Low dose latrunculin-A inhibits dexamethasone-induced changes in the actin cytoskeleton and alters extracellular matrix protein expression in cultured human trabecular meshwork cells.
We determined the effects of a low dose of the actin-disrupting agent latrunculin (LAT)-A on dexamethasone (DEX)-induced changes in actin organization, focal adhesions, and production of extracellular matrix proteins in cultured human trabecular meshwork (HTM) cells. HTM cells were cultured to a highly confluent stage with stable endothelium-like morphology and incubated with 0.1 or 0.2 microM DEX and/or 0.1 microM LAT-A. Changes in the actin cytoskeleton and vinculin-containing focal contacts were evaluated by immunofluorescence microscopy. Expression of thrombospondin-1 (TSP1) and fibronectin (FN) in HTM cells was evaluated by Western blot analysis. The results showed that DEX induced morphological changes and actin reorganization in HTM cells. The cells partly recovered after DEX withdrawal, but the addition of low dose LAT-A hastened the recovery. In addition, DEX failed to induce changes when co-incubated with LAT-A for at least 4 weeks, and for at least 2 weeks when cells were pre-treated with LAT-A for 2 weeks. HTM cells treated with 0.1 microM LAT-A only for 5 days showed mild disorganization of the actin cytoskeleton and focal adhesions, which persisted during the 4 weeks of treatment. DEX stimulated production of FN in HTM cells independent of LAT-A treatment. LAT-A and, to a lesser extent, DEX inhibited production of TSP1 by HTM cells. Although LAT-A is not a DEX receptor antagonist, it is able to prevent the effects of DEX on the actin cytoskeleton in cultured HTM cells at a dose subthreshold for increasing outflow facility in monkeys. This suggests that LAT-A at low doses may be useful in treating steroid and other glaucomas. TSP1 may be an important target of LAT-A in HTM cells and modulation of TSP may influence the actin cytoskeleton of the trabecular meshwork (TM), and consequently, intraocular pressure.
Liu
X
X
Department of Ophthalmology and Visual Sciences, University of Wisconsin, F4/328 CSC, 600 Highland Avenue, Madison, WI, USA.
Wu
Z
Z
Sheibani
N
N
Brandt
C R
CR
Polansky
J R
JR
Kaufman
P L
PL
D000199:Actins / Q000187:drug effects*; D015152:Blotting, Northern; D015153:Blotting, Western; D019086:Bridged Bicyclo Compounds, Heterocyclic / Q000008:administration & dosage* / Q000494:pharmacology; D002478:Cells, Cultured; D003599:Cytoskeleton / Q000187:drug effects*; D003907:Dexamethasone / Q000009:adverse effects*; D016326:Extracellular Matrix Proteins / Q000378:metabolism*; D005353:Fibronectins / Q000378:metabolism; D006801:Humans; D013844:Thiazoles / Q000008:administration & dosage* / Q000494:pharmacology; D053778:Thiazolidines; D019700:Thrombospondin 1 / Q000378:metabolism; D014129:Trabecular Meshwork / Q000187:drug effects*
D000199:Actins; D019086:Bridged Bicyclo Compounds, Heterocyclic; D016326:Extracellular Matrix Proteins; D005353:Fibronectins; D013844:Thiazoles; D053778:Thiazolidines; D019700:Thrombospondin 1; D003907:Dexamethasone; C037067:latrunculin A