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. 2018 Jan 30;8:1906. doi: 10.1038/s41598-018-20316-1

Figure 5.

Figure 5

Effect of clusterin deficiency on TGF-β1-induced myofibroblast differentiation and collagen deposition. Lung fibroblasts were transduced with clusterin shRNA (shCLU, open bars) and mock shRNA vectors (grey bars) or remained untransfected (black bars). αSMA mRNA was assessed via qRT-PCR (A) and clusterin and αSMA protein levels detected by western blotting (B,C quantification). 48 h following TGF-β1 stimulation (40 pM) αSMA mRNA and protein were increased. Basal and increased levels of αSMA mRNA and protein, however, did not vary between clusterin deficient, mock-transduced and control fibroblasts. Collagen mRNA (D) and deposition (E,F quantification) assessed by qRT-PCR and immunofluorescence staining were significantly increased in response to TGF-β1. Although, basal and TGF-β1 induced changes in collagen levels varied between clusterin deficient, mock and control fibroblasts, the overall fold-increase in collagen mRNA and deposition levels did not significantly change between clusterin deficient fibroblasts and control/mock fibroblasts. Data generated in A-F is representative of two individual experiments with fibroblasts derived from 2 donors. Full-length western blots are presented in Supplementary Figure e5. Scale bar in E represents 10 µm. **P < 0.01, ***P < 0.001 compared with untreated controls respectively.