Inhibitory effect of tanshinone II A on TGF II-β1-induced cardiac fibrosis

Abstract

This study examined the effect of tanshinone II A (TSN II A) on the cardiac fibrosis induced by transforming growth factor β1 (TGF-β1) and the possible mechanisms. Cardiac fibroblasts were isolated from cardiac tissues of neonatal Sprague-Dawley (SD) rats by the trypsin digestion and differential adhesion method. The cells were treated with 5 ng/mL TGF-β1 alone or pretreated with TSN II A at different concentrations (10⁻⁵ mol/L, 10⁻⁴ mol/L). Immunocytochemistry was used for cell identification, RT-PCR for detection of the mRNA expression of connective tissue growth factor (CTGF) and collagen type I (COL I), Western blotting for detection of the protein expression of Smad7 and Smad3, and immunohistochemistry and immunofluorescence staining for detection of the protein expression of phosphorylated Smad3 (p-Smad3), CTGF and COLI. The results showed that TGF-β1 induced the expression of CTGF, COL I, p-Smad3 and Smad7 in a time-dependent manner. The mRNA expression of CTGF and COL I was significantly increased 24 h after TGF-β1 stimulation (P<0.01 for all). The protein expression of p-Smad3 and Smad7 reached a peak 1 h after TGF-β1 stimulation, much higher than the baseline level (P<0.01 for all). Pretreatment with high concentration of TSN A resulted in a decrease in the expression of p-Smad3, CTGF and COL I (P<0.01). The protein expression of Smad7 was substantially upregulated after pretreatment with two concentrations of TSN II A as compared with that at 2 h post TGF-β1 stimulation (P<0.05 for low concentration of TSN I IA; P<0.01 for high concentration of TSN II A). It was concluded that TSN II A may exert an inhibitory effect on cardiac fibrosis by upregulating the expression of Smad7, suppressing the TGF-β1-induced phosphorylation of Smad3 and partially blocking the TGF-β1-Smads signaling pathway.

Keywords

tanshinone II A / transforming growth factor β1 / Smads / connective tissue growth factor

References

[1] BaudinoT.A., CarverW., GilesW., et al.. Cardiac fibroblasts: friend or foe?. Am J Physiol Heart Circ Physiol, 2006, 291(3): H1015-H1026

[2] DrobicV., CunningtonR.H., BedoskyK.M., et al.. Differential and combined effects of cardiotrophin-1 and TGF-β₁ on cardiac myofibroblast proliferation and contraction. Am J Physiol Heart Circ Physiol, 2007, 293(2): H1053-H1064

[3] FrankV., AlianM.. Transforming growth factor-beta and fibrosis. World J Gastroenterol, 2007, 13(22): 3056-3062

[4] KaniaG., BlyszczukP., SteinS., et al.. Heart-infiltrating prominin-1(+)/CD133(+) progenitor cells represent the cellular source of transforming growth factor beta-mediated cardiac fibrosis in experimental autoimmune myocarditis. Circ Res, 2009, 105(5): 462-470

[5] ZhangD.M., QinY., NiuF.L., et al.. Effects of tanshinones II A on proliferation and type I collagen synthesis of rat cardiac fibroblasts. Liaoning Zhongyi Zazhi (Chinese), 2008, 35(12): 1934-1936

[6] RosenkranzS.. TGF-beta1 and angiotensin networking in cardiac remodeling. Cardiovasc Res, 2004, 63(3): 423-432

[7] SunX.Y., CaoL., YuG.H., et al.. Effect of sodium tanshinone II A sulfonate on proliferation and cyclin E expression in human renal interstitial fibroblasts in vitro. Acta Academiae Medicinae Militaris Tertiae (Chinese), 2009, 29(7): 585-587

[8] BujakM., FrangogiannisN.G.. The role of TGF-beta signaling in myocardial infarction and cardiac remodeling. Cardiovasc Res, 2007, 74(2): 184-195

[9] MoustakasA., SouchelnytskyiS., HeldinC.K.. Smad regulation in TGF-β signal transduction. Cell Science, 2001, 114(6): 4359-4369

[10] CunningtonR.H., NazariM., DixonM.C.. C-Ski, Smurf2, and Arkadia as regulators of TGF-β signaling: new targets for managing myofibroblast function and cardiac fibrosis. Canadian J Physiol Pharmacol, 2009, 87(10): 764-772

[11] WangB., HaoJ., JonesS.C., et al.. Decreased Smad7 expression contributes to cardiac fibrosis in the infarcted rat heart. Heart Circ Physiol, 2002, 282(5): H1685-H1696

[12] HeJ.H., ChenY.L., ChenB.L., et al.. B-type natriuretic peptide attenuates cardiac hypertrophy via the transforming growth factor-beta1/smad7 pathway in vivo and in vitro. Clin Exp Pharmacol Physiol, 2010, 37(3): 283-289