Mechanism of"Zhi san zhen"inhibited hippocampal neuronal inflammatory necrosis through regulating autophagy in rats with ischemic post-stroke depression
Objective:To observe the effects of"Zhi san zhen"on on autophagy and inflammation levels in hippocampal neu-rons 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 postop-eratively 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 acupunc-ture intervention.All interventions were given once daily for 14 consecutive days.Neurological function score and sucrose prefer-ence 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 su-crose preference(P＜0.001),higher hippocampal expression of Beclin-1,BNIP3,LC3(P＜0.001),and lower expression of NL-RP3,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 de-creased 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 ex-pression was elevated(P＜0.01);Compared with the 3-MA group,the 3-MA+acupuncture group had lower neurological func-tion 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 vesi-cles 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 hip-pocampal neurons in the acupuncture and 3-MA+acupuncture group were structurally intact,and autophagic vesicles were typical-ly 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 in-flammatory necrosis,and alleviating neuroinflammation.
Ischemic post-stroke depressionZhi san zhenCellular autophagyInflammation necrosis
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