Clinical And Experimental Study Of Three-step Tuina In Intervention Of Frozen Shoulder Joint Capsular Fibrosis

Ⅰ.Clinical ResearchObjective:To evaluate the clinical efficacy of the three-step Tuina method based on MR-DTI to intervene in frozen shoulder（FS）,to investigate the mechanism of action of Tuina to intervene in FS joint capsule fibrosis,and to provide an evidence-based basis for the treatment of FS with Tuina.Methods:Using a randomized,controlled method,the 57 FS patients included were randomly divided into a treatment group（three-step Tuina method）n=29 and a control group（Intermediate frequency electrotherapy）n=28;treatment was given three times a week,once every other day,for a total of 6 weeks;at the end of treatment,data were statistically analyzed in terms of VAS score,Constant-Murley（CMS）shoulder function score,shoulder MRI,rotator cuff muscle group diffusion tensor imaging and corresponding FA and ADC values,overall clinical efficacy,shoulder pain,patient compliance and safety.Results:（1）VAS scores and CMS scores:The intra-group comparison showed that compared with the pre-treatment period,the VAS scores of both groups decreased significantly（P<0.01）and the CMS scores increased significantly（P<0.01）;the inter-group comparison showed that the treatment group was significantly more effective than the control group in reducing VAS scores and increasing CMS scores at the end of weeks 3 and6 of treatment（P<0.01）.However,there was no statistically significant difference between the two groups at the 16-week post-treatment follow-up（P>0.05）;（2）MRI characteristics:at the end of the 6th week of treatment,the swelling of the inferior glenohumeral ligament,edema outside the axillary capsule,edema of the rostro-humeral ligament,edema of the rotator cuff space,and the volume of periarticular fluid in both groups were significantly reduced compared with those before treatment（P<0.01）;the thickness of the joint capsule of the axillary humeral part and the thickness of the rostro-humeral ligament decreased.There was no significant difference in the thickness of the joint capsule of the axillary glenoid,the thickness of the joint capsule of the rotator cuff space,the height and width of the capsule before and after treatment（P>0.05）;compared with the control group,the reduction of periarthritis edema and the thickness of the joint capsule of the axillary glenoid were more significant in the treatment group（P<0.05）.（3）MR-DTI performance of rotator cuff muscles:after 6 weeks of treatment,the FA values of rotator cuff muscles（SSP,ISP,TM,SSC）in the treatment group were significantly higher than before treatment,and the difference was statistically significant（P<0.01）,while the ADC values only decreased significantly in ISP and SSC（P<0.01）,and the difference in SSP and TM was not statistically significant（P>0.05）;the FA value of rotator cuff muscles in the control group increased only in ISP and TM after treatment compared with that before treatment,and the difference was statistically significant（P<0.01）,and the difference was not statistically significant in SSP and SSC（P>0.05）;the ADC value decreased significantly before and after treatment only in ISP（P<0.05）,and the difference was not statistically significant in SSP,TM and SSC（P>0.05）;comparison between groups showed that the treatment group was significantly more effective than the control group in improving the diffusion of water molecules in the rotator cuff muscle groups（P<0.05）.（4）Overall efficacy:The effective rate of the treatment group was 89.66%,and the effective rate of the control group was 78.57%,and the short-term efficacy of the treatment group was significantly higher than that of the control group;the incidence of nocturnal shoulder pain,pain during exercise and spontaneous pain in the treatment group was also lower than that of the control group after treatment.（5）Compliance and safety:During the 6 weeks of treatment,patients in both groups had good compliance and no significant adverse reactions occurred during the treatment.Conclusion:The three-step Tuina method and IF electrotherapy both have definite efficacy in the treatment of progressive FS,but the three-step Tuina method has better short-term efficacy than IF electrotherapy in improving the symptoms of FS patients;the three-step Tuina method can rapidly relieve shoulder pain and restore the function of the affected shoulder,reduce shoulder capsule edema and fibrotic thickening,restore the function of periacetabular muscles,and shorten the natural course of FS.Ⅱ.Experimental ResearchObjective:This experimental study will observe the effects of Tuina on the histomorphology,inflammation and expression of fibrotic factors in the shoulder capsule,and initially reveal the effect mechanism of Tuina on improving fibrosis in the joint capsule of FS,which will provide experimental basis and research ideas for future research on the mechanism related to Tuina treatment of FS.Methods:Thirty-two healthy male clean grade New Zealand rabbits at 6 months of age were selected;the frozen shoulder rabbit model was prepared by using continuous strain plus ice,and 16 adult rabbits were randomly divided into 2 groups,the model group and the model tuina group,with 8 rabbits in each group;the other 16 normal rabbits were randomly divided into the blank control group and the blank tuina group,with 8 rabbits in each group.The blank control group was not intervened,and the model group was not intervened after modeling,while the blank tuina group was given tui-na intervention and the model tui-na group was given tui-na intervention after modeling.The tuina intervention was performed once/day for 20 min/time for 3 weeks.At the end of the intervention,the right shoulder joint capsule and synovial tissue specimens were collected and executed after blood sampling.The expression of IL-1β,TNF-αand PGE₂ in serum and joint capsule was detected by Elisa method,and TGF-β1,PPAR-γ,MMP-1,MMP-9,TIMP-1,COL-Ⅰ and COL-Ⅲ in joint capsule were detected by immunohistochemistry and Western blot method.protein expression levels.Results:（1）Histopathology of joint capsule and synovial membrane.Synovium:At 3weeks of intervention,HE staining of the synovial membrane of the joint capsule showed that compared with the blank control group and the Tuina group alone,the model group showed massive proliferation of synovial cells and infiltration into the fibrous layer,disorganized cell arrangement and dense structure,and massive local inflammatory cell proliferation and vascularity;compared with the model group of rabbit joint capsule,the model Tuina group showed no signs of massive proliferation and migration of synovial cells,slightly tighter arrangement,vascularity and inflammatory cell infiltration was lighter compared to the model group.Articular capsule:After 3 weeks of intervention,HE staining of the fibrous layer showed that the normal lax reticular structure of the fibrous layer of the shoulder capsule in the model group was destroyed,collagen accumulation,fibrous bundle arrangement was disordered,and a strongly eosinophilic deep-stained area appeared,and a fibrous mass-like tissue containing cell clusters was formed compared with the model group of rabbit joint capsule;compared with the model group of rabbit joint capsule,the model Tuina group showed that the reticular structure of the fibrous layer was preserved relatively intact compared with the model group,and the collagen fibrous bundle arrangement was slightly disordered,but a strongly eosinophilic deep-stained area did not appear,and no fibrous mass was formed.（2）Elisa results from animal experiments showed that:After 3 weeks of intervention,the results of serum and joint capsule Elisa assay showed that the expression of IL-1β,TNF-αand PGE₂ inflammatory factors were not significantly different between the blank control group and the Tuina-only group（P>0.05）;compared with the blank control group,the expression of IL-1β,TNF-αand PGE₂ was significantly increased in the model group（P<0.01）;compared with the model group,the The model Tuina group was able to reduce the expression of IL-1β,TNF-αand PGE₂,and the difference was statistically significant（P<0.01）.（3）Immunohistochemical and WB results of animal experiments showed that:1)Experimental animals were intervened for 3 weeks.Compared with the blank control group,the blank Tuina group could increase the expression of PPAR-γin the shoulder joint capsule,while TGF-β1,MMP-1,MMP-9,TIMP-1,COL-Ⅰ,COL-Ⅲand other proteins had no statistical significance in the shoulder joint capsule of the two groups（P>0.05）.2)Compared with the blank control group,the expression levels of TGF-β1,PPAR-γ,MMP-1,MMP-9,TIMP-1,COL-Ⅰand COL-Ⅲin the shoulder capsule tissue were increased in the model group,and the differences were statistically significant（P<0.01）;meanwhile,the MMP-1/TIMP-1,MMP-9/TIMP-1 ratios were decreased in the model group.3)Compared with the model group,the model Tuina group was able to reduce the expression levels of TGF-β1,TIMP-1,COL-Ⅰand COL-Ⅲproteins in the joint capsule tissue,and the difference was statistically significant（P<0.05）.4)Compared with the model group,the model Tuina group was able to increase the expression levels of PPAR-γ,MMP-1 and MMP-9 in joint capsule tissues,and also up-regulate the ratios of MMP-1/TIMP-1,MMP-9/TIMP-1.Conclusion:Tuina intervention may ameliorate local chronic inflammatory injury,inhibit the proliferation of fibroblasts,promote the degradation of collagen in the shoulder capsule,and reduce shoulder capsule fibrosis by inhibiting the release of inflammatory factors such as IL-1β,TNF-α,and PGE₂,down-regulating the expression of TGF-β1,up-regulating the MMPs/TIMPs ratio and PPAR-γprotein expression.