Figure 4. Increasing P2Y₁₃ expression stimulates autophagy and blocks apoA-I secretion.

HepG2 cells were transfected with either a control pCMV plasmid (pCMV) or a pCMV plasmid expressing human P2Y₁₃ (pCMV-P2Y13). Cell lysates were collected 48 h after transfection and immunoblotted for P2Y₁₃ to measure protein overexpression (Upper left panel). Histograms represent band densitometry analysis of P2Y₁₃, normalized to β-actin (inset A) and expressed as mean ± SD of 3 independent experiments. *P<0.05 vs pCMV. (A) Cell lysates were immunoblotted for LC3 and histograms represent band densitometry analysis of LC3-II, normalized to β-actin and expressed as mean ± SD of 3 independent experiments. *P<0.05 vs pCMV. (B) Transfected cells were treated with 100 µM ADP in serum-free DMEM media for 4 h, conditioned media was collected and apoA-I concentration was quantified by ELISA. ApoA-I concentration in the media is normalized to total cell protein and expressed as mean ± SD of 3 independent experiments. *P<0.05 vs pCMV Control. (C) Transfected cells were treated with 12 µM DLPC in serum-free DMEM media for 24 h, conditioned media was collected and apoA-I concentration was quantified by ELISA. ApoA-I concentration in the media is normalized to total cell protein and expressed as mean ± SD of 3 independent experiments. *P<0.01 vs pCMV Control, **P<0.01 vs pCMV+DLPC.