Notch3 signalling pathway in vascular smooth muscle cell growth and survival

Abstract

ABSTRACTUniversity of ManchesterAdem SadliFor the degree of Doctor of PhilosophySubmission date: 30/04/2013 Notch3 signalling in vascular smooth muscle cell growth and survival VSMCs are the major components of the arterial wall. Under physiological conditions VSMCs exhibit a contractile phenotype, although possessing the capacity of phenotypic transition. The plasticity of VSMCs enables them to gain proliferative features which contribute to the pathologies of common vascular diseases in response to diverse stimuli. Hence, the stability of VSMCs in cell growth and survival is essential for maintaining proper arterial function. Diverse signalling pathways and growth factors have been identified as being important in the regulation of VSMC behaviour, and the Notch signalling is a relatively new participant. Proteins of Notch family are transmembrane receptors that transduce signals from neighbouring cells. Among the four Notch receptor subtypes, Notch3 is mainly expressed in VSMCs in small arteries. NOTCH3 gene mutation causes a human genetic stroke syndrome, CADASIL which is featured by systemic VSMC degeneration, suggesting the importance of Notch3 signalling in VSMC growth and survival. Although there is some knowledge for the Notch3 signalling in the regulation of VSMC proliferation and apoptosis, the molecular mechanisms underlying such regulation are largely unknown. By overexpressing the constitutive active form of human Notch3 (N3IC), it was confirmed that Notch3 promotes VSMC proliferation and protects VSMCs against apoptosis in primary rat aortic SMCs. It was also shown that Akt mediated both Notch3-induced VSMC proliferation and protection of serum-deprivation-induced apoptosis; while Erk 1/2 only contributed to Notch3-induced VSMC proliferation. In addition, the activation of PI3K/Akt was significantly blocked by PDGFR inhibitor, even in serum-free conditions, suggesting an intrinsic effect of Notch3 on PDGFR function without the requirement of the exogenous ligand, PDGF. It was also shown that Notch3 inhibits GSK-3alpha/ by phosphorylation and this effect was not blocked by the PI3K/Akt inhibitor, suggesting an alternative Akt-independent mechanism exists by which Notch3 regulates GSK-3 activity. Furthermore, using apoptosis pathway PCR arrays a number of apoptotic genes including BCL2 that were regulated by Notch3 activation in human VSMCs were identified. Notch3 also up-regulates the expression of its ligand Jagged1 in a PI3K/Akt dependent manner. Interestingly, Notch2 was significantly down-regulated and Notch1 was mildly upregulated by Notch3. In addition, an age-dependent increase in the expression of Notch3 in SMCs of the aortic root from 8 to 16 weeks old mice was identified, however, this declined in aged mice (24 weeks). These findings suggest a model for Notch3 function which is integrally involved in multiple signalling pathways in the regulation of VSMC growth and survival. It is hoped that these findings will contribute to the current knowledge of vascular biology and provide the basis for future studies on the development of potential therapeutic targets for cardiovascular diseases.

Date of Award	31 Dec 2013
Original language	English
Awarding Institution	The University of Manchester
Supervisor	Tao Wang (Supervisor) & Cathy Holt (Supervisor)