| Objectives:To observe the effects of Buyang Huanwu Decoction(BYHWD)on neurobehavior and neuronal apoptosis in rats with vascular dementia(VD)model.The expression of insulin-like growth factor-1(IGF-1)-Phosphatidylinositol-3-kinase(PI3K)-protein kinase B(Akt)-Fox O1(Fox O1)signaling pathway related proteins was detected to explore the intervention effect and possible mechanism of Buyang Huanwu Decoction on VD.To observe the effects of Buyang Huanwu Decoction on neurobehavior and neuronal apoptosis in rats with vascular dementia(VD)model.The expression of insulin-like growth factor-1(IGF-1)-Phosphatidylinositol-3-kinase(PI3K)-protein kinase B(Akt)-Fox O1(Fox O1)signaling pathway related proteins was detected to explore the intervention effect and possible mechanism of Buyang Huanwu Decoction on VD.Methods:The animal model of vascular dementia was established by the modified bilateral common carotid artery ligation method,and the rat model was successfully established by Morris water maze screening.Sham operation(Sham)group,model(VD)group,Nimodipine 0.0126 g/kg/d(Nim)group,Buyang Huanwu Decoction 7.5g/kg/d(BYHWD-L)group,Buyang Huanwu decoction 15 g/kg/d(BYHWD-M)group and Buyang Huanwu Decoction 30 were set up g/kg/d(BYHWD-H)groups with 12 mice in each group.Administration group was given gavage at the above dosage,Sham group and model group were given gavage with equal volume of normal saline,once a day,for 30 days.Detect spatial learning and memory abilities of rats in each group through Morris water maze.Morphological changes of neurons in hippocampus CA1 and cortex were observed by HE staining and Nissl staining.The apoptosis of hippocampal CA1 and cortical neurons in rats was observed by d UTP notch end labeling(TUNEL)mediated by terminal deoxynucleotide transferase.The concentrations of IGF-1 in serum,hippocampus and cortex were determined by enzyme-linked immunosorbent assay(ELISA).The expressions of neuronal nuclear antigen(Neu N),IGF-1,insulin-like growth factor-1 receptor(IGF-1R),B-lymphoblastoma-2-associated x protein(Bax)and B-lymphoblastoma-2 protein(Bcl-2)in hippocampus and cortex were observed by immunohistochemistry.Real-time fluorescence quantitative polymerase chain reaction(RT-q PCR)was used to detect the expression of IGF-1 m RNA,PI3 K m RNA,Akt m RNA and Fox O1 m RNA in hippocampus and cortex.The expressions of IGF-1,IGF-1R,PI3 K,Akt,Fox O1,Bax and Bcl-2 in the hippocampus and cortex of rats were determined by Western blot.Results:1.Effects of Buyang Huanwu Decoction on neurobehavior,brain histomorphology and IGF-1 concentration in VD model rats: Compared with sham operation group,the average escape latency and first crossing time of rats in model group were significantly prolonged,while the number of crossing platforms and quadrant residence time of rats in model group were reduced(P < 0.05 or P < 0.01),and the swimming track showed a blanket search without goals.The number of neurons in hippocampal CA1 region and cortical region decreased,showing vacuole-like degeneration,chromatin lysis,and even cell necrosis.The Nissl staining was weak,and the cells showed hyperbasophilic behavior due to shrinkage.Neu N protein expression decreased(P < 0.01);The concentrations of IGF-1 in serum,hippocampus and cortical tissue were significantly decreased(P < 0.01).Compared with model group,Buyang Huanwu decoction groups and nimodipine groups could significantly shorten the escape latency and the time of first crossing the platform,increase the number of crossing the platform and the residence time in the target quadrant(P < 0.05 or P < 0.01),and the swimming trajectory was concentrated in the original platform quadrant,there was no significant difference in swimming speed among each group of rats.The number of neurons in hippocampal CA1 region and cortical region increased(P < 0.05 or P < 0.01),and the vacuolar degeneration and shrinkage necrosis of cells were significantly improved.The nissite stain was deep and homogeneous.The expression of CA1 protein in hippocampus and Neu N protein in cortex of rats in Buyang Huanwu Decoction medium and high dose groups and Nimodipine groups was significantly increased(P < 0.05 or P < 0.01),and the concentration of IGF-1 in serum,hippocampus and cortical tissue was significantly increased(P < 0.05 or P < 0.01).2.Effects of Buyang Huanwu Decoction on neuronal apoptosis in hippocampal CA1 region and cortical region of VD model rats: Compared with sham operation group,the number of apoptotic neurons in hippocampal CA1 and cortical regions of rats in model group was increased,the apoptotic index was significantly increased(P < 0.01),the expression of Bax protein was increased(P < 0.01),and the expression of Bcl-2 protein was significantly decreased(P < 0.01).Compared with model group,the number of apoptotic neurons in hippocampal CA1 and cortex of rats in Buyang Huanwu Decoction groups and nimodipine groups decreased,and the apoptotic index decreased to different degrees(P < 0.05 or P < 0.01).The expression level of Bax protein was significantly down-regulated(P < 0.05 or P < 0.01),while the expression level of Bcl-2 protein was significantly up-regulated(P < 0.05 or P <0.01).3.Effects of Buyang Huanwu Decoction on expression of IGF-1-PI3K-Akt-Fox O1 signaling pathway related proteins in VD model rats:Compared with sham operation group,the protein expression levels of IGF-1,IGF-1R,PI3 K,Akt,Bcl-2 and m RNA expressions of IGF-1,PI3 K and Akt in hippocampus and cortex of rats in model group were significantly down-regulated(P < 0.01).The expression levels of Fox O1 and Bax protein and Fox O1 m RNA were up-regulated(P< 0.01),while Bax/Bcl-2 was significantly increased(P < 0.01).Compared with model group,protein expression levels of IGF-1,IGF-1R,PI3 K,Akt,Bcl-2 and m RNA expressions of IGF-1,PI3 K and Akt in hippocampus and cortex of rats in Buyang Huanwu Decoction medium and high dose groups and nimodipine groups were significantly up-regulated(P < 0.05 or P < 0.01).Fox O1 and Bax protein expression levels and Fox O1 m RNA expression were significantly decreased(P <0.05 or P < 0.01),and Bax/Bcl-2 was significantly decreased(P < 0.05 or P < 0.01).Conclusions:Buyang Huanwu Decoction can improve the learning and memory ability of VD model rats to a certain extent,and its mechanism may be to inhibit neuronal apoptosis by regulating IGF-1-PI3K-Akt-Fox O1 signal pathway,thus playing a neuroprotective role. |
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