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Jeremiah Martino, Galina Bachay, Dale d Hunter, William J Brunken; Roles for Extracellular Matrix in Corneal Development and Wound Healing. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):5607.
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© ARVO (1962-2015); The Authors (2016-present)
Laminins and netrins are extracellular matrix (ECM) molecules that play an important yet not fully characterized role in corneal development and disease. Our lab has shown that laminin β2- and γ3-chains are concentrated in the limbus and the subepithelial BM/Bowman’s Layer; netrin-4 is widely distributed. Genetic ablation of these molecules affects surface innervation and epithelial cell proliferation. Here we present data on the role of these molecules in corneal innervation and wound healing.
Corneal structure and cell distribution was studied anatomically and gene exression profiles generated using qPCR using corneas from P20 wild-type (WT), Lamb2-/-, Lamc3-/-, Lamb2-/-;c3-/-, and Ntn4-/- mice. For wounding studies, adult WT, Lamc3-/- and Ntn4-/- mice were subjected to alkali burn injury. Recovery was valuated by slit-lamp exams and histology at 12, 24 and 48hrs post-wounding.
Prior work suggested a link between nerve density and epithelial proliferation: in the Ntn4-/- mice nerve density and epithelial proliferation increased; in the Lamb2-/- or Lamc3-/- mice nerve density and epithelial proliferation decreased. We examined several trophic factors that link innervation and cell proliferation. The relative expression levels of NGF and CNTF were all down-regulated in Lamb2-/-, Lamc3-/-, Lamb2-/-;c3-/-, and Ntn4-/- vs WT mice. Macrophage invasion and activation were assayed with IB4 or Iba1, and CD68 labeling; no differences were found. No changes were seen in other immune cells (CD11b+/CD45+). Current experiments are testing other linkages between epithelial proliferation and innervation density.<br /> The higher rate of epithelial proliferation in the Ntn4-/- would suggest an increase in wound recovery. We tested this directly. Wound healing data show adult Ntn4-/- mouse corneas recovering from a chemical injury at a faster rate vs. age-matched WT or Lamc3-/- mice. Innervation and neovascularization are being evaluated.
Netrin-4 is an anti-proliferative cue and current data suggest that inhibition of netrin-4 activity may improve wound healing and hyperactivity of netrin signaling should be studied in recurrent corneal erosion. Laminin β2 and γ3 appear to be proliferative cues and current data suggest they are important in maintaining the limbal germinal niche. On-going studies are aimed at elucidating the relationship between nerve terminal density and epithelial proliferation.
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