银杏叶提取物对老年大鼠肠系膜微动脉一氧化氮合酶表达和一氧化氮释放的影响

目的 探讨银杏叶提取物( GBE)对老年大鼠肠系膜微动脉内皮型一氧化氮合酶(eNOS)的表达和一氧化氮(NO)释放的影响,初步阐明其血管保护作用的机制。方法 (1)细胞学实验：体外培养人脐静脉内皮细胞( HUVEC),分成对照组、eNOS阻断剂L-硝基-精氨酸甲酯(L-NAME)组和GBE组。L-NAME组中加入100 μmol L-NAME孵育48 h;GBE组中先加入100 μmol L-NAME孵育24h,再加入GBE 20 g/L共同孵育24h。观察各组eNOS蛋白的表达。(2)动物实验：取24月龄雄性SD大鼠32只,抽签随机分为健康对照组（8只）和GBE组（分为3组分别给药3、5、7d,每组8只）,并分别测定20、40、60、80、100、120、140 mm Hg(l mm Hg=0.133 kPa)血管内压力下大鼠肠系膜一级微动脉的直径,推算血管的弹性,并检测大鼠肠系膜微动脉在15 dyn/cm2流体切应力刺激下NO的释放量以及eNOS蛋白与基因的表达。结果 (1)细胞学实验：L-NAME组HUVEC eNOS蛋白表达明显低于对照组和GBE组（0.57±0.06比0.96±0.05、0.81±0.09,均P＜0.01）。(2)GBE3、5、7d组大鼠肠系膜微动脉NO释放量(8.01±0.24、12.11 ±0.78、14.72 ±0.70) pmol·mm-2·min-1均显著高于健康对照组(5.83 ±0.75) pmol·mn-2·min-1,均P ＜0.05],且7d组最高;大鼠肠系膜微动脉eNOS蛋白的表达也明显高于健康对照组（0.59±0.20、0.86±0.02、1.09±0.13比0.41±0.16,均P＜0.05）,且7d组最高;eNOS mRNA的表达也均高于健康对照组(0.79±0.04、0.85±0.07、0.99±0.03比0.58±0.05,均P＜0.05),且7d组最高。结论 GBE可能通过增加微动脉NO的释放量以及eNOS的表达改善血管的弹性,从而发挥保护血管的作用。

Effects of Ginkgo biloba extract on the expression of eNOS and the release of NO in mesenteric arterioles of senile rats

Objective To investigate the effects of Ginkgo biloba extract (GBE) on the expression of endothelial nitric oxide synthase (eNOS) and the release of nitric oxide (NO) in mesenteric arterioles of aging rats. Methods ( 1 ) Cytologic experiment: human umbilical vein endothelial cells ( HUVEC ) were randomly divided into 3 groups: control group, NG-nitro-L-arginine methyl ester (L-NAME) group and GBE group. L-NAME group:100 μmol/L L-NAME was added into HUNEC for a 48-hour incubation. GBE group:After HUVEC was exposed to 100 μmol/L L-NAME for 24 hours, 20 g/L GBE was added for another 24-hour co-incubation. Then the expression of eNOS protein was observed in each group. (2) Animal experiment : Thirty-two 24-month-old male SD rats were randomly divided into normal control group ( n =8)and GBE group (n =24). The GBE group was further divided into 3 groups receiving an orally dosed GBE for 3, 5, 7 days respectively. Afterward the diameter of first-order mesenteric arteriole was measured under the pressures of 20, 40, 60, 80, 100, 120, 140 mm Hg(1 mm Hg =0.133 kPa) and the elasticity of blood vessels calculated. The release of NO, the expression of eNOS protein and its mRNA in mesenteric arterioles stimulated by the same shear stress ( 15 dyn/cm2 ) were evaluated respectively. Results ( 1 ) Cytological studies indicated that the expression of eNOS protein of the L-NAME group was significantly lower than those of the control and GBE groups (0.57 ±0.06 vs 0.96 ±0.05,0.81 ±0.09, both P ＜0.01 ). (2) After the dosing of GBE for 3, 5, 7 days, the release of NO was significantly higher than that of the control group (8.01 ±0.24, 12.11 ±0.78, 14.72 ±0.70 vs 5.83 ±0.75) pmol · mm-2 · min-1, all P ＜0.05] ; the expressions of eNOS protein were significantly higher than those of the control group (0.59 ± 0.20, 0.86 ±0.02, 1.09 ±0.13 vs 0.41 ±0.16, all P ＜0.05). And GBE was highest at Day 7 ; the expression levels of eNOS mRNA were significantly higher than those of the control group (0.79 ± 0.04, 0.85 ± 0.07, 0.99 ±0.03 vs 0.58 ± 0.05, all P ＜ 0.05 ). And GBE was also highest at Day 7. Conclusion GBE can improve vascular flexibility through increasing the expression of eNOS and the release of NO, protecting the functions of blood vessels.