Abstract 5778: Protein Phosphatase 1 Signalling Pathway is Involved in the Control of Vascular Smooth Muscle Cells Proliferation

Abstract

Protein phosphatase 1 (PP1), the main regulator of the transcription factor CREB, is strongly involved in the control of cell proliferation via p53 and p21. PP1 is regulated by phosphatase inhibitor-1 (I-1), which is activated by PKA phosphorylation and inhibited by protein phosphatase PP2A and PP2B (calcineurin). I-1 is expressed in vascular smooth muscle cells (VSMC), but does not appear to play a significant role in contractile or relaxant response We tested whether the I-1/PP1 pathway is involved in the control of VSMC proliferation Confocal immunofluorescence, Western blot analysis and real time PCR demonstrated that I-1 protein was expressed in the media layer of healthy human coronary and mammary arteries and down regulated in atherosclerotic vessels and proliferating human VSMC (1000 fold lower of mRNA). By contrast, PP1 expression was increased in proliferating human VSMC. Inhibition of I-1/PP1 signalling by siRNA PP1 or by adenoviral gene transfer of a constitutively active I-1 (I-1c) inhibited VSMC proliferation and migration in vitro. Promoter-reporter assay showed that in vitro I-1c gene transfer increased activity of CREB. Western blot analysis demonstrated that I-1c and siRNA PP1 increased phosphorylation of CREB, expression of p21, p53 and led to down regulation of cell cycle controlling proteins cyclin D1 and pRB. To assess the potential effect of I-1c gene transfer in the treatment of restenosis, we used a rat carotid artery injury model. I-1 was expressed in normal carotid artery but it was non-detectable two weeks after injury, which correlated with vascular remodelling during neointimal proliferation. Adenoviral gene transfer of I-1c prevented neointimal proliferation in the carotid injury model. In conclusion, the I-1/PP1 signalling pathway is involved in the control of VSMC proliferation and migration via the transcription factor CREB.