| ObjectiveThe mechanism of the specificity of acupoints in central nervous system is one of the most important fields in modern study of acupuncture. But the role of brain function in general controlling has been ignoring while the connection of acupuncture effect and target organ is the major hot spot.The hypothesis of specificity of acupoints related with brain regards that human beings as a wholly creature, the treatment effect comes from the intervention and adjustment of acupuncture which act on target organ through brain as the central process channel.Therefore it is meaningful to study on the biological mechanism of brain function changing during acupuncturefor revealing the laws of acupuncture.To construct the foundation of the hypothesis, the research take brain function image of acupuncture effect literature study in latest decade as a gateway. Summarizing general accepted brain sections of acupuncture effect through Meta-analysis to observe the screen of gene chip of phosphor-express of CREB and related signaling transduction in Wistar rats'brain sections and proposing the effective brain section and materials.MethodReviewing functional magnetic resonance imaging (fMRI) studies of acupuncture related different brain section activity in decade based on Meta-analysis, and screening related 223 papers found in VIP database and CNKI was the first step.Verified 8 papers that qualified on fMRI of healthy people acupuncture operation based on inclusive and exclusive criteria.It was regarded that sections on cerebellum and cerebral cortex would be possible to be the acupoint effective zone based on Mate-analysis.a. Selected 30 SD rats in the study of the effect on different brain section in CREB and CREB-express when needling Waiguan point and divide them into 5 groups, blank group, Waiguan point prompt group, Waiguan point 20min group, non-point prompt group, non-point 20min group. The blank group maintains its original status except catching stimulation. The Waiguan point prompt group took brains out immediately after 20 min's needling on Waiguan Point. The Waiguan point 20 min group took brains out after 20 min's needling and 20 min's rest. The non-point prompt group took brains out immediately after needling non-Waiguan point for 20 min. The non-point 20 min group took brains out after 20 min's needling and 20 min's rest.Sampled brains right after planned intervention and took normal steps in immunohistochemical study, analyzed the positive express of CREB and p-CREB in each sections of groups.b. The intervention of gene expression study to signaling pathway of cerebellum when needling the Waiguan point of the rat was the same as above, and the tonsils of cerebellum were taken out immediately after different interventions (prompt way and 20 min rest). Using Oligo Signal Transduction PathwayFinderTM Microarray from SuperArray Corp. to proceededmicroarray analysis. ObtainedX-ray exposed pictures through refining RNA, RNA quality test, tagging and combining cRNA, and DNA microarray hybridization and then save them as grey levels (8 bit or 16 bit) pictures to analyze data by GEArray Expression Analysis Suite.Results:a. The result showed that there were statistical significance in expression of CREB in cerebellum and cingulate cortex between the Waiguan group and non-point group, and the CREB expression increased in the study of CREB in different brain sections and phosphor-CREB expression when needling SD rat's Waiguanacupoint. But there were not statistical significance in expression of CREB according to acupuncture effect on acupoint and non-point. The p-CREB expression increased remarkably, in both cerebellum and cingulate cortex in the Waiguan group and had statistical significance compared with non-acupoint group meanwhile in 20 min rest groups not. The p-CREB levels were not statistical significance in tensile cerebellum between the Waiguan group and non-point group, and the phosphor-CREB expression in cingulate cortex zone in the Waiguan group had higher level than non-acupoint group.The contour analysis for p-CREB in different brain sections and time points showed that the general contours of the two brain section could be regarded as parallel based on the time and brain section interaction analysis (F=2.325, P=0.246). That mean the reaction of cerebellum and cingulate cortex in three different time points were mainly the same, and the overlap contour analysis showed that the contours of them were not overlap, and the expression in cingulate cortex was higher than cerebellum remarkably.b. The result of study of cerebellum signal channel gene expression showed that needling on Waiguan point activate multi-channels expression immediately, so that increased many gene expressions. No matter in the prompt group,20 min rest or blank group, the performance of No.7 gene is noticeable, as a member of Naip2 NLR family, anti-gene's expression was apoptosisinhibitor protein-2, its signal pathway is survival pathway, it hinted that there was connection between them.The pathways of Wnt, NFκB, Survival signals were different remarkably in acupoint stimulation in the genes'pathways Meta-analysis, and there were at least 4 kinds of related gene pathway expressions. So we can found out from above that the gene expression increased in Wnt, Survival, NFκB pathways in the Waiguan prompt group compared with the non-point prompt group and blank group, all of thismean that there were connections between the three pathways and the effect of acupuncture.Conclusionsa. The meaning of systematic review on brain function imaging study of acupuncture effect might be limited caused by different technical routes, study methods and data collections. But the cerebellum and cingulate cortex are still the focus in theacupoint effect related brain function study based on Meta-analysis. b. Needling Waiguanpoint and non-point in Waiguan nearby area increases the CREB expression in cerebellum and cingulate cortex, but the p-CREB expression increasesremarkably in effect of Waiguan point and non-point in Waiguan nearby area. Therefore the study regards that the biological process is related proteinsphosphorylation if the needling stimulation itself activated the brain section response.c. The CREB phosphorylation pathways is modified by several signal pathways and the p-CREB adjusts gene expressions and transcription of lower level cells, so the forward study focus is up level pathways and lower level targets of CREB phosphorylation in effect brain sections. It shows that there is specific expression of Survival and Wnt pathways in SD rats'cerebellum activation when needling Waiguan point and non-point based on gene chips original data analysis including clustering, factor, and principal components analysis. The literature review shows that both of Survival and Wnt pathway play roles in the adjustment of development and apoptosis of a neuron. Although there is study showed that needling could improve the expressions of the two after brain injury and protect neurons, this study is based on normal SD rats, therefore how them work in target brain section when needling acupuncture is required further research. |
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