Objective

2

To observe the effect of electroacupuncture at "Sishencong" (EX-HN1) and "Fengchi" (GB20) on hippocampal neuronal ferritinophagy mediated by the nuclear receptor coactivator 4 (NCOA4)/ferritin heavy chain 1 (FTH1) signaling pathway in vascular dementia (VD) rats

and to explore the potential mechanisms of electroacupuncture for VD.

Methods

2

A total of 60 male rats of SPF grade were randomly divided into a blank group (12 rats)

a sham surgery group (12 rats) and a modeling group (36 rats). In the modeling group

the modified 4-vessel occlusion method was used to establish the VD model. The 24 successfully modeled rats were randomly divided into a model group and an electroacupuncture group

with 12 rats in each group. In the electroacupuncture group

electroacupuncture was applied at left and right "Sishencong" (EX-HN1)

and bilateral "Fengchi" (GB20)

with continuous wave

in frequency of 2 Hz and current intensity of 1 mA

30 min a time

once daily for 21 consecutive days. The learning and memory abilities were assessed using the Morris water maze test before modeling

after modeling and after intervention

as well as the novel object recognition test after intervention. After intervention

the neuronal morphology in the hippocampus was observed by Nissl staining; the iron deposition was observed by Prussian blue staining; the reactive oxygen species (ROS) level was detected by dihydroethidium (DHE) fluorescence staining; the levels of iron

malondialdehyde (MDA) and superoxide dismutase (SOD) in the hippocampal tissue were measured by the colorimetric assay

TBA method

and WST-1 method

respectively; the positive expression of NCOA4

FTH1 and glutathione peroxidase 4 (GPX4) was detected by immunohistochemistry; the protein expression of NCOA4

FTH1

GPX4

and the ratio of microtubule-associated protein 1 light chain 3B (LC3B) Ⅱ/Ⅰ in the hippocampus were detected by Western blot.

Results

2

Compared with the sham surgery group

in the model group

the escape latency was prolonged

and the number of platform crossings reduced (

P

<

0.01)

the recognition index (RI) was decreased (

P

<

0.01); the hippocampal neurons displayed a blurred laminar structure

disorganized cellular arrangement

and the number of Nissl bodies was decreased (

P

<

0.01); the percentage of iron deposition area in the hippocampus was increased (

P

<

0.01); in the hippocampus

the levels of ROS

iron

MDA

and the protein expression of NCOA4

as well as the LC3B Ⅱ/Ⅰ ratio were increased (

P

<

0.01)

the SOD level

and the protein expression of FTH1 and GPX4 were decreased (

P

<

0.01). Compared with the model group

in the electroacupuncture group

the escape latency was shortened and the number of platform crossings was increased (

P

<

0.01)

the RI was increased (

P

<

0.01); the hippocampal neurons exhibited more regular morphology

better-organized cellular structure

and the number of Nissl bodies was increased (

P

<

0.05); the percentag

e of iron deposition area in the hippocampus reduced (

P

<

0.01); in the hippocampus

the levels of ROS

iron

MDA

and the protein expression of NCOA4

as well as the LC3B Ⅱ/Ⅰ ratio were decreased (

P

<

0.01

P

<

0.05)

the SOD level

and the protein expression of FTH1 and GPX4 were increased (

P

<

0.01).

Conclusion

2

Electroacupuncture at "Sishencong" (EX-HN1) and "Fengchi" (GB20) can improve learning and memory abilities in VD rats

and its mechanism may be associated with the regulation of the hippocampal NCOA4/FTH1 signaling pathway

inhibition of ferritinophagy

and alleviation of oxidative stress damage.