Effects Of Baicalin On CTGF And α-SMA Expressions Of Pulmonary Fibroblast And Myofibroblast In Rat

Objective: Pulmonary fibrosis is characterized by abnormally increasedmyofibroblast and excessive deposition of extracellular matrix (ECM) in lung.The α-smooth muscle actin (α-SMA) is the marker of the myofibroblast.Transforming growth factor β1(TGF-β1) is one of key cytokines, which caninduce fibroblast differentiation into myofibroblast. Connective tissue growthfactor (CTGF）is the downstream factor of TGF-β. Baicalin is a majorflavonoid extracted from the traditional Chinese medicinal herb Scutellariabaicalensis Georgi. It has been reported that baicalin has anti-fibrogenic effect.However, the effects of this drug on expressions of CTGF and α-SMA in lungfibroblast and myofibroblast in vitro have not been reported. Therefore, thepurpose of this experiment is to observe the effect of baicalin on theexpression of CTGF and α-SMA protein in lung fibroblast and myofibroblastin vitro.Methods: Lung fibroblasts from male SD rat (150-170g) were culturedin vitro. Lung myofibroblasts are obtained by culture of fibroblasts with TGF–β for48h. The3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide (MTT) was used to estimate the effects of baicalin at differentconcentrations on the survival ratio of the lung fibroblasts and themyofibroblast. Western Blot was used to determine the expressions of CTGFand α-SMA protein in lung fibroblasts and myofibroblasts.Results:1The effects of baicalin on the survival ratio of lung fibroblasts andmyofibroblastsLung fibroblasts referred to the ones cultured in normal state.Myofibroblast referred to the ones stimulated by TGF-β for48h and had highexpression of α-SMA. The control cells referred to the ones had no drug interference. Compared with the respective control cells (0μmol/L),10μmol/L,20μmol/L,40μmol/L baicalin had no effects on survival ratio of lungfibroblasts and myofibroblast (P＞0.05）, while80μmol/L,160μmol/L baicalininhibited the survival ratio of lung fibroblasts and myofibroblast (P <0.01).The above results suggested the concentrations of10-40μmol/L baicalin haveno obvious toxic effect on lung fibroblasts and myofibroblast.2The effect of baicalin on the protein expressions of CTGF and α-SMAin lung fibroblasts.Compared with control group (0μmol/L), baicalin (10,20,40, μmol/L)was of no statistical significance on the protein expressions of CTGF andα-SMA in lung fibloblasts（P＞0.05）, but the baicalin(80,160μmol/L)inhibited the protein expressions of CTGF and α-SMA. The above resultsindicate that the10-40μmol/L baicalin neither decreases the expression ofCTGF in lung fibloblasts nor induces differentiation of fibroblast intomyofibroblast, while80-160μmol/L baicalin inhibits CTGF expression andmyofibroblast differentiation (P <0.01).3The effects of baicalin on the protein expressions of CTGF and α-SMAin myofibroblastCompare with lung fibroblasts, TGF-β increased the protein expressionof CTGF and α-SMA (P <0.01), suggesting myofibroblast is succesfullyinduced in vitro. Compared with the control myofibroblast, the proteinexpressions of CTGF and α-SMA in myofibroblasts were decreased by10μmol/L,20μmol/L,40μmol/L,80μmol/L,160μmol/L Baicalin, respectively (P＜0.01), suggesting baicalin inhibits the protein expression of CTGF andpromotes the dedifferentiation of myofibroblast into fibroblast.Conclusion:1The10-40μmol/L concentrations of baicalin have no effects on thesurvival ratio of lung fibroblasts and myofibroblast (P＞0.05）, but80-160μmol/L baicalin show toxic effect.2The10-40μmol/L concentrations of baicalin neither increase theprotein expression of CTGF in lung fibroblasts nor promote the differentiation of fibroblast into myofibroblast, while80-160umol/L baicalindown-regulate the protein expression of CTGF and α-SMA in lung fibroblasts.3The baicalin (10-160μmol/L) dose-dependently inhibits the proteinexpression of CTGF and promote the dedifferentiation of myofibroblast intofibroblast.