The Research About Effection Mechanism Of Moxa Cone Moxibustion On Cerebral In Senile Mice Induced By D-galactose

ObjectiveTo explore the mechanism of moxa cone moxibustion in anti-aging on cerebral with senile mice induced by D-galactose injection.MethodsAfter two separately selecting with the Morris water maze(MWM) test,72male mice(3 months old) were randomly and equally divided into the normal sodium control,model,moxa cone moxibustion 1(acupoint ST36 and GB39),moxa cone moxibustion 2(acupoint DU20 and RN4),electroacupunctue(acupoint ST36 and GB39),Nimodipine medicated groups.The treatment and model groups were given with D-galactose injection for 42days,while the normal sodium control group was injected the same dosage of normal sodium as other groups for 42 days. From the 13^th day,the four treatment groups was treated for every other day until the model would had been finished.The six group were tested on Spatial Learning and Memory in the MWM again,and then take cerebral tissues for tissue fixation or tissue homogenate.Measured and observed the following the indexes.(1)The Spatial Learning and Memory before and after the modeled and treated with computer MWM system in six group.Including the place navigation with four targets that are escape latency,detention(platform) time,seeking (platform) frequency,percentage of platform quadrant(the percentage of swimming distant between the original platform and total distant)from the 2^th to 6^th day and the spacial probe test with two targets that are spanning (original) platform frequency and percentage of(original) platform quadrantin at the 7^th day.(2)The Pathomorphological of changes on the cerebral tissues and neurons density of cerebral cortex and hippocampal(CA3) with light-microscope in each group.(3)The ultrastructure change of the cerebral cortex in each group.(4)The Nitric Oxide(NO) content of the cerebral tissues in each group.(5)The total Nitric Oxide synthase(TNOS)activities and the induced Nitric Oxide synthase(iNOS) activities of the cerebral tissues in each group.(6)The total Superoxide dismutase(TSOD) activities and the CuZnSuperoxide dismutas(CuZnSOD) activities of the cerebral tissues in each group(7)The c-fos gene expression of the cerebral cortex in each group(8)The p16 gene expression of the cerebral cortex in each group.(9)The(Phospho- Retinoblastoma) pRb gene expression of the cerebral cortex in each group.(10)The c-fosmRNA expression of the cerebral cortex and hippocampal(CA3) in each groupResults(1)six groups mice were equal on spatial learning and memory before the experiment(P＞0.05),after modeled and treated,①)The significant difference was observed between the modeled group and the other five groups on spatial learning and memory(P＜0.01或P＜0.05),Comparing with the other five groups, the escape lantency of modeled group was prolonged and the seeking frequency, detention time,percentage of platform quadrant,the spanning(original) platform frequency,the percentage of(original) platform quadrant were decreased②There were significant difference between the Nimodipine medicated group and the normal sodium control group in escape lantency,percentage of platform quadrant(P＜0.05),the escape lantency of the moxa cone moxibustion 1,the moxa cone moxibustion 2,the Dianzhen ones were longer than that of the normal sodium control significantly(P＜0.05).The other indexes in five group are no differences.(2)With Hematoxylin and eosin(H·E) staining under the light-microscopeThe cerebral cortex and hippocampal neurons struacture of the normal sodium control group was normal,nucleus with Purple blue,rich cytoplasm with slight purple,without Swelling in Mesenchymal,on the other hand,the model group had similar change with the physio-senility mices,the cerebral cortex was thin with degeneration,the neurons become smaller,sparse,nuclei and cytoplasm are muddled,pyknosis showing dark blue.The Neuronal density in the model group was lower than that of other five group(p＜0.01).The four treated groups had the neurons degeneration in varying degrees,but the neurons degeneration was lightented than that of the model group.(3)The Ultrastructure of the cerebral cortex in normal sodium control group was better with clear neurons structure,rich organelle,showing more mitochondria,rough endoplasmic reticulum,golgi apparatus,ribosomes,etc., loose chromatin,nucleolus clearly.A small amount of phagocytic lysosome without swelling and other changes.There was axonal structure clearly, tubulin and microfilament rich neatly arranged.The model group was increased in the number of mitochondriawith extended or vacuolar,mitochondria ridge vague,detached ribosomes.Axonal was swelling and vacuolization,tubulin and microfilament were reducing in the number with scattered.The Part of myelin structure was loss,showing more phagocytic lysosomes(lipofuscin).The Part of the vascular endothelial cells have degeneration,perivascular space expansion,stellatecell vacuolar withthe foot process was damage obviously. The neurons of four treated groups had degeneration in varying degrees,with the order by lesions sorting from heavy to light being the Nimodipine medicated,the moxa cone moxibustion 2,the electroacupunctue,the moxa cone moxibustion 1.(4)There were significant difference on the NO content of cerebral tissues in six group(p＜0.01),the NO content of cerebral tissues in model group was higher than that of in other five group(p＜0.01).There was no difference between other five group.(5)There were significant difference on the TNOS,iNOS activities of cerebral tissues in six group(p＜0.01),the TNOS,iNOS activitiesof cerebral tissues in model group was increased than that of in other five group (p＜0.01).There was no difference between other five group.(6)There were significant difference on the TSOD,CuZnSOD activities of cerebral tissues in six group(p＜0.01),the TSOD,CuZnSOD activities of cerebral tissues in model group was lower than that of in other five group (p＜0.01).There was no difference between five others on TSOD activity.The CuZnSOD activities in the Nimodipine medicated were lower than that of the moxa cone moxibustion land2(p＜0.01,p＜0.05),and The CuZnSOD activity in the electroacupunctue were lower than that of the moxa cone moxibustion 2 (p＜0.05),there were no difference between others on CuZnSOD activity. (7)There were c-fos,p16,Phospho-Rb on the cerebral cortex in each group. The neurons with c-fos,Phospho-Rb protein expression on the cerebral cortex in model group was scatter and slight staining,The Cells with c-fos,Phospho-Rb protein expression were more closely and deeper staining in other fives.The neurons with p16 Protein expression on the cerebral cortex in model group was deeper staining,but in others the Cells with p16 protein expression were slightly staining.The neuronal density of c-fos,p16,Phospho-Rb on the cerebral cortex had significant difference on six groups.The neuronal density of c-fos,Phospho-Rb on the cerebral cortex on the model group were decreased compared with other groups(p＜0.01,p＜0.05),the neuronal density of p16 on the cerebral cortex was increased compared with other groups(p＜0.01, p＜0.05).The neuronal density of c-fos on the cerebral cortex of the Nimodipine medicated was significant difference compared with the normal sodium control group and the moxa cone moxibustion 1.There were no differences on other indexes between treatment groups.(8)There were c-fosmRNA expression on the cerebral cortex and hippocampal (CA3) in each group.The neurons with c-fosmRNA expression on the cerebral cortex and hippocampal(CA3) in model group was scatter and slight staining, in others group the cells with c-fosmRNA expression were more closely and deeper staining.The neuronal density of c-fosmRNA on the cerebral cortex and hippocampal on the model were decreased compared with other groups(p＜0.01, p＜0.05).There were no differences on neuronal density of the cerebral cortex c-fosmRNA between five treatment groups.The neuronal density of c-fosmRNA expression on hippocampal in the Nimodipine medicated were lower than that of in the moxa cone moxibustion 1,no differences between four other treatment groups.Conclusion(1)The moxa cone moxibustion therapy on ST36,GB39 can improve the spatial learning and memory of aging mice induced by D-galactose.(2)The moxa cone moxibustion therapy on ST36,GB39 can Protect the cerebral tissues and ultrastructural morphology of aging mice induced by D-galactose, reduce oxidative damage and protect neurons.(3)The moxa cone moxibustion therapy on ST36,GB39 can resist free radicals damage in aging brain tissue,improve brain tissue antioxidant capacity. (4) The moxa cone moxibustion therapy on ST36,GB39 can inhibit p16 protein expression on the cerebral cortex,promote c-fos,Phospho-Rb protein expression on on the cerebral cortex.(5) The moxa cone moxibustion therapy on ST36,GB39 can improve c-fosmRNA expression on the cerebral cortex and hippocampal.