Research the mechanism of Cinnamoni cortex-Aconm lateralis radix praeparaia-Achtranthis bidentatae radix combination in treatment of lumbar disc herniation based on data mining and network pharmacology

Background:This study gets a classic prescription of Song Dynasty medicine for the treatment of waist and leg pain through analyzing the inheritance of traditional Chinese medicine auxiliary platform.Further,the potential mechanism of the classic prescription was analyzed based on molecular docking and network pharmacology.Methods:Based on the frequency statistics,association rules and cluster analysis,the core herbal combination and the classic prescription was digged out.Use of network pharmacology methods and molecular docking to explore the pharmacological mechanism of classic prescriptions for treatment of lumbar disc herniation.Then gene ontology biological function annotation and Kyoto Encyclopedia of Genes and Genomes enrichment of pathways were performed.Finally,the compounds of herbs were docked with the important targets of MMP1 and CRP.Results:The high-frequency Chinese medicines for treating waist and leg pain were found and we further unearthed the “Rougui-Fuzi-Niuxi (Cinnamoni cortex-Aconm lateralis radix praeparaia-Achyranthis bidentatae radix” as the core herbal combination,and matched the classic ancient prescription of Chinese medicine Jiawei Shenzhuo decoction (CAPCMJWSZD).The targets of CAPCMJWSZD were mapped to the targets of lumbar disc herniation and 48 potential targets were obtained.The core potential targets were obtained in the protein-protein interaction network,such as CRP,IL2,FOS,MMP1,CASP3.Through the DAVID database,a total of 129 gene ontology function annotation terms (P < 0.01) and 91 Kyoto Encyclopedia of Genes and Genomes pathways (P <0.01) were obtained.Molecular docking results showed that quercetin has the lowest binding energy for docking with MMP1and CRP,and these two methods of molecular docking are most likely to occur.Conclusion:The most important bioactive components in CAPCMJWSZD can eliminate inflammation and slow disc degeneration through some potential targets,such as CRP,IL-2,MMP1,and these targets can rich in the following pathways,such as metalloendopeptidase activity,MAP kinase activity,osteoclast differentiation,et al.

Keywords:Lumbar and leg pain,Lumbar disc herniation,Data mining,Network pharmacology,Cinnamoni cortex-Aconm lateralis radix praeparaia-Achtranthis bidentatae radix,Jiawei Shenzhuo decoction

Background

Lumbar disc herniation (LDH) is the most common diagnosis among the degenerative abnormalities of the lumbar spine (affecting 2 to 3% of the population).LDH refers to that nucleus pulposus protrudes or prolapses from the rupture intervertebral disc fiber,which compresses the local adjacent near spinal nerve root and leads to waist pain,radiating pain of sciatic nerve,et al [1].The pathogenesis of the LDH [2,3,4,5] is not clear and is mainly classified into four categories: mechanical mechanics,matrix metalloproteinase (MMP) theory,immune inflammation and gene control theory.It had shown that the degenerative changes of LDH were closely related to metalloendopeptidase activity,interleukin(IL)-6,IL-17,and tumor necrosis factor (TNF)signaling pathways,et al [6,7,8,9].

There was no the term “l(fā)umbar disc herniation” in traditional Chinese medicine (TCM) theory.Based on the symptoms of pain and numbness in the lower back and legs,scholars often regard LDH as the category of“arthralgia” and “l(fā)umbar and leg pain” in Chinese medicine [10].At present,the main methods of TCM treatment for LDH include oral and external Chinese herbs treatment,acupuncture,moxibustion and bone setting manipulation.The oral Chinese herbs treatment is an important method for TCM to treat LDH [11,12].The Song Dynasty was a period of rapid development of Chinese medicine,especially in terms of bone disease.Many famous medical books in Song Dynasty,includingPeaceful Holy Benevolent Prescriptions(written by Huairen Wang,You Wang,et al.in 978 C.E.-992 C.E.),Complete Record of Benevolence(written in 1299 C.E.-1300 C.E.,unknown author) andPrescriptions from the Great Peace Imperial Grace Pharmacy(written by Great Peace Imperial Grace Pharmacy in 1148 C.E.),had recorded a large numbers of prescriptions for the treatment of waist and leg pain.This article collects and organizes these prescriptions,and the core drug combination “Rougui-Fuzi-Niuxi(Cinnamonicortex-Aconmlateralisradix praeparaia-Achyranthisbidentataeradix)” is excavated from them.Based on the core drug combination and symptoms of LDH,a classic ancient prescription of Chinese medicine (CAPCM) Jiawei Shenzhuo decoction (JWSZD) was matched.The reliable efficacy of CAPCM JWSZD in the treatment of LDH was also proved in the clinical treatment [13].However,the pharmacological mechanism has not yet been explored at the molecular level,so this study applied network pharmacology to explore the potential mechanism of CAPCM JWSZD in the treatment of LDH and used molecular docking technology to screen compounds with good affinity for MMP1 and C-reactive protein (CRP).

Data and methods

Data mining of prescriptions for treating waist and leg pain

Sources of prescriptions and criteria for inclusion and exclusion.The data came from all chapters of ancient books of the Song Dynasty,such asPeaceful Holy Benevolent Prescriptions,Complete Record of BenevolenceandPrescriptions from the Great Peace Imperial Grace Pharmacy.The inclusion criteria refered to criteria of diagnosis and therapeutic effect of diseases and syndromes in TCM [14] and diagnosis and treatment scheme of LDH in TCM [15].and the prescriptions were used for the waist pain,leg pain,weakness,urgency,cold,and numbness,which also meets the standard that the dosage form is oral prescription.The same prescription with different names was only recorded once.The invalid prescriptions were excluded according to the exclusion criteria,such as:symptoms of bones and joints were not the main symptoms; traumatic diseases or there were rare drugs and modern illegal drugs in the prescription.

The processing of data standardization and input analysis.According to the 2015 edition of thePharmacopoeia of the People's Republic of Chinaand theDictionary of traditional Chinese Medicine,the name and function classification of 450 prescriptions were standardized and corrected.Then,the standard drugs were entered into the “statistical report” and“prescription analysis” section of inheritance of TCM auxiliary platform,and the data were crosswise checked by 2 researchers.Then,the “the frequency of symptom” and “the frequency of medication” were automatically calculated by the system,and the prescription rules based on association rules and new prescription analysis based on entropy hierarchical clustering were obtained.

Mechanism analysis based on network pharmacology

Searching the targets of effective components of TCM.Tradictional Chinese Medicine Systems Pharmacology (TCMSP) database(https://tcmspw.com/tcmsp.php) was used to search the TCM of classic prescriptions and new prescriptions.According to the recommended standard of oral bioavailability ≥ 30% and drug similarity ≥ 0.18,the effective compounds with high activity were screened out.The obtained compounds were input into TCMSP database to obtain the predicted targets of main compounds.UniProt database was used to identify the targets and convert it to gene name.

Sorting out the effective targets and constructing protein-protein interaction network.Through the GeneCard database (https://www.genecards.org/),we selected the species as “Homo sapiens” and searched“l(fā)umbar disk herniation” as the key word to obtain the related targets of LDH.Venny2.1 online mapping tool was used to map the targets of drug prediction and targets of disease prediction to obtain the common targets,which are the potential targets of classic new prescription in the treatment of LDH.In the STRING database (https://string-db.org/),“Homo” was selected as the species and degree of confidence was set to be greater than 0.700.The protein-protein interaction network (PPI) network was constructed for the potential targets,and the interaction between proteins was analyzed.Finally,the core targets were obtained through network topology analysis.

Gene ontology pathway annotation and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis.Based on the core targets,the calibration of gene name was set as “offcial gene symbol” in Database for Annotation,Visualization and Integrated Discovery database,and the threshold value was set asP< 0.01,then gene ontology (Go) pathway annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis were carried out.Finally,the enrichment analysis results of Go and KEGG were visualized by rstudio 3.6.3 software.

Molecular docking analysis of core compounds and key target proteins.The crystal structures of MMP1 and CRP were obtained from RSCB database(http://www.rcsb.org/) and ZINC database(http://zinc.docking.org/),and which crystal structure of water molecules is removed and hydrotreated by using autodock tools software.Then the charge was calculated and saved as pdbqt format file.The mol2 format file of 3D structure of core compounds was downloaded from zinc database and autodock tools software was used to balance the charge of small molecules.The PyMOL software was used to process the small molecule structure of protein,and girdbox was set to determine docking pocket.Finally genetic algorithm parameters algorithm was selected for molecular docking.

Results and analysis

Drug frequency statistics

Statistics were made on the prescriptions for the treatment of lumbar and leg pain inPeaceful Holy Benevolent Prescriptions,Complete Record of BenevolenceandPrescriptions from the Great Peace Imperial Grace Pharmacy.Four hundreds and fifty prescriptions were counted,involving 281 drugs,among which 33 drugs were used more than 50 times.Among them,the top 3 herbs most frequently used wereCinnamoni cortex,Aconm lateralis radix praeparaia,Achyranthis bidentatae radix(Table 1).

Association analysis of high-frequency TCM and core herbal combination based on association rules

“Collaborative Compatibility” refers to the combination of 2 drugs with similar effects,which can enhance the efficacy of the main drug.“Assist in Compatibility” refers to the combination of drugs that have some common characteristics in terms of performance and efficacy,and taking one drug as the main one and another drug as the auxiliary one can improve the efficacy of the main drug.The association rule mining method was applied to set the support level to “70”,and 31 herbal combinations commonly used were obtained (Table 2).Among the 28 groups of herbal combinations,the ones that belong to“Collaborative Compatibility” are:Aconm lateralis radix praeparaia-Cinnamoni cortex,Achyranthis bidentatae radix-Eucommiae cortex,Rehmanniae radix praeparata-Achyranthis bidentatae radix,et al,with functions of “Warming Kidney-Yang”,“Dispels Wind and Cold”,“Strengthening Muscles and Bones”,and “Tonifying Liver and Kidney”,et al.Others mostly belong to the “Assist in Compatibility”,such as Fuling(Poria-Cinnamoni cortex）,Qianghuo (Notopterygii rhizoma et radix)-Cinnamoni cortex,et al.

The degree of confidence was set to “0.7”,after analysis,27 groups of drug combination were obtained,the top five herbal combinations are shown in Table 3,such as “Achyranthis bidentatae radix,Danggui(Angelicae sinensis radix),Fangfeng (Saposhnikoviae radix)→Cinnamoni cortex”,et al.

The Table 2 shows that the frequency for the combination of TCMs in the first three forAchyranthis bidentatae radix-Cinnamoni cortex,Aconm lateralis radix praeparaia-Cinnamoni cortex,Achyranthis bidentatae radix-Aconm lateralis radix praeparaia,and learned from Table 1 frequency tally sheet of the TCM,Cinnanmomi cortexis counted a total of 278 times;Achyranthis bidentatae radixis counted a total of 257 times;Aconiti lateralis radix praeparatais counted a total of 227 times.Therefore,it is concluded that the core herbal combination of Song Dynasty court medicine for treating waist and leg pain is“Cinnamoni cortex-Aconm lateralis radix praeparaia-Achyranthis bidentatae radix”.

Analysis of new prescriptions based on unsupervised entropy hierarchical clustering

Entropy clustering for complex systems is an unsupervised pattern discovery algorithm,which can extract the combination of the largest amount of information from massive data,and it is especially suitable for discrete data.In analysis of new prescriptions,the correlation degree was set to 14 and the penalty degree was set to 7.According to the principle of reducing toxicity and increasing effectiveness,8 new prescriptions for drug clustering was obtained (Table 4).It is worth noting that the new prescriptions ranked first also contains the core herbal combination of “Cinnamoni cortex-Aconm lateralis radix praeparaia-Achyranthis bidentatae radix”.

Mining of traditional classic prescription

Table 1 Herbs with frequency of more than 50 times in the prescriptions for treating waist and leg pain in Song Dynasty

Table 2 Correlation analysis of high-frequency Chinese medicines with support below 70

Table 2 Correlation analysis of high-frequency Chinese medicines with support below 70 (Continued)

Table 3 Herbal combination with confidence level above 0.7

Table 4 New method combination mining based on entropy hierarchical clustering

After the above data mining,the core Chinese hebal combination of “Cinnamoni cortex-Aconiti lateralis radix praeparata-Achyranthis bidentatae radix” for the treatment of lumbar and leg pain was obtained,based on the drug frequency and correlation analysis of TCM.Compared with the prescriptions related to diseases of lumbar and leg pain,which are listed inDictionaryofTraditionalChineseMedicine Prescriptions,the potential prescriptions containing the core drug combination and the minimal amount of herbs was found.The results showed that CAPCM JWSZD was the most suitable classic prescription.CAPCM JWSZD is derived from volume 10 ofSynopsis of the Medical World(written by Fu Wang in 1758 C.E.) in the Qing Dynasty.It is mainly used to treat waist gangrene exposed to “Cold and Dampness”.CAPCM JWSZD is composed of CAPCM Shenzhuo decoction (Zhigancao (Glycyrrhizae radix et rhizoma),Ganjiang (Zingiberis rhizoma),Poria,Baizhu(Atractylodis macrocephalae rhizoma)),added with herbs such asAconm lateralis radix praeparaia,,Cinnamoni cortex,Zexie (Alismatis rhizoma),Duzhong (Eucommiae cortex),Achyranthis bidentatae radix,et al.CAPCM Shenzhuo decoction originated from the CAPCM GanJiang Lingzhu decoction of theSynopsis of the Golden Chamber(written by Zhongjing Zhang in 219 C.E.) in Eastern Han Dynasty,where the original description of CAPCM Shenzhuo decoction was:“Kidney Disease,The Body Weight,Waist Cold,Like Sitting In Water.Cold Pain From The Waist Down”.The original meaning of “Shen Zhuo” is the pathogenesis of “Cold and Wet” in the waist and causing “Cold” pain below the waist.In CAPCM Shenzhuo decoction,Atractylodis macrocephalae rhizoma,Poria,Glycyrrhizae radix et rhizomawere used to “Invigorate the Spleen” andZingiberis rhizomais used to “Warm Cold and Damgness”.CAPCM JWSZD added 4 core drugs to “Warm Kidney” and“Help Yang”,“Dispel Cold” and “Remove Dampness”.The application of CAPCM JWSZD in the treatment of LDH with “Cold and Dampness” can significantly relieve the symptoms of waist and 1eg pain,fear of co1d,lower limb pain and numbness,insomnia and anxiety,et al,and the overall effect is satisfactory [13].Therefore,the Clinical efficacy is consistent with the data mining prediction.

Network pharmacological analysis

Construction of herb-compound-target network and screening of core compounds.

The effective chemical components and predicted targets of the drugs in CAPCM JWSZD were obtained through TCMSP database.A total of 125 active compounds and 283 targets were screened according to oral bioavailability ≥ 30% and drug similarity ＞ 0.18.The network diagram of herb-compound-target was drawn by Cytoscape software (Figure 1),which consisted of 369 nodes and 1,643 edges.Based on network topology analysis,the top five core compounds in higher degree value were cinchonan-9-al,6’-methoxy-,(9R)-,quercetin,kaempferol,glyasperins M,tabernemontanine,and β-sitostero1.The detailed information of the top 15 compounds with degree value are listed in Table 5.

Table 5 Basic information of active compounds of Jiawei Shenzhuo decoction

Effective targets of CAPCM JWSZD and construction of PPI

Through the GeneCard database,291 targets related to LDH were obtained.With the help of Venny 2.1 online software,the targets in CAPCM JWSZD were mapped to the targets of LDH,and 48 common targets were obtained (Table 6,Figure 2),which was the effective targets of CAPCM JWSZD in the treatment of LDH.The 48 common targets were imported into the STRING database,the species was selected as “Homo sapiens”,and the confidence was set to be greater than 0.700.Then the exported TSV file was imported into Cytoscape software to analyze the network topology and PPI network was constructed (Figure 3).The PPI network consists of 48 nodes and 372 edges,with an average degree of 15.5 and an average centrality of 0.663.The core targets that are greater than the average degree value and average median centrality were screened out,and 5 core targets in the protein interaction network were obtained:CRP,IL-2,fos proto-oncogene,MMP1,and caspase-3 (Table 7).

Figure 1 Jiawei Shenzhuo decoction herb-compound-target network

Figure 2 Jiawei Shenzhuo decoction targets and LDH targets venny diagram.LDH,lumbar disc herniation;MSZD,modified Shenzhuo decoction.

Figure 3 Diagram of protein-protein interaction network

GO enrichment analysis and KEGG pathway enrichment analysis

The Database for Annotation,Visualization and Integrated Discovery database was used to perform GO enrichment analysis on the 48 common targets,a total of 92 biological process terms,23 molecular function terms and 14 cellular component terms were obtained (P< 0.01).The first 20 biological process information was visualized (Figure 4),including collagen catabolic process,immune response,negative regulation of IL-6 production,positive regulation of IL-17 production,positive regulation of angiogenesis,et al.

To further reveal the potential mechanism of CAPCM JWSZD on the effect of LDH,we conducted KEGG pathway enrichment analysis on 48 targets (P<0.01) and the top 20 pathways was visualized by Rstudio software (Figure 5).The first 20 pathways with strong significance were enriched,such as pathways in cancer,TNF signaling pathway,rheumatoid arthritis,hypoxia-inducible factor-1 signaling pathway,toll-like receptor signaling pathway,et al.

Construction of compound-target-pathway network diagram

The main pathways and targets obtained from the GO and KEGG enrichment analysis of CAPCM JWSZD were imported into Cytoscape software for visualization processing (Figure 6),where the green rectangle represents the pathway,the red diamond represents the target,and the yellow node represents the compound.Through topological analysis,the targets with higherP-value in GO enrichment analysis are TGFB1,NOS2,IL1A,IFNG,IL10,TNF,IL-2,MMP2,MMP1,et al.The targets with higherP-value in the KEGG target-pathway network are JUN,MAPK1,MAPK8,TNF,TGFB1.

The results of molecular docking

According to the prediction of core targetsin Table 8,MMP1 and CRP are the key proteins of CAPCM JWSZD in the treatment of LDH.The core compound in Table 4 is molecularly docked with the above 2 key proteins (some compounds have not been found in the National Center for Biotechnology Information database in MOL2 format chemical structure information).The binding energy is less than 0,which means that the compound and protein can bind spontaneously,the lower the binding energy,the greater the possibility of action.With binding energy ≤-5 kJ/mol as the screening criterion,the docking mode of the molecule with the smallest binding energy was selected for display (Figure 7).The binding energy of all docking methods is shown in Table 8.The results show that the binding energy of quercetin and MMP1 is the lowest,and that of quercetin and CRP is the lowest,the molecules of these 2 docking methods are the easiest to combine.

Table 6 Forty-eight potential targets of Jiawei Shenzhuo decoction in the treatment of lumbar disc herniation

Table 7 Five core targets of the protein-protein interaction network

Table 8 The binding ability of main compounds with HIF-1a and VEGFA protein

Figure 4 Histogram of gene ontology function enrichment of Jiawei Shenzhuo decoction

Figure 5 Diagram of Kyoto Encyclopedia of Genes and Genomes enrichment analysis of Jiawei Shenzhuo decoction

Figure 6 Compound-target-pathway network diagram of Jiawei Shenzhuo decoction for treating lumbar disc herniation

Figure 7 A:The docking mode diagram of quercetin and matrix metalloproteinase 1.The type of binding bond is H bond,the yellow-red compound is quercetin,and the docking site of cyan compound (MMP1) is ARG-127.B,the structural formula of Quercetin; C,diagram of the docking between quercetin and C-reactive protein.The type of bonding bond is H bond,the yellow compound is quercetin,and the docking site of light blue compound (CRP) is LYS-12; D,The structural formula of quercetin.

Discussion

Based on the excavation and analysis of the prescriptions for treating waist and leg pain in books such asPeaceful Holy Benevolent Prescriptions,Complete Record of BenevolenceandPrescriptions from the Great Peace Imperial Grace Pharmacyin Song Dynasty,the core combination of Chinese herbal medicine for waist and leg pain is obtained.“Cinnamonicortex-Aconmlateralisradix praeparaia-Achyranthis bidentatae radix”,and the CAPCM JWSZD was mined through the core combination.Finally,we analyzed the signal pathway and molecular mechanism of CAPCM JWSZD in the treatment of LDH through network pharmacology research.

TheComplete Record of Benevolencebelieves that waist and leg pain is related to the “Liver and Kidneys”.Through frequency analysis,among the top 15 herbs,the herbs that “Warm the Kidney” and“Boost Yang” and “Strengthen the Kidney and Bone”are the main ones,such asCinnamoni cortex,Aconm lateralis radix praeparaia,Achyranthis bidentatae radix,Duzhong (Eucommiae cortex),Shudihuang(Rehmanniaeradixpraeparata),Roucongrong(Cistanches herba),and it also contains herbs for“Anti-Wind” and“Anti-Dampness” such asSaposhnikoviae radix,Notopterygii rhizoma et radix,Duhuo (Angelicae pubescentis radix).In addition,the medicines of “Nourishing Blood” and “Promoting Blood Circulation” includeAngelicae sinensis radix,Chuanxiong(Chuanxiongrhizoma).Through high-frequency Chinese medicine correlation analysis,the core Chinese medicine combination “Cinnamoni cortex-Aconm lateralis radix praeparaia-Achyranthis bidentatae radix” was obtained,and the CAPCM for the treatment of waist and leg pain-JWSZD was obtained by matching the core Chinese medicine prescriptions.

The main compounds such as quercetin,kaempferol,glyasperins M,tabernemontanine,β-sitosterol,cinnamaldehyde,et al,were obtained by the compounds-targets network topology analysis.According to the average degree value and the average intermediate value centrality,5 core targets in the PPI network are obtained:CRP,IL-2,fos proto-oncogene,MMP1,caspase-3.Among them,MMP1,CRP,IL-2 were closely related to LDH.High-sensitivity CRP can reflect the degree of inflammation in the lumbar intervertebral disc tissue and evaluate postoperative recovery [16].Mohammadi [17] clinical experiments showed that quercetin could significantly reduce the serum CRP of patients.

From the results of GO function annotation analysis and KEGG pathway enrichment,the pathways related to LDH include MMP activity,negative regulation of IL-6 production,positive regulation of IL-17 production,TNFs,rheumatoid arthritis,MAP kinase activity,and osteoclast differentiation,et al.

The MMP activity signal pathway mainly includes 4 targets:MMP9,MMP3,MMP2,and MMP1.MMPs are activated to produce a cascade of amplification effect,which accelerates the degeneration of the intervertebral disc by promoting the degradation of the extracellular matrix.MMPs are a type of matrix proteolytic enzymes,and they are the key enzymes that cause the degradation of intervertebral disc collagen.Sally [6] and others found that there are a large number of MMPs in the herniated disc specimens,and the degree of disc degeneration is closely related to the expression of MMP-2,MMP-3,and MMP-9.Bachmeire [18] conducted CPR polymerase chain reaction analysis on intervertebral disc tissue and found that the expression of MMP-3 enzyme was the most obvious.The active ingredients ofLicoriceandAchyranthis bidentatae radixin CAPCM JWSZD can act on MMPs,which is found through experiments [19]that isoliquiritin can down-regulate the expression of MMP-2,MMP-9 mRNA and protein in glioma stem cells,inhibit their migration and invasion.Ke Hui found [7]Achyranthis bidentatae radixcan treat knee osteoarthritis by inhibiting the expression of MMP-3 and MMP-13.

Immune inflammation is also an important mechanism in the development of LDH.The nerve roots or ganglia that are compressed by the intervertebral disc will have an inflammatory reaction and cause varying degrees of pain,and even destroy the tissue structure of the intervertebral disc.IL-6 is an important inflammation-causing factor.Moreover,Nemoto [20] found that IL-6 caused the degradation of collagen and elastin in the intervertebral disc tissue by reducing the production and activity of MMP inhibitor-1,leading to the herniation and degeneration of the intervertebral disc.Cheng et al,found that [21]cinnamaldehyde can reduce the expression of IL-6 in the serum of rats with collagen-induced arthritis.Li Xin [22] experiments showed that quercetin could inhibit the expression of IL-6mRNA in neutrophils induced by LPS,and inhibit the secretion and synthesis of IL-6.Chen YC [23] found that wogonin might inhibit the expression of necrosis factor alpha (TNF-α)and COX-2,thereby affecting the transcription and expression of IL-6.Therefore,in CAPCM JWSZD,quercetin inGlycyrrhizae radix et rhizomaandEucommiae cortex,cinnamaldehyde inCinnamoni cortex,and wogonin inAchyranthis bidentatae radixcan reduce the inflammation of local compressive nerve tissue of intervertebral disc herniation by reducing the level of IL-6 in the serum.

In addition,IL-17 is the main pro-inflammatory factor in autoimmunity or chronic inflammation [8],which can promote the synthesis of inflammatory mediators in the intervertebral disc,the rapid apoptosis of intervertebral disc cells and the abnormal metabolism of the extracellular matrix of the intervertebral disc.Tanigawa [24] confirmed that IL-17F promotes cartilage degradation by increasing the expression of MMP-1,3,and 13 in chondrocytes and reducing the expression of TIMP-2,4,and type II collagen.IL-17 can also promote the massive increase of inflammatory mediators (PGE2,IL-6 and NO) and ICAM-1 on the surface of degenerated intervertebral disc cells through the NF-κB and MAPK pathways [9].

TNF is an important cytokine in the inflammatory response of intervertebral disc tissues.Zhang SQ [25]found that the pain degree of patients with LDH was positively correlated with the level of TNF,and Li JM[26] also found that the degree of disc herniation and fibrous annulus.The degree of damage is positively correlated with tumor TNF-α.Studies have shown [27]that TNF-α blockers can effectively reduce inflammation and delay the process of intervertebral disc degeneration.The process that TNF-α induces bone cells to produce IL-6 and monocyte chemoattractant protein-1 can be inhibited by kaempferol [28],and both quercetin and kaempferol[29] can strongly inhibited NF-κB ligand induced osteoclast differentiation marker.Quercetin [28] has the ability to regulate the production and gene expression of the pro-inflammatory cytokine TNF-α in peripheral blood mononuclear cells.Therefore,the effective compounds of kaempferol and quercetin inEucommiae cortexandAchyranthis bidentatae radixcan play a role in resisting the inflammation of lumbar intervertebral disc tissues through the TNF signaling pathway.

This study sorts out the prescriptions of the Song Dynasty court medicine for treating waist and leg pain,excavates the core herbal combination of TCMCinnamonicortex-Aconmlateralisradix praeparaia-Achyranthis bidentatae radixand the CAPCM JWSZD.The network pharmacology was used to explore the molecular mechanism of CAPCM JWSZD in the treatment of lumbar intervertebral disc herniation.It was found that the main compounds in CAPCM JWSZD,quercetin,kaempferol,cinnamaldehyde,and wogonin,interfere with the development of LDH by acting on MMP activity,negative regulation of IL-6 production,positive regulation of IL-17 production,and TNF pathways.