Date of Completion
Citrullination, Gliosis, Retina, GFAP, Vimentin, Soluble, Intermediate Filament
Dr. Royce Mohan
Dr. Elisa Barbarese
Dr. Stephen Crocker
Dr. Betty Eipper
Dr. Zhao-Wen Wang
Field of Study
Doctor of Philosophy
Visual disorders are a massive economic and personal burden. Two leading causes of blindness in the United States, Glaucoma and Age-Related Macular Degeneration (AMD), currently have no cure. A major secondary issue in these diseases, often untreated, is the development and ongoing presence of scars in the eye. While scarring is a byproduct of the wound healing process, this scarring in the eye can lead to blindness. In the retina the scarring process is a result of ongoing retinal gliosis, mediated by the structural Müller glial cells. Two well-characterized markers of this process are the cytoskeletal Intermediate Filament (IF) proteins vimentin and the glial fibrillary acidic protein (GFAP), which become overexpressed as a stress response. Importantly, interfering with the soluble, non-cytoskeletal forms of these two proteins with the small molecule withaferin A (WFA) has anti-proliferative and anti-migratory effects, two cellular processes integral to scarring. As with many other proteins, GFAP and vimentin can be regulated by a number of different post-translational modifications (PTM). One PTM which the IF proteins are subjected to is citrullination.
Here, I have identified citrullination as an injury-induced response in the retina. Increases in citrullination are seen as early as 1 day after injury. This citrullination occurs along GFAP and vimentin filaments. I have identified two sites on GFAP that are citrullinated after retinal injury. The enzyme responsible for this injury-induced citrullination in the retina was PAD4. The citrullinated form of GFAP was targeted initially in a retinal explant system by the pan-PAD inhibitor Cl-amidine using an intravitreal injection model. The treatment decreased the levels of GFAP protein indicating a reduction of the gliotic phenotype. These findings mimicked what was seen in a mouse model of glaucoma, the DBA/2J mouse line. In this system, changes in citrullination within the glaucomatous eyes preceded changes in GFAP protein levels. These findings identify citrullination of GFAP as a major response to injury in the retina, mimicking the disease pathology of glaucoma. The ability to target these proteins within the retina, initially, and in the whole eye with an intravitreal injection to reduce a major gliotic marker has implications for the future development of therapeutics for treating the often undiagnosed gliosis associated with retinal disorders.
Wizeman, John, "The Role of Citrullination and the Type III Intermediate Filaments in Retinal Gliosis" (2016). Doctoral Dissertations. 1295.