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目的从胚胎肝激酶-1(Flk-1)因子表达变化揭示脑脉通促进老龄大鼠脑缺血/再灌注血管生成的作用机制。方法将SD雄性老龄大鼠随机分为假手术组、模型组、尼莫地平组、脑脉通组,采用免疫组化和原位杂交等方法测定脑微血管密度(MVD)、血管场面积以及Flk-1的蛋白与mR-NA表达。结果脑脉通组缺血3 h和缺血/再灌注1、3、12 d的MVD,缺血3 h和缺血/再灌注3、6、12 d血管场面积,均较模型组升高(P<0.05,P<0.01);与尼莫地平组比较,脑脉通组缺血/再灌注12 d MVD增加(P<0.01),缺血3 h血管场面积增大(P<0.01)。模型组MVD自缺血3 h持续下降至缺血/再灌注12 d;血管场面积缺血/再灌注1 d达到峰值,然后迅速下降,至12 d降至最低。脑脉通组MVD缺血3 h明显增加,缺血/再灌注1 d达到峰值,然后开始下降,缺血/再灌注6 d降至最低,至12 d又明显增加;血管场面积缺血3 h有明显增加,缺血/再灌注1 d降至最低,3 d时有所升高,至6 d再次降低,随后再次升高,至12 d恢复至缺血/再灌注3 d时水平。脑脉通组各时间点Flk-1表达均较模型组增强(P<0.05,P<0.01);与尼莫地平组比较,脑脉通组缺血/再灌注1、6、12 d Flk-1表达增强(P<0.05,P<0.01),缺血/再灌注3 d Flk-1表达减弱(P<0.01)。模型组Flk-1表达于缺血3 h开始增强,缺血/再灌注1 d达到峰值,1~12 d表达迅速减弱;脑脉通组Flk-1于缺血3 h有较强表达,缺血/再灌注3 d达到峰值,此后表达稍有减弱,至12 d表达仍处于较高水平。脑脉通组各时间点Flk-1 mRNA表达水平均较模型组增强(P<0.01);与尼莫地平组比较,脑脉通组缺血/再灌注6、12 d Flk-1 mRNA表达水平增强(P<0.01)。模型组Flk-1 mRNA表达于缺血/再灌注1 d最强,1~12 d迅速下降;脑脉通组Flk-1 mRNA缺血3 h即有较高表达,缺血/再灌注1 d达峰值,此后逐渐下降,至12 d表达仍能维持一个较高水平。结论脑脉通可促进老年大鼠脑缺血/再灌注微血管生成,其机制可能与提高Flk-1及其mRNA表达有关。
Objective To investigate the mechanism of Naomaitong promoting cerebral ischemia / reperfusion angiogenesis in aged rats from the change of embryonic liver kinase-1 (Flk-1) expression. Methods SD male aged rats were randomly divided into sham operation group, model group, nimodipine group and Naomaitong group. Immunohistochemistry and in situ hybridization were used to determine the microvessel density (MVD), vascular field and Flk -1 protein and mR-NA expression. Results The MVD at 3 h and the 1,3,12 d after ischemia / reperfusion in the Naomaitong group were higher than those in the model group at 3 h and at the 3, 6 and 12 d after ischemia / reperfusion (P <0.05, P <0.01). Compared with nimodipine group, the MVD of cerebral ischemia-reperfusion group was increased (P <0.01) . The MVD of model group decreased from ischemia to reperfusion for 12 days, and reached the peak at 1 day after ischemia / reperfusion, then decreased rapidly and reached the lowest level at 12 days. MVD in Naotometuon group increased significantly 3 h after ischemia and peaked at 1 d after ischemia / reperfusion, then began to decrease and decreased to the lowest level at 6 d after ischemia / reperfusion, and then increased significantly after 12 d. MVD 3 h after ischemia / reperfusion was reduced to the lowest one day after ischemia / reperfusion, increased on the third day, decreased again on the sixth day, and then increased again until the 3rd day after ischemia / reperfusion. Compared with nimodipine group, the expression of Flk-1 in Naomaitong group was significantly higher than that in model group (P <0.05, P <0.01) 1 (P <0.05, P <0.01), and the expression of Flk-1 was decreased 3 days after ischemia / reperfusion (P <0.01). The expression of Flk-1 in the model group began to increase 3 h after ischemia, peaked at 1 d after ischemia / reperfusion, and rapidly decreased from 1 to 12 d. Flk-1 in Naomaitong group was strongly expressed at 3 h after ischemia, After 3 days of blood / reperfusion, the expression peaked and then weakened slightly. The expression was still high at 12 days. Compared with nimodipine group, the expression level of Flk-1 mRNA in Naomaitong group at 6 and 12 days after ischemia / reperfusion was significantly higher than that in model group (P <0.01) Enhanced (P <0.01). The expression of Flk-1 mRNA in the model group was the strongest at 1 d after ischemia / reperfusion and decreased rapidly at 1 to 12 d. The expression of Flk-1 mRNA in Naomaitong group was higher at 3 h after ischemia, Reached its peak value, then decreased gradually, and maintained a high level by the 12th day. Conclusion Naomaitong can promote cerebral ischemia / reperfusion microvessel formation in aged rats. The mechanism may be related to the increase of Flk-1 and its mRNA expression.