Abstract 17568: Phospholipase C Gamma-1 Mediated Akt-notch1 Signaling is Essential for Intima Formation

Abstract

Introduction: Vascular smooth muscle cell (VSMC) proliferation, migration and dedifferentiation are critical for vascular diseases, such as atherosclerosis and restenosis after angioplasty. Recently, it is demonstrated that after vascular injury Notch 1and Notch 3 knockout (KO) mice have decreased intima formation whereas Notch 2 KO mice have increased one. Therefore, it is important to investigate the specific regulatory pathways that activate the different Notch receptors. Methods and Results: There was a time and dose dependent activation of Notch1, as determined by western blot of N1-ICD but not Notch 2 and 3 in VSMC. As Akt activates Notch signaling in endothelial cells and phospholipase C gamma-1 (PLCγ1) binds to and activates Akt in tumor cells, we therefore propose that PLCγ1 mediated Akt -Notch1 pathway is critical for intima formation by increasing VSMC proliferation, migration and dedifferentiation. To test our hypothesis, we measured PLCγ1 and Akt interaction in VSMC. We observed an association between PLCγ1 and Akt, which was increased upon AngII or PDGF stimulation. Depletion of PLCγ1 by siRNA significantly inhibited Akt and Notch1 activation. As the major target gene of Notch1, Hey2 protein expression was increased 1.8 fold and 2.3 fold respectively after AngII and PDGF stimulation. PLCγ1 siRNA treatment inhibited it by 68.3% and 64.7%. Depletion of PLCγ1 significantly inhibited PDGF induced VSMC proliferation, migration and dedifferentiation but increased apoptosis. In vivo, PLCγ1 and control siRNA were delivered periadventitially in pluronic gel after carotid ligation. Morphometric analysis 21days after ligation demonstrated that PLCγ1 depletion attenuated intima area (2.2 ± 0.7 vs 4.3 ± 1.0, *104 μm2) and intima/media ratio (0.21 ± 0.09 vs 0.43 ± 0.12) compared to the control group. Conclusions: In conclusion, PLCγ1 mediated Akt-Notch1 signaling is crucial for intima formation. The PLCγ1 and Akt interaction will be a promising target for therapeutic stratergies that inhibit vascular remodeling.