Objective To observe the effects of hydrogen peroxide on the expression of transforming growth factorβ1(TGF-β1) and Smad3 protein in A549 cells. Methods A549 cells were cultured with different concentrations of hydrogen peroxide. MTT assay was used to determine the cell growth and survival rates. The level of TGF-β1 and p-Smad3 protein were detected by western blotting. Results It was observed that hydrogen peroxide significantly inhibit proliferation of A549 cells. When the concentration of hydrogen peroxide was 1.0 mmol/L, the inhibition ratio reaches 46.34%, and the level of TGF-β1 and p-Smad3 protein were increased in a time-dependence manner and reached a peak after 24 h, then decreased a little but also remained at high level. Conclusions In the early oxidative damage, A549 cells express high level of TGF-β1 and p-Smad3 protein. It may be relevant to tissue repair and remodeling after lung injury.
【Abstract】 Objective To summarize the recent progress in related research on transforming growth factor β1 （TGF-β1）/Smad3 signal transduction pathway and post-traumatic scar formation. Methods Recent related literature at home and abroad on TGF-β1/Smad3 signal transduction pathway and post-traumatic scar formation was reviewed and summarized. Results TGF-β1 is an important influence factor of fibrotic diseases, and it plays biological effects by TGF-β1/Smad3 signal transduction pathway. The pathway is regulated by many factors and has crosstalk with other signal pathways at cellular and molecular levels. The pathway is involved in the early post-traumatic inflammatory response, wound healing, and late pathological scar formation. Intervening the transduction pathway at the molecular level can influence the process of fibrosis and extracellular matrix deposition. Conclusion TGF-β1/Smad3 signal transduction pathway is an important way to affect post-traumatic scar formation and extracellular matrix deposition. The further study on the pathway will provide a theoretical basis for promotion of wound healing, as well as prevention and treatment of pathological scar formation.
The transforming growth factor-β1 (TGF-β1)/Smad3 signal pathway is related to mutiple physiological and pathological generation mechanism of human being. Up to date, however, the spacial and time information on the phosphorylated Smad3 is still unclear. In this study, the process of Smad3 phosphorylation was observed under the physiological state in the living cells. Firstly, the ECFP-Smad3-Citrine (Smad3 biosensor) fusion protein expression vector was constructed and identified. Then the Smad3 biosensor was transfected into 293T cells. The transfection efficiency and the expressions of fusion proteins were observed in 24 hours. Thirdly, Smad3 biosensor flurorescence resonance energy transfer (FRET) was observed with the inversion fluorescence microscope and measured by the MetaFlour FRET 4.6 software. Smad3 biosensor transfection efficiency was nearly 40% and the fusion protein was seen under the fluorescence microscope. The FRET ratio of Smad3 biosensor in living 293T cells was decreased after 10 minutes incubation with the ligand of TGF-β1. The period of decreasing CFP and enhancing Citrine signals was about 300 seconds. With the technology of FRET, the TGF-β1/Smad3 signal pathway could be real time monitored dynamically under the physiological condition in living cells.
ObjectiveTo explore the effect of naringenin on the expression of vascular endothelial growth factor (VEGF) released by human fetal lung fibroblasts. MethodsHuman fetal lung fibroblast cells were divided into a control group,a cigarette smoke extract (CSE) group and a naringenin group. Cells in the naringenin group were incubated with different doses of naringenin for 2h. Then the naringenin group and the CSE group were incubated with CSE to adjust the final concentration of naringenin (5 μmol/L,10 μmol/L,and 20 μmol/L) and of CSE(5%). The concentration of VEGF was measured in human fetal lung fibroblasts after cultured for 24h,48h and 72h by enzyme-linked immunoabsorbent assay. Smad3 and p-Smad3 levels were detected by Western blot. ResultsELISA results showed that the CSE can significantly increase the VEGF expression,and naringenin can inhibit the increasing of the VEGF expression. Western blot results showed that the CSE can increase the p-Smad3 expression,and the naringenin can inhibit the increasing of the p-Smad3 expression. ConclusionNaringenin can inhibit phosphorylation of Smad3,and decrease the expression of VEGF released by human fetal lung fibroblasts.