Original Article Utilizing Network Pharmacology to Explore the Underlying Mechanism of Cinnamomum Cassia Presl in Treating Osteoarthritis

Int J Clin Exp Med 2019;12(12):13359-13369 www.ijcem.com /ISSN:1940-5901/IJCEM0100310

Original Article Utilizing network pharmacology to explore the underlying mechanism of Cinnamomum cassia Presl in treating osteoarthritis

Guowei Zhou, Ruoqi Li, Tianwei Xia, Chaoqun Ma, Jirong Shen

Affiliated Hospital of Nanjing University of Chinese Medicine, Nanjing 210029, Jiangsu, China Received July 30, 2019; Accepted October 8, 2019; Epub December 15, 2019; Published December 30, 2019

Abstract: Objective: The network pharmacology method was adopted to establish the relationship between “Ingre- dients-Disease Targets-Biological Pathway”, screen the potential target of Cinnamomum cassia Presl in treating osteoarthritis (OA), and explore the underlying mechanism. Methods: Chemical components and selected targets related to Cinnamomum cassia Presl were retrieved from BATMAN-TCM. OA disease targets were searched from TTD, DrugBank, OMIM, GAD, PharmGKB, and DisGeNET databases. Canonical SMILES were obtained from Pub- chem, and targets were obtained from SwissTargetPrediction. We constructed a target interaction network graph by using the STRING database and Cytoscape 3.2.1, and screened key targets by using the MCC algorithm of cytoHubba. Enrichment analysis of the GO function and KEGG pathway was conducted using the DAVID database. Results: Eighteen active compounds derived from Cinnamomum cassia Presl and 40 intersections between Cin- namomum cassia Presl and OA were obtained. Ten key targets were screened by the MCC algorithm, including PTGS2, MAPK8, MMP9, ALB, MMP2, MMP1, SERPINE1, TGFB1, PLAU, and ESR1. The GO analysis consisted of 33 enrichment results, including biological processes (e.g., collagen catabolic process and inflammatory response), cell composition (e.g., extracellular space and extracellular matrix), and molecular function (e.g., heme binding and aromatase activity). A total of 32 pathways were enriched by KEGG, including osteoclast differentiation, arachidonic acid metabolism, hypoxia-inducible factor (HIF)-1, nuclear factor кB (NF-кB), Toll-like receptors (TLRs), and tumor necrosis factor (TNF). Conclusion: This study predicted the main possible mechanism of Cinnamomum cassia Presl in treating OA, including anti-inflammation, regulating cellular proliferation, differentiation and apoptosis, and an- tioxidation, which provided a theoretical basis for exploring active ingredients and experimental research about Cinnamomum cassia Presl against OA.

Keywords: Network pharmacology, Cinnamomum cassia Presl, osteoarthritis, mechanism

Introduction Cinnamomum Cassia is the dry bark of Cin- namomum cassia. Modern pharmacologic stu- Osteoarthritis (OA) is a chronic disease that dies have found that cinnamaldehyde and other occurs in articular cartilage and impacts the monomers in Cinnamomum Cassia have the ligaments, synovium, articular capsule and following properties: anti-inflammatory; regulat- subchondral bone, leading to joint deformity ing cell proliferation; antioxidant and other ther- and loss of function [1]. OA is a common dis- apeutic effects on OA [5, 6]. The toxicity is rela- ease in middle-aged and elderly patients, and tively small. These studies indicated that Cin- the disability rate is > 50% [2]. The etiology of namomum Cassia have the potential ability to OA is still unclear [3]. Modern medicine can treat OA; however, Cinnamomum Cassia has only relieve symptoms and many of them also multi-component characteristics, and its mech- have side effects [4]. It is of great value to exca- anism is relatively complex. vate potential active ingredients for the treatment of OA from traditional Chinese medicine. As a new research method, network pharmacol- OA is often referred to as “bi zheng” in ancient ogy breaks away from the traditional research Chinese medical books. Traditional Chinese mode of “one target, one disease, one drug” medicine often used Cinnamomum Cassia to [7]. The systematism of the strategy resonates treat OA. well with the holistic view of traditional Chinese Underlying mechanism of Cinnamomum cassia Presl against OA

Figure 1. Flowchart of utilizing network pharmacology to explore the underlying mechanism of Cinnamomum cassia Presl in treating osteoarthritis.

medicine (TCM), as well as the mechanism of base, can be used for target prediction of tradi- multi-ingredient, multi-pathway, and multi-tar- tional Chinese medicine components, function- get synergy in TCM [8]. In recent years, many al analysis of targets, visualization of related related research reports indicate that exploring networks, and comparative analysis of tradi- the molecular mechanism of TCM efficacy will tional Chinese medicine. The active compounds become possible [9-11]. This study analyzed in Cinnamomum Cassia were collected from the active ingredients, targets, and possible the BATMAN-TCM (http://bionet.ncpsb.org/ba- mechanisms of Cinnamomum cassia approach tman-tcm) database (with “ROUGUI” as the network pharmacology to uncover the pharma- search term) [12]. cological mechanism of Cinnamomum Cassia acting on OA. The workflow of the study on Target fishing of cinnamomum cassia Cinnamomum Cassia against OA based on network pharmacology was shown in Figure 1. All active compounds in Cinnamomum Cassia were input to the Pubchem database (ht tp s:// Methods pubchem.ncbi.nlm.nih.gov/) to obtain the Can- Active compounds screening of cinnamomum onical SMILES. SwissTargetPrediction (http:// cassia www.swisstargetprediction.ch/) is a network platform that can predict small molecule tar- BATMAN-TCM (http://bionet.ncpsb.org/batman- gets on the basis of the 2D and 3D similarity tcm), an online bioinformatics analysis data- measurements of ligands [13]. First, the Can-

13360 Int J Clin Exp Med 2019;12(12):13359-13369 Underlying mechanism of Cinnamomum cassia Presl against OA onical SMILES of compounds were submitted anthocyanidin B2, coumarin, melilotocarpan A, to the platform. Then, the property was set to cinnamyl benzoate, cinnamyl alcohol, anethole, “Homo Sapiens” to obtain the predicted tar- procyanidin B1, procyanidin B5, protocatechuic gets. The active compounds obtained from BA- acid, cinnamyl acetate, procyanidin C1, (-)-epi- TMAN-TCM were uploaded to the GeneCards catechin-3-o-gallate, trans-cinnamic acid, ethyl database (https://www.genecards.org/), the cinnamate, cinnamic ccid, styrene, and procy- human gene compendium [14], with the prop- anidin B7. erty set to “Homo Sapiens”. The intersections of GeneCards and SwissTargetPrediction were Prediction and analysis active compound tar- the targets of Cinnamomum Cassia. gets of Cinnamomum cassia

Disease targets database building and col- Eight hundred twenty-one targets were ob- lection of cinnamomum cassia-OA common tained, which were reduced to 335 after dupli- target cation removal. A total of 3247 targets were screened from the GeneCards database, and The OA-associated disease targets were sea- 1396 after banishing duplication. There were rched from TTD (ht tp s://db.idrblab.org/ttd/), 169 targets in GeneCards and SwissTarget- DrugBank (https://www.drugbank.ca/), OMIM Prediction (Figure 2). Then, the “compound-pre- (http://www.omim.org/), GAD (https://geneti- dicted” network was built by Cytoscape 3.2.1 cassociationdb.nih.gov/), PharmGKB (https:// software (Figure 3). www.pharmgkb.org/), and DisGeNET (http:// www.disgenet.org/web/DisGeNET/menu/home) Collection of OA targets and acquisition of cin- 6 databases with “osteo-arthritis” as a key namomum cassia (RG)-OA common targets word. The common targets of Cinnamomum Cassia and OA were obtained. Six hundred four targets were obtained from six databases, and the number was reduced to Construction of network and analysis 531 after removing duplication. Forty common targets were obtained that intersected with The active compounds and common targets Cinnamomum Cassia (RG)-associated targets were put into Cytoscape 3.2.1 to draw “Drug- (Figure 4). They are ABCC1, C1R, AKR1C1, CA3, Compounds-Targets Network”. The common BCHE, CA2, MMP13, CTSK, PLAU, MMP1, targets were also uploaded to the STRING data- SERPINE1, ADAM17, BCL2, TGFB1, MMP2, MM- base (http://string-db.org/) for building a pro- P9, AR, NFKB1, MAPK8, ALB, PTGS2, ESR1, tein interaction network. Next, the MCC algo- TLR9, ESR2, NOS2, CYP19A1, CYP1A1, ACHE, rithm of the cytoHubba plug-in of the Cytoscape CYP1A2, CYP2C19, ABCB1, PTGS1, CES1, AK- 3.2.1 was used to screen key targets [15]. R1C3, AKR1B1, PLA2G2A, ALOX5, MPO, TTR, and TDO2. Enrichment analysis of the go function and kegg pathway Forty targets were uploaded to the STRING database to draw the Protein-Protein Interaction Database for Annotation, Visualization and (PPI) network (Figure 5A). There were 39 nodes Integrated Discovery (DAVID, https://david.ncif- (isolated nodes were removed) and 184 sides crf.gov/, v6.8) was used for gene ontology (GO) (network nodes represent targets, and lines enrichment analysis. Pathway enrichment anal- represent the interactions of targets). The aver- ysis was accomplished using Kyoto Ency- age node degree was 9.2. “Cinnamomum Ca- clopedia of Genes and Genomes (KEGG, http:// ssia (RG)-Compounds-Targets Network” was www.kegg.jp/) data obtained from DAVID [16]. drawn by Cytoscape 3.2.1 (Figure 5B). The in- Results formation on protein interactions was uploaded to the Cytoscape 3.2.1, and the MCC algorithm Active compounds of cinnamomum cassia (the higher the score, the higher the signifi- cance) of the cytoHubba plug-in was used to A total of 18 active herbal ingredients were screen out 10 key targets (Figure 5C). Key tar- retrieved from the BATMAN-TCM (Table 1). They gets that rank from more importance-to-less are cinnamic aldehyde (cinnamaldhyde), pro- importance are PTGS2, MAPK8, MMP9, ALB,

13361 Int J Clin Exp Med 2019;12(12):13359-13369 Underlying mechanism of Cinnamomum cassia Presl against OA

Table 1. Basic information of 18 candidate compounds of Cinnamomum cassia Compound Pubchem ID Canonical SMILES Cinnamic Aldehyde/Cinnamaldehyde 637511 C1=CC=C(C=C1)C=CC=O Proanthocyanidin B2 122738 C1C(C(OC2=C1C(=CC(=C2C3C(C(OC4=CC(=CC(=C34)O)O) C5=CC(=C(C=C5)O)O)O)O)O)C6=CC(=C(C=C6)O)O)O Coumarin 323 C1=CC=C2C(=C1)C=CC(=O)O2 Melilotocarpan A 4484393 COC1=CC2=C(C=C1)C3COC4=C(C3O2)C=CC(=C4O)OC Cinnamyl benzoate 5705112 C1=CC=C(C=C1)C=CCOC(=O)C2=CC=CC=C2 Cinnamyl alcohol 5315892 C1=CC=C(C=C1)C=CCO Anethole 637563 CC=CC1=CC=C(C=C1)OC Procyanidin B1 11250133 C1C(C(OC2=C1C(=CC(=C2C3C(C(OC4=CC(=CC(=C34)O)O) C5=CC(=C(C=C5)O)O)O)O)O)C6=CC(=C(C=C6)O)O)O Procyanidin B5 124017 C1C(C(OC2=CC(=C(C(=C21)O)C3C(C(OC4=CC(=CC(=C34)O) O)C5=CC(=C(C=C5)O)O)O)O)C6=CC(=C(C=C6)O)O)O Protocatechuic Acid 72 C1=CC(=C(C=C1C(=O)O)O)O Cinnamyl acetate 5282110 CC(=O)OCC=CC1=CC=CC=C1 Procyanidin C1 169853 C1C(C(OC2=C1C(=CC(=C2C3C(C(OC4=C(C(=CC(=C34)O) O)C5C(C(OC6=CC(=CC(=C56)O)O)C7=CC(=C(C=C7)O)O)O) C8=CC(=C(C=C8)O)O)O)O)O)C9=CC(=C(C=C9)O)O)O (-)-Epicatechin-3-O-Gallate 107905 C1C(C(OC2=CC(=CC(=C21)O)O)C3=CC(=C(C=C3)O)O) OC(=O)C4=CC(=C(C(=C4)O)O)O Trans-Cinnamic Acid 444539 C1=CC=C(C=C1)C=CC(=O)O Ethyl cinnamate 637758 CCOC(=O)C=CC1=CC=CC=C1 Cinnamic Acid 444539 C1=CC=C(C=C1)C=CC(=O)O Styrene 7501 C=CC1=CC=CC=C1 Procyanidin B7 474541 C1C(C(OC2=CC(=C(C(=C21)O)C3C(C(OC4=CC(=CC(=C34)O) O)C5=CC(=C(C=C5)O)O)O)O)C6=CC(=C(C=C6)O)O)O

Enrichment analysis of the go function and kegg pathway

A total of 48 GO enrichment results were obtained during this study, and 33 results were screened out according to P < 0.05. The top 20 results were shown in Table 2. They are collagen catabolic process, heme binding, extracellular space, extracellular matrix, aromatase activity, and so on.

Similarly, 36 KEGG enrichment results were obtained in this analysis, and 28 results were collected based on the threshold of P < 0.05. The Top Figure 2. Venn diagram of Cinnamomum cassia targets. The blue circle rep- 20 results were shown in resents the targets predicted by the Genecards, and the red circle represents the targets predicted by the SwissTargetPrediction. Figure 6. They are pathways in cancer, toxoplasmosis, Chag- as disease, MicroRNAs in can- MMP2, MMP1, SERPINE1, TGFB1, PLAU, and cer, tuberculosis, ovarian steroidogenesis, Lei- ESR1. shmaniasis, prolactin signaling pathway, hepa-

13362 Int J Clin Exp Med 2019;12(12):13359-13369 Underlying mechanism of Cinnamomum cassia Presl against OA

Figure 3. Compound-predicted targets network. The square nodes represent the compounds of Cinnamomum Cassia, the round nodes represent the targets. The node color changes from orange-to-blue reflect the degree value changes from low-to-high in the network.

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that the active compounds of Cinnamomum Cassia have more than one targets. Such targets can also interact with multiple compounds. More- over, 10 key targets can be further obtained from the MCC algorithm of the cytoHub- ba plug-in of Cytoscape 3.2.1. There are PTGS2, MAPK8, MMP9, ALB, MMP2, MMP1, SERPINE1, TGFB1, PLAU, and ESR1. The results of KEGG pathway enrichment showed that pathways in cancer, To- xoplasmosis, Chagas disease, Figure 4. Venn diagram of the common targets both in Cinnamomum Cas- microRNAs in cancer, and tu- sia (RG) and OA targets network construction and analysis. The blue circle berculosis were the most sig- represents the 169 targets with RG, and the yellow circle represents the 531 nificant signaling pathways targets with OA. (Figure 3). These pathways are complex and many down- titis B, small cell lung cancer, NF-kappaB signal- stream pathways are included. For example, ing pathway, estrogen signaling pathway, HIF-1 downstream pathways of cancer signaling pa- signaling pathway, proteoglycans in cancer, toll- thways include the Wnt signaling pathway, like receptor signaling pathway, TNF signaling Hedgehog signaling pathway, Notch signaling pathway, bladder cancer, Sphingolipid signaling pathway, MAPK signaling pathway, and vas- pathway, tryptophan metabolism, and osteo- cular endothelial growth factor signaling pa- clast differentiation. thways.

Discussion In combination with the common OA signaling pathways and after the exclusion of cancer- According to the investigations, it was suggest- associated pathways and other broad path- ed that the global age-standardized prevalence ways, Cinnamomum Cassia can be predicted to of knee OA is 3.8% [1]. With the development of have effects in treating OA by an anti-inflamma- molecular biology, disease-modifying osteoar- tory effect, and mediate cell proliferation, dif- thritis drugs have become a major focus of ferentiation, and apoptosis, thus promoting the research. The positive effects and relatively balance between osteoblast (OB) and osteo- small toxicity of Cinnamomum Cassia in treat- clast (OC) and the antioxidant effects. ing OA have attracted the attention of scien- tists and researchers [5, 6]. Anti-inflammatory

Eighteen compounds of Cinnamomum Cassia Enrichment analysis results showed that the were retrieved from the BATMAN-TCM database inflammation-associated signaling pathways in this study. The targets of such compounds mainly enrich NF-кB, TLRs, TNF, arachidonic were further predicted by Swisstarget predic- acid metabolism-associated signaling path- tion and were obtained in combination with ways and other signaling pathways. Inflam- GeneCards database retrieval, thus 169 Cinna- matory factors, such as IL-1β, play an important momum Cassia-associated targets were ob- role in the pathogenesis of OA [17]. IL-1β is an tained. OA-associated targets were retrieved important inflammatory factor causing OA and from the 6 online databases. A total of 40 in- can aggravate OA-associated inflammation by tersection targets between OA targets and activating NF-κB and other signaling pathways, Cinnamomum Cassia-associated targets were leading to an increase in protease expression obtained. It can be seen from the PPI diagram in the downstream of inflammatory reaction, and the “drug-compounds-targets” diagram accelerating the degradation of proteoglycan

13364 Int J Clin Exp Med 2019;12(12):13359-13369 Underlying mechanism of Cinnamomum cassia Presl against OA

Figure 5. A. Protein-protein interaction. B. Drug- Compound-Target Network. The red squares represent the drug, Cinnamomum Cassia (RG); green triangles represent compounds; and yellow rounds represent targets. C. Key Target Proteins. The node color changes from yellow-to-red reflect the MCC value changes from low-to-high in the network.

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Table 2. GO entries of targets (P < 0.05) Category GO ID Discription Genes P Value BP GO:0030574 collagen catabolic process 5 8.18E-09 MF GO:0020037 heme binding 8 4.07E-08 CC GO:0005615 extracellular space 12 1.23E-06 CC GO:0031012 extracellular matrix 5 7.00E-05 MF GO:0070330 aromatase activity 3 1.54E-04 MF GO:0004222 metalloendopeptidase activity 5 2.26E-04 MF GO:0004601 peroxidase activity 3 6.68E-04 BP GO:0045893 positive regulation of transcription, DNA-templated 5 0.001653109 MF GO:0005496 steroid binding 3 0.001704889 MF GO:0008270 zinc ion binding 9 0.003388197 MF GO:0005506 iron ion binding 4 0.005819584 MF GO:0003990 acetylcholinesterase activity 2 0.006506693 BP GO:0010469 regulation of receptor activity 2 0.007896778 MF GO:0004497 monooxygenase activity 3 0.008190211 BP GO:0006979 response to oxidative stress 3 0.008534181 CC GO:0072562 blood microparticle 3 0.009127845 BP GO:0001666 response to hypoxia 3 0.009166887 MF GO:0004666 prostaglandin-endoperoxide synthase activity 2 0.009744585 BP GO:0019371 cyclooxygenase pathway 2 0.010515617 BP GO:0006954 inflammatory response 4 0.012832271 and collagen, and damaging articular cartilage patients [28]. Wu et al. found that administra- [18-23]. The preliminary study of this research tion of cinnamaldehyde promotes osteogenesis group showed that cinnamaldehyde can pro- in ovariectomized rats and differentiation of tect cartilage by mediating signal pathways osteoblast in vitro [29]. The results of western such as NF-κB to resist IL-1β-induced inflamma- blot supported that cinnamaldehyde promotes tion on chondrocytes [6]. In addition, type-A the maturation and differentiation of osteo- procyanidine polyphenols extracted from the blasts for cinnamaldehyde and apparently en- bark of Cinnamomum zeylanicum was also hances the expression of runx2, ocn and col1a1 reported to have disease-modifying potential in [29]. Moreover, with the treatment of cinnamal- animal models of inflammation and arthritis in dehyde, the number of osteoclasts in ovariecto- rats [24]. Youn et al. have reported that cinnam- mized rats is gradually decreased, so bone aldehyde suppressed the activation of NF-κB resorption activity was inhibited [29]. Zhang et and IRF3 induced by LPS, leading to the de- al. found that cinnamaldehyde inhibits prolifer- creased expression of target genes, such as ation and bone resorption of osteoclast induced cyclooxygenase-2 (COX-2) and Interferon-β by dexamethasone, which may be mediated by (IFN-β), in RAW264.7 macrophages [25]. down-regulation of receptor activator of NF kappaB (RANK) and nuclear factor of activating Mediate cell proliferation, differentiation, T cells 1 (NFATc1) mRNA [30]. apoptosis, and promote the balance between OB and OC Antioxidant effects

In the enrichment results, such OC differentia- Previous studies have shown that the mito- tion-related pathways, such as HIF-1, and the chondrial dysfunction of chondrocytes and downstream pathways of cancer signaling pa- synovial cells in patients with OA produces a thways, such as the Wnt [26] and MAPK signal- large amount of NO and reactive oxygen spe- ing pathways, are also closely correlated to cies, which can cause oxidative damage. Accor- such effects [27]. Wang et al. found that a high ding to the previous study, cinnamaldehyde concentration of cinnamaldehyde significantly can increase the activity of catalase, superox- inhibits the growth of synovial fibroblasts in OA ide dismutase and glutathione peroxidase, and

13366 Int J Clin Exp Med 2019;12(12):13359-13369 Underlying mechanism of Cinnamomum cassia Presl against OA

Figure 6. KEGG pathway entries of targets. inhibit the oxidation of chondrocytes. Cinna- network pharmacology to provide thoughts and momum Cassia polyphenols also have strong a basis for future studies. Due to the limitation reducibility [31]; and the higher the content, the of certain conditions, this study had the follow- stronger the antioxidant ability [32]. In addition, ing shortcomings. First, TCM often uses Cin- enrichment analysis results showed that the namomum Cassia as a decoction, but this estrogen, prolactin, sphingolipid, and other sig- study did not consider the absorption and naling pathways indirectly affect the growth metabolism of monomers in the body. Second, and repair of articular cartilage, thus playing a the content of different compounds in Cin- role in treating OA. namomum Cassia and the interactions (syner- Conclusion gism or antagonism) between monomers after oral administration were not taken into account. In conclusion, the complex mechanism of Cin- Third, the number of Cinnamomum Cassia-OA namomum Cassia in treating OA with multiple intersection targets obtained was small. We components, targets, and pathways was stud- believe that with the development of technolo- ied with a theoretical approach in this study by gy and in-depth study, the underlying mecha-

13367 Int J Clin Exp Med 2019;12(12):13359-13369 Underlying mechanism of Cinnamomum cassia Presl against OA nisms of Cinnamomum Cassia in treating OA in England. Rheumatology (Oxford) 2015; 54: will be more adequately revealed. 2051-2060. [5] Kim ME, Na JY and Lee JS. Anti-inflammatory Acknowledgements effects of trans-cinnamaldehyde on lipopolysaccharide-stimulated macrophage activation We are grateful to all the coworkers and part- via MAPKs pathway regulation. Immunophar- macol Immunotoxicol 2018; 40: 219-224. ners in this study. This work was supported by [6] Xia T, Gao R, Zhou G, Liu J, Li J and Shen J. Jiangsu Province Hospital of Chinese Medicine Trans-cinnamaldehyde inhibits IL-1beta-stimu- Project (Y2018CX54). lated inflammation in chondrocytes by - sup pressing NF-kappaB and p38-JNK pathways Disclosure of conflict of interest and exerts chondrocyte protective effects in a rat model of osteoarthritis. Biomed Res Int None. 2019; 2019: 4039472. [7] Hopkins AL. Network pharmacology: the next Abbreviations paradigm in drug discovery. Nat Chem Biol 2008; 4: 682-690. OA, Osteoarthritis; TCM, traditional Chinese me- [8] Wan Y, Xu L, Liu Z, Yang M, Jiang X, Zhang Q dicine; NF-Κb, nuclear factor-kappa B; MAPKs, and Huang J. Utilising network pharmacology mitogen-activated protein kinase; HIF, hypoxia- to explore the underlying mechanism of Wu- inducible factor-1; NF-кB, nuclear factor кB; mei Pill in treating pancreatic neoplasms. BMC Complement Altern Med 2019; 19: 158. TLRs, Toll-like receptors; TNF, tumor necrosis [9] Gao L, Wang XD, Niu YY, Duan DD, Yang X, Hao factor; LPS, lipopolysaccharide; COX-2, cyclo- J, Zhu CH, Chen D, Wang KX, Qin XM and Wu oxygenase-2; IFN-β, interferon-β; OB, osteo- XZ. Molecular targets of Chinese herbs: a clini- blast; OC, osteoclast; RANK, receptor activator cal study of hepatoma based on network phar- of NF kappaB; NFATc1, nuclear factor of activat- macology. Sci Rep 2016; 6: 24944. ing T cells 1. [10] Zhang B, Wang X and Li S. An integrative platform of TCM network pharmacology and its ap- Address correspondence to: Chaoqun Ma and plication on a herbal formula, Qing-Luo-Yin. Jirong Shen, Affiliated Hospital of Nanjing University Evid Based Complement Alternat Med 2013; of Chinese Medicine, Nanjing 210029, Jiangsu, 2013: 456747. China. E-mail: [email protected] (CQM); joint- [11] Li H, Zhao L, Zhang B, Jiang Y, Wang X, Guo Y, Liu H, Li S and Tong X. A network pharmacology [email protected] (JRS) approach to determine active compounds and References action mechanisms of ge-gen-qin-lian decoction for treatment of type 2 diabetes. Evid [1] Cross M, Smith E, Hoy D, Nolte S, Ackerman I, Based Complement Alternat Med 2014; 2014: Fransen M, Bridgett L, Williams S, Guillemin F, 495840. Hill CL, Laslett LL, Jones G, Cicuttini F, Osborne [12] Liu Z, Guo F, Wang Y, Li C, Zhang X, Li H, Diao L, R, Vos T, Buchbinder R, Woolf A and March L. Gu J, Wang W, Li D and He F. BATMAN-TCM: a The global burden of hip and knee osteoarthri- bioinformatics analysis tool for molecular tis: estimates from the global burden of dis- mechANism of traditional chinese medicine. ease 2010 study. Ann Rheum Dis 2014; 73: Sci Rep 2016; 6: 21146. 1323-1330. [13] Gfeller D, Grosdidier A, Wirth M, Daina A, Mich- [2] Hootman JM, Helmick CG, Barbour KE, Theis ielin O and Zoete V. SwissTargetPrediction: a KA and Boring MA. Updated projected preva- web server for target prediction of bioactive lence of self-reported doctor-diagnosed arthri- small molecules. Nucleic Acids Res 2014; 42: tis and arthritis-attributable activity limitation W32-38. among US adults, 2015-2040. Arthritis Rheu- [14] Stelzer G, Rosen N, Plaschkes I, Zimmerman S, matol 2016; 68: 1582-1587. Twik M, Fishilevich S, Stein TI, Nudel R, Lieder [3] Martel-Pelletier J, Barr AJ, Cicuttini FM, I, Mazor Y, Kaplan S, Dahary D, Warshawsky D, Conaghan PG, Cooper C, Goldring MB, Gold- Guan-Golan Y, Kohn A, Rappaport N, Safran M ring SR, Jones G, Teichtahl AJ and Pelletier JP. and Lancet D. The genecards suite: from gene Osteoarthritis. Nat Rev Dis Primers 2016; 2: data mining to disease genome sequence 16072. analyses. Curr Protoc Bioinformatics 2016; 54: [4] Yu D, Peat G, Bedson J and Jordan KP. Annual 1.30.1-1.30.33. consultation incidence of osteoarthritis esti- [15] Chin CH, Chen SH, Wu HH, Ho CW, Ko MT and mated from population-based health care data Lin CY. cytoHubba: identifying hub objects and

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