Fetal and adult fibroblasts have similar TGF-β-mediated, Smad-dependent signaling pathways

Background: The scarless fetal skin-healing mechanism is mediated in part by the fibroblast and involves differential expression of transforming growth factor (TGF)-beta isoforms 1 and 3. The authors hypothesized that fetal and adult fibroblasts respond differently to TGF-beta isoform-specific stimulation, which may influence whether wounds scar. Connective tissue growth factor (CTGF), Smad3, and Smad7 are TGF-beta target genes. Expression of these targets was quantitated after TGF-beta 1 and -beta 3 stimulation of fetal and adult fibroblasts. Methods: Primary mouse fibroblast cultures at gestational day 16.5 (E17), 18.5 (E19), and 6 weeks (adult) were stimulated with TGF-beta 1 or TGF-beta 3. Quantitative polymerase chain reaction was performed for CTGF, Smad3, and Smad7 expression. Results: CTGF was reduced four-fold in E17 and E19 compared with adult fibroblasts (p < 0.005). After TGF-beta 1 stimulation, CTGF expression increased more than 60-fold in both E17 and E19 (p < 0.01), which was three-fold greater than that in adult fibroblasts (p < 0.005). TGF-beta 3 induced more than 70-fold, 50-fold, and 20-fold increases in CTGF expression in E17, E19, and adult fibroblasts, respectively (p < 0.01 for each). Both TGF-beta 1 and -beta 3 decreased Smad3 expression and increased Smad7 expression in each fibroblast type, suggesting that intact TGF-beta-mediated signaling pathways were present. Conclusions: Fetal (E17 and E19) fibroblasts have lower CTGF expression compared with adult fibroblasts. However, fetal fibroblasts have larger increases in CTGF expression after TGF-beta 1 or -beta 3 stimulation. Fetal and adult mouse fibroblasts have similar TGF-beta 1 and TGF-beta 3 transcriptional regulation of Smad3 and Smad7. This suggests that scarless healing is likely not mediated by different Smad-dependent transcriptional responses to TGF-beta isoforms in the fetal E17 fibroblast.