论文部分内容阅读
目的探讨不同疗程高压氧(HBO)对缺氧缺血脑损伤(HIBD)新生大鼠神经干细胞(NSCs)增殖和分化的影响。方法7日龄SD大鼠随机分为正常对照组(CON组)、HIBD组、HBO组,其中HBO组又分为HBO1疗程组[HBO-1,在HIBD2 h开始HBO治疗,HBO压力为0.1 MPa(2 ATA),稳压60 min,1次/d,7次为1疗程],2疗程组(HBO-2,予2个疗程HBO)和3疗程组(HBO-3,予3个疗程HBO)。疗程结束后(鼠龄35 d)取脑组织标本,采用5-溴-2-脱氧嘧啶核苷(BrdU)/神经巢蛋白(Nestin)免疫荧光双标法检测缺血侧侧脑室室管膜下区(SVZ区)和海马齿状回区(DG区)的内源性NSCs的变化。Western blot法检测缺血侧大脑半球Nes-tin蛋白变化。BrdU/神经元特异性烯醇化酶(NSE)和BrdU/胶质纤维酸性蛋白(GFAP)免疫荧光双标法检测大脑皮层神经元和星形胶质细胞数量;TUNEL法检测皮层神经细胞凋亡;并进行T迷宫试验、放射臂迷宫测试、足错误等行为学测试。结果1.HBO治疗后SVZ区BrdU+/Nestin+细胞随疗程增加而逐渐增加,以HBO-3组的阳性细胞数最多;2.各组大鼠的海马DG区均仅有极少量的BrdU+/Nestin+细胞表达,且随着HBO疗程的增加,BrdU+/Nestin+细胞表达无增加;3.大脑皮层BrdU+/NSE+细胞表达随HBO疗程增加逐渐增多,其中以HBO-3组的阳性细胞数最多;4.HIBD组可见较多GFAP+细胞,高于CON组和HBO组;且各组均仅有个别BrdU+/GFAP+细胞表达;5.Western blot显示HBO-3组Nestin蛋白表达高于其余各组;6.T迷宫、放射臂迷宫和足错误测试结果均表明随着HBO疗程的增加,疗效愈加显著。结论增加HBO疗程可促进HIBD新生大鼠SVZ区内源性NSCs皮层神经元细胞的增殖和分化。
Objective To investigate the effects of hyperbaric oxygen (HBO) in different courses on the proliferation and differentiation of neural stem cells (NSCs) of neonatal rats with hypoxic-ischemic brain damage (HIBD). Methods 7-day-old SD rats were randomly divided into normal control group (CON group), HIBD group and HBO group. HBO group was further divided into HBO-1 group [HBO-1, HBO treatment at HBO 2 h, HBO pressure was 0.1 MPa (2 ATA) for 60 min, once / d for 7 times for 1 course of treatment, 2 courses of HBO-2 for 2 courses of HBO and 3 courses of HBO-3 for 3 courses of HBO ). At the end of the treatment period (35 days old), the brain tissue samples were taken and the brains were examined by immunofluorescence double labeling with BrdU / Nestin (SVZ zone) and hippocampal dentate gyrus (DG zone) endogenous NSCs changes. Western blot was used to detect the change of Nes-tin protein in the ischemic hemisphere. BrdU / neuron specific enolase (NSE) and BrdU / glial fibrillary acidic protein (GFAP) immunofluorescence were used to detect the number of cortical neurons and astrocytes; TUNEL method was used to detect the apoptosis of cortical neurons; And T maze test, radioactive maze test, foot errors and other behavioral tests. BrdU + / Nestin + cells in SVZ increased gradually with HBO-3 treatment, and HBO-3 group had the highest number of positive cells.2. There were only a few BrdU + / Nestin + cells Expression of BrdU + / Nestin + cells increased with the increase of HBO treatment; 3. The expression of BrdU + / NSE + cells in the cerebral cortex increased gradually with the increase of HBO treatment, and the positive cells in HBO-3 group were the most; The expression of Nestin protein in HBO-3 group was higher than that in the other groups. 6.T Labyrinths, the number of BrdU + / GFAP + cells in each group were higher than that in CON group and HBO group Radiofrequency maze and foot error test results show that with the HBO treatment increased, the effect is more significant. Conclusion Increased HBO treatment can promote the proliferation and differentiation of neuronal cells in the cortex of NSCs in SVZ of HIBD neonatal rats.