白藜芦醇通过单磷酸腺苷活化的蛋白激酶信号促进颅脑创伤远期神经功能恢复

Resveratrol promotes neural function recovery after traumatic brain injury through adenosine monophosphate activated protein kinase signaling

目的观察白藜芦醇通过调节单磷酸腺苷活化蛋白激酶(AMPK)信号活性对创伤性脑损伤远期神经修复的作用。方法将SPF级健康成年雄性C57BL/6小鼠随机将小鼠分为白藜芦醇组、生理盐水组和白藜芦醇+Compound C组,每组10只,建立创伤性脑损伤(TBI)后1~14 d、1~28 d模型。免疫荧光染色方法检测TBI后各组梗死周边皮层GAP-43的表达;改良神经功能评分(mNSS)评分量表评价白藜芦醇对神经功能影响;蛋白质印迹法(Western blot)检测p-AMPK的表达;mNSS评分采用重复测量,组间采用t检验。结果神经功能分析,在1 d白藜芦醇组mNSS评分[(12.20±1.03)分]与对照组[(12.10±1.28)分]比较,差异无统计学意义(P>0.05);在14 d[(5.50±0.85)分]、28 d[(3.30±0.95)分]时间点白藜芦醇组的mNSS评分均低于对照组14 d[(10.60±1.51)分]、28 d[(10.00±1.33)分]时间点,差异有统计学意义(14 d:t=-9.329,P<0.05;28 d:t=-12.948,P<0.05)。利用免疫荧光染色方法检测了TBI后各组脑损伤周边皮层GAP-43的表达,治疗组(14 d:39.58±1.44;28 d:46.62±0.99)较对照组(14 d:26.34±1.22;28 d:26.93±1.30)GAP-43表达提高,差异有统计学意义(14 d:t=22.236,P<0.05;28 d:t=38.217,P<0.05);抑制AMPK活性对白藜芦醇治疗TBI后神经功能的影响,在1 d白藜芦醇组mNSS评分(12.20±1.03)与白藜芦醇+Compound C组(12.40±1.34)差异无统计学意义(P>0.05);在14 d和28 d mNSS评分白藜芦醇组[(5.50±0.85)分]、28 d[(3.30±0.95)分]均低于白藜芦醇+Compound C组14 d[(7.70±1.64)分]、28 d(7.90±1.52)分],差异有统计学意义(14 d:t=-3.773,P<0.05;28 d:t=-8.104,P<0.05)。Western blot结果显示,白藜芦醇组14、28 d(0.528±0.011、0.618±0.010)p-AMPK表达增强较对照组(0.407±0.013、0.406±0.011),差异有统计学意义(t=20.266、44.227,P<0.05)。Compound C(0.501±0.012)抑制p-AMPK表达(0.704±0.017),差异有统计学意义(t=30.914,P<0.05)。结论白藜芦醇通过调节AMPK信号活性促进创伤性脑损伤远期神经修复。

Objective To investigate the effect of resveratrol on long-term neural repair of traumatic brain injury by regulating the activity of adenosine monophosphate-activated protein kinase(AMPK)signaling.Methods The experiment was divided into resveratrol group,normal saline group and resveratrol+compound C group,and the models were established 1-14 d and 1-28 d after TBI.Immunofluorescence staining was used to detect the expression of GAP-43 in the peri-infarct cortex of each group after TBI.The modified neurological severity scores(mNSS)score scale was used to evaluate the effect of resveratrol on neurological function.Western blotting was used to detect the expression of p-AMPK.The SPSS 25.0 statistical software was used to analyze and quantify.The data were expressed as±s.The mNSS score was repeatedly measured,and the t test was used.Results Neurological function analysis showed there was no statistical difference in the mNSS score between the resveratrol group(12.20±1.03)and the control group(12.10±1.28)on the 1st day(P>0.05).The mNSS scores in the resveratrol group at the time points of 14 d(5.50±0.85)and 28 d(3.30±0.95)were significantly lower than those in the control group at 14 d(10.60±1.51)and 28 d(10.0±1.33)(14 d:t=-9.329,P<0.05;28 d:t=-12.948,P<0.05).Immunofluorescence staining was used to detect the expression of GAP-43 in the peripheral cortex of brain injury in each group after TBI.The expression of GAP-43 in the treatment group(14 d:39.58±1.44;28 d:46.62±0.99)was significantly increased as compared with control group(14 d:26.34±1.22;28 d:26.93±1.30)(14 d:t=22.236,P<0.05;28 d:t=38.217,P<0.05).For the effect of inhibiting AMPK activity on the neurological function after the treatment of TBI with resveratrol,there was no significant difference in the mNSS score between the resveratrol group(12.20±1.03)and the resveratrol+compound C group(12.40±1.34)on the 1st day(P>0.05).The mNSS scores on 14 d and 28 d in resveratrol group(5.50±0.85)and 28 d(3.30±0.95)were significantly lower than those in resveratrol+compound C group on 14 d(7.70±1.64)and 28(7.90±1.52)(14 d:t=-3.773,P<0.05;28 d:t=-8.104,P<0.05).Western blotting results showed that the expression of p-AMPK in the resveratrol group on 14 d and 28 d(0.528±0.011,0.618±0.010)was significantly enhanced as compared with the control group(0.407±0.013,0.406±0.011)(t=20.266,44.227,P<0.05).Compound C(0.501±0.012)inhibited the expression of p-AMPK(0.704±0.017),and the difference was statistically significant(t=30.914,P<0.05).Conclusion Resveratrol promotes long-term nerve repair after TBI by regulating the AMPK signaling activity.