| Objective:In this study,S100A8/A9 was screened as a key differential protein for Tuina therapy in MPS through proteomics experiments,and the S100A8/A9/TLR4 signaling pathway was selected as the starting point.Animal experiments and clinical trials were conducted to study and analyze the therapeutic effects of Tuina on MPS and its related biological mechanisms in resolving adhesions and relieving pain in traditional Chinese medicine,in order to provide high-quality research evidence for Tuina therapy in treating MPS.Method:(1)A study on the effect of Jiejie Chubi Tuina on the repair quality of skeletal muscles and spontaneous electrical activity in MPS rats: after 7 days of adaptive feeding,6 rats with poor swimming ability or inability to swim were selected as the blank group,and the remaining rats were used for animal modeling.After modeling,18 rats were randomly selected and divided into the model group,control group,and Tuina group for inclusion in the experimental study,with 6 rats in each group.MPS modeling was performed using blunt contusion combined with exhaustive exercise.After successful modeling,the blank group and model group were fed routinely without any treatment,the Tuina group was treated with massage therapy for 10 minutes once a day for a total of 7 days,and the control group was treated with fasciotome therapy for 5 minutes once every other day for a total of 7 days.The same intensity and time of touch stimulation were given to the Tuina group and the control group during each treatment;the blank group was only given the same intensity and time of touch stimulation as the Tuina group every day.After 24 hours of modeling,the rats in each group were subjected to electromyography(EMG)testing to verify the presence of spontaneous electrical activity(SEA)after modeling.After 7 days of intervention,SEA wave amplitude was collected from each group of rats to evaluate the spontaneous electrical potential of the pain points after treatment.HE staining and Masson staining were used to observe the modeling and muscle repair in each group of rats.(2)Study on the effect of Jiejie Chubi Tuina on skeletal muscle proteomics in MPS rats:after 7 days of adaptive feeding,6 rats with poor swimming ability or inability to swim were selected as the blank group,and the remaining rats were used for animal modeling.After modeling,18 rats were randomly selected and divided into model A group(immediately sampled after successful modeling),model B group(no intervention after successful modeling and continued feeding),and Tuina group(intervention for 7 days after successful modeling)for experimental research.The modeling method and massage treatment method are the same as(1).After 7 days of intervention,proteomic technology was used to detect the differentially expressed proteins and key molecular pathways focusing on differential proteins and key molecular pathways through GO enrichment analysis,KEGG pathway enrichment,and protein structure domain enrichment,and proteins related to MPS were selected for PRM verification.(3)Study on the effect of Jiejie Chubi Tuina on S100A8/A9/TLR4 signaling pathway in MPS rats: the modeling and intervention methods are the same as(1).After 7 days of intervention,Western Blot was used to detect the protein content of S100A8,S100A9,TLR4,CD14,p38,p-p38,JNK and p-JNK in the skeletal muscle of the four groups of rats.PCR was used to detect the m RNA expression of S100A8,S100A9,TLR4,and CD14 in the skeletal muscle of the four groups of rats.ELISA was used to detect the content of inflammatory factors S100A8,S100A9,TNF-α and IL-6 in the serum of the four groups of rats.(4)Clinical exploratory study on the therapeutic effect of Jiejie Chubi Tuina on myofascial pain of nape: 72 patients diagnosed with myofascial pain syndrome in the upper back were randomly divided into a Tuina group and a control group using a random number table.The control group received myofascial release combined with Mc Kenzie therapy for20 minutes per session,three times a week,once every other day for a total of 2 weeks.The Tuina group received only massage therapy for 20 minutes per session,once a day,for a total of 2 weeks.The simplified Mc Gill pain questionnaire,Neck Disability Index(NDI),and cervical range of motion were used to evaluate the pain and neck function of the two groups of patients before treatment(-1-0d),after the first course of treatment(1-2d),and after the second course of treatment(1-2d).The expression level of serum inflammatory factors in patients was observed and tested before treatment(-1-0d),after the first course of treatment(1-2d),and after the second course of treatment(1-2d).Results:(1)Study on the effect of Jiejie Chubi Tuina on the repair quality of skeletal muscle and spontaneous potential in MPS rats: after 7 days of intervention,HE staining results of each group of rats showed that in the model group,there were more muscle cells necrosis,nuclear fragmentation,dissolution,presenting acidophilic homogeneous substance,and accompanied by a large number of inflammatory cell infiltration at the edge of muscle tissue.In the control group,local muscle cells were irregularly arranged with irregular shapes,limited amount of interstitial collagen fiber hyperplasia and inflammatory cell infiltration.A limited number of newly generated muscle cells were visible,with small cell volume,multiple nuclei in cytoplasm,and nuclei located in the center.In the Tuina group,there was minute quantities of interstitial collagen fiber hyperplasia and inflammatory cell infiltration.A great quantity of newly generated muscle cells were visible,with small cell volume,multiple nuclei in cytoplasm,and nuclei located in the center.Masson staining results of each group of rats showed that compared with the blank group,the integral optical density(IOD)and area ratio of collagen fibers in the model group increased significantly(P<0.05,P<0.01).Compared with the model group,the IOD and area ratio of collagen fibers in the Tuina group decreased significantly(P<0.05,P<0.01).Compared with the model group,the area ratio of collagen fibers in the control group decreased significantly(P<0.01).The IOD of collagen fibers decreased,but there was no significant difference(P>0.05).Compared with the control group,the IOD and area ratio of collagen fibers in the Tuina group decreased,but there was no significant difference(P>0.05).Electromyogram results of each group of rats showed that compared with the model group,the amplitude of SEA wave in the Tuina group and the control group decreased significantly(P<0.01).Compared with the control group,the amplitude of SEA wave in the Tuina group significantly decreased(P<0.01).(2)Study on the effect of Jiejie Chubi Tuina on the proteomics of skeletal muscle in MPS rats: through proteomics research,a total of 3597 proteins were screened,and 2879 proteins were quantifiable.733 differential proteins were screened out in the Tuina group and model B group with a 1.5-fold threshold,including 498 up-regulated proteins and 235down-regulated proteins.The biological processes mainly included neutrophil-mediated immunity,neutrophil activation,and granulocyte activation.Abnormal activation of related signaling pathways mainly enriched in inflammation protein response,immune-related pathways,complement,coagulation cascade signaling pathways and cell adhesion molecules were observed.S100A8 and S100A9 inflammatory key proteins were selected for verification.(3)Study on the effect of Jie Jie Chu Bi Tuina on S100A8/A9/TLR4 signaling pathway in MPS rats: after 7 days of intervention,Western blot results of each group of rats showed that compared with the blank group,the protein content of S100A8,S100A9,TLR4,CD14,p-JNK and p-p38 in the skeletal muscle of model group rats increased significantly(P<0.01).Compared with the model group,the protein content of S100A8,S100A9,TLR4,CD14,p-JNK and p-p38 in the skeletal muscle of Tuina group rats decreased significantly(P<0.01).Compared with the control group,the protein content of S100A8 and p-JNK in the skeletal muscle of control group rats decreased significantly(P<0.05).Compared with the control group,the protein content of S100A8 and p-p38 in the skeletal muscle of Tuina group rats decreased significantly(P<0.05).After 7 days of intervention,PCR results of each group of rats showed that compared with the blank group,the m RNA expression levels of S100A8,S100A9,CD14 and TLR4 in the skeletal muscle of model group rats increased significantly(P<0.05,P<0.01).Compared with the model group,the m RNA expression levels of S100A8,S100A9,CD14 and TLR4 in the skeletal muscle of Tuina group rats decreased significantly(P<0.05,P<0.01).(4)A clinical exploratory study on the therapeutic effect of resolving muscle adhesions and promoting blood circulation massage on neck and back myofascial pain: there was no significant difference in Mc Gill score between the Tuina group and the control group before treatment(P>0.05).However,both groups showed a significant reduction in Mc Gill score during and after treatment compared to the ones before treatment(P<0.01).Compared with the control group,the Mc Gill score in the Tuina group was significantly lower after treatment(P<0.01).Both the Tuina group and the control group showed a significant decrease in NDI score during and after treatment compared to before treatment(P<0.01).Compared with the control group,the NDI score in the Tuina group was significantly lower(P<0.05).Both the Tuina group and the control group showed an increase in the angle of cervical spine rotation,lateral flexion,and extension after treatment compared to before treatment(P<0.01).Compared with the control group,the angle of cervical spine flexion in the Tuina group was significantly increased after treatment(P<0.01).Compared with before treatment,the Tuina group showed a significant increase in right rotation and left lateral flexion during treatment(P<0.01),while the control group showed a significant increase in left rotation(P<0.05)and left lateral flexion(P<0.01).After treatment,compared with during treatment,the Tuina group showed a significant increase in both left and right lateral flexion(P<0.05),while the control group showed a significant increase in left rotation(P<0.05)and left lateral flexion(P<0.01).Compared with the control group,the Tuina group showed a greater increase in cervical spine mobility,but the difference was not statistically significant(P>0.05).The Tuina group was significantly better than the control group in improving cervical spine mobility.Conclusion:(1)Jiejie Chubi Tuina intervention in MPS rats can promote the increase of skeletal muscle cells,repair skeletal muscles,reduce skeletal muscle fibrosis,improve the abnormality of motor end-plates,and reduce the amplitude of spontaneous potentials.(2)Jiejie Chubi Tuina intervention in MPS rats can reduce the content of S100A8,S100A9,CD14,TLR4,p-p38,and p-JNK proteins,as well as the expression levels of S100A8,S100A9,CD14,and TLR4 m RNA and the content of S100A8,S100A9,TNF-α,and IL-6 in rat serum on the injured side of all groups.The therapy regulates the S100A8/A9/TLR4 signaling pathway,reduces the inflammatory response of rats,and verifies that the S100A8/A9/TLR4 signaling pathway is the mechanism of action of Jiejie Chubi Tuina therapy for MPS.(3)Jiejie Chubi Tuina therapy for thoracolumbar fasciitis patients can significantly reduces the patients’ level of pain,improve their cervical spine function,increase their activity,and reduce the serum content of inflammatory factors S100A8,S100A9,TNF-α,and IL-6,proving that Jiejie Chubi Tuina therapy has clinical effectiveness for fasciitis of nape muscle.(4)Jiejie Chubi Tuina therapy for MPS has significant clinical efficacy,and its mechanism of action is related to the inhibition of the inflammatory response,which has clinical value for promotion and application. |
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