端粒酶激活对缺氧缺血大鼠神经干细胞生物学行为的影响  

Effect of telomerase activation on biological behaviors of neural stem cells in rats with hypoxic-ischemic insults

作　　者：孟俊杰[1] 李世平[1] 赵凤艳[1] 童煜[1] 母得志[1] 屈艺[1] 

机构地区：[1]四川大学华西第二医院儿科/出生缺陷与相关妇儿疾病教育部重点实验室/发育与妇儿疾病四川省重点实验室,四川成都610041

出　　处：《中国当代儿科杂志》2017年第2期229-236,共8页Chinese Journal of Contemporary Pediatrics

基　　金：国家自然科学基金(81330016;81270724;81501302);四川省科技厅基金(2014SZ0149;2016TD0002);国家临床重点专科(儿科新生儿专业)建议项目(1311200003303)

摘　　要：目的探索神经干细胞端粒酶激活后,在缺氧缺血条件下其增殖与存活能力的变化。方法将传代培养神经干细胞分为对照组、氧糖剥夺(OGD)组、OGD+高浓度环黄芪醇(CAG)组和OGD+低浓度CAG组。高、低浓度CAG组分别加入终浓度为25μM和10μM的CAG;除对照组外,其余各组进行OGD处理。Western blot检测细胞端粒酶逆转录酶(TERT)表达水平;TRAP法检测端粒酶活性;显微镜下计数细胞数目、测量神经球直径;化学发光法检测乳酸脱氢酶(LDH)活性;流式细胞术检测细胞增殖和凋亡情况。结果与对照组相比,OGD处理后,神经干细胞数目减少,神经球直径缩小,凋亡细胞增多,细胞增殖率下降,存活率下降(P<0.05);CAG作用后,神经干细胞TERT表达和端粒酶活性较对照组显著升高(P<0.05);与OGD组比较,CAG作用后神经干细胞减少和神经球直径缩小程度明显减轻(P<0.05),细胞增殖率升高,存活率升高(P<0.05);不同浓度CAG作用效果无明显差异(P>0.05)。在正常培养的神经干细胞中,与对照组相比,CAG作用能够提高神经干细胞TERT表达水平(P<0.05),促进细胞数目增加和神经球直径增大(P<0.05)。结论缺氧缺血状态下,端粒酶激活能够显著促进神经干细胞的增殖与存活,CAG有望用于缺氧缺血后脑损伤的修复与治疗。Objective To investigate the effect of telomerase activation on biological behaviors of neural stem cells after hypoxic-ischemic insults. Methods The neural stem cells passaged in vitro were divided into four groups： control, oxygen-glucose deprivation（OGD）, OGD＋cycloastragenol（CAG） high concentration（final concentration of 25 μM）, and OGD＋CAG low concentration（final concentration of 10 μM）. The latter three groups were subjected to OGD. Telomerase reverse transcriptase（TERT） expression level was evaluated by Western blot. Telomerase activity was detected by telomerase repeat amplification protocol（TRAP）. Cell number and neural sphere diameter were measured under a microscope. The activity of lactate dehydrogenase（LDH） was examined by chemiluminescence. Cell proliferation rate and apoptosis were detected by flow cytometry. Results After OGD insults, obvious injury of neural stem cells was observed, including less cell number, smaller neural sphere, more dead cells, lower proliferation rate and decreased survival rate. In CAG-treated groups, there were higher TERT expression level and telomerase activity compared with the control group（P〈0.05）. In comparison with the OGD group, CAG treatment attenuated cell loss（P〈0.05） and neural sphere diameter decrease（P〈0.05）, promoted cell proliferation（P〈0.05）, and increased cell survival rate（P〈0.05）. Low and high concentrations of CAG had similar effects on proliferation and survival of neural stem cells（P〈0.05）. In the normal cultural condition, CAG treatment also enhanced TERT expression（P〈0.05） and increased cell numbers（P〈0.05） and neural sphere diameter（P〈0.05） compared with the control group. Conclusions Telomerase activation can promote the proliferation and improve survival of neural stem cells under the state of hypoxic-ischemic insults, suggesting telomerase activators might be potential agents for the therapy of hypoxic-ischemic brain injury.

关 键 词：细胞增殖 细胞凋亡 端粒酶 神经干细胞 

分 类 号：R742[医药卫生—神经病学与精神病学]