Research Of Smad4 Mutant Gene Inhibiting Abnormal Scar Formation

Objective:To exploer the effects of mutant Smad4 (Smad4ΔM4) on inhibition of fibroblasts proliferation and expressions of collagenⅠ.Ⅲ. extracellular matrix.the anti-scar role in vivo while using animal model,to study the role of Smad4ΔM4 in scar formation and offer theory on which new clinical theatment of abnormal scar bases.Methods:To Construct recombinant adenovirus comaining Smad4ΔM4 (wild-type Smad4 removing the sequence 275-322 amino acid. Ad-Smad4ΔM4. abbreviated as Ad-AM4), Mouse fibroblast (NIH3T3) were transfected whith different concentrations of Ad-ΔM4(MOI=1,10,100), then the cell were stimulated with TGF-β1. The NIH3T3 proliferation were determined by MTT assay. And expression of collagenⅠ,ⅢmRNA were analyzed by RT-PCR method. Expression ofⅠ,Ⅲcollagen protein were detected with Western-Blot.At the same time scar model mice were set up.then treated the wounds with Ad-ΔM4 (1010pfu,109pfu). Histopathological methods. mage-ProPlus6.0 software were used to analysed the impact of Ad-AM4 on scar formation in mice.Results:The experiment in vitro demonstrate:Ad-ΔM4 can effectively inhibit NIH3T3 proliferation, down-regulated expression levels of collagenⅠ,ⅢmRNA and protein. This effect was positively correlated with Ad-AM4(MOI=1,10,100) (P<0.05). The experiment in vivo showed that::Ad-ΔM4 was effective in reducing scar formation. Relative scar areas of the experimental group were 36%(1010pfu group) and 47%(109 pfu group)compared with the blank control group.Conclusions:Ad-AM4 can inhibit proliferation of fibroblast, down-regulate over-expression of collagen and extracellular matrix, effectively inhibit scar formation. which may be one of the mechanisms why mutant Smad4 resists TGF-β1-mediated fibrosis.