汉黄芩素增强乙酰胆碱对大鼠胸主动脉血管的舒张作用研究
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摘要
目的探讨汉黄芩素增强乙酰胆碱对大鼠胸主动脉血管的舒张作用及其作用机制。方法实验分为对照组和加药组,加药组使用含有不同浓度汉黄芩素的DMEM/F12培养基作用;对照组使用含有与加药组相同体积二甲基亚砜(DMSO)的DMEM/F12培养基作用。汉黄芩素孵育大鼠心脏微血管内皮细胞24 h,MTT法检测细胞活力;硝酸还原酶法检测细胞上清NO含量;ELISA法检测细胞内皮型一氧化氮合酶(eNOS)蛋白表达。汉黄芩素孵育大鼠胸主动脉环24h,去甲肾上腺素(1×10~(-6) mol/L)收缩血管后应用乙酰胆碱(1×10~(-8)、1×10~(-7)、1×10~(-6)、1×10~(-5)mol/L)舒张血管,检测血管张力变化。结果与对照组比较,汉黄芩素处理细胞后,各组细胞活力没有显著变化。与对照组比较,汉黄芩素(5、20μmol/L)显著促进NO的产生(P<0.01),但亚硝基左旋精氨酸甲酯(L-NAME)可以抑制汉黄芩素的这种作用(P<0.01)。汉黄芩素(0.5、5、20μmol/L)可以浓度相关性地促进心脏微血管内皮细胞eNOS蛋白表达,与对照组比较,汉黄芩素产生的eNOS蛋白水平差异非常显著(P<0.01)。乙酰胆碱(1×10~(-8)、1×10~(-7)、1×10~(-6)、1×10~(-5)mol/L)能够浓度相关性地舒张离体大鼠胸主动脉环。与对照组比较,汉黄芩素(20μmol/L)孵育血管24h,能显著增加乙酰胆碱对大鼠胸主动脉血管环的舒张程度,降低胸主动脉血管环的收缩率(P<0.01、0.05)。结论汉黄芩素可能通过促进血管内皮细胞eNOS蛋白表达和NO产生这一途径增强乙酰胆碱对血管的舒张作用。
Objective To study the effect of wogonin induced by acetylcholine on the vasodilation of rat thoracic aorta and explore its mechanism. Method The experiments were divided into control group and medication group. The medication group was treated with DMEM/F12 medium containing different concentrations of wogonin, while the control group was treated with DMEM/F12 medium containing the same volume of DMSO as the medication group. Rat cardiac microvascular endothelial cells were incubated with wogonin treated for 24 h. Cell viability was measured by MTT assay, NO content was measured by nitrate reductase assay, and eNOS protein expression was detected by ELISA method. The thoracic aortic rings of rats were incubated with wogonin treated for 24 h, NE(1×10~(-6) mol/L) was used to constrict blood vessels, and the acetylcholine(1×10~(-8), 1×10~(-7), 1×10~(-6), and 1×10~(-5) mol/L) was used to relax blood vessels to detect the changes of vascular tension. Results Compared with the control group, there was no significant changes of the cell viability in each group after treatment with wogonin. Compared with the control group, wogonin(5, 20 μmol/L) significantly promoted NO production(P < 0.01), but L-NAME could inhibit the effect of wogonin(P < 0.01). Wogonin(0.5, 5, 20 μmol/L) could promote eNOS protein expression in cardiac microvascular endothelial cells in a concentration-dependent manner. Compared with the control group, the level of eNOS protein produced by wogonin was significantly different(P < 0.01). Acetylcholine(1×10~(-8), 1×10~(-7), 1×10~(-6), and 1×10~(-5) mol/L) could dilate isolated rat thoracic aortic rings in a concentration-dependent manner. Compared with the control group, wogonin(20 μmol/L) incubated for 24 h significantly increased the relaxation of the thoracic aortic rings and decreased the contraction rate of the thoracic aortic rings in rats(P < 0.01, 0.05). Conclusion Wogonin may enhance vasodilation of acetylcholine by promoting expression of eNOS protein and NO production in vascular endothelial cells.
Objective To study the effect of wogonin induced by acetylcholine on the vasodilation of rat thoracic aorta and explore its mechanism. Method The experiments were divided into control group and medication group. The medication group was treated with DMEM/F12 medium containing different concentrations of wogonin, while the control group was treated with DMEM/F12 medium containing the same volume of DMSO as the medication group. Rat cardiac microvascular endothelial cells were incubated with wogonin treated for 24 h. Cell viability was measured by MTT assay, NO content was measured by nitrate reductase assay, and eNOS protein expression was detected by ELISA method. The thoracic aortic rings of rats were incubated with wogonin treated for 24 h, NE(1×10~(-6) mol/L) was used to constrict blood vessels, and the acetylcholine(1×10~(-8), 1×10~(-7), 1×10~(-6), and 1×10~(-5) mol/L) was used to relax blood vessels to detect the changes of vascular tension. Results Compared with the control group, there was no significant changes of the cell viability in each group after treatment with wogonin. Compared with the control group, wogonin(5, 20 μmol/L) significantly promoted NO production(P < 0.01), but L-NAME could inhibit the effect of wogonin(P < 0.01). Wogonin(0.5, 5, 20 μmol/L) could promote eNOS protein expression in cardiac microvascular endothelial cells in a concentration-dependent manner. Compared with the control group, the level of eNOS protein produced by wogonin was significantly different(P < 0.01). Acetylcholine(1×10~(-8), 1×10~(-7), 1×10~(-6), and 1×10~(-5) mol/L) could dilate isolated rat thoracic aortic rings in a concentration-dependent manner. Compared with the control group, wogonin(20 μmol/L) incubated for 24 h significantly increased the relaxation of the thoracic aortic rings and decreased the contraction rate of the thoracic aortic rings in rats(P < 0.01, 0.05). Conclusion Wogonin may enhance vasodilation of acetylcholine by promoting expression of eNOS protein and NO production in vascular endothelial cells.
引文
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[9]杨宝林,伍校琼,罗明英,等.L-NAME对人脐静脉内皮细胞VE-cadherin表达的影响[J].中国临床解剖学杂志,2012,30(4):412-416.
[10]Oche B,Chen L,Ma Y K,et al.Cryptotanshinone and wogonin up-regulate eNOS in vascular endothelial cells via ERαand down-regulate i NOS in LPS stimulated vascular smooth muscle cells via Erβ[J].Arch Pharm Res,2016,39(2):249-258.
[2]Huang Y,Luo X,Li X,et al.Wogonin inhibits LPS-induced vascular permeability via suppressing MLCK/MLC pathway[J].Vascul Pharmacol,2015(72):43-52.
[3]Lee W,Ku S K,Bae J S.Anti-inflammatory effects of baicalin,baicalein,and wogonin in vitro and in vivo[J].Inflammation,2015,38(1):110-125.
[4]Ku S K,Bae J S.Antithrombotic activities of wogonin and wogonoside via inhibiting platelet aggregation[J].Fitoterapia,2014(98):27-35.
[5]Qi Q,Peng J,Liu W,et al.Toxicological studies of wogonin in experimental animals[J].Phytother Res,2009,23(3):417-422.
[6]Qu J T,Zhang D X,Liu F,et al.Vasodilatory effect of wogonin on the rat aorta and its mechanism study[J].Biol Pharm Bull,2015,38(12):1873-1878.
[7]Akinyi M,Gao X M,Li Y H,et al.Vascular relaxation induced by eucommiae ulmoides Oliv.and its compounds oroxylin A and wogonin:implications on their cytoprotection action[J].Int J Clin Exp Med,2014,7(10):3164-3180.
[8]薛瑾,屈会化,赵琰,等.三草降压汤含药血清对人脐静脉内皮细胞活性及NO分泌的影响[J].中医学报,2014,29(6):763-765.
[9]杨宝林,伍校琼,罗明英,等.L-NAME对人脐静脉内皮细胞VE-cadherin表达的影响[J].中国临床解剖学杂志,2012,30(4):412-416.
[10]Oche B,Chen L,Ma Y K,et al.Cryptotanshinone and wogonin up-regulate eNOS in vascular endothelial cells via ERαand down-regulate i NOS in LPS stimulated vascular smooth muscle cells via Erβ[J].Arch Pharm Res,2016,39(2):249-258.