title: Inhibition of TGFBIp expression by lithium: implications for TGFBI-linked corneal dystrophy therapy.
abstract: PURPOSE. The purpose of this study was to investigate the effects and molecular mechanisms of lithium on inhibition of TGFBIp expression as a potential therapy for TGFBI-linked corneal dystrophy. METHODS. Primary culture corneal fibroblasts were isolated from the corneas of healthy subjects and patients with granular corneal dystrophy type 2 (GCD2) with a homozygous mutation in TGFBI R124H. Levels of TGFBIp and its mRNA in corneal fibroblasts treated with various lithium (LiCl) concentrations were analyzed by Western blot, RT-PCR, and quantitative real-time PCR. RESULTS. LiCl treatment reduced the expression levels of normal and mutant TGFBIp in a dose- and a time-dependent manner. Furthermore, TGF-β1-induced TGFBIp expression decreased by 35% and 67% after treatment with 5 mM and 10 mM LiCl, respectively. LiCl decreased the level of pSmad3 (S423/425) in a dose-dependent manner. Furthermore, LiCl increased the level of pGSK-3α/β (S21/9) in a dose-dependent manner. Also observed was the interaction between GSK-3β and Smad3, which was enhanced by lithium. In addition, Western blot analysis showed that the ratio of LC3-II/LC3-I in corneal fibroblasts increased after LiCl treatment. Cell viability at different doses was greater than 98%, indicating that LiCl did not induce significant corneal fibroblast death. Finally, the observed attenuating effects of LiCl on TGFBIp expression were not the results of cell death. CONCLUSIONS. The accumulation of mutant TGFBIp ultimately leads to the histopathologic and clinical manifestations associated with TGFBI-linked corneal dystrophy. These data strongly suggest that lithium may be used for the prevention or treatment of this disease.
authors.affiliation: Corneal Dystrophy Research Institute, Yonsei University College of Medicine, Seoul, South Korea.
authors.forename: Seung-Il
authors.initials: SI
authors.lastname: Choi
authors.forename: Bong-Yoon
authors.initials: BY
authors.lastname: Kim
authors.forename: Shorafidinkhuja
authors.initials: S
authors.lastname: Dadakhujaev
authors.forename: James V
authors.initials: JV
authors.lastname: Jester
authors.forename: Hyunmi
authors.initials: H
authors.lastname: Ryu
authors.forename: Tae-Im
authors.initials: TI
authors.lastname: Kim
authors.forename: Eung Kweon
authors.initials: EK
authors.lastname: Kim
mesh_terms: D001343:Autophagy / Q000187:drug effects; D015153:Blotting, Western; D002478:Cells, Cultured; D003315:Cornea / Q000166:cytology; D003317:Corneal Dystrophies, Hereditary / Q000188:drug therapy* / Q000473:pathology; D004305:Dose-Response Relationship, Drug; D016326:Extracellular Matrix Proteins / Q000235:genetics* / Q000378:metabolism; D005347:Fibroblasts / Q000187:drug effects* / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects*; D038362:Glycogen Synthase Kinase 3 / Q000378:metabolism; D000071679:Glycogen Synthase Kinase 3 beta; D006801:Humans; D018021:Lithium Chloride / Q000494:pharmacology*; D010766:Phosphorylation; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D051900:Smad3 Protein / Q000378:metabolism; D013997:Time Factors; D016212:Transforming Growth Factor beta / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D000099222:betaIG-H3 Protein
chemical_list: D016326:Extracellular Matrix Proteins; D038362:Glycogen Synthase Kinase 3; D000071679:Glycogen Synthase Kinase 3 beta; D018021:Lithium Chloride; D012333:RNA, Messenger; D051900:Smad3 Protein; D016212:Transforming Growth Factor beta; D000099222:betaIG-H3 Protein; C497790:SMAD3 protein, human; C000605506:GSK3B protein, human; C448021:glycogen synthase kinase 3 alpha