Abstract 16194: Nucleostemin Induced by Pim-1 Kinase is Critical to Inhibit Senescence, Maintain Multipotency and Enhance Regenerative Potential of Cardiac Progenitor Cells
Myocardial regeneration and repair after injury are governed in part by cell survival, proliferation and pluripotency. Proliferation and survival of c-kit+ cardiac progenitor cells (CPCs) are mediated by Pim-1, a serine threonine kinase, and nucleostemin (NS), a nucleolar stress sensor protein that is associated with stem cell pluripotency and regeneration. The role of NS in regulating CPC multipotency and mechanisms of NS induction are unknown, prompting our hypothesis that NS induced by Pim-1 mediated stabilization of transcription factor c-Myc maintains CPC multipotency and inhibits senescence. NS and c-Myc are regulated by Pim-1 as indicated by increased expression of NS and c-Myc in cultured CPCs overexpressing Pim-1 (3.1 and 5.5 fold, p<0.01). Knockdown of Pim-1 using sh-RNA decreases c-Myc and NS expression (-60%, -54%, p<0.05), consistent with pharmacological inhibition of Pim-1. c-Myc is necessary and sufficient for NS regulation, as indicated by increase (3.1 fold, p<0.01) or decrease (-60.2%, p<0.01) in NS expression following either overexpression or knockdown of c-Myc, respectively. Regulation of NS promoter by c-Myc is evident following knock down of c-Myc in CPCs isolated from transgenic mice created with a NS-GFP reporter construct. NS regulates CPC phenotypic characteristics , as silencing of NS induces flattening of cells, decreases expression of stem cell marker c-Kit (-55%, p<0.05), up-regulates cell cycle inhibitors p53 and p16 (4.2, 3.8 fold, p<0.01), decreases expression of “Yamanaka pluripotency factor” proteins KLF4, c-Myc and Sox2 (p<0.05) and decreases proliferation (p<0.05) compared to control CPCs. Collectively these phenotypic and molecular characteristics are consistent with acquisition of cellular senescence. NS-mediated regulation of CPC senescence is p53 dependent, as silencing p53 rescues the phenotype induced by NS-silencing. In conclusion, NS which is induced downstream of Pim-1 kinase, maintains multipotency and inhibits senescence in CPCs, consistent with cumulative evidence that Pim-1 induced cardiac regeneration is mediated in part by NS, thereby providing an additional mechanistic basis for benefits of genetic engineering with Pim-1.
- © 2012 by American Heart Association, Inc.