Relaxin-Expressing Adenovirus Decreases Collagen Synthesis and Up-Regulates Matrix Metalloproteinase Expression in Keloid Fibroblasts

Abstract

The hormone relaxin has been shown to affect the extracellular matrix by inhibiting collagen synthesis and expression in fibroblasts stimulated with a profibrotic agent. It also increases matrix metalloproteinase (MMP) expression. To investigate its effect on expression of collagen and MMPs in keloid fibroblasts and human dermal fibroblasts, the authors introduced a relaxin-expressing adenovirus (dE1-RGD/lacZ/RLX) into a human dermal fibroblast cell line and keloid fibroblasts. Both fibroblasts were infected with dE1-RGD/lacZ/RLX or control virus, and protein levels of relaxin and secreted transforming growth factor (TGF)-β1 were assessed by enzyme-linked immunosorbent assay, and mRNA levels of collagen type I, collagen type III, MMP-1, and MMP-3 were assessed by real-time reverse-transcriptase polymerase chain reaction and enzyme-linked immunosorbent assay. Expression of Smad3 and phosphorylated Smad3 was also examined, and relaxin's effect on Smad2/3 complex localization was evaluated. When human dermal fibroblasts and keloid fibroblasts were transduced with dE1-RGD/lacZ/RLX or dE1-RGD/lacZ (control), mRNA expression of type I and type III collagen was markedly decreased by relaxin regardless of TGF-β (10 ng/ml) treatment. Expression of Smad3 and phosphorylated Smad3 was reduced in keloid fibroblasts and decreased translocation of Smad 2/3 complex from cytosols to the nucleus of the human dermal fibroblasts with TGF-β after dE1-RGD/lacZ/RLX transduction, suggesting that relaxin reduces collagen synthesis by blocking TGF-β signaling. Analyses revealed that MMP-1 and MMP-3 expression were significantly increased in human dermal fibroblasts and keloid fibroblasts after dE1-RGD/lacZ/RLX transduction. These results suggest that the antifibrotic effect of relaxin-expressing adenovirus may have therapeutic effects on keloids.