The Clinical Efficacy Of Guizhi Plus Gegen Decoction In The Treatment Of Cervical Spondylosis Of Cervical Type(Wind-cold-dampness Type) And The Mechanism Of Regulating PI3K/AKT/mTOR Autophagy Pathway

BackgroundThe cervical type cervical spondylosis(wind-cold-dampness type)is one of the most common types of cervical spondylosis,and is recognized as the representative of early cervical spondylosis.Its incidence increases year by year and presents a trend of younger,which seriously affects the life quality of patients.The cervical type cervical spondylosis(wind-cold-dampness type)belongs to the category of "xiangbi disease"in the Traditional Chinese Medicine.Guizhi plus Gegen decoction is a classic prescription for the treatment of "xiangbi disease".It is widely used to treat all kinds of diseases and has obtained good clinical effect.However,the molecular mechanism of Guizhi plus Gegen decoction in treating cervical type cervical spondylosis(wind-cold-dampness type)remains unclear.In this paper,the molecular mechanism of Guizhi plus Gegen decoction in treating cervical type cervical spondylosis(wind-cold-dampness type)will be discussed from three aspects of clinical research,network pharmacological analysis and animal experiments,which provides a new therapeutic idea for TCM prevention and treatment of cervical type cervical spondylosis(wind-cold-dampness type).Part 1:Retrospective study on the treatment of cervical type cervical spondylosis(wind-cold-dampness type)with Guizhi plus Gegen decoctionObjective:To observe the clinical effect of Guizhi plus Gegen decoction on cervical type cervical spondylosis(wind-cold-dampness type).Methods:A retrospective analysis was performed on 51 patients diagnosed as cervical type cervical spondylosis(wind-cold-dampness type)admitted to the orthopedics Department of Hubei Provincial Hospital of Traditional Chinese Medicine from December 2021 to December 2022.In group A,a total of 26 cases were treated with neck traction+far-infrared therapy;in group B,a total of 25 cases were treated with Guizhi plus Gegen decoction+neck traction+far-infrared therapy.The treatment continued for 2 weeks.After treatment,VAS score,NDI score,neck range of motion and TCM syndrome score were compared between the two groups.The patients were followed up by telephone 6 weeks after the end of treatment,and the recurrence rate of the two groups was compared.P<0.05 was considered statistically significant.Results:There was no significant difference in gender,age and course of disease between the two groups(P>0.05).After treatment,neck pain and range of motion were improved in both groups,while VAS score,NDI score and TCM syndrome score were decreased,the difference was statistically significant(P<0.05).Compared with group A after treatment,VAS score,NDI score and TCM syndrome score of group B were lower,and the recurrence rate was lower,the difference was statistically significant(P<0.05).Conclusions:Far-infrared therapy+neck traction can reduce the VAS score and NDI score of patients and improve the range of motion of the neck.Moreover,after the intervention of Guizhi plus Gegen decoction on this basis,the clinical efficacy is better and the recurrence rate is lower.Part 2:Mechanism study of Guizhi plus Gegen decoction in treating cervical spondylosis based on network pharmacologyObjective:To explore the molecular mechanism of Guizhi plus Gegen decoction in treating cervical spondylosis.Methods:TCMSP database was used to search the active ingredients of Guizhi plus Gegen decoction and screen their molecular targets.Molecular targets of cervical spondylosis were obtained from OMIM,Genecards and DisGeNET databases.The intersection of the molecular targets of drugs and disease was performed for PPI network analysis.Finally,GO and KEGG enrichment analysis was performed.Results:There were 173 active ingredients and 954 molecular targets in Guizhi plus Gegen decoction,and 7095 molecular targets in cervical spondylosis,among which 610 were common molecular targets.The top 20 key molecular targets were AKT1,TP53,TNF,ALB,CTNNB1,EGFR,SRC,VEGFA,HSP90AA1,STAT3,MAPK3,JUN,HRAS,CASP3,ESR1,HIF1A,MTOR,ERBB2,IL-6 and PP ARG.In GO enrichment analysis,molecular targets mainly involved membrane regions,membrane microdomains,neuron cell bodies and other cell components.In terms of molecular function,these molecular targets mainly involved protein serine/threonine kinase activity,protein tyrosine kinase activity,peptide chain endonuclease activity,etc.In terms of biological processes,these molecular targets mainly involve tyrosine phosphorylation,tyrosine modification,and serine modification.In KEGG enrichment analysis,these molecular targets mainly involved MAPK signaling pathway,PI3K-Akt signaling pathway,Ras signaling pathway,etc.These signaling pathways are mainly involved in such biological processes as autophagy,cell senescence,cell apoptosis and inflammatory response.Conclusions:Guizhi Jiagen Decoction plays a multi-target and multi-signal regulatory role in cervical spondylosis by inhibiting inflammatory response,cell autophagy and other signaling pathways.Part 3:The molecular mechanism of Guizhi plus Gegen decoction regulating cervical type cervical spondylosis(wind-cold-dampness type)through PI3K/AKT/mTOR autophagy pathway of Cervical annulus fibrosus cellObjective:To study the effect of Guizhi plus Gegen decoction on the inflammatory factors and PI3K/AKT/mTOR autophagy pathway of intervertebral disc annulus fibrosus cells.Methods:New Zealand rabbits were divided into normal control group,cervical type cervical spondylosis(wind-cold-dampness type)model group and Guizhi plus Gegen decoction low,medium and high dose drug group with 8 rabbits in each group.In addition to the normal control group,the other groups underwent cervical type cervical spondylosis(wind-cold-dampness type)modeling.After successful modeling,the normal control group and the model group were given normal saline intragastric administration,and the drug group was given low,medium and high dose Guizhi plus Gegen decoction intragastric administration respectively.All animals were given continuous gavage for 4 weeks.After treatment,HE staining and Safranin O staining were used to observe the morphological changes of intervertebral disc tissue.The expression levels of inflammatory factors in serum were detected by ELISA assay.The expression changes of autophagy genes and the activity of autophagy pathway were detected by RT-PCR and Western Blot assay.Results:Compared with the normal control group,the number of intervertebral disc cells in the model group was reduced,the arrangement was disordered and the tissue boundary was blurred.The gene and protein expression levels of ATG5 and ATG12 were down-regulated.The expression levels of TNF-α and IL-1β were up-regulated.PI3K/AKT/mTOR autophagy pathway activity was activated.Compared with the model group,after the intervention of Guizhi plus Gegen decoction,the number of intervertebral disc cells increased,the arrangement was relatively regular and the boundary was clear.The gene and protein expression levels of ATG5 and ATG12 were up-regulated,the expression levels of TNF-α and IL-1β were down-regulated,and PI3K/AKT/mTOR autophagy pathway activity was suppressed,which was dose-dependent.Conclusions:Guizhi plus Gegen decoction can promote autophagy and inhibit inflammation by inhibiting the activity of PI3K/AKT/mTOR autophagy pathway,which plays a role in delaying the occurrence of cervical type cervical spondylosis(wind-cold-dampness type).