Reducing P4HA2 expression or inhibiting its activity impairs collagen deposition. (A) Western blot analysis of collagen I and IV in conditional media generated from the same amount of control and P4HA2-silenced (sh-P4HA2-1) T4-2 cells; bar graph shows quantification of Western blot results (*p < 0.05; **p < 0.01) (B) Immunofluorescence staining of collagen I (green) and IV (green) in control and P4HA2-silenced T4-2 cells. P4HA2 knockdown (sh-P4HA2-1) T4-2 cells showed impaired collagen I and IV protein deposition. (C) Immunofluorescence staining of collagen I (green) and IV (green) in control and 1,4-DPAC treated T4-2 cells in 3D culture. T4-2 cells treated with 1,4-DPAC showed lower expression of collagen I and IV (scale bar, 50 μm).