Experimental Study Of RNAi Silencing TIEG On The Functions Of Keloid Fibroblasts

PART ONE The differences of normal human fibroblasts and keloid fibroblasts in the TGF-β/Smads signaling pathwayObjective:To discuss the different expression of normal human fibroblasts and keloid fibroblasts cultivated in vitro in the TGF-β/Smads signaling pathway.Methods:To Culture normal human fibroblasts and keloid fibroblasts by using tissue culture method, expression condition of TIEG,Smad2,Smad7,α-SMA and collagen I were detected by the way of reverse transcription polymerase chain reaction(RT-PCR).Results:(1)The expression of TIEG mRNA was lower in normal human, while the expression of TIEG mRNA was higher in keloid fibroblasts. (2)Compared with normal human fibroblasts and keloid fibroblasts, the expression of Smad2 mRNA was increased and Smad7 mRNA was reduced in keloid fibroblasts. (3) The expression of a-SMA mRNA and Collagen I mRNA were enhanced.Conclusions:TIEG can be detected increasing in keloid fibroblasts, it can activate TGF-β/Smads signaling pathway by downward smad7 expression and raising expression of smad2, TIEG can promote fibroblast transdifferentiation and ECM synthetic. PART TWO Influence of pSUPER.gfp/TIEG on TGF-β/Smads signal gene expression of keloid fibroblast.Objective:To discuss the influence of pSUPER.gfp/TIEG on TGF-β/ Smads signal gene expression of keloid fibroblasts.Methods:pSUPER.gfp/TIEG was adoped to transfect the primary cultured human keloid fibroblasts(KFB)by the interfere technology of micromolecule RNA and carrier of liposome. Expression condition of TIEG,Smad2,Smad7,α-SMA and collagen I in KFB were detected by the way of RT-PCR on condition that endogenous TGF-β/Smads signal transducfion has been impeded.Results:pSUPER.gfp/TIEG could effectively interfere with expression of TIEG in KFB. The expression of Smad2 mRNA was weaken and Smad7 mRNA was enhanced, at the same time, the expression ofα-SMA and Collagen I mRNA were weaken.Conclusions:Fibroblast transdifferentiation and ECM synthetic were markedly decreased after TIEG mRNA expression level was inhibited and the TGF-β/Smads signaling pathway was regulated by RNAi. This indicated that TIEG could be acandidated target for gene therapy.