The effect of electroacupuncture at

doi:10.11958/20220727

Abstract

Abstract:

Objective To investigate the effects of electroacupuncture (EA) stimulation at “Zusanli” and “Neiguan” points on cerebral ischemia and the mTOR signaling pathway in rats. Methods A total of 75 rats were divided into the sham group, the middle cerebral artery occlusion (MCAO) model 3-d group, the EA intervention 3-d group, the MCAO model 7-d group, and the EA intervention 7-d group by random number table method, with 15 rats in each group. Except for the sham group, the permanent MCAO rat model was constructed using the Longa thread embolization method. The EA intervention group was treated at "Neiguan" and "Zusanli" points for 20 min daily after surgery. The neurological deficits were scored at the corresponding time points in each group. 2,3,5-triphenyltetrazolium (TTC) staining was used to analyze the cerebral infarct volume. HE and Nissler staining were used to assess the post-ischemic pathological changes. Western blot assay was used to detect the expression levels of mammalian target of rapamycin (mTOR) pathway-related factors protein kinase B (AKT), p-mTOR and t-mTOR protein in cerebral cortex on the infarct side. Real-time fluorescence quantitative PCR was used to detect the expression levels of LC3B, Beclin-1 and B lymphocytoma-2-associated protein X (Bax) mRNA in cerebral cortex on the infarct side. Results Compared with the sham group, neurological deficit scores at all time points increased in the MCAO group (P<0.05), large necrotic and inflammatory infiltrates in cortical areas, massive neuronal necrosis, disturbed cell arrangement and significantly reduced intact neurons (P<0.05). There were significantly decreased LC3B and Beclin-1 mRNA expression (P<0.05), and Bax mRNA expression was significantly increased in the MCAO model 7-d group (P<0.05). The phosphorylation level of mTOR (p-mTOR/t-mTOR) was significantly higher in the MCAO model 7-d group than that in the MCAO model 3-d group (P<0.05). Compared with the MCAO group, the neurological deficit score was significantly reduced in the EA intervention 7-d group, and the percentage of brain infarct volume was significantly reduced in the EA intervention 3-d group and 7-d group (P<0.05). The necrotic foci were significantly reduced in cortical area, the inflammatory infiltration was improved, the cell arrangement was inregular and the number of intact neurons was significantly increased (P<0.05). Compared with the MCAO model 7-d group, LC3B and Beclin-1 mRNA expression levels were significantly increased (P<0.05), and Bax mRNA, p-mTOR/t-mTOR expression levels were significantly decreased (P<0.05) in the EA intervention 7-d group (P<0.05). There was no significant difference in AKT protein expression between the different groups. Conclusion EA stimulation at “Zusanli” and “Neiguan” points has a neuroprotective effect on cerebral ischemic injury and induces autophagy and inhibits apoptotic responses, which may be related to the inhibition of mTOR signaling pathway activation.

Key words: brain ischemia, electroacupuncture, POINT ST36 (ZUSANLI), POINT PC6 (NEIGUAN), autophagy, Beclin-1, bcl-2-associated X protein, TOR serine-threonine kinases

CLC Number:

R743，R245

Figures/Tables 9

References 29

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基因名称	引物序列（5'→3'）	产物大小（bp）
LC3B	上游：AACACAGCCACCTCTCGACCT	125
	下游：ACACAACCCACACACGGCAG	125
Beclin-1	上游：AGGAGTTGCCGTTGTACTGTTCT	178
	下游：GTGTCTTCAATCTTGCCTTTCTCC	178
Bax	上游：GGGTGGTTGCCCTTTTCTACTT	104
	下游：GAAGTCCAGTGTCCAGCCCAT	104
β-actin	上游：TGCTATGTTGCCCTAGACTTCG	240
	下游：GTTGGCATAGAGGTCTTTACGG	240

基因名称	引物序列（5'→3'）	产物大小（bp）
LC3B	上游：AACACAGCCACCTCTCGACCT	125
	下游：ACACAACCCACACACGGCAG	125
Beclin-1	上游：AGGAGTTGCCGTTGTACTGTTCT	178
	下游：GTGTCTTCAATCTTGCCTTTCTCC	178
Bax	上游：GGGTGGTTGCCCTTTTCTACTT	104
	下游：GAAGTCCAGTGTCCAGCCCAT	104
β-actin	上游：TGCTATGTTGCCCTAGACTTCG	240
	下游：GTTGGCATAGAGGTCTTTACGG	240

组别	术后1 d	术后3 d	术后7 d
Sham组	0.00±0.00	0.00±0.00	0.00±0.00
MCAO模型组	2.70±0.48^a	2.50±0.52^a	2.30±0.48^a
EA干预组	2.80±0.42^a	2.60±0.51^a	1.60±0.51^b
F	184.135^＊＊	119.571^＊＊	83.400^＊＊

组别	术后1 d	术后3 d	术后7 d
Sham组	0.00±0.00	0.00±0.00	0.00±0.00
MCAO模型组	2.70±0.48^a	2.50±0.52^a	2.30±0.48^a
EA干预组	2.80±0.42^a	2.60±0.51^a	1.60±0.51^b
F	184.135^＊＊	119.571^＊＊	83.400^＊＊

组别	脑梗死面积百分比（%）	神经元数量（个/视野）
Sham组	—	542.00±43.13
MCAO模型3 d组	68.32±5.78	245.27±48.84^a
EA干预3 d组	52.91±0.47^b	356.13±82.28^b
MCAO模型7 d组	60.25±2.86	406.27±39.40^a
EA干预7 d组	36.15±1.66^c	500.87±36.23^c
F	84.053^＊＊	75.273^＊＊