Caspase-3,-9,-12及c-jun在VEGF介导的神经保护作用中的调控机制

目的探讨血管内皮生长因子(VEGF)对兔脑缺血再灌注损伤半胱氨酸蛋白酶(caspase)-3、-9、-12和c-jun的影响,从多角度探讨VEGF介导神经保护作用的调控机制。方法健康雄性新西兰白兔26只,体重2.6±0.2kg。随机分成假手术组(Sham组,n=6)、缺血/再灌注损伤组(I/R组,n=10)和VEGF治疗组(V组)(n=10)。实验3d后,各组行神经功能缺损评分、TTC染色测梗死体积、免疫组织化学方法检测缺血半暗带区caspase-3、-9、-12和c-jun表达、用DNA原位末端缺口标记法(TUNEL法)检测相应区域的神经细胞凋亡。结果Sham组未见梗死体积及神经功能缺损,caspase-3、-9、-12和c-jun呈低表达,TUNEL染色见少量阳性细胞;与Sham组比较,I/R组和V组可见左侧半球梗死,神经功能缺损,caspase-3、-9、-12和c-jun表达增加(I/R组:0.49±0.05、0.38±0.02、0.36±0.02、0.37±0.04;V组:0.24±0.05、0.18±0.02、0.15±0.03、0.17±0.03;Sham组:0.09±0.03、0.08±0.03、0.07±0.02、0.08±0.02,P<0.01),TUNEL阳性细胞数增多(32.8±2.6,9.3±0.9 vs.0.7±0.2,P<0.01);与I/R组比较,V组梗死体积减少(367.0±15.7)mm3 vs.468.6±29.7mm3,P<0.01),神经功能恢复更好,caspase-3、-9、-12和c-jun表达明显下降(0.24±0.05、0.18±0.02、0.15±0.03、0.17±0.03 vs.0.49±0.05、0.38±0.02、0.36±0.02、0.37±0.04,P<0.01),TUNEL阳性细胞数明显减少(32.8±2.6 vs.9.3±0.9,P<0.01)。结论VEGF可通过下调凋亡基因的表达,抑制反应性凋亡途径的激活来发挥其神经保护作用。

The regulated mechanism of caspase-3,-9,-12 and c-jun involved in neuroprotective effect mediatiated by VEGF

Objective To observe the regulated mechanism of VEGF on the caspase-3, -9, -12 and c-jun in cerebral ischemia-reperfusion, and elucidate the protective mechanism of VEGF on cerebral tissue underwent ischemia-reperfusion in rabbit. Methods Twenty-six healthy male New Zealand rabbits weighting (2.6±0.2)kg were randomly divided into sham operation group, ischemia-reperfusion group and VEGF group. At 3d after operation, we evaluated neurological function and measured infarct volumes by 2,3,5-triphenyltetrazolium chloride (TTC) staining. In the meantime, the tissues of ischemic penumbra region were taken to detect the expression of caspase-3, -9, -12 and c-jun by immunohistochemistry and apoptosis by TUNEL staining methods. Results In sham operation group there was no infarct volume and neurological deficit, the caspase-3, -9, -12 and c-jun expressed in a low level, a few apoptotic cells were detected by TUNEL staining. As compared with that in sham operation group, there were left infarct and light limbs deficit, the expression of caspase-3, -9, -12 and c-jun were higher in I/R group and VEGF group (I/R group: 0.49±0.05, 0.38±0.02, 0.36±0.02, 0.37±0.04;VEGF group: 0.24±0.05, 0.18±0.02, 0.15±0.03, 0.17±0.03;vs. Sham group: 0.09±0.03, 0.08±0.03, 0.07±0.02, 0.08±0.02, P<0.01); the apoptotic cells detected by TUNEL methods were significantly increased, as compared with those of sham operation group (32.8±2.6, 9.3±0.9 vs. 0.7±0.2, P<0.01). As compares with those of I/R group, infarct volume was lower (367.0±15.7)mm3 vs 468.6±29.7 mm3, P<0.01)and neurological function was better in VEGF group. The levels of caspase-3, -9, -12 and c-jun in VEGF group were lower than that of I/R group(0.24±0.05, 0.18±0.02, 0.15±0.03, 0.17±0.03 vs. 0.49±0.05, 0.38±0.02, 0.36±0.02, 0.37±0.04, P<0.01), the number of apoptotic cells was lower than that of I/R group (9.3±0.9 vs. 32.8±2.6, P<0.01). Conclusion VEGF can down-regulate the expression of caspase-3, -9, -12 and c-jun, and performs neuroprotective effect by inhibiting apoptosis pathway.