中医正骨2025，Vol.37Issue(3)：14-22,38,10.
中医整脊棘突旁指压法干预大鼠膝骨关节炎的效果及作用机制研究
Efficacy and mechanism of traditional Chinese medicine chiropractic paraspinous thumb-pressing therapy against knee osteoarthritis in rats:an experimental study
摘要
Abstract
Objective:To observe the effects of traditional Chinese medicine(TCM)chiropractic paraspinous thumb-pressing therapy against knee osteoarthritis(KO A)in rats,and to explore its underlying mechanism.Methods:Forty SD rats were selected and randomized into blank group,model group,low-intensity group,medium-intensity group and high-intensity group,with 8 ones in each group.All rats but the ones in blank group were modeled by using bilateral knee intra-articular injection of sodium iodoacetate for inducing KOA,while the ones in blank group by using injection of the same dosage of normal saline at the corresponding sites.Fourteen days after successful model-ing,the rats in the low-,medium-,and high-intensity groups were subjected to 4 N,8 N,and 16 N vertical downward pressures,respectively,beside the L3-L5 spinous processes by using a force-measuring finger pressure device for small animals to simulate the paraspinous thumb-pressing manipulation with different intensities,once every other day,2 seconds per time and 2 minutes per site for consecutive 28 days;while the ones in blank group and model group were not given any intervention.The paw withdrawal mechanical threshold(PWMT)of rats were tested using a Von Frey electronic algometer after 1-,7-,14-,21-and 28-day intervention,respectively.After 28-day intervention,the levels of nerve growth factor(NGF)and calcitonin gene related peptide(CGRP)in serum were detected by using ELISA kit,and the Ca2+flux in L3 dorsal root ganglion(DRG)neurons was detected by using Calcium imaging.Furthermore,the protein expression levels of transient receptor potential vanilloid 1(TRPV1),protein phosphatases 2B(PP2B)and calmodulin(CaM)in the L3 DRG tissues were detected using Western blot.After that,the rat knee synovial tissues were harvested and stained with HE staining kit,Masson staining kit,and Sirius red staining kit,respectively,to observe the inflammatory cell infiltration and fibrosis,and the protein expression levels of interleukin(IL)-1β,tumor necrosis factor-α(TNF-α),transforming growth factor(TGF)-β,and α-smooth muscle actin(α-SMA)in the knee synovial tissues were detected using Western blot.Results:①The measuring results of PWMT.The PWMT remained essentially unchanged over time in blank group,model group,and low-intensity group(F=0.885,P=0.487;F=0.339,P=0.849;F=0.555,P=0.697),while that presen-ted an upward trend over time in medium-and high-intensity groups(F=8.222,P=0.000;F=9.844,P=0.000).After 21-and 28-inter-vention,the PWMT was higher in medium-and high-intensity groups compared to model group and low-intensity group(21-intervention:P=0.002,P=0.001;P=0.022,P=0.011;28-intervention:P=0.003,P=0.000;P=0.044,P=0.003),while,it was not significantly dif-ferent from each other between high-intensity group and medium-intensity group(P=0.774,P=0.273).②The serum level of pain medi-ator.The serum levels of NGF and CGRP were lower in medium-and high-intensity groups compared to model group(NGF:P=0.003,P=0.000;CGRP:P=0.000,P=0.000).The serum level of CGRP was lower in medium-intensity group compared to low-intensity group(P=0.000),while,there was no statistical difference in the serum level of NGF between the 2 group(P=1.000).The serum levels of NGF and CGRP were lower in high-intensity group compared to low-intensity group(P=0.027,P=0.000),while,they were not significantly differ-ent from each other between high-intensity group and medium-intensity group(P=1.000,P=1.000).③The Ca2+flux in L3 DRG neu-rons.The average fluorescence intensity of Ca2+in L3 DRG neurons was higher in low-,medium-,and high-intensity groups compared to model group(P=0.001,P=0.000,P=0.000),and was higher in medium-and high-intensity groups compared to low-intensity group(P=0.003,P=0.016),while,it was not significantly different from each other between high-intensity group and medium-intensity group(P=0.361).④The protein expression levels of TRPV1,PP2B and CaM in the L3 DRG tissues.The relative protein expression level of TRPV1 in the L3 DRG tissues was higher,while that of PP2B and CaM was lower in medium-and high-intensity groups compared to model group and low-intensity group(model group:P=0.002,P=0.000;P=0.011,P=0.000;P=0.000,P=0.000;low-intensity group:P=0.047,P=0.014;P=0.043,P=0.033;P=0.000,P=0.000),while,there was no statistical difference in the relative protein expression levels of TRPV1,PP2B,and CaM between high-intensity group and medium-intensity group(P=1.000,P=1.000,P=1.000).⑤Patho-logical observation of knee synovial tissues.HE staining showed that,in model group,there was obvious inflammatory cell infiltration in the synovial tissues of rats,which was improved in low-,medium-,and high-intensity groups compared to model group,with a more pronounced improvement in medium-and high-intensity groups.Masson staining revealed that,compared to blank group,the collagen deposition in the synovial tissues significantly increased in rats of model group;compared to the model group,the collagen deposition decreased in low-,me-dium-,and high-intensity groups,with a more marked reduction in medium-and high-intensity groups.Sirius red staining showed that the type Ⅱ collagen in the synovial tissues significantly decreased in rats of model group,while,compared to the model group,the type Ⅱ collagen increased in low-,medium-,and high-intensity groups,with a more significant increase in medium-and high-intensity groups.⑥The protein expression levels of IL-1β,TNF-α,TGF-β and α-SMA in knee synovial tissues.The relative protein expression levels of IL-1β,TNF-α,TGF-β and α-SMA in knee synovial tissues were lower in medium-and high-intensity groups compared to model group and low-intensity group(model group:P=0.000,P=0.000,P=0.000,P=0.000;P=0.000,P=0.000,P=0.000,P=0.000;low-intensity group:P=0.004,P=0.002,P=0.003,P=0.000;P=0.005,P=0.008,P=0.007,P=0.004),while,they were not significantly different from each other between high-intensity group and medium-intensity group(P=0.116,P=0.083,P=0.224,P=0.341).Conclusion:The TCM chiropractic paraspinous thumb-pressing therapy can alleviate mechanical pain sensitization and synovial inflammation in KOA rats,and the effect exhibits intensity-dependence.It may work by inducing the desensitization of TRPV1 in DRG neurons and influencing the crosstalk between DRG neurons and synovial fibroblasts.关键词
骨关节炎,膝/中医整脊/指压法/疼痛/敏化/瞬时受体电位香草素受体1/神经节,脊/神经元/滑膜/成纤维细胞/串话/大鼠Key words
osteoarthritis,knee/traditional Chinese medicine chiropractic/thumb pressing manipulation/pain/sensitization/transient recep-tor potential vanilloid 1/ganglia,spinal/neurons/synovial membrane/fibroblasts/crosstalk/rats引用本文复制引用
张力,张华,李平,高嵩,柯广娟,屈留新..中医整脊棘突旁指压法干预大鼠膝骨关节炎的效果及作用机制研究[J].中医正骨,2025,37(3):14-22,38,10.基金项目
东南大学附属中大医院中西医协同医学科研项目(2023zxyxt03) （2023zxyxt03）
