Effect Of Acupuncture Twisting Reinforcing And Draining Manipulation On Differential Protein Expression In The Parietal Cortex Of SH

ObjectiveIn this study,we selected SHR as the research object,and used the acupuncture twirling reinforcing and twirling reducing method on Taichong(LR3)point.We want to discussed the possible central mechanism of twirling reinforcing and twirling reducing method in treating hypertension from four aspects of general situation,path morphology,molecular biology and proteomics.We observed the effects of twirling reinforcing and twirling reducing method on the blood pressure,pathological of parietal cortex,the expression of IL-6(Interleukin-6)、TNF-α(Tumor necrosis factor alpha)and NGF(Nerve growth factor)in parietal cortex and the influence of differential protein changes.We hoped to provide ideas and references for twirling reinforcing and twirling reducing method in treating hypertension.MethodFifty-six SHR(9 weeks old,male)were selected and randomly divided into model group,twirling reinforcing group,twirling reducing group,electroacupuncture group,14 rats in each group,and 14 WKY(9 weeks old,male)rats as blank group.The model group and the blank group were only grasped and fixed once a day for a total of 14 days.The other three groups respected treated with twirling reinforcing,twirling reducing,electroacupuncture stimulation once a day for a total of 14 days.We used the constant temperature non-invasive blood pressure meter to observe the tail pressure of rats in each group on the day before acupuncture and the 2nd,6th,10th and 14th days after acupuncture.The parietal cortex of rats was taken on the 15th day to make the HE staining to observe the pathological changes of parietal cortex.ELISA was used to detect the changes of IL-6,TNF-α and NGF levels in parietal cortex.Non-labeled quantitative proteomics(label free)technology combined with Parallel Reaction Monitoring(PRM)technology were used to carry out the quantitative and qualitative proteomics research on proteins in parietal cortex of rats in each group,then screen differential proteins and analyze their bioinformatics in parietal cortex of rats in each group.Results1.The systolic blood pressure changes After acupuncture for 14 days:Compared with the blank group,the systolic blood pressure of rats were significantly increased(P<0.01)in model group.Compared with the model group,the systolic blood pressure of rats in the twirling reinforcing group,the twirling reducing group,the electroacupuncture group were significantly decreased(P<0.01),and the twirling reducing group had the fastest antihypertensive effect and the largest antihypertensive amplitude.2.The pathological changes of parietal cortex:In the blank group,the parietal cortex was evenly colored,none abnormality neurons,and blood vessels were not hyperemia.In the model group and the twirling reinforcing group,neurons had different degrees of damage occurred.In the twirling reducing group and the electroacupuncture group,the morphology of neurons was normal and the arrangement of cells were regularly.3.The changes of IL-6,TNF-α and NGF expression in parietal cortex:Compared with the model group,in the twirling reducing group,the twirling reinforcing group,the electroacupuncture group,the levels of IL-6 and TNF-α were significantly decreased(P<0.05)after acupuncture,and the levels of NGF was significantly increased(P<0.01).4.The label free test results of parietal cortex:Compared with the blank group,the model group expressed 1641 differentially proteins(601 up and 1040 down).Compared with the model group,there were 72 differential proteins(44 up and 28 down)in the twirling reinforcing group,there were 1043 differential proteins(616 up and 427 down)in the twirling reducing group,there were 1286 differential proteins(800 up and 486 down)in the electroacupuncture group.The results of biological information analysis showed that the pathogenesis of hypertension and the mechanism of acupuncture antihypertensive action are mainly closely related to biological processes such as cellular processes,biological regulation,metabolic processes,cellular components,cellular parts,organelles,junctions and protein binding.The enrichment of KEGG pathway showed that 42 pathways were enriched by differential proteins in the blank group and the model group,4 pathways were enriched by differential proteins in the twirling reinforcing group and the model group,56 pathways were enriched by differential proteins in the twirling reducing group and the model group,and 43 pathways were enriched by differential proteins in the electroacupuncture group and the model group.5.The PRM validation results of key protein in parietal cortex:The PRM validation results of 10 key proteins were consistent with the label free expression trends.Conclusion1.Acupuncture at LR3 combined with twirling reinforcing and twirling reducing manipulation has obvious antihypertensive effect on SHR rats.2.Hypertension can bring certain damage on parietal cortex of rats,while different acupuncture methods can reduce brain injury.3.The mechanism of acupuncture on antihypertensive may be closely related to inhibiting neuroinflammatory reaction and promoting neuronal repair.4.Acupuncture at LR3 combined with twirling reinforcing and twirling reducing manipulation can change protein expression in parietal cortex,and its mechanism may be related to Ras signal pathway,PI3K-Akt signal pathway,mTOR signal pathway and MAPK signal pathway.