| ObjectiveTo investigate the EA (electro-acupuncture, EA) combined with mouse nerve growth factor (nerve growth factor, NGF) on cerebral ischemia and reperfusion injury in rat endogenous neural stem cell proliferation, migration and differentiation. Protection and development of drugs for the brain to provide experimental evidence for clinical application while EA and mouse nerve growth factor promote neurological function after ischemic stroke recovery provide a theoretical basis.MethodsSPF male SD rats 150 prepared by thread embolism of middle cerebral artery occlusion model (middle cerebral artery occlusion, MCAO), were randomly divided into sham operation group (Sham group), model group (MCAO group) EA group (EA group), group of mouse nerve growth factor (NGF group) and EA combined with mouse nerve growth factor group (EA+NGF group), post-infarction and reperfusion injury in neurological deficit scores by tris benzene tetrazolium (TTC) staining cerebral infarct volume; the number of hippocampal neurons Nissl staining; dynamic brain detected by immunohistochemistry in rats after ischemia Article Id,7d and 14d three different time, derived neural stem cell marker protein associated Nestin different brain regions, cells immunoreactive PSA-NCAM, BrdU, GFAP, NeuN and other markers of expression; immunofluorescence double labeled anti-detection marker Nestin/BrdU cells, NeuN/BrdU cells, GFAP/BrdU cells, PSA-NCAM/BrdU cell proliferation BrdU-labeled cells in order to determine, differentiation and migration.Results1, neurologic impairment score:packet ANOVA, F=67.619, P= 0.00＜0.05, illustrate the significant difference between the groups. Time multiple comparisons in 1d,3d,7d between the three time points, in addition to a significant difference between Id and 3d outside (p=0.016＜0.05), no statistically significant difference between the rest of the time point (p＞ 0.05). Multiple comparisons between groups, each group except the model group and EA group (p=0.345), EA group and nerve growth factor group (p=1.000), nerve growth factor group and EA plus NGF group (p=no significant difference between the 0.181), the rest were significant differences between the groups (p＜0.05). Behavior in rats in each group before the experiment normal capacity. Sham group no behavioral abnormalities, neurological score was 0; model group rats eat less, weight loss, reduced activity. Observed to 7d, rat neurological deficits than before symptoms have improved.2, TTC infarct volume Detection:The grouping analysis of variance, P= 0.00＜0.05, illustrate the significant difference between the groups. Multiple comparisons between groups are shown between the groups were statistically significant (p＜0.05). Between each measurement point average pairwise comparison group (ie, were compared Id,3d,7d data among each group) showed P＜0.05, significant difference between the groups. EA group and NGF treatment group after injury score was significantly lower than the model group (P<0.01). Ischemic brain function can cause serious deficiencies, acupuncture and nerve growth factor in cerebral ischemia have a role in recovery.3,The number of Nissl staining neurons:the packet ANOVA, F= 32.184, P =0.000＜0.05, illustrate the significant difference between the groups. Each time points between groups (ie, were compared 1d,7d,14d between the data in each group) showed P＜0.05, significant difference between the groups. Multiple comparisons between groups among the groups model group and nerve growth factor group (p=0.018), electro-acupuncture plus NGF group and the other groups (p=0.00＜0.05) were significantly different between the outside, the rest of the group There was no significant difference between (p＞0.05). Ischemic brain Nissl staining showed that sham group hippocampal CAl area cellular level clear, complete cell structure, uniform cytoplasm and large round nuclei. Model group, ischemic hippocampal CAl area cell disorder, irregular, cell loss, nuclear condensation. EA and NGF treatment group ischemic region corresponding cells arranged relatively regular, normal morphology, cell slightly lost. Compared with cerebral infarction group, EA group, NGF group to reduce the number of hippocampal neurons was significantly (P＜0.05)4, Immunohistochemistry, Immunofluorescence:(1) Nestin positive cells:the packet analysis of variance, P=0.00＜0.05, illustrate the significant difference between the groups. Multiple comparisons between groups were statistically significant among the groups (p＜0.05) between the groups were compared at each time point (ie, were compared Id,7d,14d between the data in each group) showed P＜0.05, each there are significant differences between the groups. The results showed that in the sham group Nestin almost unexpressed; ischemia and over time,7d its positive cells were detected the expression of positive cells found many more (P＜0.05), 14d expression is the best, maximum; Nestin/BrdU-labeled cells was slightly expressed in the sham group, ischemia 7d expression increased (P＜0.05),14d expression peaked. EA group, NGF group, EA+NGF group and model group at 7d, 14d when the differences were statistically significant (P＜0.05);(2) NeuN positive cells:the packet analysis of variance, P=0.00＜0.05, illustrate the significant difference between the groups. Multiple comparisons between groups were statistically significant among the groups (p＜0.05) between the groups were compared at each time point (ie, were compared 1d,7d,14d between the data in each group) showed P＜0.05, each there are significant differences between the groups. NeuN expression in the sham group more ischemia 7d positive cells increased their expression; NeuN/ BrdU-labeled cells in sham operation group was more increased the most obvious of EA group, NGF group, EA+NGF group also had significantly increased model group increased the least;(3) GFAP positive cells:the packet analysis of variance, P=0.00＜0.05, illustrate the significant difference between the groups. Packet analysis of variance, P=0.00＜0.05, illustrate the significant difference between the groups. Each time points between groups (ie, were compared 1d,7d,14d between the data in each group) showed P＜0.05, significant difference between the groups. GFAP expression in the sham group was also a lot; continue to track testing, found after ischemia 7d the positive cells increased (P＜0.05), and is distributed in clusters around a lot longer short of projections,14d start expression decline; GFAP/BrdU-labeled cells are more expressed in the sham group, ischemia 7d expression increased (P＜0.05),14d decreased expression. Compared with model group, EA group, NGF group and EA+NGF group at 7d,14d when decreased expression differences were statistically significant (P ＜0.05);(4)PSA-NCAM positive cells:the packet analysis of variance, P= 0.00＜0.05, illustrate the significant difference between the groups. Multiple comparisons between groups were statistically significant (p＜0.05) between groups. Each time points between groups (ie, were compared Id,3d,7d data among each group) showed P＜0.05, significant difference between the groups. PSA-NCAM expression in the sham group was not obvious, but it is not obvious dyeing; Also in 7d after ischemia detected, the number of the positive cells also increased (P＜0.05), are clustered distribution, there are around long little short of projections,14d expression started to decline; PSA-NCAM/ BrdU-labeled cells was slightly expressed in the sham group, ischemia 7d expression increased (P＜0.05),14d decreased expression. EA group, NGF group, EA+NGF group and model group at 7d,14d when the differences were statistically significant (P＜0.05);Conclusion:1,cerebral ischemia and reperfusion injury in rat brain is cell regeneration and proliferation inducing factor, acupuncture combined with nerve growth factor after the intervention can induce brain cell regeneration, and accelerate the proliferation of new cells and promotes the differentiation of new cells;2, The method of acupuncture and nerve growth factor used in the treatment of ischemic area or endogenous stem cells within the region becomes mature neurons can promote, such treatment of neurological recovery mechanism has important effect. |
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