IKKβ is a protein kinase critical for the transmission of inflammatory cytokine signals, such as TNFα, to activation of transcription factor NF-κB and regulation of gene expression. Genetic studies in mice, however, show that IKKβ is not only involved in inflammation, but also plays a pivotal role in tissue homeostasis and stress responses. Our previous studies show that IKKβ is essential for maintaining redox homeostasis, but the underlying mechanisms and biological consequences have not been understood. The central theme of this thesis is to investigate how IKKβ modulates homeostasis and what roles it plays in biological processes. The thesis is composed of three main components.
First, mouse fibroblasts were used to investigate the molecular signatures and functions of IKKβ. I showed that genetic knockout of Ikkβ gene in fibroblasts reduced the expression of anti-oxidants thus increased the intracellular ROS level. The increased ROS in turn activated AP-1, leading to transcriptional activation of the TGFβ gene promoter and its expression, while increased TGFβ can act through NOX4 to further stimulate ROS production. Removal of IKKβ therefore triggered the autocrine amplification of the ROS-TGFβ loop that led to progressive ROS accumulation. The IKKβ-deficient fibroblasts ultimately displayed SMAD activation, leading to the overproduction of extracellular matrices and remodeling enzymes, accelerated cell motility, and enhanced markers for myofibroblast transformation and cellular senescence. These results suggested that IKKβ safeguarded the homeostasis of fibroblasts by repressing a ROS-AP-1-TGFβ autocrine regulatory loop.
Second, the in vivo roles of IKKβ were assessed in keratocytes, the fibroblasts in corneal stoma of the eye. I generated mice with Ikkβ ablation specifically in corneal keratocytes. While the knockout mice had normal corneal appearance, suggesting that IKKβ was dispensable for corneal keratocytes development and maintenance, they exhibited aggravated infiltration of inflammatory cells, cellular senescence and severe scar formation compared to normal mice in response to alkali burn injury. The wounded corneas of the knockout mice also exhibited enhanced activation of stress response and fibrogenic pathways, including JNK/AP-1 and TGFβ/SMAD cascade. Hence, similar to fibroblasts, keratocytes in corneal stroma utilize IKKβ for maintaining homeostasis and repressing ROS and TGFβ during injury responses.
Finally, the roles of IKKβ in epithelial cells of the cornea were investigated by generating and using the corneal epithelial-specific Ikkβ knockout mice. These mice had normal cornea development and maintenance, but significantly delayed healing of corneal epithelium wounds. Furthermore, delayed wound healing in the knockout cornea was not due to reduced cell proliferation or increased apoptosis, but was correlated to a slower epithelial cell migration.
Taken together, results presented in this thesis reveal that IKKβ has highly cell type-specific functions apart from its well-recognized roles in inflammatory responses. While IKKβ promotes corneal epithelial cell migration at the surface of ocular tissues, it prevents senescence, fibrosis and scar tissue formation in corneal stromal fibroblasts in response to injuries. Correspondingly, IKKβ represses a ROS-TGFβ regulatory loop in fibroblasts, thereby preventing ROS, cell migration, senescence and fibrosis.
|Commitee:||Kao, Winston Whei Yang, Lang, Richard, Liu, Chia-Yang, Puga, Alvaro|
|School:||University of Cincinnati|
|Department:||Toxicology (Environmental Health)|
|School Location:||United States -- Ohio|
|Source:||DAI-B 73/08(E), Dissertation Abstracts International|
|Subjects:||Molecular biology, Cellular biology, Environmental Health|
|Keywords:||Cell migration, Cell senescence, Cornea, Fibrosis, IkappaB kinase beta, Wound healing|
Copyright in each Dissertation and Thesis is retained by the author. All Rights Reserved
The supplemental file or files you are about to download were provided to ProQuest by the author as part of a
dissertation or thesis. The supplemental files are provided "AS IS" without warranty. ProQuest is not responsible for the
content, format or impact on the supplemental file(s) on our system. in some cases, the file type may be unknown or
may be a .exe file. We recommend caution as you open such files.
Copyright of the original materials contained in the supplemental file is retained by the author and your access to the
supplemental files is subject to the ProQuest Terms and Conditions of use.
Depending on the size of the file(s) you are downloading, the system may take some time to download them. Please be