摘要
目的基于GSK3β/Nrf2/HO-1及NF-κB信号通路探讨青藤碱(Sinomenine,SIN)对帕金森病(Parkinson’s disease,PD)小鼠的干预作用及机制。方法将C57BL/6小鼠,随机分为6组:正常组、模型组、阳性药组(左旋多巴,75 mg·kg^(-1))及青藤碱低、中、高剂量组(20、40、80 mg·kg^(-1)),每组8只。腹腔注射20 mg·kg^(-1)1-甲基-4-苯基-1,2,3,6-四氢吡啶(MPTP),每天1次,共造模5 d。在注射MPTP后1 h进行灌胃给药,每天1次,共12 d。在给药第11天对小鼠进行爬杆实验,第12天进行转棒实验,测试小鼠行为学变化。采用ELISA法检测血清肿瘤坏死因子α(TNF-α)、白细胞介素1β(IL-1β)、IL-6含量;RT-qPCR法检测脑组织中TNF-α、IL-1β、IL-6 mRNA表达水平;Western Blot法检测脑组织中TH、Nrf2、HO-1、p-GSK3β、GSK3β、p-IκB、IκB、p-NF-κB、NF-κB蛋白表达水平。结果与正常组比较,模型组小鼠的自动转身潜伏期(T-turn)明显延长(P<0.05),掉落次数显著增多(P<0.001);脑组织中TH蛋白表达水平显著降低(P<0.01),IL-1β、TNF-α、IL-6 mRNA表达水平显著升高(P<0.001);血清TNF-α、IL-1β、IL-6水平明显升高(P<0.05,P<0.01);脑组织中p-GSK3β/GSK3β、Nrf2、HO-1蛋白表达水平显著降低(P<0.05,P<0.001),p-IκB/IκB、p-NF-κB/NF-κB蛋白表达比值显著升高(P<0.001)。与模型组比较,青藤碱中、高剂量组小鼠的T-turn明显缩短(P<0.05,P<0.001),掉落潜伏期明显延长(P<0.05,P<0.01),掉落次数显著减少(P<0.001),脑组织中TH蛋白表达水平显著升高(P<0.01,P<0.001),血清TNF-α水平明显降低(P<0.05);各给药组小鼠脑组织中IL-1β、TNF-α、IL-6 mRNA表达水平均显著降低(P<0.001),血清IL-1β、IL-6水平显著降低(P<0.01,P<0.001),脑组织中p-GSK3β/GSK3β、Nrf2、HO-1蛋白表达水平显著升高(P<0.05,P<0.01,P<0.001),p-IκB/IκB、p-NF-κB/NF-κB蛋白表达比值显著降低(P<0.001)。结论青藤碱可通过调节帕金森病小鼠脑内GSK3β/Nrf2/HO-1和NF-κB通路,增强抗氧化应激能力和抑制神经炎症,从而发挥神经保护作用。
Objective To investigate the intervention effect and mechanism of sinomenine(SIN)on Parkinson’s disease(PD)mice based on GSK3β/Nrf2/HO-1 and NF-κB signaling pathways.Methods C57BL/6 mice were randomly divided into 6 groups:normal group,model group,positive drug group(Levodopa,75 mg·kg^(-1))and SIN low-,medium-and high-dose groups(20,40,80 mg·kg^(-1)),with 8 mice in each group.Mice were intraperitoneally injected with 20 mg·kg^(-1)1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine(MPTP)once a day for 5 days.Intragastric administration was performed 1 hour after injection of MPTP,once a day for 12 days.On the day 11 of administration,the mice were subjected to a pole-climbing experiment,and on the day 12,a rotating rod experiment was performed to test the behavioral changes of the mice.The levels of serum tumor necrosis factor-α(TNF-α),interleukin-1β(IL-1β)and IL-6 were detected by ELISA.The mRNA expression levels of TNF-α,IL-1βand IL-6 in brain tissue were detected by RT-qPCR.The protein expression levels of TH,Nrf2,HO-1,pGSK3β,GSK3β,p-IκB,IκB,p-NF-κB and NF-κB in brain tissue were detected by Western Blot.Results Compared with the normal group,the automatic turning latency(T-turn)of the model group was significantly prolonged(P<0.05),and the number of falls was significantly increased(P<0.001).The expression level of TH protein in brain tissue was significantly decreased(P<0.01),and the mRNA expression levels of IL-1β,TNF-αand IL-6 were significantly increased(P<0.001).The serum levels of TNF-α,IL-1βand IL-6 were significantly increased(P<0.05,P<0.01).The protein expression levels of p-GSK3β/GSK3β,Nrf2 and HO-1 in brain tissue were significantly decreased(P<0.05,P<0.001),and the protein expression ratios of p-IκB/IκB and p-NF-κB/NF-κB were significantly increased(P<0.001).Compared with the model group,the T-turn in the Sin medium-and high-dose groups was significantly shortened(P<0.05,P<0.001),the falling latency was significantly prolonged(P<0.05,P<0.01),the times of falls was significantly reduced(P<0.001),the expression level of TH protein in brain tissue was significantly increased(P<0.01,P<0.001),and the level of serum TNF-αwas significantly decreased(P<0.05).The mRNA expression levels of IL-1β,TNF-αand IL-6 in brain tissue of mice in each administration group were significantly decreased(P<0.001),the serum levels of IL1βand IL-6 were significantly decreased(P<0.01,P<0.001),the protein expression levels of p-GSK3β/GSK3β,Nrf2 and HO-1 in brain tissue were significantly increased(P<0.05,P<0.01,P<0.001),and the protein expression ratios of p-IκB/IκB and p-NF-κB/NF-κB were significantly decreased(P<0.001).Conclusion SIN can enhance anti-oxidative stress and inhibit neuroinflammation by regulating GSK3β/Nrf2/HO-1 and NF-κB pathways in the brain of Parkinson’s disease mice,thereby exerting neuroprotective effects.
作者
张玲玉
何长宏
赵忠正
倪浩杰
易浪
董燕
ZHANG Lingyu;HE Changhong;ZHAO Zhongzheng;NI Haojie;YI Lang;DONG Yan(Science and Technology Innovation Center,Guangzhou University of Chinese Medicine,Guangzhou 510405 Guangdong,China;Dongping County People’s Hospital,Tai’an 271500 Shandong,China)
出处
《中药新药与临床药理》
CAS
CSCD
北大核心
2024年第5期607-614,共8页
Traditional Chinese Drug Research and Clinical Pharmacology
基金
国家自然科学基金项目(81973545)
中药质量研究国家重点实验室(澳门科技大学)开放课题[QRCM(MUST)-2020-2022/2R2102]。
