新生大鼠缺氧缺血性脑损伤模型中葡萄糖转运蛋白1表达及神经元凋亡的研究

[The studies of GLUT1 expression and neuron apoptosis in neonatal hypoxic-ischemia brain damage rat model].

作者信息

Zhang Xiao-Lan, Li De-Yuan, Zhao Feng-Yan, Qu Yi, Mu De-Zhi

机构信息

Department of Pediatrics, West China Second Hospital, Sichuan University, Chengdu, China.

出版信息

Sichuan Da Xue Xue Bao Yi Xue Ban. 2009 Sep;40(5):829-33.

PMID:19950593

Abstract

OBJECTIVE

To investigate the relationship between the expression of glucose transporter protein 1 and the apoptosis of neuron during hypoxic-ischemia brain damage in neonatal rats.

METHODS

Total 120 10-day old SD rats were divided into normal group, sham control group and hypoxic-ischemia (HI) group. In HI group, hypoxic-ischemia brain damage (HIBD) were generated according to Rice-Vannucci method, brain tissues were harvested at 2, 8, 24, 48 and 72 h after HI. The brain samples were also collected at the same time points in normal group and sham control group. The pathological changes was observed by hematoxylin-eosin (HE) staining, the mRNA expression of glucose transporter 1 (GLUT1) was detected by reverse transcriptase-polymerase chain reaction (RT-PCR), the protein expressions of GLUT1 and cleaved caspase-3 (CC3) were detected by immunohistochemistry, the apoptosis of neuron was measured by TdT-mediated dUTP-biotin nick end labeling (TUNEL) staining.

RESULTS

HE staining showed that the degree of brain cell damage increased with time after HI, the loss of neuronal cells peaked at 48 h, while the cells in control group apperanted in an orderly and normal morphology. The mRNA and protein expressions of GLUT1 were increased after HI, which began to increase at 2 h, and reach the peak at 24 h. and the expression levels at each time points were statistically higher (P< 0.01) than those in control group. CC3 protein expression also began to increase at 2 h, peaked at 48 h after HI, which was higher than that of control group (P<0.01). The number of positive cells was significantly increased after HI,with a peak at 48 h.

CONCLUSION

The mRNA and protein expression of GLUT1 in brain tissue increased significantly after hypoxic-ischemia, and the peak time was earlier than that of CC3 protein and cellular apoptosis. This suggests that GLUT1 expression upregulation may be a certain degree of inhibition on neuronal apoptosis.

摘要

目的

探讨新生大鼠缺氧缺血性脑损伤时葡萄糖转运蛋白1表达与神经元凋亡的关系。

方法

将120只10日龄SD大鼠分为正常组、假手术对照组和缺氧缺血（HI）组。HI组采用Rice-Vannucci法制备缺氧缺血性脑损伤（HIBD）模型，于HI后2、8、24、48和72 h取脑组织。正常组和假手术对照组在相同时间点取脑样本。采用苏木精-伊红（HE）染色观察病理变化，逆转录聚合酶链反应（RT-PCR）检测葡萄糖转运蛋白1（GLUT1）的mRNA表达，免疫组织化学检测GLUT1和裂解的半胱天冬酶-3（CC3）的蛋白表达，TdT介导的dUTP生物素缺口末端标记（TUNEL）染色检测神经元凋亡。

结果

HE染色显示，HI后脑细胞损伤程度随时间增加，神经元细胞丢失在48 h达到高峰，而对照组细胞形态正常、排列有序。HI后GLUT1的mRNA和蛋白表达增加，2 h开始升高，24 h达到峰值，各时间点表达水平均高于对照组（P<0.01）。CC3蛋白表达也在2 h开始增加，HI后48 h达到峰值，高于对照组（P<0.01）。HI后阳性细胞数显著增加，48 h达到高峰。