Cryptotanshinone Attenuated Pathological Cardiac Remodeling In Vivo and In Vitro Experiments

Figure 6

CTS treatment inhibited TGF-β-induced proliferation and myofibroblast activation. (a) CCK-8 determined the proliferation of isolated neonatal CFs and CFs was treated with PBS, TGF-β (10 μM), or CTS (12 μM) for 48 hours as indicated in the pictures; (b) immunofluorescence staining indicated that TGF-β treatment (10 μM, 48 hours) induced α-SMA expression; CTS treatment (12 μM) significantly inhibited TGF-β induced α-SMA expression; (c) assessing the α-SMA fluorescence intensity; (d) calculating the cell surface area (CSA), about 100 NRCMs were used for calculating CSA; relative quantification expression of mRNA of (e) CCN2, (f) COL1A1, and (g) COL3A1; CFs were incubated with CTS or TGF-β for 24 hours before collecting for RNA extraction; the mRNA expression of these target genes was normalized to GAPDH before relative quantification; (h) representative Western blots of p-STAT3, T-STAT3, α-SMA, p-SMAD3, T-SMAD3, p-SMAD2, T-SMAD2, p-SMAD1/5, T-SMAD1/5, and GAPDH; CFs were transfected with adenovirus for GFP or STAT3 overexpression during 24 h and then were incubated with CTS or TGF-β as indicated in picture for another 12 hours before collecting for total protein extraction; (i) relative quantification of p-STAT3/T-STAT3; (j) relative quantification of TGF-β, p-SMAD3/T-SMAD3, p-SMAD2/T-SMAD2, and p-SMAD1/5/T-SMAD1/5; all of these proteins were normalized to GAPDH before the relative quantification; cell experiments were repeated three times independently. Compared with the PBS-treated group (); ^#compared with the TGF-β-treated group ().

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