丁香酚与甲基丁香酚的抗焦虑作用机制研究

Anti-anxiety mechanisms of eugenol and methyleugenol

目的探究丁香酚及甲基丁香酚的抗焦虑作用及机制。方法 90只雄性SPF级SD大鼠,随机分为对照组,模型组,地西泮(1 mg/kg)组,丁香酚低、中、高剂量(10、20、40 mg/kg)组和甲基丁香酚低、中、高剂量(10、20、40 mg/kg)组,每组10只,对照组和模型组给予0.5%CMC-Na溶液,每只大鼠给药1 mL,ig给药,给药7 d后,除对照组外,进行高架十字迷宫实验。采用酶联免疫法(ELISA)测定焦虑大鼠血清中沉默信息调节因子2相关酶1(SIRT1)、单胺氧化酶A(MAO-A)、儿茶酚-O-甲基转移酶(COMT)、乙醛脱氢酶(ALDH)及醛糖还原酶(AR)的表达量,同时通过逆转录-聚合酶链反应(RT-PCR)检测各组海马体组织中相应的基因表达水平。结果丁香酚高剂量组大鼠进入开放臂次数(OE)百分比和开放臂停留时间(OT)百分比显著高于模型组(P<0.01),作用呈剂量相关性;随着剂量的增加,甲基丁香酚表现出抗焦虑作用趋势,但差异无统计学意义。与对照组比较,模型组大鼠SIRT1和MAO-A酶含量及基因表达显著升高(P<0.05、0.01),丁香酚与甲基丁香酚均发挥显著回调作用(P<0.01)。与对照组比较,模型组大鼠血清中ALDH与AR含量显著下降,COMT含量显著升高(P<0.01);海马组织中COMT与ALDH的基因表达显著上调,AR的基因表达显著下调(P<0.01)。与模型组比较,丁香酚可显著回调血清COMT、ALDH及AR含量(P<0.05、0.01),甲基丁香酚显著回调血清COMT、AR含量(P<0.05、0.01);丁香酚与甲基丁香酚能够显著抑制焦虑大鼠海马体COMT与ALDH的基因表达上调(P<0.01),甲基丁香酚显著抑制AR的基因表达下调(P<0.05)。结论丁香酚和甲基丁香酚抗焦虑活性良好,可通过调控SIRT1-MAO-A信号通路进而影响单胺类神经递质5-羟色胺(5-HT)的代谢以及儿茶酚胺代谢通路中COMT、ALDH及AR酶的活性和基因表达发挥作用。

Objective To explore the anti-anxiety effect of eugenol and methyleugenol and its possible mechanisms. Methods Totally ninety male Sprague-Dawley Rats(SPF) were randomly divided into nine groups: control group, model group, diazepam(1 mg/kg) group, eugenol low, middle and high-dose(10, 20, and 40 mg/kg) group, methyleugenol low, middle and high-dose(10,20, and 40 mg/kg) group. Then the coments of NAD-dependent deacetylase sirtuin-1(SIRT1), monoamine oxidase-A(MAO-A),catechol-o-methyl transferase(COMT), acetaldehyde dehydrogenase(ALDH) and aldose reductase(AR) in serum of anxiety rats were determined by enzyme-linked immunosorbent assay(ELISA), at the same time, the corresponding gene expression level in hippocampus was detected by Reverse Transcription-Polymerase Chain Reaction(RT-PCR). Results The percentage of OE and OT in eugenol high-dose group were significantly higher than those in model group(P < 0.01);With the increase of dosage, methyl eugenol showed the trend of anti anxiety effect, but the difference was not statistically significant. Compared with the control group,SIRT1 and MAO-A enzyme contents and mRNA levels in the anxiety model rats were significantly increased(P < 0.05 and 0.01),and eugenol and methyleugenol treatment reversed this trend(P < 0.01). The contents of ALDH and AR in serum of anxiety rats decreased significantly, while the contents of COMT increased(P < 0.01);The mRNA levels of COMT and ALDH were upregulated, while AR gene expression was down-regulated(P < 0.01). Compared with model group, eugenol reversed the change trend of COMT, ALDH and AR(P < 0.05, 0.01), and methyl eugenol significantly reversed the change trend of serum COMT and AR(P < 0.05, 0.01);Eugenol and methyl eugenol significantly inhibited the up regulation of COMT and ALDH gene expression in hippocampus of anxiety rats(P < 0.01), while methyl eugenol significantly inhibited the down regulation of AR gene expression(P < 0.05). Conclusion Eugenol and methyleugenol have good anti-anxiety activity. They can adjust the metabolism of 5-hydroxytryptamine(5-HT) by regulating SIRT1-MAO-A signal pathway and affect the enzyme activity and gene expression of COMT, ALDH and AR in the metabolic pathway of catecholamine.