| Background and ObjectiveOf the various growth factors, transforming growth factor-13 appears to play a key role intissue fibrosis. Three isoforms including TGFβ1,β2 andβ3 are found in mammalian cells andare highly conservative during different genera. TGFβis secreted by most cells involved inwound healing including macrophages and fibroblasts, etc. Three isoforms play different roles inwound healing. Both TGFβ1 and TGFβ2 can induce scar formation, while TGFβ3 may have ananti-scarfing effect. To obtain the transferred fibroblasts with pcDNA3.1(-)/TGFβ3, we transferthe recombinant vector-pcDNA3.1(-)/TGFβ3 into Wistar rat skin fibroblasts. In addition, wedetect the level of intracellular TGFβ1,β3 before and after transfection with FCM. To do someelementary rearch on biological behaviour(release of TGFβ,etc.)of the transferred fibroblasts andprovide experiment foundation for clinical application.Methods1 To culture dermal fibroblasts derived from Wistar rat skin with tissue masses andmonolayer cultures.2 The stable transfection of primary culture fibroblasts with recombinant plasmid expressingTGF3 was established by using LipofectamineTM2000 and G418 selection. To obtain monoclonalcell, and amplify the transferred fibroblasts.3 To detect the level ofintracellular TGFβ1,β3 with FCM.Results1 It is successful of obtaining stable expression cell clone.2 The FCM results show that the total level of intracellar TGFβis 60.86 and TGFβ1/TGFβ3is 1.38 in normal fibroblasts, while one ofintracellar TGFβis 225.23 and TGFβ1/TGFβ3 is 0.09in transferred cells. Above-mentioned results show that the level of intracellar TGFβ3 intransferred cells obviously higher than one of normal cells, while TGFβ1 in the oppsite direction. ConclusionThe transgenic fibroblasts can express TGFβ3 proteinum by using LipofectamineTM2000 andG418 selection, inhibiting the expression of TGFβ1 proteinum, which provides theory for thefeasibility of applying the transferred fibroblasts on wound.
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