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Sharad Mittal, Afsaneh Amouzegar, Masahiro Omoto, Sunil Chauhan; Mesenchymal stem cells restore corneal transparency via secretion of hepatocyte growth factor. Invest. Ophthalmol. Vis. Sci. 201657(12):.
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© ARVO (1962-2015); The Authors (2016-present)
Mesenchymal stem cells (MSCs) have been shown to promote repair in corneal injury and inflammation. The purpose of this study was to investigate the underlying mechanisms by which MSCs inhibit stromal fibrosis and restore transparency in corneal injury.
Mesenchymal stem cells were generated from bone marrow of wild-type C57BL/6 mice, and characterized by flow cytometry for the expression of CD29+SCA1+CD45−CD34−. Corneal injury was induced by mechanical removal of the corneal epithelium and anterior stroma. Real-time PCR was performed on injured corneas as well as on resting and IL1β-stimulated MSCs to quantify anti-inflammatory and growth factors, including interleukin-10, TNFα–stimulated gene/protein 6 (TSG6), transforming growth factor beta (TGFβ) and hepatocyte growth factor (HGF). HGF in MSCs was knocked down using HGF-specific siRNA. Control or HGF-silenced MSCs were intravenously injected into mice 1h after corneal injury. Corneal epithelial injury and transparency were evaluated using corneal fluorescein staining and biomicroscopy. Immunohistochemistry was performed to examine the effect of HGF on TGFβ-induced α-smooth muscle actin (αSMA; a marker of fibrosis) expression in a corneal fibroblast cell line (MK/T1).
Real-time PCR analysis showed that IL1β-stimulated MSCs expressed significantly higher levels of HGF compared to unstimulated resting MSCs (p=0.0035). Following injury, corneas of MSC-treated mice showed a 2-fold increase in the expression of HGF compared to MSC-untreated injured cornea (p=0.01). siRNA-mediated silencing of HGF expression in MSCs substantially abrogated their wound repair function. Quantification of corneal opacity scores demonstrated that HGF-silencing of MSCs led to a significant 2-fold reduction in their capacity to prevent the development of stromal fibrosis compared to wild type MSCs (p=0.016). Immunohistochemistry data demonstrated that HGF completely suppressed TGFβ-induced α-SMA expression in corneal fibroblasts (p=0.0001).
These findings indicate that mesenchymal stem cells exert their anti-fibrotic function primarily via secretion of hepatocyte growth factor.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.
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