FIGURE 6.

Down-regulation of Cnpy2/Msap inhibits the effect of FGF21 on LDLR and lipid uptake. Raw cells were transfected with 4 μg shRNA against Cnpy2/Msap for 24 h and half of the cells were further stimulated for 20 h with 50 ng/ml FGF21 followed by immunoblotting. Values are means ± S.D., n = 3. *, p < 0.05 and **, p < 0.01 for treated versus control. A, Cnpy2/Msap. Left, immunoblots. Right, quantification. Note down-regulation of Cnpy2/Msap by shRNA. The increase in Cnpy2/Msap by FGF21 was blocked by the shRNA. B, LDLR. Left, immunoblots; Right, quantification. Down-regulation of Cnpy2/Msap by shRNA decreased LDLR levels and reduced the increase in LDLR by FGF21. C, control experiment. Cells were transfected with 4 μg of scrambled shRNA (sc), and immunoblotting was done as above. FGF21 increased LDLR levels in sc-treated cells. D, Mylip/Idol down-regulation. Cells were transfected with 4 μg of shRNA against Mylip/Idol. Note a decrease in Mylip/Idol and an increase in LDLR by the shRNA. Treatment with 50 ng/ml FGF21 for 24 h further reduced Mylip/Idol levels. E, lipoprotein uptake. Cnpy2/Msap was down-regulated using shRNA for 24 h, and cells were incubated with 50 ng/ml FGF21 for an additional 20 h. 5 μg/ml DiI-labeled LDL was then added for 6 h, and the uptake was determined as described under “Experimental Procedures.” Values are means ± S.D., n = 3. **, p < 0.01 and ***, p < 0.001 and for treated versus C.