Abstract
Purpose: :
Cornea is readily exposed to environmental influences and is susceptible to injuries. For these reasons, cornea serves as an ideal in vivo system to study gene-environment interactions. IKKβ is responsible for transmitting cytokine signals to the activation of NF-ΚB, which regulates gene expression and inflammatory responses. Using mice loss of IKKβ specifically in corneal epithelial cells, we showed previously that IKKβ is required for optimal healing of corneal epithelial debridement wound by mediating NF-ΚB and p38 activation. The role of IKKβ in corneal keratocytes has never been studied.
Methods: :
Mouse embryonic fibroblasts (MEF) of Ikkβ(-/-), p65(-/-) and Ikkβflox with or without ad-Cre infection were used for in vitro studies. They were examined for gene expression profiles, cell proliferation and migration. The expression of TGFβ, its receptors and target genes, and the Tgfβ promoter activity were also evaluated. A double transgenic mouse line with IKKβ ablated specifically in corneal keratocytes was used for in vivo studies. Corneal alkali burn injury was performed and scar formation was examined at different stages after the wound.
Results: :
Loss of IKKβ or NF-ΚB p65 in MEFs resulted in higher promoter activity and expression of Tgfβ2 and Tgfβ3, higher SMAD activity and faster cell migration. Reconstitution of IKKβ expression in Ikkβ(-/-) cells significantly reversed Tgfβ level. Infection of Ikkβflox MEFs with ad-Cre also led to the same outcome; however, it only occurred 3 months after Cre-mediated Ikkβ gene ablation. Loss of IKKβ also caused increased expression of α-smooth muscle actin (α-SMA), a known consequence of TGFβ pathway activation. α-SMA is also an indicator of myofibroblast transformation that underlies fibrosis and scar tissue formation. Consistent with this observation, IKKβ deletion in corneal keratocytes resulted in more severe scar tissue formation after alkali burn injury compared to that in wild type mice.
Conclusions: :
IKKβ negatively regulates TGFβ expression and suppresses the TGFβ pathway activity. Loss of IKKβ in corneal keratocytes may potentiate scar formation after alkali burn injury. The molecular mechanism underlying the crosstalk between IKKβ and TGFβ signaling pathways requires further investigation.
Keywords: cornea: basic science • signal transduction • transcription