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Alessandra Micera, Bijorn Omar Balzamino, Loredana Mastrella, Paolo Lauretti, Stefano Bonini; Cyclosporin A Suppresses Tgfβ1-induced Collagen Typei/iv Expression In Both Primary And Cell Line Of Conjunctival Fibroblasts: A Possible Implication In Modulation Of Fibrosis. Invest. Ophthalmol. Vis. Sci. 2011;52(14):2014.
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© ARVO (1962-2015); The Authors (2016-present)
To explore in vitro the influence of Cyclosporin A (CsA), a powerful immunosuppressive agent, on fibrosis in term of proliferation, differentiation, ECM metabolism and contraction of primary (from n=3/biopsies and from ScienCell Res. Lab, Carlsbad, CA) cultures of conjunctival fibroblasts. CsA effects were compared with those of TGFβ1.
Both fibroblasts (FBs) and in vitro induced myofibroblasts (myoFBs) were used for CsA (Restasis, Allergan Inc., Irvine, CA) at scalar doses (50, 200, 400 and 800 ng/mL). Cells were also exposed to 10ng/mL TGFβ1, 800ng/mL CsA either alone or combined. Sister untreated cells were used as control. After 24 hours, cell viability and proliferation were assessed. Extracellular metabolism was investigated by measuring collagen typeI/IV (csELISA) and MMP9/TIMP1 (zymography, ssELISA) protein expression. Extracellular matrix contraction was monitored according to the 3D gel assay. Statistical analysis was assessed by parametric ANOVA-Tukey Kramer post hoc assays.
Increasing CsA doses did not influence either proliferation nor αSMA expression in both FBs and myoFBs. CsA did not modulate significantly collagen typeI/IV nor MMP9 expression, while TGFβ1 exposure resulted in a drastic increase of collagen typeI/IV in both cell types (p<.05). A trend toward a MMP9 increase was detected upon CsA exposure while a trend to a decrease was observed for TIMP1 (p>.05). Addition of 800ng/mL CsA to 10ng/mL TGFβ1 abolished this typeI/IV increase in both FBs and myoFBs (p<.05). Merely to MMP9/TIMP1, addition of 800ng/mL CsA to 10ng/mL TGFβ1 resulted in a suppression of TGFβ1-induced MMP9/TIMP1 expression in both cell types. At low doses, CsA triggered FB mediated contraction of a 3D gel with no effects at higher doses, as compared to those of TGFβ1.
CsA eye drops are therapeutically used for severe allergic conjunctivitis, but the mechanism of action is actually unknown. Herein, CsA appears to modulate profibrogenic effects induced by TGFβ1, a well-known factor involved in FBs/myoFBs activation, suggesting new potential mechanisms of CsA in ocular tissue remodeling/fibrosis.
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