N-乙酰半胱氨酸预防硝酸酯耐药的实验效果及机制探讨

Experimental effect and mechanism of N-acetylcysteine in preventing nitrate resistance

目的通过建立单硝酸异山梨醇酯(ISMN)耐药动物模型,观察N-乙酰半胱氨酸(NAC)对预防硝酸酯耐药的实验效果,分析相关机制。方法取成年雄性新西兰大白兔32只,体质量(2500±200)g,随机分为对照组、常规组、耐药组和NAC组,每组8只。对照组不予用药;常规组经胃管灌服ISMN 25 mg,1/d;耐药组灌服ISMN 25 mg,1/12 h;NAC组同时灌服ISMN 25 mg和NAC 100 mg,1/12 h。7 d后处死所有实验兔,取腹主动脉,每5 mm剪为数段,分别检测血管环舒张反应以及血管组织总巯基(T-SH,微量酶标法)、超氧阴离子(O_2^-,化学比色法)、超氧化物歧化酶(SOD,黄嘌呤氧化酶法)、丙二醛(MDA,硫代巴比妥酸法)、内皮素-1(ET-1,双抗体夹心法)和亚硝酸盐(NO_2^-,硝酸还原酶法)水平。结果随着硝酸甘油(GTN)浓度的递增,各组腹主动脉血管环的舒张幅度都呈现不同程度的增加。耐药组血管环的舒张幅度均明显小于其他3组,差异有统计学意义(P均<0.01);而对照组、常规组和NAC组之间的差异无统计学意义(P均>0.05)。当GTN浓度增至10^(-4)mol/L时,对照组、常规组和NAC组的舒张幅度分别达到(98.91±1.99)%、(95.59±3.97)%和(93.18±6.36)%而耐药组仅达到(63.62±6.03)%,明显低于其他3组。T-SH含量和SOD活力由高到低依次为对照组、NAC组、常规组和耐药组,差异均有统计学意义(P均<0.05);与之相反,O_2^-、MDA、ET-1和NO_2^-含量由高到低则依次为耐药组、常规组、NAC组和对照组,差异均有统计学意义(P均<0.05)。结论口服NAC可通过增加血管组织的巯基含量、抑制ISMN诱发的氧化应激和缩血管物质的生成来预防硝酸酯的耐药性,进而充分发挥其扩血管效应。

Objective To observe the experimental effect of N-acetylcysteine(NAC)in preventing nitrate resistance and analyze relevant mechanism through establishing an animal model of isosorbide-5-mononitrate(ISMN)resistance.Methods Male adult New Zealand rabbits(n=32)with body mass=(2500±200)g were randomly divided into control group,routine group,drug-resistance group and NAC group(each n=8).The control group was not given any drugs,routine group was intragastrically given ISMN(25mg/time)once a day,drugresistance group,ISMN(25mg/time)once every12h,and NAC group,ISMN(25mg/time)and NAC(100mg)once every12h.All rabbits were executed after7d and the abdominal aorta was collected and cut into segments(5mm)for detecting vascular ring relaxation response,and levels of total sulfhydryl(T-SH,micro enzyme standard method),superoxide anion(O2-,chemical colorimetry),superoxide dismutase(SOD,xanthine oxidase method),malondialdehyde(MDA,thiobarbituric acid method),endothelin-1(ET-1,double antibody sandwich method),and nitrite(NO2-,nitric acid reductase method).Results As the concentration of nitroglycerin(GTN)increased progressively,vascular ring relaxation amplitude of abdominal aorta increased in varying degrees in all groups.The vascular ring relaxation amplitude was lower in drug-resistance group than that in other3groups(all P<0.01),and the difference among control group,routine group and NAC group had no statistical difference(all P>0.05).When the concentration of GTN increased to10-4mol/L,the vascular ring relaxation amplitude reached to(98.91±1.99)%in control group,(95.59±3.97)%in routine group,(93.18±6.36)%in NAC group,and only(63.62±6.03)%in drug-resistance group.The level of T-SH and SOD vitality showed a descending order in control group,NAC group,routine group and drug-resistance group(all P<0.05).While the levels of O2-,MDA,ET-1and NO2-showed a descending order in drug-resistance group,routine group,NAC group and control group(all P<0.05).Conclusion Orally administrated NAC can prevent nitrate resistance and exert the effect of vascular dilation through increasing sulfhydryl level and inhibiting oxidative stress and generation of vasoconstrictive substance induced by ISMN.