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Satoshi Inagaki, Kazuhide Kawase, Michinori Funato, Junko Seki, Chizuru Kawase, Kazuki Ohuchi, Tsubasa Kameyama, Shiori Ando, Arisu Sato, Wataru Morozumi, Shinsuke Nakamura, Masamitsu Shimazawa, Daisuke Iejima, Takeshi Iwata, Tetsuya Yamamoto, Hideo Kaneko, Hideaki Hara; Effect of Timolol on Optineurin Aggregation in Transformed Induced Pluripotent Stem Cells Derived From Patient With Familial Glaucoma. Invest. Ophthalmol. Vis. Sci. 2018;59(6):2293-2304. doi: 10.1167/iovs.17-22975.
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To determine a chemical agent that can reduce the aggregation of optineurin (OPTN) in cells differentiated from induced pluripotent stem cells obtained from a patient with normal-tension glaucoma (NTG) caused by an E50K mutation in the OPTN gene (OPTNE50K-NTG).
Retinal ganglion cells (RGCs) were created from induced pluripotent stem cells derived from a healthy individual (wild-type [WT]-iPSCs) and from a patient with NTG due to OPTNE50K (E50K-iPSCs) mutation. The death of the induced RGCs was evaluated by counting the number of TUNEL- and ATH5-positive cells. Axonal growth was determined by measuring the axonal length of TUJ1-positive cells. OPTN aggregation was assessed by measuring the OPTN-positive area by immunofluorescence and by Western blotting. Autophagic flux assay was investigated by determining the light chain 3 (LC3)B-II/LC3B-I ratio and p62 expression by Western blotting.
The results showed OPTNE50K aggregation, activation of astrocytes, reduction in the number of RGCs, and enhancement of apoptotic cell death in the in vitro OPTNE50K model of NTG. Timolol was found to reduce the OPTNE50K-positive area and decreased the insoluble OPTNE50K, suggesting that it has the potential of reducing the OPTNE50K aggregation. Timolol also increased the ATH5-positive cells, decreased TUNEL-positive cells, increased the LC3B-II/LC3B-I ratio, and decreased the expression of p62. These findings suggest that timolol might enhance autophagic flux, leading to reduced OPTNE50K aggregation.
Timolol should be considered a potential therapeutic agent specific to OPTNE50K-NTG because it can reduce the OPTNE50K aggregation in E50K-iPSCs-RGCs by enhancing autophagic flux and neuroprotective effects.
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