AMPA谷氨酸受体亚基-2在大鼠脊髓损伤急性期对少突胶质前体细胞凋亡的影响

Effect of AMPA-GluR2 on apoptosis of oligodendrocyte precursor cells in acute stage of spinal cord injury in rats

中文关键词:  脊髓损伤  AMPA受体  谷氨酸受体亚基2  NBQX  JSTx  少突胶质前体细胞  凋亡

英文关键词:Spinal cord injury  AMPA receptor  Glutamate receptor subunit 2  NBQX  JSTx  Oligodendrocyte precursor cells  Apoptosis

中文摘要:

  【摘要】 目的：探讨AMPA谷氨酸受体亚基-2（AMPA-GluR2）在大鼠脊髓损伤急性期对少突胶质前体细胞（oligodendrocyte precursor cells，OPCs）凋亡的影响。方法：60只SD雌性大鼠随机分为假手术组（Sham组，n=15），脊髓损伤组（SCI组，n=15），AMPA受体拮抗剂NBQX组（n=15）和Ca2+通透性AMPA受体拮抗剂JSTx组（n=15）。SCI组、NBQX组和JSTx组应用Allen′s打击法建立大鼠脊髓（T9）损伤模型。采用BBB评分评估各组大鼠脊髓损伤后运动功能恢复情况。HE染色观察脊髓损伤后病理学改变。免疫组化和免疫蛋白印迹（Western blot）法检测AMPA-GluR2在各组大鼠脊髓组织中的表达情况。免疫荧光标记OPCs并应用TUNEL法检测其在各组中的凋亡情况。结果：SCI组BBB评分较假手术组降低（P＜0.05），脊髓损伤后3d NBQX组（3.60±0.65）和JSTx组（3.80±0.76）BBB评分较SCI组（1.50±0.35）高（P＜0.05），NBQX组和JSTx组之间无差异（P>0.05）。HE染色结果显示SCI组和两个拮抗剂组的脊髓组织均存在损伤，脊髓组织腹侧和腹外侧白质病理学损伤评分显示NBQX组（1.60±0.42）与JSTx组（1.50±0.35）评分较SCI组（2.30±0.20）低（P＜0.05）。AMPA-GluR2免疫组化结果显示，脊髓损伤后3d SCI组阳性细胞数（6.15±0.52）较假手术组（13.25±0.21）明显减少（P＜0.05），NBQX组（2.10±0.42）和JSTx组（4.45±0.54）阳性细胞数较SCI组少（P＜0.05），NBQX组阳性细胞数最少，与JSTx组比较有显著性差异（P＜0.05）。Western blot结果显示，脊髓损伤后3d SCI组和两个拮抗剂组AMPA-GluR2表达水平均较假手术组（0.94±0.07）降低（P＜0.05），NBQX组（0.37±0.07）及JSTx组（0.54±0.12）较SCI组（0.69±0.03）低（P＜0.05），且NBQX组AMPA-GluR2表达水平最低（P＜0.05）。免疫荧光显示，脊髓损伤后3d各组大鼠脊髓组织均存在免疫荧光标记的OPCs；TUNEL法检测OPCs凋亡指数表明，NBQX组（0.21±0.02）和JSTx组（0.17±0.01）较SCI组（0.42±0.02）均降低（P＜0.05），且JSTx组较NBQX组降低（P＜0.05）。结论：大鼠脊髓损伤急性期OPCs凋亡与脊髓组织中AMPA-GluR2表达下降有关。

英文摘要:

  【Abstract】 Objectives: To investigate the effect of AMPA-glutamate receptor subunit-2(AMPA-GluR2) on the apoptosis of oligodendrocyte precursor cells(OPCs) in the acute phase of spinal cord injury in rats. Methods: 60 female SD rats were randomly divided into sham operation group(Sham group, n=15), spinal cord injury group(SCI group, n=15), AMPA receptor antagonist NBQX group(n=15) and Ca2+ permeability AMPA receptor antagonist JSTx group(n=15). Allen′s method was used in SCI group, NBQX group and JSTx group to establish a rat spinal cord(T9) injury model using. BBB score was used to evaluate the recovery of motor function after spinal cord injury in rats. The pathological changes after spinal cord injury were observed by HE staining. The expression of AMPA-GluR2 in spinal cord tissue of each group was detected by immunohistochemistry and Western blot. The OPCs was labeled with immunofluorescence and its apoptosis in each group was detected by TUNEL method. Results: The BBB score of SCI group was lower than that of Sham group(P<0.05), the BBB scores of NBQX group(3.60±0.65) and JSTx group(3.80±0.76) were higher than that of SCI group(1.50±0.35) on the 3rd day after spinal cord injury(P<0.05) and there was no difference between NBQX group and JSTx group(P>0.05). The results of HE staining showed that there were spinal cord injuries in SCI group and two antagonist groups, the ventral and ventrolateral white matter pathological damage scores of spinal cord tissue showed that the scores of NBQX group(1.60±0.42) and JSTx group(1.50±0.35) were lower than that of SCI group(2.30±0.20)(P<0.05). The results of AMPA-GluR2 immunohistochemistry showed that the number of positive cells in SCI group(6.15±0.52) was significantly lower than that in Sham group(13.25±0.21) 3 days after spinal cord injury(P<0.05), the number of positive cells in NBQX group(2.10±0.42) and JSTx group(4.45±0.54) was lower than that in SCI group(P<0.05) and the number of positive cells in NBQX group was the least, the number of positive cells in JSTx group was higher than that in the NBQX group(P<0.05). Western blot results showed that the expression of AMPA-GluR2 in SCI group and two antagonist groups was lower than that in Sham group(0.94±0.07) 3 days after spinal cord injury(P<0.05), the expression level of AMPA-GluR2 in NBQX group(0.37±0.07) was lower than that in JSTx group(0.54±0.12)(P<0.05), and the expression level of AMPA-GluR2 was the lowest in the NBQX group. Immunofluorescence showed that positive-labeled OPCs were presented in the spinal cord of rats in each group 3 days after spinal cord injury, the apoptosis index of OPCs in NBQX group and JSTx group was lower than that in SCI group(0.42±0.02) by TUNEL method(P<0.05), and the apoptotic index of JSTx group(0.17±0.01) was lower than that of NBQX group(0.21±0.02)(P<0.05). Conclusions: The apoptosis of OPCs in the acute stage of spinal cord injury in rats is related to the decrease of AMPA-GluR2 expression in spinal cord.