摘要
目的:观察“智三针”对卒中后抑郁(PSD)模型大鼠海马神经元自噬、炎症水平的影响。方法:63只雄性SD大鼠随机分为7组,每组9只。除假手术组外,其余各组先构建大脑中动脉阻塞再灌注(MCAO/R)模型,术后第4天起采用孤养联合CUMS 21 d构建缺血性PSD模型。造模24 h后,予针刺组“智三针”针刺干预,帕罗西汀组盐酸帕罗西汀(10 mg/kg)灌胃给药,假针刺组非穴不透皮假针干预,3‑MA组自噬抑制剂3‑MA(2.5 mg/kg)腹腔注射,3‑MA+针刺组3‑MA注射后针刺干预。1次/d,共14 d。神经功能缺损评分和糖水偏好率评估大鼠行为学,Western blot检测海马组织Beclin‑1、BNIP3、LC3、NLRP3、NF‑κB、IL‑1β、IL‑18蛋白表达,HE染色观察海马组织病理形态,透射电镜观察海马神经元超微结构和自噬小体。结果:造模后,与假手术组相比,模型组神经功能缺损加重(P<0.001),糖水偏好上升(P<0.01),海马Beclin‑1、BNIP3、LC3、NLRP3、NF‑κB、IL‑1β、IL‑18表达均升高(P<0.05)。干预后,与模型组相比,针刺组神经功能缺损改善(P<0.001),糖水偏好上升(P<0.001),海马Beclin‑1、BNIP3、LC3表达升高(P<0.001),NLRP3、NF‑κB、IL‑1β、IL‑18表达降低(P<0.01);帕罗西汀组神经功能缺损无显著差异(P>0.05),糖水偏好率上升(P<0.001),Beclin‑1、BNIP3、LC3表达升高(P<0.01),NLRP3、NF‑κB、IL‑1β、IL‑18表达降低(P<0.01);假手术组及假针刺组各项指标均无显著差异(P>0.05);3‑MA组神经功能缺损加重(P<0.05),糖水偏好下降(P<0.05),海马Beclin‑1、BNIP3、LC3表达降低(P<0.01),NLRP3、NF‑κB、IL‑1β、IL‑18表达均显著升高(P<0.01);与3‑MA组相比,3‑MA+针刺组神经功能缺损改善(P<0.001),糖水偏好上升(P<0.001),Beclin‑1、BNIP3、LC3表达升高(P<0.001),NLRP3、NF‑κB、IL‑1β、IL‑18表达下降(P<0.01);HE染色及透射电镜观察可见,假手术组海马神经元结构完整,未见自噬小体;PSD模型组和假针刺组海马神经元病理变化明显,自噬小体散在;药物组神经元细胞排列相对整齐,可见自噬小体;3‑MA组海马神经元形态和结构破坏,可见单个自噬小体;针刺组和3‑MA+针刺组海马神经元结构较完整,自噬小体典型可见。结论:“智三针”对缺血性PSD模型大鼠的神经功能损伤和抑郁行为有明显改善作用,其作用机制与激活海马神经元自噬,抑制神经元细胞炎性坏死,改善神经元结构有关。
Objective:To observe the effects of"Zhi san zhen"on on autophagy and inflammation levels in hippocampal neurons of rats with post‑stroke depression(PSD).Methods:63 male SD rats were randomly divided into seven groups,nine in each group.The middle cerebral artery occlusion reperfusion(MCAO/R)model was constructed first in all groups except the sham‑operated group,and the ischaemic PSD model was constructed using solitary support combined with CUMS for 21 d postoperatively onwards from the 4th postoperative day.After the model was successfully constructed,the acupuncture group was treated with acupuncture at"Zhi san zhen"acupoints.The drug group received paroxetine hydrochloride(10 mg/kg)by gavage.The sham acupuncture group received non‑acupoint transdermal sham needle intervention.The 3‑MA group received the autophagy inhibitor 3‑MA(2.5 mg/kg)by intraperitoneal injection.The acupuncture+3‑MA group received 3‑MA injection followed by acupuncture intervention.All interventions were given once daily for 14 consecutive days.Neurological function score and sucrose preference test of rats were detected after intervention;Western blot was used to detect the protein expression levels of Beclin‑1,LC3,BNIP3,NF‑κB,NLRP3,IL‑1β,IL‑18 in hippocampal tissue.HE staining was used to observe the histopathological morphology of the hippocampus,and observe the hippocampal neurons and autophagic vesicles by transmission electron microscopy.Results:The model group had elevated neurological function scores(P<0.001),decreased sucrose preference(P<0.01),and elevated hippocampal expression of Beclin‑1,BNIP3,LC3,NLRP3,NF‑κB,IL‑1β,IL‑18(P<0.05)compared with the sham‑operated group;Compared with the model group,the acupuncture group had lower neurological function scores(P<0.001),higher sucrose preference(P<0.001),higher hippocampal expression of Beclin‑1,BNIP3,LC3(P<0.001),and lower expression of NLRP3,NF‑κB,IL‑1β,IL‑18(P<0.01);the paroxetine group had no significant difference in neurological function scores(P>0.05),while had elevated sucrose preference(P<0.001),elevated expression of Beclin‑1,BNIP3,LC3(P<0.01),and decreased expression of NLRP3,NF‑κB,IL‑1β,IL‑18(P<0.01);No significant difference was observed in all indexes of the sham acupuncture group(P>0.05);the neurological function scores in the 3‑MA group was elevated(P<0.05),sucrose preference rate was reduced(P<0.05),Beclin‑1,BNIP3,LC3 expression was reduced(P<0.01),and NLRP3,NF‑κB,IL‑1β,IL‑18 expression was elevated(P<0.01);Compared with the 3‑MA group,the 3‑MA+acupuncture group had lower neurological function scores(P<0.001),higher sucrose preference(P<0.001),higher expression of Beclin‑1,BNIP3,LC3(P<0.001),and lower expression of NLRP3,NF‑κB,IL‑1β,IL‑18(P<0.01);HE staining and transmission electron microscopy showed that the hippocampal neurons in the sham operation group were structurally intact,and no autophagic vesicles were seen;pathological changes of the hippocampal neurons in the PSD model group and the sham acupuncture group were obvious,and autophagic vesicles were scattered;the neuronal cells in the drug group were arranged in a relatively neat manner,and autophagic vesicles could be seen;morphological and structural disruption of the hippocampal neurons in the 3‑MA group,and single autophagic vesicles could be seen;hippocampal neurons of the acupuncture and the 3‑MA+acupuncture group were more structurally intact.The hippocampal neurons in the acupuncture and 3‑MA+acupuncture group were structurally intact,and autophagic vesicles were typically seen.Conclusion:"Zhi san zhen"could improve neurological dysfunction and depressive behavior in ischemic PSD rats.Its effect might be related to upregulating the level of autophagy in hippocampal neurons of ischemic PSD rats,inhibiting neuronal cell inflammatory necrosis,and alleviating neuroinflammation.
作者
武莉华
高静
王孟雨
傅文
史册
赖名殷
WU Lihua;GAO Jing;WANG Mengyu;FU Wen;SHI Ce;LAI Mingyin(Seventh Clinical Medical College of Guangzhou University of Chinese Medicine,Guangzhou 510006,China;Rehabilitation Center,The First Affiliated Hospital of Henan University of Chinese Medicine,Zhengzhou 450000,China;College of Rehabilitation Medicine,Henan University of Chinese Medicine,Zhengzhou 450046,China;Rheumatology Department,The First Affiliated Hospital of Henan University of Chinese Medicine,Zhengzhou 450000,China;The Third Clinical Medical College,Henan University of Traditional Chinese Medicine,Zhengzhou 450003,China;School of Traditional Chinese Medicine,Hainan Medical University,Haikou 571199,China)
出处
《海南医学院学报》
CAS
北大核心
2024年第13期961-969,共9页
Journal of Hainan Medical University
基金
国家自然科学基金区域科学基金项目(82260968)
海南省自然科学高层次人才基金项目(821RC744)。
关键词
缺血性卒中后抑郁
智三针
细胞自噬
炎性坏死
Ischemic post‑stroke depression
Zhi san zhen
Cellular autophagy
Inflammation necrosis
