MK-801对新生大鼠脑缺氧缺血后Caspase-3激活及其对凋亡的影响

Effect of MK-801 on Caspase-3 Activation and Apoptosis after Cerebral Hypoxic Ischemic Injury in Neonatal Rats

摘要:

目的：探讨N-甲基-D-天冬氨酸(NMDA)受体拮抗剂MK-801对新生大鼠缺氧缺血(HI)后半胱天冬酶-3(Caspase-3)激活及凋亡的影响。方法：7日龄新生大鼠在HI后即刻给予腹腔注射MK-801 0.5 mg/kg，在HI后24 h取脑制作脑组织连续切片进行微管相关蛋白 2(MAP-2)，Caspase-3免疫组化染色及发夹寡核苷酸探针(Hairpin Probe, HPP)原位杂交，计算脑损伤面积及Caspase-3，HPP阳性细胞数。结果：①MK-801干预组脑损伤面积为(23±5)%，较生理盐水对照组(52±12)%明显降低，P<0.01；②MK-801干预组Caspase-3及HPP阳性细胞数(65±8)/高倍视野，(61.6±11.5)/高倍视野，与对照组(40±6.7)/高倍视野，(12.6±5.2)/高倍视野相比均明显减少，其中HPP阳性细胞数减少比Caspase-3阳性细胞数减少更明显。结论：MK-801能明显抑制Caspase-3的激活，减少神经元的凋亡，减轻缺氧缺血性脑损伤(HIBD)的程度。

Abstract:

OBJECTIVE: To study the effect of MK-801, the antagonist of NMDA receptor, on Caspase-3 activation and apoptosis after cerebral hypoxic ischemic injury in neonatal rats. METHODS: Sevendayold rat pups were injected with either 0.5 mg/kg MK-801 or normal saline immediately after cerebral hypoxicischemic injury (HI). The pups were killed 24 h after the injection. Brain damage was evaluated using MAP-2 immunostaining. Caspase-3 activation was examined using antip17 subunit antibody. Apoptosis was examined by in situ labeling of hairpin probe (HPP). The infarction area was calculated according to MAP-2 staining. Active Caspase-3 cells and HPP positive cells were counted in the MAP-2 negative area of the cortex. RESULTS: The infarction area was reduced from (52±12)% to (23±5)% after the treatment with MK-801. A few Caspase-3 and HPP positive cells were also found in the cortex of normal rat brains. The number of positive cells reached a peak at 24 h after HI. Both the number of Caspase-3 (from 65±8 to 40±6.7 per high power visual field) and HPP (from 61.6±11.5 to 12.6±5.2 per high power visual field) positive cells were decreased after the treatment with MK-801. CONCLUSIONS: MK-801 inhibits Caspase-3 activation and reduces the number of apoptotic cells. NMDA is involved in the activation of the apoptotic process in the immature brain after HI.