Human ENPP1 over-expression inhibits DMI-induced adipocyte maturation in 3T3-L1 cells.

<p>Human ENPP1 was over-expressed in 3T3-L1 using retrovirus transfection with CMV promoter. Panel A. ENPP1 protein content in cells containing vector only and in cells over-expressing human ENPP1 K121K or K121Q. ENPP1 was identified at 130 kDa using antibody from Imegex Corp. (San Diego, CA). Panel B. Oil-red-O stained 3T3-L1 cells after induction of adipogenesis. Cells were 100% confluent at the time of assay. Cellular triglyceride content is presented as fold changes from the triglyceride content of cells containing vector. Results are reported as mean and SD from measurements performed three times (8 wells measured in each experiment). Panel C. Cells were plated on 6-well plate and induced to differentiation with DMI. At different days, as indicated, the cell number was counted. Results are reported as mean and SD from measurements repeated three times (3 wells measured in each experiment). Open squares represent the data for the vector 3T3-L1. Open circles represent the data for 3T3-L1 over-expressing human ENPP1 K121K. Dark circles represent the data for 3T3-L1 over-expressing human ENPP1 K121Q. Cell death was measured at day 2 and day 4 of differentiation with DMI. Results are shown as mean and SD of percentage of dead cells relative to total cells. Measurements were performed for number of viable and dead cells in three wells for each cell type. These experiments were repeated 3 times. * p<0.05 for t-test between 3T3-L1 over-expressing human ENPP1 K121K and the vector group. † p<0.05 for t-test between 3T3-L1 over-expressing human ENPP1 K121Q and the vector group.