Participation of the SMAD2/3 signalling pathway in the down regulation of megalin/LRP2 by transforming growth factor beta (TGF-ß1)
Fig 10
Albumin decreases megalin expression and this effect is counteracted by TGF-βRI inhibition and SMAD2/3 silencing.
LLC-PK1 cells were treated with albumin concentrations ranging from 0.01 to 20 mg/ml for 24 h. Megalin (A) and PAI-1 mRNA levels (B) were measured by qPCR. (C) Megalin and (D) PAI-1 mRNA levels in LLC-PK1 cells that were exposed to 20 mg/ml of albumin and/or the inhibitor SB for 20 h. The results are expressed as the average RQ ± standard deviation (SD). (E) LLC-PK1 cells were transfected with the megalin promoter construct (1500PromBasic) and a ß-galactosidase vector (pCMVß) and incubated in the presence or absence of 20 mg/ml of albumin for 24h. Later, the cells were lysed, and luciferase activity was measured. (F) LLC-PK1 cells were transfected with the 1500PromBasic plasmid and SMAD2/SMAD3- siRNAs, after 24h the cells were treated with 20 mg/ml of albumin for another 24h. Luciferase activity was determined using a Luciferase Reporter Assay System, and the ß-galactosidase signal was used to normalize the results. Data are expressed as the means ± SD. Significant differences with controls are indicated by: *P ≤ 0.05; **P ≤ 0.01; ***P ≤ 0.001. Significant differences between treatments are indicated by # P≤0.05, ## P ≤ 0.01.