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Invest Ophthalmol Vis Sci. 2019 Aug 1;60(10):3669-3679. doi: 10.1167/iovs.19-26983.

Induction of Fibroblast Senescence During Mouse Corneal Wound Healing.

Author information

1
Medical College, Qingdao University, Qingdao, China.
2
State Key Laboratory Cultivation Base, Shandong Provincial Key Laboratory of Ophthalmology, Shandong Eye Institute, Shandong First Medical University & Shandong Academy of Medical Sciences, Qingdao, China.
3
School of Medicine and Life Sciences, University of Jinan-Shandong Academy of Medical Sciences, Jinan, China.
4
Department of Integrative Medical Biology and Department of Clinical Sciences, Ophthalmology, Umeå University, Umeå, Sweden.

Abstract

Purpose:

To investigate the presence and role of fibroblast senescence in the dynamic process of corneal wound healing involving stromal cell apoptosis, proliferation, and differentiation.

Methods:

An in vivo corneal wound healing model was performed using epithelial debridement in C57BL/6 mice. The corneas were stained using TUNEL, Ki67, and α-smooth muscle actin (α-SMA) as markers of apoptosis, proliferation, and myofibroblastic differentiation, respectively. Cellular senescence was confirmed by senescence-associated β-galactosidase (SA-β-gal) staining and P16Ink4a expression. Mitogenic response and gene expression were compared among normal fibroblasts, H2O2-induced senescent fibroblasts, and TGF-β-induced myofibroblasts in vitro. The senescence was further detected in mouse models of corneal scarring, alkali burn, and penetrating keratoplasty (PKP).

Results:

The apoptosis and proliferation of corneal stromal cells were found to peak at 4 and 24 hours after epithelial debridement. Positive staining of SA-β-gal was observed clearly in the anterior stromal cells at 3 to 5 days. The senescent cells displayed P16Ink4a+ vimentin+ α-SMA+, representing the major origin of activated corneal resident fibroblasts. Compared with normal fibroblasts and TGF-β-induced myofibroblasts, H2O2-induced senescent fibroblasts showed a nonfibrogenic phenotype, including a reduced response to growth factor basic fibroblast growth factor (bFGF) or platelet-derived growth factor-BB (PDGF-BB), increased matrix metalloproteinase (MMP)1/3/13 expression, and decreased fibronectin and collagen I expression. Moreover, cellular senescence was commonly found in the mouse corneal scarring, alkali burn, and PKP models.

Conclusions:

Corneal epithelial debridement induced the senescence of corneal fibroblasts after apoptosis and proliferation. The senescent cells displayed a nonfibrogenic phenotype and may be involved in the self-limitation of corneal fibrosis.

PMID:
31469894
DOI:
10.1167/iovs.19-26983

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