Functional analysis of platelet-derived growth factor-β receptor in self-renewal of neural stem cells

Abstract

Objective To investigate the role of platelet-derived growth factor-β receptor (PDGFR-β) in self-renewal of neural stem cells (NSCs). Methods In this study, NSCs of subventricular zone were isolated and cultured from PDGFR-β knockout (PDGFR-β–/–) mice of postnatal day 1 (P1) and P28; the number and diameters of secondary neurospheres were calculated; using 5-bromo-2-deoxyuridine (BrdU) incorporation assay and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay, cell proliferation and survival rates were analyzed; gene expression profiles were determined by PCR-array analyses; the effect of brain-derived neurotrophic factor (BDNF) on secondary neurospheres formation was examined in these cells. Results In PDGFR-β–/– NSCs, stem cell activities, such as number (/well; P1: 25.9±1.3 vs 117.6±3.6, t=4.236, P<0.01; P28: 13.8± 0.7 vs 19.8±0.6, t=2.116, P<0.01) and diameters (μm; P1: 67.7±1.9 vs 69.1±2.0, t=3.211, P<0.01; P28: 33.4±0.8 vs 37.8±0.8, t=2.354, P<0.01) of secondary neurospheres, cell proliferation (%; P1: 73.3±2.7 vs 88.7±3.6, t=2.773, P<0.05; P28: 28.6±9.6 vs 68.2±4.5, t=6.302, P<0.05) and survival rates (%; P1: 14.5±2.1 vs 9.3±1.3, t=7.222, P<0.05), were significantly lower as compared with age-matched controls. In comparison of the same genotypic NSCs, the decrease of secondary neurosphere formation was more striking in P28 NSCs than in P1 NSCs. PCR Array analyses demonstrated that expressions of fibroblast growth factor-2 and BDNF were decreased (–2.04±0.25, t=2.653, P<0.05; –3.24±0.37, t=1.324, P<0.05), and Noggin (2.31±0.37, t=2.749, P<0.05) was increased in P1 PDGFR-β–/– NSCs as compared with P1 controls. Addition of BDNF rescued the number and diameter of secondary neurospheres in P1 PDGFR-β–/– NSCs to similar levels as controls. Conclusions PDGFR-β signaling may play a role in the self-renewal and proliferation of NSCs. BDNF may be involved in the effects of PDGFR-β signaling in these cells. Key words: Receptor, platelet-derived growth factor beta; Neural stem cells; Mice, knockout