Dexamethasone disrupts intercellular junction formation and cytoskeleton organization in human trabecular meshwork cells.
OBJECTIVE
Patients reproduce symptoms of primary open-angle glaucoma (POAG) when treated with glucocorticoids (GCs) topically on the eyes. Here we investigated the effects of GCs on junctional protein expression and cytoskeleton organization in primary human trabecular meshwork (TM) cultures to understand the molecular pathologies of POAG.


METHODS
Human TM cells from POAG (GTM) and age-matched nondiseased (NTM) individuals were obtained by standard surgical trabeculectomy. Some of the cultures were treated with dexamethasone (DEX), a synthetic GC, at 1-5 x 10(-7) mol/l for 1-7 days. The expression levels of zonula occluden-1 (ZO-1) and connexin43 (Cx43) in TM cells with or without DEX treatment were measured using reverse transcription (RT)-PCR, immunocytochemistry, and western blot analysis.


RESULTS
mRNA and proteins of ZO-1 and Cx43 were found in both NTM and GTM cells. ZO-1 and Cx43 were located on the plasma membrane, especially along the border of adjacent cells. ZO-1 had no marked changes in localization in NTM and GTM cells after treatment with 10(-7) mol/l DEX for 48 h, whereas Cx43 appeared to increase in the cytoplasm. mRNA of two ZO-1 isoforms, alpha+ and alpha-, were present in TM cells, and the former was expressed less than the latter. Only ZO-1 alpha- isoform protein was expressed in NTM cells, whereas proteins of both isoforms were found in GTM cells. DEX increased the protein levels of ZO-1 and Cx43 in both NTM and GTM cells. DEX also altered the F-actin architecture and promoted cross-linked actin network formation, the effects of which were more pronounced in GTM cells.


CONCLUSIONS
Our findings not only provide molecular insights to the pathogenesis of GC-induced glaucoma but also suggest that junctional proteins ZO-1 and Cx43 as well as F-actin are targets for developing new modalities in glaucoma therapy.
Zhuo
Ye Hong
YH
State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University, Guangzhou, China.
He
Yuan
Y
Leung
Kar Wah
KW
Hou
Fei
F
Li
Yi Qing
YQ
Chai
Fang
F
Ge
Jian
J
D000199:Actins / Q000378:metabolism; D000293:Adolescent; D000328:Adult; D002452:Cell Count; D048430:Cell Shape / Q000187:drug effects; D002478:Cells, Cultured; D018031:Connexin 43 / Q000235:genetics / Q000378:metabolism; D003432:Cross-Linking Reagents / Q000494:pharmacology; D003599:Cytoskeleton / Q000187:drug effects* / Q000378:metabolism*; D003907:Dexamethasone / Q000494:pharmacology*; D005786:Gene Expression Regulation / Q000187:drug effects; D005902:Glaucoma, Open-Angle / Q000378:metabolism / Q000473:pathology; D006801:Humans; D007365:Intercellular Junctions / Q000187:drug effects* / Q000473:pathology*; D008297:Male; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D008875:Middle Aged; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism; D014129:Trabecular Meshwork / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology*; D055815:Young Adult; D062826:Zonula Occludens-1 Protein
D000199:Actins; D018031:Connexin 43; D003432:Cross-Linking Reagents; D008565:Membrane Proteins; D010750:Phosphoproteins; C569042:TJP1 protein, human; D062826:Zonula Occludens-1 Protein; D003907:Dexamethasone