Omi／HtrA2在缺氧缺血性脑损伤新生大鼠顶叶皮质中的表达变化

Expressions of Omi/HtrA2 in parietal cortex of neonatal rats with hypoxic-ischemic brain damage

目的 观察缺氧缺血性脑损伤（HIBD）新生大鼠顶叶皮质中Omi／HtrA2的表达变化，进而探讨其在HIBD中的可能作用。方法 将新生7d龄SD大鼠按完全随机数字表法分为假手术组和模型组（制作新生大鼠HIBD模型），并按术后观察时间点不同进一步分为6h、12h、24h、72h4个亚组。分别于术前及缺氧后1h对大鼠进行行为学检查：不同时间点处死后观察每只大鼠的脑组织大体形态．并采用免疫组化染色和Westem blotting检测病变侧脑组织中Omi／HtrA2蛋白的表达变化。结果（1）行为学检查：术前每只大鼠翻身能力、夹尾右旋测定均正常。术后假手术组测定结果同前。模型组大鼠中18只不能翻身，20只出现夹尾右旋，还有14只表现为不能翻身和夹尾右旋同时存在。（2）脑大体形态观察：模型组大鼠结扎侧半球脑组织受损严重，24h时苍白、水肿明显，体积明显大于对侧。假手术组大鼠大脑半球左右对称，未见任何病理变化。（3）Western blotting结果：模型组Omi／HtrA2蛋白的表达水平在各时间点均较假手术组明显升高，差异均有统计学意义俨〈0．05）。Omi／HtrA2在HIBD后6h开始表达，24h达高峰，随后开始下降。（4）免疫组化染色结果：模型组病变侧大脑皮层Omi／HtrA2阳性细胞数在HIBD后24h达到高峰．随后降低，与假手术组在各时问点比较差异均有统计学意义（P〈0．05）。结论新生大鼠HIBD后病变侧大脑皮层Omi／HtrA2的表达水平明显升高，并具有时间差异性，表明其参与了HIBD的病理过程，可能在HIBD诱导的细胞凋亡中发挥重要作用。

Objective To study the dynamic changes of Omi/HtrA2 in the cytoplasm of cerebral parietal cortex in neonatal rats with hypoxic-ischemic brain damage （HIBD）, and explore the role of Omi/HtrA2 in HIBD. Methods Seven-day-old SD rats were randomly assigned to sham-operated group and vehicle group; and HIBD models were established by the Rice and CHEN Hui-jin methods. Each group was further divided into 4 subgroups （n=8） according to the observation time points （6, 12, 24 and 72 h）. In each subgroup, the turnover ability and dextrorotatory ability when their tails were gripped were observed before the experiment and 1 h after hypoxic-ischemia （HI）. The appearance of the brain was observed, and the protein levels of Omi/HtrA2 in the brain tissues or homogenates were examined by immunohistochemistry and Western blotting. Results （1） Changes of behavior of the rats： before the experiment, all rats were healthy; 1 h after HI, the rats in sham-operated group were still healthy, while out of 32 rats which bared HI injury, 18 could not turn themselves over, 20 became dextrorotatory when their tails were gripped and 14 exhibited both behavior problems. （2） General examination of the brain： the ligated brain hemisphere in the vehicle group showed obvious pallor and edema at 24 h; the two brain hemispheres in sham-operated group showed no pathological change. （3） Western blotting results showed that the protein levels of Omi/HtrA2 in the vehicle group were significantly higher than those in the sham-operated group at each observation time points （P〈0.05）; the expressions of Omi/HtrA2 in the vehicle group began to increase at 6 h, peaked at 24 h after HI insult and decreased thereafter. （4） By immunohistochemistry, it showed that the Omi/HtrA2 positive cells in cytoplasm and membrane of cerebral cortex in the vehicle group peaked at 24 h after HI insult and decreased thereafter; the quantity of positive cells in the vehicle group was larger than that in the sham-operated group （P〈0.05）. Conclusion The levels of Omi/HtrA2 in the cerebral cortex increase after HI insult in neonatal rats, showing time-difference expression, which may play important roles in cell apoptosis induced by HIBD.