Figure 5.

Effect of AdCLU on the levels and distribution ECM and pro‐fibrotic proteins. Protein expression analysis by western blot showed significant decrease in ‐ (a) ECM proteins‐COL1A, FN, ELN, and pro‐fibrotic TGFβ2 and TSP‐1 in WCL, (b) secretory FN and pro‐fibrotic PAI‐1 and TSP‐1 in CM showed ‐ in AdCLU compared with AdMT. GAPDH was used as a loading control. Values represent the mean ± SEM, where n = 4 (biological replicates). *p ≤ 0.05 was considered significant. (d) Immunofluorescence imaging shows increase in clusterin distribution in AdCLU treatment (Alexa Flour 568/red) (first column in bottom panel) and a concomitant reduction in actin stress fiber distribution stained by phalloidin (green) (second column in bottom panel) compared to AdMT (top panel). AdCLU treatment reduced COL1A (Alexa Flour 568/red) (second column bottom panel), and FN (Alexa Flour 488/green) (third column bottom panel) distribution compared to AdMT. The nucleus was stained with DAPI in blue. Images were captured in z‐stack in a confocal microscope, and stacks were orthogonally projected. Scale bar 50 microns. AdCLU, adenovirus‐mediated constitutive expression of clusterin; ECM, extracellular matrix; DAPI, 4′,6‐diamidino‐2‐phenylindole; SEM, standard error of mean; WCL, whole‐cell lysate