[Transcriptome data mining combined with clinical and animal experiments for identification of syndrome element marker genes during entire course of steroid-induced osteonecrosis of femoral head].

A research strategy combining transcriptome data mining and experimental verification was adopted to identify the marker genes characterizing the syndrome elements of phlegm, stasis, and deficiency in steroid-induced osteonecrosis of the femoral head(SONFH). Firstly, the common differentially expressed gene sets of SONFH with the syndromes of phlegm-stasis obstructing collaterals, vessel obstruction, and liver-kidney deficiency were obtained from the clinical transcriptomic analysis of a previous study. The differential expression trend analysis and functional gene mining were then employed to predict the candidate marker gene sets representing phlegm, stasis, and deficiency. The whole blood samples from SONFH patients, whole blood samples from SONFH rats, and affected femoral head tissue samples were collected for qPCR, which aimed to determine the expression levels of the candidate marker genes mentioned above. Furthermore, the receiver operating characteristic curve(ROC) was established to objectively evaluate the syndrome differentiation effectiveness of the candidate marker genes mentioned above. The transcriptome data analysis results showed that the candidate marker genes for phlegm was ELOVL fatty acid elongase 6(ELOVL6), and those for stasis were ankyrin 1(ANK1), glycophorin A/B(GYPA/B), and Rh-associated glycoprotein(RHAG). The candidate marker genes for deficiency were solute carrier family 2 member 1(SLC2A1) and stomatin(STOM). The qPCR results showed that compared with that in the non-SONFH group, ELOVL6 had the lowest expression level in the peripheral blood of the SONFH patients with the syndrome of phlegm-stasis obstructing collaterals(P<0.05). Compared with that in the normal control group, ELOVL6 had the lowest expression level in the peripheral blood and affected femoral head tissue of SONFH rats modeled for 4 weeks(P<0.01), and it showed better syndrome differentiation effectiveness of rats modeled for 4 weeks(AUC=0.850, P=0.006) than at other modeling time points(8, 12, 16, and 21 weeks, AUC of 0.689, 0.766, 0.588, and 0.662, respectively). Compared with that in the non-SONFH group, the expression levels of ANK1, GYPA, and RHAG were the lowest in the peripheral blood of SONFH patients with the vessel obstruction syndrome(P<0.05). The expression levels of the three genes were the lowest in the peripheral blood and affected femoral head tissue of SONFH rats modeled for 12 weeks(P<0.05, P<0.01), and their syndrome differentiation effectiveness in the rats modeled for 12 weeks(GYPA: AUC=0.861, P=0.012; ANK1: AUC=0.855, P=0.006; RHAG: AUC=0.854, P=0.009) was superior to that for 4, 8, 16, and 21 weeks(GYPA: AUC=0.646, 0.573, 0.691, and 0.617, respectively; ANK: AUC1=0.630, 0.658, 0.657, and 0.585, respectively; RHAG: AUC=0.592, 0.511, 0.515, and 0.536, respectively). Compared with the non-SONFH group, both SLC2A1 and STOM had the lowest expression levels in the peripheral blood of patients with the syndrome of liver and kidney deficiency(P<0.05). Compared with the normal control group, their expression levels were the lowest in the peripheral blood and affected femoral head tissue of SONFH rats modeled for 21 weeks(P<0.05, except STOM in the peripheral blood of rats). Moreover, the syndrome differentiation effectiveness of SLC2A1 in the rats modeled for 21 weeks(AUC=0.806, P=0.009) was superior to that for 4, 8, 12, and 16 weeks(AUC=0.520, 0.580, 0.741, 0.774, respectively), and STOM was meaningless in syndrome differentiation. In summary, the candidate marker gene for phlegm in SONFH is ELOVL6; the candidate marker genes for stasis are GYPA, RHAG, and ANK1; the candidate marker gene for deficiency is SLC2A1. The results help to reveal the biological connotations of phlegm, stasis, and deficiency in SONFH at the genetic level.

[Characteristic changes of blood stasis syndrome in rat model of steroid-induced femoral head necrosis based on the combination of disease, syndrome, and symptom].

The approach combining disease, syndrome, and symptom was employed to investigate the characteristic changes of blood stasis syndrome in a rat model of steroid-induced osteonecrosis of the femoral head(SONFH) during disease onset and progression. Seventy-two male SD rats were randomized into a healthy control group and a model group. The rat model of SONFH was established by injection of lipopolysaccharide(LPS) in the tail vein at a dose of 20 µg·kg~(-1)·d~(-1) on days 1 and 2 and gluteal intramuscular injection of methylprednisolone sodium succinate(MPS) at a dose of 40 mg·kg~(-1)·d~(-1) on days 3-5, while the healthy control group received an equal volume of saline. The mechanical pain test, tongue color RGB technique, gait detection, open field test, and inclined plane test were employed to assess hip pain, tongue color, limping, joint activity, and lower limb strength, respectively, at different time points within 21 weeks of modeling. At weeks 2, 4, 8, 12, 16, and 21 after modeling, histopathological changes of the femoral head were observed by hematoxylin-eosin(HE) staining and micro-CT scanning; four coagulation items were measured by rotational thromboelastometry; and enzyme-linked immunosorbent assay(ELISA) was employed to determine the levels of six blood lipids, vascular endothelial growth factor(VEGF), endothelin-1(ET-1), nitric oxide(NO), tissue-type plasminogen activator(t-PA), plasminogen activator inhibitor factor-1(PAI-1), bone gla protein(BGP), alkaline phosphatase(ALP), receptor activator of nuclear factor-κB(RANKL), osteoprotegerin(OPG), and tartrate-resistant acid phosphatase 5b(TRAP5b) in the serum, as well as the levels of 6-keto-prostaglandin 1α(6-keto-PGF1α) and thromboxane B2(TXB2) in the plasma. The results demonstrated that the pathological alterations in the SONFH rats were severer over time. The bone trabecular area ratio, adipocyte number, empty lacuna rate, bone mineral density(BMD), bone volume/tissue volume(BV/TV), trabecular thickness(Tb.Th), trabecular number(Tb.N), bone surface area/bone volume(BS/BV), and trabecular separation(Tb.Sp) all significantly increased or decreased over the modeling time after week 4. Compared with the healthy control group, the mechanical pain threshold, gait swing speed, stride, standing time, and walking cycle of SONFH rats changed significantly within 21 weeks after modeling, with the greatest difference observed 12 weeks after modeling. The time spent in the central zone, rearing score, and maximum tilt angle in the open field test of SONFH rats also changed significantly over the modeling time. Compared with the healthy control group, the R, G, and B values of the tongue color of the model rats decreased significantly, with the greatest difference observed 11 weeks after modeling. The levels of total cholesterol(TC), total triglycerides(TG), low-density lipoprotein-cholesterol(LDL-C), and apoprotein B(ApoB) in the SONFH rats changed significantly 4 and 8 weeks after modeling. The levels of VEGF, ET-1, NO, t-PA, PAI-1, 6-keto-PGF1α, TXB2, four coagulation items, and TXB2/6-keto-PGF1α ratio in the serum of SONFH rats changed significantly 4-16 weeks after modeling, with the greatest differences observed 12 weeks after modeling. The levels of BGP, TRAP5b, RANKL, OPG, and RANKL/OPG ratio in the serum of SONFH rats changed significantly 8-21 weeks after modeling. During the entire onset and progression of SONFH in rats, the blood stasis syndrome characteristics such as hyperalgesia, tongue color darkening, gait abnormalities, platelet, vascular, and coagulation dysfunctions were observed, which gradually worsened and then gradually alleviated in the disease course(2-21 weeks), with the most notable differences occurred around 12 weeks after modeling.

Sulforaphane, an NRF2 agonist, alleviates ferroptosis in acute liver failure by regulating HDAC6 activity.

OBJECTIVE: Acute liver failure (ALF) is characterized by severe liver dysfunction, rapid progression and high mortality and is difficult to treat. Studies have found that sulforaphane (SFN), a nuclear factor E2-related factor 2 (NRF2) agonist, has anti-inflammatory, antioxidant and anticancer effects, and has certain protective effects on neurodegenerative diseases, cancer and liver fibrosis. This paper aimed to explore the protective effect of SFN in ALF and it possible mechanisms of action. METHODS: Lipopolysaccharide and D-galactosamine were used to induce liver injury in vitro and in vivo. NRF2 agonist SFN and histone deacetylase 6 (HDAC6) inhibitor ACY1215 were used to observe the protective effect and possible mechanisms of SFN in ALF, respectively. Cell viability, lactate dehydrogenase (LDH), Fe2+, glutathione (GSH) and malondialdehyde (MDA) were detected. The expression of HDAC6, NRF2, glutathione peroxidase 4 (GPX4), acyl-CoA synthetase long-chain family member 4 (ACSL4) and solute carrier family 7 member 11 (SLC7A11) were detected by Western blotting and immunofluorescence. RESULTS: Our results show that NRF2 was activated by SFN. LDH, Fe2+, MDA and ACSL4 were downregulated, while GSH, GPX4 and SLC7A11 were upregulated by SFN in vitro and in vivo, indicating the inhibitory effect of SFN on ferroptosis. Additionally, HDAC6 expression was decreased in the SFN group, indicating that SFN could downregulate the expression of HDAC6 in ALF. After using the HDAC6 inhibitor, ACY1215, SFN further reduced HDAC6 expression and inhibited ferroptosis, indicating that SFN may inhibit ferroptosis by regulating HDAC6 activity. CONCLUSION: SFN has a protective effect on ALF, and the mechanism may include reduction of ferroptosis through the regulation of HDAC6. Please cite this article as: Zhang YQ, Shi CX, Zhang DM, Zhang LY, Wang LW, Gong ZJ. Sulforaphane, an NRF2 agonist, alleviates ferroptosis in acute liver failure by regulating HDAC6 activity. J Integr Med. 2023; 21(5): 464-473.

[Identification and validation of novel biomarkers for cold-dampness syndrome of rheumatoid arthritis based on integration of multiple bioinformatics methods].

This study aims to identify the novel biomarkers of cold-dampness syndrome(RA-Cold) of rheumatoid arthritis(RA) by gene set enrichment analysis(GSEA), weighted gene correlation network analysis(WGCNA), and clinical validation. Firstly, transcriptome sequencing was carried out for the whole blood samples from RA-Cold patients, RA patients with other traditional Chinese medicine(TCM) syndromes, and healthy volunteers. The differentially expressed gene(DEG) sets of RA-Cold were screened by comparison with the RA patients with other TCM syndromes and healthy volunteers. Then, GSEA and WGCNA were carried out to screen the key DEGs as candidate biomarkers for RA-Cold. Experimentally, the expression levels of the candidate biomarkers were determined by RT-qPCR for an independent clinical cohort(not less than 10 cases/group), and the clinical efficacy of the candidates was assessed using the receiver operating characteristic(ROC) curve. The results showed that 3 601 DEGs associated with RA-Cold were obtained, including 106 up-regulated genes and 3 495 down-regulated genes. The DEGs of RA-Cold were mainly enriched in the pathways associated with inflammation-immunity regulation, hormone regulation, substance and energy metabolism, cell function regulation, and synovial pannus formation. GSEA and WGCNA showed that recombinant proteasome 26S subunit, ATPase 2(PSMC2), which ranked in the top 50% in terms of coefficient of variation, representativeness of pathway, and biological modules, was a candidate biomarker of RA-Cold. Furthermore, the validation results based on the clinical independent sample set showed that the F1 value, specificity, accuracy, and precision of PSMC2 for RA-Cold were 70.3%, 61.9%, 64.5%, and 81.3%, respectively, and the area under the curve(AUC) value was 0.96. In summary, this study employed the "GSEA-WGCNA-validation" integrated strategy to identify novel biomarkers of RA-Cold, which helped to improve the TCM clinical diagnosis and treatment of core syndromes in RA and provided an experimental basis for TCM syndrome differentiation.

[Mechanism of Panlongqi Tablets intervening in vertebral artery type of cervical spondylosis in rats through PI3K/AKT signaling pathway based on network pharmacology and experimental verification].

This study aimed to further explore the relevant mechanism of action by network pharmacology integrated with animal experimental verification based on previous proven effective treatment of vertebral artery type of cervical spondylosis(CSA) by Panlongqi Tablets. Bionetwork analysis was performed to establish drug-disease interaction network, and it was found that the key candidate targets of Panlongqi Tablets were enriched in multiple signaling pathways related to CSA pathological links, among which phosphatidylinositol 3-kinase(PI3 K)/serine-threonine kinase(AKT/PKB) signaling pathway was the most significant. Further, mixed modeling method was used to build the CSA rat model, and the rats were divided into normal, model, Panlongqi Tablets low-, medium-and high-dose(0.16, 0.32, 0.64 g·kg~(-1)) and Jingfukang Granules(positive drug, 1.35 g·kg~(-1)) groups. After successful modeling, the rats were administered for 8 consecutive weeks. Pathological changes of rat cervical muscle tissues were detected by hematoxylin-eosin(HE) staining, and the content of interleukin-1ß(IL-1ß), tumor necrosis factor-α(TNF-α), vascular endothelial cell growth factor(VEGF) and chemokine(C-C motif) ligand 2(CCL2) in rat serum and/or cervical tissues was determined by enzyme-linked immunosorbent assay(ELISA). Western blot was employed to detect the protein expression levels of chemokine(C-C motif) receptor 2(CCR2), PI3 K, AKT, phosphorylated AKT(p-AKT), I-kappa-B-kinase beta(IKK-beta/IKKß), nuclear factor kappa B(NF-κB P65) and phosphorylated nuclear factor kappa B(NF-κB p-P65) in rat cervical tissues, and positive expression of p-NF-κB P65 in rat cervical muscle tissues was detected by immunofluorescence. The results showed that Panlongqi Tablets at different doses improved the degree of muscle fibrosis and inflammation in cervical muscle tissues of CSA rats, and reduced the content of inflammatory factors IL-1ß, TNF-α, VEGF, CCL2 and CCR2 in serum and/or cervical tissues. The protein expression levels of PI3 K, p-AKT, IKKß and p-NF-κB P65 as well as the nuclear entry of p-NF-κB P65 in cervical tissues were down-regulated. These findings suggest that Panlongqi Tablets can significantly inhibit the inflammatory response of CSA rats, and the mechanism of action may be related to the down-regulation of the activation of PI3 K/AKT signaling pathway.

[Mechanism of Tianhe Zhuifeng Ointment in treating rheumatoid arthritis with syndrome of internal obstruction and cold-dampness and compatibility principles based on "disease-syndrome-formula" association network].

This study aims to explore the mechanism of Tianhe Zhuifeng Ointment in treating rheumatoid arthritis(RA) with syndrome of internal obstruction and cold-dampness and the compatibility characteristics based on the "disease-syndrome-formula" association network. A gene set associated with the clinical symptoms of RA was collected from Integrative Pharmacology-based Research Platform of Traditional Chinese Medicine v2.0(TCMIP v2.0). The different expression gene set of RA with syndrome of internal obstruction and cold-dampness was screened out by transcriptomic expression profile detection and bioinformatics data mining of the comparison of RA patients with syndrome of internal obstruction and cold-dampness and healthy volunteers. The chemical composition information of 35 Chinese medicines from Tianhe Zhuifeng Ointment was collected from TCMIP v2.0 and Traditional Chinese Medicine Bank(TCMBank). The candidate targets were predicted based on the similarity principle of compounds structure. The interactive network of "related gene of RA with syndrome of internal obstruction and cold-dampness-candidate target of Tianhe Zhuifeng Ointment" was constructed. The core network targets were screened out by topological characteristics of calculating network, and the functional exploration was carried out based on Kyoto Encyclopedia of Genes and Genomes(KEGG) and Reactome Pathway Database. The compatibility mechanisms of various efficacy groups of Tianhe Zhuifeng Ointment were further explored. The results showed that the candidate targets of Tianhe Zhuifeng Ointment were mainly involved into the modules of "immune-inflammation" regulation, nervous system function, cell function, and substance and energy metabolism, etc. The mechanisms of various efficacy groups emphasized on different aspects. The group of dispelling wind and removing dampness-dredging channels and activating collaterals, the group of extinguishing wind and stopping convulsions, and the group of pungent analgesia regulated "immune-inflammation" system by warming meridians and dissipating cold. The group of activating blood and resolving stasis and the group of strengthening sinews and bones regulated "immune-inflammation" system by activating blood and dredging channels. The group of dispelling wind and removing dampness-dredging channels and activating collaterals, the group of extinguishing wind and stopping convulsions, the group of activating blood and resolving stasis, the group of strengthening sinews and bones, and the group of clearing heat and draining water affected the nervous system by invigorating Qi-blood and benefiting spirit. The group of dispelling wind and removing dampness-dredging channels and activating collaterals and the group of extinguishing wind and stopping convulsions regulated cell function and substance and energy metabolism by dispelling wind and eliminating cold-dampness. The group of activating blood and resolving stasis and the group of strengthening sinews and bones regulated the cell function and substance and energy metabolism by activating blood and strengthening sinews and bones. The results showed that Tianhe Zhuifeng Ointment exerted the comprehensive efficacy of dispelling wind, removing dampness, activating blood, removing stasis, warming meridians, dredging channels, and strengthening sinews and bones through adjusting the imbalance of "immune-inflammation", regulating nervous system, cell function, and interfering with substance and energy metabolism, thus improving the syndrome of internal obstruction and cold-dampness. The findings of this study laid foundations for clarifying the therapeutic characteristics and clinical orientation of Tianhe Zhuifeng Ointment.

[Biological connotation of four traditional Chinese medicine syndromes of rheumatoid arthritis based on "disease-syndrome-symptom" association network].

The present study explored the biological connotation of traditional Chinese medicine(TCM) syndromes of rheumatoid arthritis(RA) from the "disease-syndrome-symptom" association network. RA patients with four TCM syndromes(dampness-heat obstruction, phlegm-stasis obstruction, Qi-blood deficiency, and liver and kidney deficiency), three for each type, were assigned as the RA TCM syndrome group, and three healthy volunteers as the normal control group. The differential gene sets of four syndromes were screened out through transcriptome expression profiling and bioinformatics mining. The relevant gene sets of syndrome-related clinical symptoms were collected from TCMIP v2.0(http://www.tcmip.cn/). The "disease-syndrome-symptom" association networks of four RA syndromes were established by using the intersection genes of syndrome-related differential genes and symptom-related genes, and the key network target genes of each syndrome were screened out and the corresponding biological functions were mined through topological feature calculation and enrichment analysis. The genes associated with clinical symptoms such as vasculitis, joint pain, and fever in the damp-heat obstruction syndrome ranked the top, and the key network target genes of this syndrome were most significantly enriched in the pathways related to material and energy metabolism and thermal reaction biological processes. The clinical symptom-related genes of the phlegm-stasis obstruction syndrome were most significantly enriched in the pathways related to "immunity-inflammation", nervous system regulation, and sensory response. The clinical symptoms such as hypoglycemia, hypotension, weight loss, palpitation, and arrhythmia in Qi-blood deficiency syndrome were predominant, and its key network target genes were most significantly enriched in the pathways related to the nervous system and "immunity-inflammation" response. The abnormal symptoms in the liver and kidney in the liver and kidney deficiency syndrome were commonly seen, and its key network target genes were most significantly enriched in the "immunity-inflammation" regulatory pathways, and liver and kidney development and metabolic response. In conclusion, the differences and connections of the biological basis between different TCM syndromes of RA are in line with the theoretical interpretation of TCM on the etiology and pathogenesis of RA. This study summarized the objective essence of syndromes to a certain extent from the "disease-syndrome-symptom" association network and is expected to provide a theoretical basis for the discovery of serum biomarkers of RA syndromes.

[Exploration on scientific connotation of TCM syndromes and recommended prescriptions against COVID-19 based on TCMTP V2.0].

Coronavirus disease 2019(COVID-19) has attracted great attentions from the whole world. Traditional Chinese medicine(TCM) has been widely used and shown satisfying efficacies in treating all stages of COVID-19. In this study, the molecular interaction networks of different stages of COVID-19(the early, severe, critical and recovery stage) were constructed using the links among symptoms-related genes collected from TCMIP V2.0(http://www.tcmip.cn/), an integrated pharmacology network-computing platform for TCM. Following the network topological feature calculation and functional enrichment analysis, we found that the molecular targets and pathways related with the "immune-inflammation system" were involved throughout all the stages of COVID-19. The severe stage and the critical period of COVID-19 were occupied by a large proportion of inflammatory factors and pathways, suggesting that there might be a cytokine storm in these periods, along with respiratory disorders, cardiopulmonary dysfunction, nervous system disorders, etc. Accordingly, the therapeutic targets and pathways hit by the recommended prescriptions against COVID-19 were also aimed to regulate the balance of immune-inflammation system, nutrient absorption and metabolism, abnormal energy metabolism, the cardio-pulmonary function, nerve system function, etc., which may be related to the therapeutic effects of these prescriptions in terms of several clinical symptoms, such as expiratory dyspnea, chest tightness and shortness of breath, abdominal distension and constipation, sweating and limb cold, dizziness, and irritability, etc. The above findings reflect the integrative actions of TCM characterizing by multiple-components, multiple-targets, multiple-pathways, and multiple-effects. This study systematically constructed the molecular networks of different TCM syndromes during the development and progression of COVID-19 and uncovered the biological basis for symptomatic treatment of TCM. Furthermore, our data revealed the pharmacological mechanisms and the scientific connotation of recommended prescriptions, which may provide supports for the prevention and treatment of COVID-19 using TCM.

[Systematic reviews of effects of Tripterygium Glycosides Tablets on pro-inflammatory factors in rheumatoid arthritis].

To systematically evaluate the effects of Tripterygium Glycosides Tablets alone or in combination with methotrexate(MTX) and leflunomide(LEF) on the levels of pro-inflammatory cytokines in patients or animal models with rheumatoid arthritis(RA), and to provide reference for clinical application and related basic research, this study systematically searched databases of CNKI, VIP, WanFang, PubMed, Embase and Cochrane Library, collected relevant clinical or animal experimental studies, used risk assessment tools to evaluate the quality of research, and used Revman 5.3 software to conduct Meta-analysis or descriptive analysis of the outcome indicators included in the literatures. Of the 1 709 papers retrieved, 3 clinical studies and 12 animal experiments were included. The results showed that compared with MTX alone, Tripterygium Glycosides Tablets combined with MTX could further reduce the expression levels of peripheral blood TNF-α(SMD=-8.88,95%CI[-10.77,-6.99],P<0.000 01),IL-1ß(P<0.000 01) and IL-6(SMD=-8.63, 95%CI[-10.57,-6.69], P<0.000 01) in RA patients. Compared with LEF alone, the combination of Tripterygium Glycosides Tablets and LEF could not further reduce the expression levels of TNF-α(P=0.20), IL-1ß(P=0.17), IL-6(P=0.31). In RA animal model, compared with model group, Tripterygium Glycosides Tablets could reduce the expression levels of peripheral blood IL-1ß(SMD=-6.29,95%CI[-9.64,-2.93],P<0.000 2)in peripheral blood(SMD=-1.39,95%CI[-1.77,-1.02],P<0.000 01), joint fluid(P<0.000 01) and paw plasma(P=0.02), and also reduce the expression levels of TNF-α in RA animal model group. Compared with MTX alone, Tripterygium Glycosides Tablets alone reduced the same levels of TNF-α(P=0.42) and IL-6(P=0.08) in joint fluid, while Tripterygium Glycosides Tablets combined with MTX could further reduce the levels of IL-6(P=0.000 1) in joint fluid; compared with LEF alone, Tripterygium Glycosides Tablets have the similar effects on reducing the expression levels of peripheral blood TNF-α(P=0.16), IL-1ß(P=0.32), IL-6(P=0.12), while Tripterygium Glycosides Tablets combined with LEF could further reduce the expression levels of TNF-α(P=0.008), IL-1ß(P=0.02), IL-6(P<0.000 1) in peripheral blood. Therefore, Tripterygium Glycosides Tablets combined with MTX could further reduce the expression levels of pro-inflammatory cytokines in peripheral blood of RA patients. Tripterygium Glycosides Tablets alone could reduce the expression levels of pro-inflammatory cytokines in peripheral blood and local joint of RA animal models. Tripterygium Glycosides Tablets combined with MTX or LEF could further reduce the express levels of pro-inflammatory cytokines in peripheral blood of RA animal models. Due to the limitation of literature, this conclusion needs to be further validated.

[Meta-analysis of RCT studies on clinical efficacy of single administration of Tripterygium Glycosides Tablets or combined administration with methotrexate against rheumatoid arthritis].

To evaluate the clinical efficacy of single administration of Tripterygium Glycosides Tablets(TGT) or combined administration with methotrexate(MTX) against rheumatoid arthritis(RA) based on American College of Rheumatology(ACR) efficacy standard. Six databases, namely CNKI, WanFang, VIP, PubMed, Embase and Cochrane Library, were retrieved for randomized controlled trials(RCT), and clinical trials were screened out according to the preset inclusion and exclusion criteria. Then, the study quality was evaluated by the risk assessment tools. Data extraction and analysis were performed by using RevMan 5.3 software for Meta-analysis. Sensitivity analysis and publication bias analysis were made to test the stability and reliability of results. Until December 2018, a total of 1 709 articles were obtained, and finally 10 clinical RCT studies with a total of 1 184 patients were included. As a result, the single administration of TGT showed a significantly better ACR efficiency(RR=1.31, 95%CI[1.15, 1.49], P<0.000 1) than methotrexate(MTX). The combined administration of TGT and MTX showed a significantly better ACR efficiency(RR=1.28, 95%CI[1.20, 1.38], P<0.000 01) than the single administration of MTX. In conclusion, the single administration of TGT and the combined administration of TGT and MTX were more effective in achieving ACR20, ACR50, ACR70 compliance than the single administration of MTX. Further validations based on more RCT studies with high-quality are required.

Clinical Practice Guideline for Tripterygium Glycosides/Tripterygium wilfordii Tablets in the Treatment of Rheumatoid Arthritis.

Tripterygium wilfordii Hook F (TwHF) is one of the most commonly used and effective traditional Chinese herbal medicines against rheumatoid arthritis (RA). Both Tripterygium Glycoside Tablets (TGT) and Tripterygium wilfordii Tablets (TWT) are the representative TwHF-based agents enrolled into the 2019 edition of Medicine Catalog for National Basic Medical Insurance, Injury Insurance, and Maternity Insurance. However, individual differences in TGT/TWT response across patients usually exist in the process of treating RA, implying that the clinical application of the two agents may not be standardized leading to the ineffective treatment and the risk of side effects. Growing evidence show that the bioactive constituents of TwHF may often have toxicity, the package insert of TGT and TWT may not be described in detail, and the therapeutic windows of the two agents are narrow. Thus, it is an urgent task to develop a standardized clinical practice guideline for TGT and TWT in the treatment of RA. In the current study, a group of clinical experts of traditional Chinese medicine and Western medicine in the research field of rheumatism diseases, pharmacists, and methodologists of evidence-based medicine were invited to select the clinical questions, to determine the levels of the evidence and the strength of the recommendations, and to develop the recommendations and good practice points. The guideline is formed based on the combination of clinical research evidence and expert experience (evidence-based, consensus, supplemented by experience). The clinical problems which are supported by clinical evidence may form recommendations, and the clinical problems without clinical evidence may form experts' suggestions. Both recommendations and experts' suggestions in this guideline summarized the clinical indications, usage, dosage, combined medication, and safety of TGT and TWT against RA systematically and comprehensively, which may offer a professional guidance in the context of the clinical application of the two TwHF-based agents.

[Identification of biomarkers to response of Tripterygium Glycosides Tablets acting on rheumatoid arthritis by integrating transcriptional data mining and biomolecular network analysis].

Growing clinical evidence shows that a partial rheumatoid arthritis( RA) patient treated with Tripterygium Glycosides Tablets( TGT) may fail to achieve clinical improvement. It is of great clinical significance to predict the therapeutic effect of TGT in RA. Therefore,the aim of the current study was to identify potential biomarkers for TGT treatment in RA. Affymetrix EG1.0 arrays were applied to detect gene expression in peripheral blood mononuclear cells obtained from 6 RA patients( 3 responders and 3 non-responders) treated with TGT. By integrating differential expression data analysis and biomolecular network analysis,360 mRNAs( 185 up-regulated and 175 down-regulated) and 24 miRNAs( 7 up-regulated and 17 down-regulated) which were differentially expressed between TGT responder and non-responder groups were identified. A total of 206 candidate target genes for the differentially expressed miRNAs were obtained based on miRanada and Target Scan databases,and then the miRNA target gene coexpression network and miRNA-mediated gene signal transduction network were constructed. Following the network analyses,three candidate miRNAs biomarkers( hsa-miR-4720-5 p,hsa-miR-374 b-5 p,hsa-miR-185-3 p) were identified as candidate biomarkers predicting individual response to TGT. Partialleast-squares( PLS) was applied to construct a model for predicting response to TGT based on the expression levels of the candidate gene biomarkers in RA patients. The five-fold cross-validation showed that the prediction accuracy( ACC) of this PLS-based model efficacy was 100.00%,100.00%,100.00%,66.67% and 66.67% respectively,and all the area under the receiver operating characteristic curve( AUC) were 1.00,indicating the highly predictive efficiency of this PLS-based model. In conclusion,the integrating transcription data mining and biomolecular network investigation show that hsa-mir-4720-5 p,hsa-mir-374 b-5 p and hsa-mir-185-3 p may be candidate biomarkers predicting individual response to TGT. In addition,the PLS model based on the expression levels of these candidate biomarkers may be helpful for the clinical screen of RA patients,which potentially benefit individualized therapy of RA in a daily clinical setting.

[Meta-analysis of laboratory index of Tripterygium Glycosides Tablets in treatment of rheumatoid arthritis].

The aim of this study was to systematically evaluate the clinical efficacy of Tripterysium Glycosides Tablets( TGT) alone or in combination with methotrexate( MTX) in the treatment of rheumatoid arthritis( RA) based on the laboratory index criteria and to provide a basis for the clinical application of TGT against RA. Six databases including CNKI,Wan Fang,VIP,PubMed,EMbase and Cochrane were retrieved for randomized controlled trials( RCT) about TGT alone or combination with MTX in the treatment of RA.Then risk assessment tools were used for quality evaluation of the studies,and data extraction and analysis were conducted by using Rev Man 5.3 software for Meta-analysis. A total of 1 709 articles were retrieved,and finally 25 studies were included,with a total sample size of 2 507 cases. Meta-analysis results showed that between TGT alone and TGT alone,MDESR=-2. 66,95%CI[-8.17,2.86],P = 0.35; MDCRP=-2.38,95%CI[-9.01,4.24],P = 0.48; between TGT combined with MTX and MTX alone,MDESR= 8.74,95%CI[6.72,10.76],P<0.000 01; MDCRP= 5.37,95%CI[3.71,7.03],P<0.000 01; SMDRF= 1.05,95%CI[0.51,1.60],P = 0.000 1.The effect of TGT on decreasing CRP and ESR in RA patients was similar to the MTX. In addition,TGT combined with MTX were more effective in decreasing CRP,ESR,RF than MTX alone. However,due to the potential bias in the included studies,more and high-quality randomized controlled trials would be needed to improve the level of evidence.

[Clinical symptoms effect of Tripterygium Glycosides Tablets alone or combined with methotrexate in treatment of rheumatoid arthritis: a Meta-analysis].

To systematically review the improvement effects of Tripterygium Glycosides Tables( TGT) alone or in combination with methotrexate( MTX) on the clinical signs and symptoms of rheumatoid arthritis( RA),and provide a basis for the rational use of TGT in clinic,in the current study,six literature databases including CNKI,Wan Fang,VIP,PubMed,EMbase,and Cochrane Library,were systematically searched,according to the inclusion and exclusion criteria. Review Manager 5.3 software was used to input the literatures,and we assessed the risk bias on the level of outcome indicators for each included literature. A total of 18 literatures were included,and the classification results showed that: compared with MTX,TGT alone can reduce the number of joint swelling( MD =0. 18,95%CI[-1.06,1.42],P = 0.78) and joint tenderness( MD =-0.06,95% CI[-1.69,1.56],P = 0.94) in RA patients with the same effect as MTX. In terms of drug combination,TGT combined with MTX had an advantage over MTX alone in lessening the morning stiffness time( MD = 18. 24,95% CI[12. 64,23. 84],P < 0. 000 01) of RA,joint tenderness( MD = 2. 65,95% CI[1. 85,3. 44],P<0.000 01) and joint swelling( MD = 3.01,95% CI[2.09,3.39],P< 0.000 01). In conclusion,this Meta-analysis suggest that TGT alone was superior to MTX in improving joint swelling and tenderness in RA patients,TGT combined with MTX may improve the clinical manifestation of RA patients better than MTX alone.

[Current research analysis and prospects on sensitization effect of artesunate on anti-cancer radiotherapy and chemotherapy].

The wide application of artemisinins in the treatment of multiple cancers reflects the advantages of traditional Chinese medicine used in this field. The existing basic and clinical studies have revealed that artesunate can effectively suppress the malignant progression of breast cancer,colon cancer,leukemia,melanoma,ovarian cancer,prostate cancer,kidney cancer and various tumors in central nervous system. The pharmacological mechanisms of artesunate against cancers are reflected in many aspects,such as inhibiting tumor cell proliferation,invasion and metastasis,inducing tumor cell apoptosis and autophagy,regulating cell signal transduction and inhibiting tumor angiogenesis. Meanwhile,growing experimental evidences have indicated that artesunate has been used for the sensitization of radiotherapy with X-ray,ß-ray,γ-ray and~(60)Co γ-ray,as well as chemotherapy with cisplatin,carboplatin and doxorubicin.This review collected basic and clinical studies on the sensitization effect of artesunate on anti-cancer radiotherapy and chemotherapy published on PubMed and CNKI during April 2000 and February 2019,and summarized the clinical positioning and application of artesunate,with the aim to provide a more comprehensive explanation on the sensitization effect of artesunate on anti-cancer radiotherapy and chemotherapy,and offer the inspiration and ideas for the development of radiotherapy and chemotherapy sensitizers,as well as cancer resistance reversal agents.

ETCM: an encyclopaedia of traditional Chinese medicine.

Traditional Chinese medicine (TCM) is not only an effective solution for primary health care, but also a great resource for drug innovation and discovery. To meet the increasing needs for TCM-related data resources, we developed ETCM, an Encyclopedia of Traditional Chinese Medicine. ETCM includes comprehensive and standardized information for the commonly used herbs and formulas of TCM, as well as their ingredients. The herb basic property and quality control standard, formula composition, ingredient drug-likeness, as well as many other information provided by ETCM can serve as a convenient resource for users to obtain thorough information about a herb or a formula. To facilitate functional and mechanistic studies of TCM, ETCM provides predicted target genes of TCM ingredients, herbs, and formulas, according to the chemical fingerprint similarity between TCM ingredients and known drugs. A systematic analysis function is also developed in ETCM, which allows users to explore the relationships or build networks among TCM herbs, formulas,ingredients, gene targets, and related pathways or diseases. ETCM is freely accessible at http://www.nrc.ac.cn:9090/ETCM/. We expect ETCM to develop into a major data warehouse for TCM and to promote TCM related researches and drug development in the future.

Molecular mechanisms of the analgesic action of Wu-tou Decoction on neuropathic pain in mice revealed using microarray and network analysis.

Wu-tou Decoction (WTD) is a classic herbal formula in traditional Chinese medicine for the treatment of joint diseases, neuropathic pain (NP) and inflammatory pain. In this study we investigated whether WTD produced analgesic action in a mouse spinal nerve ligation (SNL) model and elucidated the underlying molecular mechanisms. Mice were subjected to SNL and orally treated with WTD (3.15, 6.30 or 12.60 g·kg-1·d-1) for 21 d. SNL induced mechanical hyperalgesia and heat hyperalgesia characterized by rapid and persistent pain hypersensitivity. In addition, the expression levels of IL-1ß, TNF-α, CCL2 and CXCL1 in the spinal cord dorsal horn were dramatically increased on the 10th d post-surgery. Oral administration of WTD dose-dependently suppressed both mechanical and heat hyperalgesia as well as the expression levels of inflammatory cytokines in the spinal cord dorsal horn on the 21st d post-surgery. Then whole-genome microarray analyses were conducted to detect the gene expression profiles of spinal cord dorsal horn in SNL mice with or without WTD treatment. After construction of the WTD-SNL-network and topological analysis, a list of candidate target genes of WTD acting on SNL-induced NP was identified and found to be functionally enriched in several glial cell activation-related pathways and neuroinflammatory pathways. Our data have clarified the gene expression patterns in the mouse spinal cord under the NP condition. We also demonstrate the analgesic action of WTD through suppression of glial cell activation and neuroinflammation, which suggest the potential of WTD as a promising candidate for the treatment of NP.

Main active constituent identification in Guanxinjing capsule, a traditional Chinese medicine, for the treatment of coronary heart disease complicated with depression.

Guanxinjing capsules (GXJCs) are used in traditional Chinese medicine as a common therapy for coronary heart disease (CHD) complicated with depression. In this study, we aimed to identify the main active constituents in GXJCs and to investigate the mechanisms of GXJC action on CHD complicated with depression. The chemical constituent profile of the GXJC was identified by UHPLC-LTQ-Orbitrap assay, and oral bioavailability was evaluated to screen the GXJC drug-like chemical constituents. A total of 16 GXJC drug-like chemical constituents were identified. Then, putative targets of the GXJC drug-like chemical constituents were predicted using MedChem Studio, with 870 genes found to be the putative targets of these molecules. After that, a GXJC putative target-known CHD/depression therapeutic target network was constructed, and four topological features, including degree, betweenness, closeness and K-coreness, were calculated. According to the topological feature values of the GXJC putative targets, 14 main active constituents were identified because their corresponding putative targets had topological importance in the GXJC putative target-known CHD/depression therapeutic target network, which were defined as the candidate targets of GXJC against CHD complicated with depression. Functionally, these candidate targets were significantly involved in several CHD/depression-related pathways, including repairing pathological vascular changes, reducing platelet aggregation and inflammation, and affecting patient depression. This study identified a list of main active constituents of GXJC acting on CHD complicated with depression using an integrative pharmacology-based approach that combined active chemical constituent identification, drug target prediction and network analysis. This method may offer an efficient way to understand the pharmacological mechanisms of traditional Chinese medicine prescriptions.

[Exploiture and application of an internet-based Computation Platform for Integrative Pharmacology of Traditional Chinese Medicine].

Recently, integrative pharmacology(IP) has become a pivotal paradigm for the modernization of traditional Chinese medicines(TCM) and combinatorial drugs discovery, which is an interdisciplinary science for establishing the in vitro and in vivo correlation between absorption, distribution, metabolism, and excretion/pharmacokinetic(ADME/PK) profiles of TCM and the molecular networks of disease by the integration of the knowledge of multi-disciplinary and multi-stages. In the present study, an internet-based Computation Platform for IP of TCM(TCM-IP, www.tcmip.cn) is established to promote the development of the emerging discipline. Among them, a big data of TCM is an important resource for TCM-IP including Chinese Medicine Formula Database, Chinese Medical Herbs Database, Chemical Database of Chinese Medicine, Target Database for Disease and Symptoms, et al. Meanwhile, some data mining and bioinformatics approaches are critical technology for TCM-IP including the identification of the TCM constituents, ADME prediction, target prediction for the TCM constituents, network construction and analysis, et al. Furthermore, network beautification and individuation design are employed to meet the consumer's requirement. We firmly believe that TCM-IP is a very useful tool for the identification of active constituents of TCM and their involving potential molecular mechanism for therapeutics, which would wildly applied in quality evaluation, clinical repositioning, scientific discovery based on original thinking, prescription compatibility and new drug of TCM, et al.

Deciphering the pharmacological mechanism of the Chinese formula huanglian-jie-du decoction in the treatment of ischemic stroke using a systems biology-based strategy.

AIM: Huanglian-jie-du decoction (HLJDD) is an important multiherb remedy in TCM, which is recently demonstrated to be effective to treat ischemic stroke. Here, we aimed to investigate the pharmacological mechanisms of HLJDD in the treatment of ischemic stroke using systems biology approaches. METHODS: Putative targets of HLJDD were predicted using MetaDrug. An interaction network of putative HLJDD targets and known therapeutic targets for the treatment of ischemic stroke was then constructed, and candidate HLJDD targets were identified by calculating topological features, including 'Degree', 'Node-betweenness', 'Closeness', and 'K-coreness'. The binding efficiencies of the candidate HLJDD targets with the corresponding compositive compounds were further validated by a molecular docking simulation. RESULTS: A total of 809 putative targets were obtained for 168 compositive compounds in HLJDD. Additionally, 39 putative targets were common to all four herbs of HLJDD. Next, 49 major nodes were identified as candidate HLJDD targets due to their network topological importance. The enrichment analysis based on the Gene Ontology (GO) annotation system and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway demonstrated that candidate HLJDD targets were more frequently involved in G-protein-coupled receptor signaling pathways, neuroactive ligand-receptor interactions and gap junctions, which all played important roles in the progression of ischemic stroke. Finally, the molecular docking simulation showed that 170 pairs of chemical components and candidate HLJDD targets had strong binding efficiencies. CONCLUSION: This study has developed for the first time a comprehensive systems approach integrating drug target prediction, network analysis and molecular docking simulation to reveal the relationships between the herbs contained in HLJDD and their putative targets and ischemic stroke-related pathways.