3-HB协同bFGF诱导促进神经干细胞向神经元方向分化
3-hydroxybutyrate and fibroblast growth factor-2 synergistically promote the in vitro neuronal differentiation of rat cortical neural stem cells

摘要：目的 研究3-羟基丁酸(3-HB)与bFGF或EGF协同作用对神经干细胞(NSCs)活力及分化的影响。方法 分离培养孕14～15d大鼠胚胎脑皮质NSCs，以不加处理的实验组为对照，在bFGF(10ng/mL)或EGF(20ng/mL)处理条件下分别以不同浓度3-HB(0、0.02、0.2、2mmol/L)处理P3代NSCs。分别检测细胞活力，免疫细胞化学染色MAP2观察NSCs 向神经元方向的分化情况，RT-qPCR检测转录因子SOX2、PAX6和神经元核定位因子NeuN的mRNA表达水平。结果 3-HB协同bFGF处理NSCs能够提高NSCs的细胞活力。0.02mmol/L 3-HB处理组NSCs分化的MAP2阳性细胞比例增加，PAX6和NeuN的mRNA水平升高。外源添加bFGF的条件下，0.02mmol/L处理组3-HB促进NSCs向神经元方向分化的效果更明显。结论 低浓度3-HB能够促进NSCs向神经元方向的分化，0.02mmol/L的3-HB与bFGF协同作用对NSCs向神经元方向分化的促进作用更明显，其可能通过影响SOX2、PAX6及NeuN的表达而发挥此调控作用。
ABSTRACT: Objective ?To explore the synergistic effects of 3-hydroxybutyrate (3-HB) combined with fibroblast growth factor-2 (bFGF) or epidermal growth factor (EGF) on the in vitro proliferation and differentiation of rat cortical neural stem cells (NSCs). Methods ?NSCs were isolated from the cerebral cortex of rat embryos on embryonic day 14-15. The P3 NSCs were treated with different concentrations of 3-HB (0.02, 0.2, and 2mmol/L) plus bFGF (10ng/mL) or EGF (20ng/mL), respectively. The proliferation and viability of NSCs were detected by CCK-8; the neuronal differentiation of NSCs was analyzed by immunocytochemistry staining of MAP2; relative mRNA expression levels of SOX2, PAX6 and NeuN were quantified by RT-qPCR. Results? ?3-HB plus bFGF could synergistically enhance the cell viability. The percentage of MAP2-positive cells was significantly increased in 0.02mmol/L 3-HB treatment group, and it was even higher when bFGF was added. 3-HB regulated the mRNA expressions of SOX2, PAX6 and NeuN. Conclusion? ?3-HB treatment promoted the neuronal differentiation of NSCs and its effect was further enhanced when supplemented with bFGF. The possible mechanism might invovle the regulation of SOX2, PAX6 and NeuN expressions by 3-HB