Abstract 5474: Crucial Role of Nrf3 in Smooth Muscle Cell Differentiation From Stem Cells and Vessel Injury-Induced Neointima Formation
Nuclear factor erythroid 2-related factor 3 (Nrf3), a member of the cap’n’collar family of transcription factors that binds to the DNA-antioxidant responsive elements, have been reported to be involved in muscle precursor migration during early embryo development. The present study aimed to investigate the functional role of Nrf3 in stem cell differentiation into smooth muscle cells (SMCs). When embryonic stem cells were cultivated on collagen IV-coated plates, Nrf3 was up-regulated significantly following 1–6 days of cell differentiation. Knockdown of Nrf3 resulted in down-regulation of smooth muscle specific markers including smooth muscle actin, SM22, calponin and smooth muscle myosin heavy chain, while enforced expression of Nrf3 enhanced SMC differentiation in a dose-dependent manner. SMC-specific transcription factor myocardin, but not serum response factor, was significant upregulated by Nrf3 overexpression. Strikingly, the binding of serum response factor and myocardin to the promoter of smooth muscle differentiation genes was dramatic increased by Nrf3 overexpres-sion, and Nrf3 can direct bind to the smooth muscle gene promoters as demonstrated by chromatin immunoprecipitation (CHIP) assay. Moreover, NADPH-derived ROS production during SMC differentiation was further enhanced by Nrf3 overexpression through upregualtion of NADPH oxidase (NOX4) and inhibition of anti-oxidant signalling pathway. Furthermore, overexpression of Nrf3 resulted in increase of plasma phospholipase A2 (Pla2g7) production, while knockdown Nrf3 ablated the Pla2g7 gene expression and protein activity. Additionally, both Nrf3 and Pla2g7 were up-regulated in adult SMC phenotype swiching from proliferating to contractile phenotype upon serum starvation. Importantly, in vivo knockdown of Nrf3 in the vessel wall significant increased wire-injury-induced neointima formation in ApoE-deficient mice. Our findings demonstrated for the first time that Nrf3 has a crucial role in SMC differentiation from stem cells and important protective function in vessel injury-induced neointima formation, indicating that they could be potential new therapeutic targets for intervention of SMC proliferative-related vascular diseases.