Human Optic Nerve Head Glial Cell Activation Following Biomechanical Compression

Glaucoma is the most prevalent neurodegenerative disease and the worldâ s second leading cause of blindness resulting from the slow and painless neuropathy of the optic nerve head, and death of the retinal ganglion cells. While very little is known about the onset and pathology of the disease, activated astrocytes are found in the optic nerve heads of early glaucoma patients. Activation of glial cells, including astrocytes, is known to be associated with several neurodegenerative diseases and believed to contribute to their pathophysiology. Glial cell activation has successfully been induced using biomechanical stretch, which was predicted to be experienced by the lamina cribrosa under elevated intra ocular pressure conditions. However, biomechanical compression was also predicted to be even more prevalent, although no good models existed to replicate this condition in vitro. It is for this reason that I have developed a novel model for replicating cellular compression by compressing the medium to which cells adhere in an equi-axial manner. Through careful calibration this model uses a device that is capable of generating specific and consistent amounts of compression by pre-stretching a sylastic membrane upon which cells are then seeded and cultured. The membranes are then removed from the device allowing them to return to their un-stretched state and compressing all cells that are attached. In this research I show how the model was designed and calibrated as well as its usefulness as an in vitro model for glial cell activation using biomechanical compression on human astrocytes and lamina cribrosa glial cells. Using advanced proteomic techniques, nearly 350 unique proteins of interest were observed to be regulated as a result of activation giving us significant insight into the pathways and networks involved in this process. Of particular interest are PEA15, RhoA, IL6, MMP2, HSP90, and FLNB. This model was also successfully used to target and block the actin stress fibre formation pathway, which is normally present in activated astrocytes. The results presented in this research give a powerful new model for examining the effects of cellular compression. It also provides insight into the networks and pathways associated with glial cell activation allowing us to better understand its role in the pathophysiology of glaucoma. This information will assist in efforts to understand the disease and develop better methods for detection, treatment and potential cure for this debilitating disease.Ph.D