凝血酶和血红蛋白对大鼠水通道蛋白的影响及与脑积水的相关性研究

摘要

Objective To investigate the effect of thrombin and hemoglobin on aquaporin (AQP) and the correlation between AQP and hydrocephalus.Methods Eighty-four clean grade healthy male SD rats were randomly divided into 3 groups:a control group,a thrombin group,and a hemoglobin group using the random number table method.A hydrocephalus model was induced by injecting isotonic saline (0.3 ml),thrombin (0.3 ml[10,U/ml]) and hemoglobin (0.3 ml[150 mg/ml]),respectively into the cisterna magna.According to the deficiency and complement way,each group maintained 24 rats.The relative area of the lateral ventricles,the expression of AQP1 and AQP4,and the correlation between AQP and the area of the lateral ventricles were observed at 1,3,7,and 14 d after molding.Results (1) Compared with the control group,both the thrombin group and hemoglobin group showed hydrocephalus at 1 ,3 ,7 and 14 d,and they were most obvious at 1 day (6.94±0.19% and 6.58±0.15% vs.3.40±0.13%,6.06±0.12% and 5.79±0.09% vs.3.55±0.15%,5.80±0.13% and 5.58±0.08% vs.3.78±0.18%,5.66±0.14% and 5.47±0.13% vs.3.52±0.18 %,respectively).There were significant differences (all P<0.01).(2) The increase of AQP1 was mainly in the basal membrane and apical membrane of ventricular choroid plexus epithelial cells,and the increase of AQP4 was mainly in the ependymal cell of ventricle.The relative expression levels of AQP1 and AQP4 at 1,3,7,and 14 d in the control group were 1.09±0.07 and 1.30±0.15,0.91±0.06 and 1.18±0.12,1.33±0.17 and 1.16±0.08,1.22±0.11 and 1.00±0.10,respectively;the thrombin group were 4.40±0.14 and 3.69±0.11,3.88±0.11 and 3.17±0.07,3.55±0.07 and 2.86±0.13,and 3.36±0.07 and 2.70±0.07,respectively,the hemoglobin group were 4.24±0.07 and 3.55±0.10,3.77±0.08 and 3.04±0.09,3.46±0.07 and 2.76±0.08,and 3.31±0.10 and 2.62±0.08,respectively;the relative expression levels of AQP1 and AQP4 of the thrombin group and hemoglobin group at each time point were significantly higher than those of the control group.There were significant differences among the groups (all P<0.01).There were no significant differences in the relative expression levels of AQP1 and AQP4 mRNAs in the hemoglobin group at each time point (P>0.05);in the thrombin group and hemoglobin group,compared with those at 1 d,the expression levels of AQP1 and AQP4 at 3,7,and 14 d were significantly decreased (all P<0.01);compared with those at 3 d,AQP1 was decreased significantly at 7 and 14 d (P<0.05).The differences were statistically significant (P<0.05).(3) The relative expression levels of AQP1 (r=0.983,P<0.01) and AQP4 (r=0.987,P<0.01) in the thrombin group at each time point were positively correlated with the contralateral ventricular area;and the relative expression levels of AQP1 (r=0.964,P<0.01) and AQP4 (r=0.962,P<0.01) in the hemoglobin group at each time point were positively correlated with the contralateral ventricular area Conclusions After injecting thrombin and hemoglobin into subarachnoid space,it could cause the increased expression levels of AQP1 and AQP4 of ventricles and their surrounding areas.Thrombin and hemoglobin may be the important mediating factors of hydrocephalus after subarachnoid hemorrhage.%目的 探讨凝血酶和血红蛋白对水通道蛋白(AQP)的影响及AQP与脑积水的相关性.方法 取清洁级健康雄性SD大鼠84只,采用随机数字表法分为对照组、凝血酶组及血红蛋白组,分别通过枕大池注射等渗盐水(0.3 ml)、凝血酶[0.3 ml(10 U/ml)]及血红蛋白[0.3 ml(150 mg/ml)]制作脑积水模型,按照缺失补足的方式,保持各组大鼠24只.观察造模后1、3、7、14 d时的相对侧脑室面积、AQP1和AQP4的表达及AQP与相对侧脑室面积的相关性.结果 (1)相对于对照组,凝血酶组与血红蛋白组在1、3、7、14 d均显示脑积水,且在1 d时最明显,分别为[(6.94±0.19)%和(6.58±0.15)%比(3.40±0.13)%、(6.06±0.12)%和(5.79±0.09)%比(3.55±0.15)%、(5.80±0.13)%和(5.58±0.08)%比(3.78±0.18)%、(5.66±0.14)%和(5.47±0.13)%比(3.52±0.18)%],差异均有统计学意义(P<0.01).(2)AQP1的增加主要在脑室脉络丛上皮细胞基底膜与顶膜,AQP4的增加主要在脑室室管膜细胞.1、3、7、14 d时AQP1与AQP4的相对表达量,对照组分别为1.09±0.07和1.30±0.15、0.91±0.06和1.18±0.12、1.33±0.17和1.16±0.08、1.22±0.11和1.00±0.10,凝血酶组分别为4.40±0.14和3.69±0.11、3.88±0.11和3.17±0.07、3.55±0.07和2.86±0.13、3.36±0.07和2.70±0.07,血红蛋白组分别为4.24±0.07和3.55±0.10、3.77±0.08和3.04±0.09、3.46±0.07和2.76±0.08、3.31±0.10和2.62±0.08;凝血酶组与血红蛋白组在各时点的AQP1与AQP4的相对表达量均明显高于对照组,组间差异均有统计学意义(均P<0.01);血红蛋白组各时点的AQP1与AQP4 mRNAs的相对表达量与凝血酶组的差异均无统计学意义(均P>0.05);在凝血酶组和血红蛋白组中,相对于1 d时,AQP1与AQP4的表达在3、7、14 d有明显下降(均P<0.01);相对于3 d时,AQP1在7、14 d均有明显下降(均P<0.05),AQP4在14 d时下降,差异均有统计学意义(均P<0.05).(3)凝血酶组各时点AQP1(r=0.983,P<0.01)及AQP4(r=0.987,P<0.01) 相对表达量与相对侧脑室面积呈正相关;血红蛋白组各时点AQP1(r=0.964,P<0.01)及AQP4(r=0.962,P<0.01)相对表达量与相对侧脑室面积呈正相关.结论 蛛网膜下腔注射凝血酶与血红蛋白后可引起脑室及其周围区域AQP1与AQP4的表达增加,凝血酶与血红蛋白可能是蛛网膜下腔出血后脑积水的重要介导因素.