TGFβ2 may phosphorylate SMAD2, SMAD3, ERK1/2, JNK and p38 in human umbilical vein endothelial cells

Sammanfattning: Introduction The dysfunction of human endothelial cells caused by exposure to inflammatory mediators, such as TGFβ, leads to development of diseases, such as atherosclerotic diseases. TGFβ mediated endothelial dysfunction may involve MMP-9. MMP9 is highly expressed in atherosclerotic plaques. The aim of this study was to assess, in a comprehensive manner, the signaling pathways induced by TGFβ2 in vein endothelial cells. In addition, this study also aimed to determine whether TGFβ2 induces MMP-9 expression in human umbilical vein endothelial cells. Method Western blot, Zymography, sandwich ELISA and qPCR were performed on the cells, lysate and medium of cultured human umbilical vein endothelial cells, which were treated with TGFβ2 at different doses and were incubated at different time-points. The induction of MMP9 by TGFβ2 on protein-level and gene expression-level of human umbilical vein endothelial cells were studied using Zymography and qPCR respectively. Results TGFβ2 may phosphorylate SMAD2 in a dose-dependent manner, which seemed to peak at 10 ng/ml. A time-course experiment indicated that TGFβ2 may phosphorylate SMAD2 as early as 10-15 min after stimulation and the phosphorylation seemed to peak at 30min. TGFβ2 may phosphorylate SMAD3 as early as 10-15 min after stimulation and the phosphorylation seemed to peak at 30min. TGFβ2 may phosphorylate ERK1/2 at 2 hours and 4 hours only. TGFβ2 may not phosphorylate AKT. TGFβ2 may phosphorylate p38, which seemed to peak at 1 hour. TGFβ2 may phosphorylate JNK as early as 15-30 min after stimulation and the phosphorylation seemed to peak at 1 hour. MMP9 was not induced by TGFβ2 both on the protein-level and gene expression-level. Conclusion In conclusion, TGFβ2 may phosphorylate SMAD2, SMAD3, ERK1/2, JNK and p38 in human umbilical vein endothelial cells. MMP9 was not induced by TGFβ2 both on the protein-level and gene expression-level.