摘要
目的基于NR/CaMKII通路探讨壮骨止痛解郁方改善类风湿性关节炎并发抑郁症(Rheumatoid arthritis-related depression,RAD)大鼠海马神经元突触损伤的作用机制。方法复制RAD大鼠动物模型及模拟RAD环境的体外细胞模型,实验设正常组、RA组、模型组、NR阻断剂组和壮骨止痛解郁方组。采用足趾容积测定仪检测大鼠足肿胀程度;旷场试验和水迷宫试验检测大鼠抑郁样行为;高尔基染色和透射电镜检测大鼠海马神经元树突、树突棘及突触亚微结构;细胞成像分析检测体外大鼠滑膜细胞和海马神经元形态;尼氏染色检测体外海马神经元树突及树突棘形态结构;免疫荧光染色联合激光共聚焦显微镜检测大鼠海马组织和体外海马神经元中谷氨酸受体2A(NR2A)、谷氨酸受体2B(NR2B)、钙调蛋白激酶II(CaMKII)、脑源性神经营养因子(BDNF)、突触后致密蛋白95(PSD-95)表达情况。结果动物实验结果表明,壮骨止痛解郁方能有效改善RAD大鼠类风湿性关节程度和抑郁样行为,抑制谷氨酸NR2A和NR2B受体异常活化,上调突触相关蛋白CaMKII、BDNF及PSD-95表达,显著减轻大鼠海马神经元树突、树突棘及突触亚微结构损伤。体外细胞实验亦显示,壮骨止痛解郁方含药血清能明显保护大鼠滑膜细胞和海马神经元,同时改善海马神经元突触损伤,并抑制NR2A、NR2B蛋白上调,CaMKII、BDNF及PSD-95蛋白下调。结论从“体内-体外两方面,行为-形态-功能-分子四水平”发现壮骨止痛解郁方可能是通过调控NR/CaMKII信号通路,抑制海马神经元突触损伤,进而改善大鼠抑郁样行为,这可能是其防治RAD的重要机制。
Objective This study aimed to investigate the mechanism of Zhuanggu Zhitong Jieyu Formulation on alleviating hipp⁃ocampal neuron synapsis impairment based on NR/CaMKII pathway in rheumatoid arthritis-related depression.Methods RAD model in vivo and in vitro was both established.The rats or the cultured cells were randomly divided into control group,model group,NR blocker group,and Zhuanggu Zhitong Jieyu Formulation group.The arthritis severity of rats was evaluated by hind paw volume.Depressive-like behaviors of rats was assessed by open field test and morris water maze test.The impairment of hippocampal neuron synaptic structure in rats,including dendrites,dendritic spines and synaptic submicrostructure,was evaluated by Golgi staining as well as transmission electron microscopic analysis.The cell morphology of synovial cells and hippocampal neu⁃rons were analyzed by cell imaging analysis.The morphology and structure of dendrites and dendritic spines of hippocampal neu⁃rons in vitro was tested by Nissl's staining.The protein expression of NR2A,NR2A,CaMKII,BDNF,and PSD-95 were detec⁃ted by immunofluorescent staining and laser scanning confocal microscope.Results Our results shown that Zhuanggu Zhitong Jieyu Formulation obviously ameliorated the arthritis severity and depressive-like behaviors(P<0.05,0.01),and significantly restrained the abnormal activation of glutamate NR2A and NR2B receptors(P<0.05、0.01)and up-regulated the synapsis-re⁃lated CaMKII,BDNF and PSD-95 protein expression(P<0.05,0.01),and further relieved the impairment of hippocampal neuron synaptic dendrites,dendritic spines and synaptic submicrostructure in RAD rats(P<0.01).Additionally,our results in vitro also indicated that Zhuanggu Zhitong Jieyu Formulation exhibited a protective effects against damage of synovial cells and hip⁃pocampal neurons,markedly alleviated hippocampal neuron synapsis dysfunction,and finally suppressed the aberrant expression of synapsis-related protein NR2A,NR2ACaMKII,BDNF and PSD-95(P<0.05,0.01).Conclusion Collectively,the current findings revealed that Zhuanggu Zhitong Jieyu Formulation could ameliorated the impairment of hippocampal neuron synapsis and further relieved the depressive-like behaviors via regulating the NR/CaMKII pathway,which might be the important mechanism of Zhuanggu Zhitong Jieyu Formulation for the prevention and treatment of RAD.
作者
刘检
蔺晓源
赵洪庆
唐林
杨蕙
王宇红
刘平安
LIU Jian;LIN Xiao-yuan;ZHAO Hong-qing;TANG Lin;YANG Hui;WANG Yu-hong;LIU Ping-an(First Hospital of Hunan University of Chinese Medicine,Changsha 410007,China;Hunan Key Labora-tory of Traditional Chinese Medicine Prevention&Treatment of Depressive Diseases,Changsha 410208,China;National Key Laboratory Cultivation Base of Chinese Medicinal Powder&Innovative Medicinal Jointly Estab-lished by Province and Ministry,Changsha 410208,China;Science&Technology Innovation Center,Hunan University of Chinese Medicine,Changsha 410208,China;Hunan Academy of Chinese Medicine,Changsha 410006,China)
出处
《时珍国医国药》
CAS
CSCD
北大核心
2023年第8期1803-1809,共7页
Lishizhen Medicine and Materia Medica Research
基金
国家自然科学基金青年项目(82104793)
湖南省自然科学基金面上项目(2022JJ30451)
湖南省发展和改革委创新研发项目(201930)
湖南省中药粉体与创新药物省部共建国家重点实验培育基地开放基金资助项目(21PTKF1008)
湖南中医药大学中医学一流学科开放基金重点项目(2022ZYX16)
湖南省三尖计划青年科技人才项目(2022RC1226)。
关键词
壮骨止痛解郁方
类风湿性关节炎
抑郁症
突触
NR/CaMKII通路
Zhuanggu Zhitong Jieyu Formulation
Rheumatoid arthritis
Depression
Synapsis
N-methyl-D aspartate receptors-calmodulin-dependent protein kinase II pathway
