The pterygium fibroblasts were cultured on glass chamber slides and pretreated with rosiglitazone (75 μM) in serum-free DMEM, followed by TGF-β1 (2.5 ng/mL). After 48 hours of incubation, the cells were fixed with 4% paraformaldehyde (PFA) followed by peroxidase and protein blocking. The slides were then incubated with primary antibodies against α-SMA and fibronectin overnight at 4°C and then with Alexa Fluor labelled secondary antibodies (Thermo Fisher Scientific, Rockford, IL, USA) and incubated at room temperature in the dark for 2 hours. The nuclei were counterstained with 4′,6-diamidino-2-phenylindole (DAPI), and immunofluorescence was visualized and digitally captured using a confocal microscope (Leica Microsystems, GmbH, Wetzlar, Germany). Internal control was performed using PBS and secondary antibodies without the primary antibodies. The proportion of α-SMA positive cells was quantified using indirect flow cytometry. Briefly, cell suspensions (approximately 1 × 105 cells/100 μL) prepared in stain buffer (ice cold PBS, 2% FBS, 5% sodium azide) were fixed for 10 minutes with 1% PFA. After washing, the cells were suspended in 500 μL permeability buffer (stain buffer + 0.2% saponin), incubated in the dark for 5 minutes, and washed again. The cell pellets obtained after centrifugation at 1500 rpm for 5 minutes were resuspended in permeability buffer and incubated with primary antibodies against α-SMA for 30 minutes on ice in the dark. After washing, the cells were then incubated with an Alexa Fluor 488-conjugated secondary antibody for another 30 minutes, washed, and analyzed by flow cytometry.