, HAIXIA ZHANG^1,2, LIN LI^1,2,*

1 School of Biomedical Engineering, Capital Medical University, Beijing, 100069, China
2 Beijing Key Laboratory of Fundamental Research on Biomechanics in Clinical Application, Capital Medical University, Beijing, 100069, China

* Corresponding Author:* Address correspondence: Lin Li,

(This article belongs to the Special Issue: Cellular Biomechanics in Health and Diseases)

BIOCELL 2021, 45(4), 1023-1032. https://doi.org/10.32604/biocell.2021.014873

Received 20 November 2020; Accepted 10 January 2021; Issue published 22 April 2021

Abstract

Corneal collagen crosslinking (CXL) has revolutionized the treatment of keratoconus in the past decade. In order to evaluate the 3-month effects of CXL on corneal fibroblasts, a longitudinal study at the tissue and cellular level was carried out with a total of 16 rabbits that underwent CXL, deepithelialization (DEP), or non-treatment (control) and kept for 1 to 3 months. The duration of corneal stromal remodeling after CXL was determined by examining the differentiation, apoptosis, and number changes of keratocytes in tissue sections from animals 1, 2, or 3 months post-treatment. Upon the finish of tissue remodeling, separate rabbits were used to extract keratocytes and set up cell culture for vimentin immunofluorescence staining. The same cell culture was used for (1) migration measurement through the wound-healing assay; (2) elastic modulus measurement by atomic force microscope (AFM); (3) the proliferation, apoptosis, cytoskeleton and α-SMA expression tests through EdU (5-ethynyl-2’-deoxyuridine) assay, TUNEL (TdT-mediated dUTP Nick-End Labeling) assay, phalloidin and α-SMA (alpha-smooth muscle actin) immunofluorescence analysis, respectively. Results showed that the migratory activity, elastic modulus, and α-SMA expression of the corneal fibroblasts increased after CXL treatment, while apoptosis, proliferation, and morphology of F-actin cytoskeleton of the fibroblasts had no significant change after 3 months. In contrast, measured cellular parameters (migratory, elastic moduli, α-SMA expression, apoptosis, proliferation, and morphology of F-actin cytoskeleton of fibroblasts) did not change significantly after DEP. In conclusion, the dynamic changes of keratocytes were nearly stable 3 months after CXL treatment. CXL has an impact on corneal fibroblasts, including migration, elastic modulus and α-SMA expression, while epithelialization may not alter the biological behavior of cells significantly.

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