| ObjectivesIn this experiment,3,6,and 10-month-old rapidly aging mice SAMP8 were selected as the experimental objects,combined with the idea of"acupuncture and moxibustion for preventing disease",to observe the influence of acupuncture and moxibustion at "BL23","GV20" and "HT7"points on behavior,expression of Aβ1-42 in hippocampal dentate gyrus,expression of hippocampal Tau protein,number of hippocampal dentate gyrus neurons,and hippocampal dentate gyrus neuronal dysplasia,and explore the characteristics of hippocampal dentate gyrus neuronal dysplasia under AD pathology,expected to provide a new entry point for the study of the mechanism of neuronal dysplasia in the formation and development of AD pathology,and provide a new experimental basis for acupuncture as an effective treatment method to improve neuron regeneration barriers to prevent AD.MethodsSAMP8 mice aged 3,6,and 10 months and SAMR1 mice with the same age and the same genetic background were selected as experimental subjects.Three-month-old SAMP8 mice were randomly divided into 3M acupuncture group and 3M model group,each with 6 mice;3M normal group was 63-month-old SAMR1 mice.The 6-month-old SAMP8 mice were randomly divided into 6M acupuncture group and 6M model group,each with 6 mice;6M normal group was 6 6-month-old SAMR1 mice.The 10-month-old SAMP8 mice were randomly divided into a 10M acupuncture group and a 10M model group,with 6 in each group;the 10M normal group was composed of 6 10-month-old SAMR1.3M acupuncture group,6M acupuncture group,and 10M acupuncture group were treated with acupuncture at "GV20" and bilateral "HT7" acupoints,and moxibustion at bilateral "BL23"cupoints,one course of treatment lasted for 6 days and the interval between courses was 1 day,4 consecutive courses were performed.After the course of treatment,the MWM experiment,open field test,and NOR experiment were used to detect the behavioral changes of each group of mice;immunohistochemical method was used to detect the expression of Aβ1-42 and NeuN(mature neuron marker protein)in the hippocampus of each group of mice,and the expression of Tau protein in each group of mice was detected by Western blotting;the hippocampus of each group of mice was detected by immunofluorescence double-labeling method PCNA(proliferating cell nuclear antigen)/NeuN,PCNA/DCX(immature neuron marker protein)positive cells expressed in the dentate gyrus.Results1.Behavioral changes of mice in each group(1)The MWM experiment showed that in the water maze experiment,the average escape latency of the 3M acupuncture group was shorter than that of the 3M model group(P<0.01),and the number of crossing platforms was more than that of the 3M model group(P<0.05).The average escape latency of the 6M acupuncture group was shorter than that of the 6M model group(P<0.01),and the number of crossing platforms was more than that of the 6M model group(P<0.05).The average escape latency of the 10M acupuncture group was shorter than that of the 10M model group,and the number of crossing platforms was more than that of the 10M model group,but the difference was not statistically significant(2)The open filed test showed that the 3M acupuncture group had longer activity time in the central area than the 3M model group(P<0.05).The 6M acupuncture group had longer activity time in the central area than the 6M model group(P<0.01).The activity time of the 10M acupuncture group in the central area was longer than that of the 10M model group(P>0.05).(3)The NOR experiment showed that the novel object recognition coefficient of the 3M acupuncture group was higher than that of the 3M model group(P<0.05).The novel object recognition coefficient in the 6M acupuncture group was higher than that in the 6M model group(P<0.05).The novel object recognition coefficient of the 10M acupuncture group was higher than that of the 10M model group(P>0.05).2.Aβ1-42,NeuN expression in hippocampal dentate gyrus and Tau protein expression in hippocampus of mice in each groupThe results of immunohistochemistry showed that:compared with the 3M normal group,the 3M model group mouse hippocampal dentate gyrus Aβ1-42 average optical density value(MOD)increased(P<0.05),while the NeuN average optical density value decreased(P<0.05);The average optical density value(MOD)of the hippocampal dentate gyrus Aβ1-42 of the 3M acupuncture group was lower than that of the 3M model group(P<0.05),while the average value of NeuN was higher than the 3M model group(P<0.05).Compared with the 6M normal group,the average optical density value(MOD)of Aβ1-42 in the hippocampal dentate gyrus of the 6M model group increased(P<0.01),while the average optical density value of NeuN decreased(P<0.01);The average optical density value(MOD)of Aβ1-42 of the 6M acupuncture group in the dentate gyrus of the mouse hippocampus was lower than that of the 6M model group(P<0.01),while the average optical density value of NeuN was higher than that of the 6M model group(P<0.05).Compared with the 10M normal group,the average optical density value(MOD)of Aβ1-42 in the hippocampal dentate gyrus of the 10M model group increased(P<0.05),while the average optical density value of NeuN decreased(P<0.05);The average optical density value(MOD)of the mouse hippocampal dentate gyrus Aβ1-42 of 10M acupuncture group was lower than the 10M model group(P>0.05),while the NeuN average optical density value was higher than the 10M model group(P>0.05),and the difference was not statistically significant Comparing the aging changes of Aβ1-42 average optical density(MOD)in mice of each month,the results showed:3M normal group<6M normal group<10M normal group;3M model group<6M model group<10M model group;3M acupuncture group<6M acupuncture group<10M acupuncture group.Comparing the aging changes of NeuN mean optical density(MOD)in mice of each month,the results showed:10M normal group<6M normal group<3M normal group;10M model group<6M model group<3M model group;10M Acupuncture group<6M acupuncture group<3M acupuncture group.The results of Western blot experiments showed that,compared with the 3M normal group,the relative expression of Tau protein in the hippocampus of the 3M model group increased(P<0.01);the relative expression of Tau protein in the hippocampus of the 3M acupuncture group was lower 3M model group(P<0.01).Compared with the 6M normal group,the relative expression of Tau protein in the hippocampus of the 6M model group increased(P<0.01);the relative expression of Tau protein in the hippocampus of the 6M acupuncture group was lower than that of the 6M model group(P<0.05).Compared with the 10M normal group,the relative expression of Tau protein in the hippocampus of the 10M model group increased(P<0.01);the relative expression of Tau protein in the hippocampus of the 10M acupuncture group was lower than that of the 10M model group(P<0.05).3.Expression of PCNA/NeuN and PCNA/DCX positive cells in the hippocampal dentate gyrus of each group of miceThe results of immunofluorescence double-label detection showed that the 3M model group had a lower expression of PCNA/NeuN positive cells in the hippocampal dentate gyrus of mice compared with the 3M normal group(P<0.01),while the expression of PCNA/DCX positive cells was increased(P<0.01);compared with the 3M normal group,the expression of PCNA/NeuN positive cells in the hippocampal dentate gyrus of the 3M acupuncture group was higher than that of the 3M model group(P<0.01),while the expression of PCNA/DCX positive cells was higher than that of the 3M model group(P<0.05).Compared with the 6M normal group,the expression of PCNA/NeuN positive cells in the hippocampal dentate gyrus of the 6M model group was lower(P<0.05),while the expression of PCNA/DCX positive cells increased(P<0.01);the 6M acupuncture group had higher expression of PCNA/NeuN positive cells in the hippocampus dentate gyrus than the 6M model group(P<0.01),while the expression of PCNA/DCX positive cells was higher than that of the 6M model group(P<0.01).Compared with the 10M normal group,the expression of PCNA/NeuN positive cells in the hippocampal dentate gyrus of the 10M model group decreased(P<0.01),while the expression of PCNA/DCX positive cells in the hippocampal dentate gyrus of the 10M model group increased(P<0.05));the expression of PCNA/NeuN positive cells in the hippocampal dentate gyrus of the 10M acupuncture group was higher than that of the 10M model group(P>0.05),and the expression of PCNA/DCX positive cells was higher than that of the 10M model group(P>0.05),the difference was not statistically significant.Comparing the aging changes of PCNA/NeuN positive cell expression in mice of each month,the results showed:10M normal group<6M normal group<3M normal group;10M model group<6M model group<3M model Group;10M acupuncture group<6M acupuncture group<3M acupuncture group.Comparing the aging changes of PCNA/DCX positive cell expression in mice of each month,the results showed:10M normal group<6M normal group<3M normal group;10M model group<6M model group<3M model Group;10M acupuncture group<6M acupuncture group<3M acupuncture group.4.Comparison of neurogenesis function of hippocampal dentate gyrus neurons in each group of miceThe positive rate of PCNA/NeuN is recorded as A,the positive rate of PCNA/DCX is recorded as B,C=B/(A+B),the C value represents the function of neurogenesis of the hippocampal dentate gyrus.C3M normal group>C3M acupuncture group>C3M model group;C6M normal group>C6M acupuncture group>C6M model group;C10M normal group>C10M acupuncture group>C10M model group.Comparing the aging changes of C value in each month-old mice,the result showed:C3M normal group>C6M normal group>C10M normal group;C3M model group>C6M model group>C10M model group;C3M acupuncture group>C6M acupuncture group>C 10M acupuncture group.ConclusionWith the increase of months of age,in terms of behavior,the spatial learning and memory ability and working memory ability of SAMP8 mice gradually decreased,and negative emotions such as anxiety and depression appeared.In terms of pathology,the expression of Aβ1-42 in the hippocampal dentate gyrus of SAMP8 mice increased,the increase in the expression of Tau protein in the hippocampus and the decrease in the expression of NeuN in the hippocampal dentate gyrus,thus it was revealed that the pathological changes of SAMP8 mice regarding AD,namely,Aβ plaques,neurofibrillary tangles,and neuronal loss all showed aggravation with aging.The expression of PCNA/NeuN and PCNA/DCX positive cells in the dentate gyrus of the mouse hippocampus decreased with the increase of months,which further indicated that the number of neurons in the dentate gyrus of the hippocampus decreased,and there was a progressive neuron regeneration disorder,revealing the characteristics of the neurogenesis disorder of the hippocampal dentate gyrus under AD pathology,that was,there were too few new neurons to compensate for the lost neurons.Acupuncture treatment could up-regulate the expression of NeuN and DCX in the hippocampal dentate gyrus of SAMP8 mice,increase the number of new neurons,improve hippocampal neuron regeneration,delay the pathological process of AD,and improve the learning and memory ability of SAMP8 mice.And according to the statistical results,it could be determined that the therapeutic effect of acupuncture on SAMP8 mice at the age of 3 months and 6 months was better than that of SAMP8 mice at 10 months. |
PDF Full Text Request