Pharmacological regulation of SPARC by lovastatin in human trabecular meshwork cells.
OBJECTIVE
Statins have been shown to increase aqueous outflow facility. The matricellular protein SPARC (secreted protein acidic and rich in cysteine) is a critical mediator of aqueous outflow and intraocular pressure (IOP). Here, we examine the effects of lovastatin on SPARC expression in trabecular meshwork (TM) cells, exploring the molecular mechanisms involved.


METHODS
Primary cultured human TM cells were incubated for 24, 48, and 72 hours with 10 μM lovastatin. In separate cultures, media was supplemented with either farnesyl pyrophosphate (FPP) or geranylgeranyl pyrophosphate (GGPP) for the duration of the 72-hour time point experiment. Trabecular meshwork cells were also pretreated for 24 hours with lovastatin followed by 24-hour stimulation with 3 ng/mL TGF-β2. Cell lysates and media were harvested and relative mRNA and protein level changes were determined. Krüppel-like factor 4 (KLF4) localization in normal human anterior segments was examined by immunofluorescence. Adenovirus expressing human KLF4 was used and relative changes in SPARC mRNA and protein levels were assessed.


RESULTS
Incubating TM cells with lovastatin suppressed SPARC mRNA and protein levels. This effect was reversed upon media supplementation with GGPP but not FPP. Pretreating cells with lovastatin inhibited TGF-β2 induction of SPARC. The KLF4 transcription factor was expressed throughout the TM and the inner and outer walls of Schlemm's canal. Lovastatin treatment upregulated KLF4 mRNA and protein levels. Overexpression of KLF4 downregulated SPARC expression.


CONCLUSIONS
Collectively, our data identify lovastatin as an important pharmacological suppressor of SPARC expression in TM cells, and provide further insight into the molecular mechanisms mediating statin enhancement of aqueous outflow facility.
Department of Ophthalmology, Massachusetts Eye & Ear Infirmary, Harvard Medical School, Boston, Massachusetts.
Guadalupe
G
Villarreal
Ayan
A
Chatterjee
Sarah S
SS
Oh
Dong-Jin
DJ
Oh
Douglas J
DJ
Rhee
D000328:Adult; D000368:Aged; D002478:Cells, Cultured; D005786:Gene Expression Regulation / Q000187:drug effects*; D005901:Glaucoma / Q000188:drug therapy / Q000235:genetics / Q000378:metabolism; D006801:Humans; D019161:Hydroxymethylglutaryl-CoA Reductase Inhibitors / Q000494:pharmacology; D015151:Immunoblotting; D000090062:Kruppel-Like Factor 4; D008148:Lovastatin / Q000494:pharmacology*; D008875:Middle Aged; D015676:Osteonectin / Q000096:biosynthesis / Q000235:genetics*; D012313:RNA / Q000235:genetics*; D060888:Real-Time Polymerase Chain Reaction; D014129:Trabecular Meshwork / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism
D019161:Hydroxymethylglutaryl-CoA Reductase Inhibitors; C000715467:KLF4 protein, human; D000090062:Kruppel-Like Factor 4; D015676:Osteonectin; C577241:SPARC protein, human; D012313:RNA; D008148:Lovastatin
KLF4; SPARC; TGF-β2; glaucoma; statins; trabecular meshwork