It is now well established that genetic factors contribute to glaucoma, and several glaucoma-associated genes have been identified. The first identified and the most studied gene is MYOCILIN, which is highly expressed in the trabecular meshwork, one of the key components in the aqueous humor outflow system of the eye. Although MYOCILIN is expressed in both ocular and non-ocular tissues, the function of the encoded protein, myocilin, is still not fully understood. We are continuing our investigations on the role of myocilin in the central (optic nerve) and peripheral (sciatic nerve) nervous systems. We have demonstrated that myocilin affects myelination in both CNS and PNS but acts through different signaling pathways. In the sciatic nerve, myocilin is expressed in Schwann cells, binds to ErbB2/ErbB3, activates these receptors, and affects the downstream PI3K-AKT signaling pathway. In the optic nerve, myocilin is expressed in astrocytes. We have demonstrated that secreted myocilin interacts with the Lingo-1/Nogo receptor complex. This interaction affects differentiation of oligodendrocytes and induces elongation of oligodendrocyte processes in the optic nerve. We have demonstrated that myocilin plays an important role regulating cell growth/survival. Myocilin-containing conditioned medium increased cell proliferation and myocilin-expressing cells were more resistant to serum starvation-induced apoptosis. Myocilin-defective mesenchymal stem cells exhibited reduced proliferation and enhanced susceptibility to serum starvation-induced apoptosis. To find the signaling pathway(s) induced by myocilin, we screened phospho-MAPK antibody arrays and identified increased phospho ERK1/2 in myocilin-expressing cells. We confirmed that phosphorylation of ERK1/2 and its upstream kinases, c-Raf and MEK, was increased in myocilin-expressing cells. In addition, the elevated phosphorylation of ERK1/2 was observed in the trabecular meshwork of transgenic mouse expressing 15-fold higher levels of myocilin. Our results suggest that myocilin promotes cell proliferation and apoptosis resistance via ERK1/2 MAPK signaling pathways. Our data also implies that myocilin may have functions not only in cells of the eye, but also in adult stem cells outside the eye. In addition to studying the pathophysiology of glaucoma, we are also interested in potential treatments for this disease. Glaucoma is associated with impairment in retrograde transport of neurotrophic factors to retinal ganglion cell bodies. Mesenchymal stem cell (MSC) transplantation appears to be protective in a variety of neurodegenerative disorders of the brain and spinal cord, in part, via neurotrophic factor secretion. We identified several factors that were secreted by MSCs and provided protection of retinal ganglion cells (RGCs) in retinal explants. Among identified factors, the strongest effect was seen for members of the PDGF family of proteins, which increased RGC survival. Blockade of PDGF signalling with anti-PDGF antibody or with small molecule inhibitors of PDGF receptor kinase or downstream phosphatidylinositol 3 kinase eliminated RGC neuroprotection conferred by MSC co-culture. We expanded these observations to animal model of glaucoma. In collaboration with Dr. Keith Martin (Cambridge University, UK), we demonstrated that intravitreal injection of PDGF-AA or -AB led to profound optic nerve neuroprotection in vivo following experimental induction of elevated intraocular pressure. These data suggest that targeting PDGF signalling pathways may represent a new approach to RGC neuroprotection and glaucoma treatment. Another possible approach for myocilin-induced glaucoma treatment was developed in collaboration with Dr. Chad Dickey (University of South Florida, Tampa). We have previously shown that myocilin is a secreted protein that is typically transported through the endoplasmic reticulum/Golgi network. Mutations in myocilin leading to severe forms of juvenile glaucoma lead to its misfolding and aggregation within trabecular meshwork cells, and ultimately, cell death. It was shown that glucose-regulated protein 94 (Grp94) specifically recognizes mutant myocilin, triaging it through endoplasmic reticulum-associated degradation. However, depletion of Grp94 led to efficient mutated myocilin degradation through an alternative autophagic clearance pathway. These data suggest that selective inhibition of Grp94 could be beneficial for patients suffering from myocilin-induced glaucoma.

Agency
National Institute of Health (NIH)
Institute
National Eye Institute (NEI)
Type
Investigator-Initiated Intramural Research Projects (ZIA)
Project #
1ZIAEY000318-16
Application #
8737619
Study Section
Project Start
Project End
Budget Start
Budget End
Support Year
16
Fiscal Year
2013
Total Cost
$1,276,516
Indirect Cost
Name
U.S. National Eye Institute
Department
Type
DUNS #
City
State
Country
Zip Code
Kwon, Heung Sun; Nakaya, Naoki; Abu-Asab, Mones et al. (2014) Myocilin is involved in NgR1/Lingo-1-mediated oligodendrocyte differentiation and myelination of the optic nerve. J Neurosci 34:5539-51
Joe, Myung Kuk; Kwon, Heung Sun; Cojocaru, Radu et al. (2014) Myocilin regulates cell proliferation and survival. J Biol Chem 289:10155-67
Johnson, Thomas V; DeKorver, Nicholas W; Levasseur, Victoria A et al. (2014) Identification of retinal ganglion cell neuroprotection conferred by platelet-derived growth factor through analysis of the mesenchymal stem cell secretome. Brain 137:503-19
Kwon, Heung Sun; Johnson, Thomas V; Tomarev, Stanislav I (2013) Myocilin stimulates osteogenic differentiation of mesenchymal stem cells through mitogen-activated protein kinase signaling. J Biol Chem 288:16882-94
Kwon, Heung Sun; Johnson, Thomas V; Joe, Myung Kuk et al. (2013) Myocilin mediates myelination in the peripheral nervous system through ErbB2/3 signaling. J Biol Chem 288:26357-71
Johnson, Thomas V; Tomarev, Stanislav I (2010) Rodent models of glaucoma. Brain Res Bull 81:349-58
Kwon, Heung-Sun; Lee, Hee-Sheung; Ji, Yun et al. (2009) Myocilin is a modulator of Wnt signaling. Mol Cell Biol 29:2139-54