Figure 3. TGFBI was degraded by autophagy. (A) The levels of intracellular TGFBI and APP [amyloid β (A4) precursor protein] in primary WT, HE and HO corneal fibroblasts were examined by immunoblot analysis following bafilomycin A₁ (Baf A₁) and MG132 treatment. At approximately 60–80% confluence, cells were treated with inhibitors for 12 h. (B) Quantification of the TGFBI levels in (A) from three independent experiments using image analysis software. Intracellular TGFBI levels were highly increased after Baf A₁ (0.1 μM) treatment, whereas the levels were decreased by MG132 treatment (0.1 μM). (C) Primary corneal fibroblasts were incubated with Baf A₁ (0.1, 0.3, 0.6, 1 or 2 μM) for 12 h and harvested. TGFBI levels were determined by immunoblot analysis with an anti-TGFBI antibody. The levels of intracellular TGFBI increased in a dose-dependent manner. (D) Primary corneal fibroblasts were incubated with Baf A₁ (0.1 μM) for 1, 3, 6 and 12 h, and the TGFBI levels were determined by immunoblot as shown in (C). Intracellular TGFBI increased in a time-dependent manner. (E) Primary corneal fibroblasts were incubated with different doses of leupeptin (100 and 300 μg/ml) for 12 h and TGFBI levels were determined by immunoblot analysis with an anti-TGFBI antibody. Vertical bars: SD *p < 0.05; NS, not significant.